dna colonic hiv rna  (Qiagen)


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    Name:
    QIAamp Viral RNA Mini Kit
    Description:
    For isolation of viral RNA from cell free body fluids Kit contents Qiagen QIAamp Viral RNA Mini Kit 50 preps 140L Sample 50L Elution Volume Liquid Media Sample Viral RNA Purification Silica Technology Spin Column Format Manual Processing 90 Yield 20 to 40 min Time Run Ideal for PCR qPCR Real time PCR For Isolation of Viral RNA from Cell free Body Fluids Includes 50 QIAamp Mini Spin Columns Carrier RNA 2mL Collection Tubes RNase free Buffers Benefits Rapid isolation of high quality ready to use RNA No organic extraction or alcohol precipitation Consistent high yields Complete removal of contaminants and inhibitors
    Catalog Number:
    52904
    Price:
    257
    Category:
    QIAamp Viral RNA Mini Kit
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    Structured Review

    Qiagen dna colonic hiv rna
    QIAamp Viral RNA Mini Kit
    For isolation of viral RNA from cell free body fluids Kit contents Qiagen QIAamp Viral RNA Mini Kit 50 preps 140L Sample 50L Elution Volume Liquid Media Sample Viral RNA Purification Silica Technology Spin Column Format Manual Processing 90 Yield 20 to 40 min Time Run Ideal for PCR qPCR Real time PCR For Isolation of Viral RNA from Cell free Body Fluids Includes 50 QIAamp Mini Spin Columns Carrier RNA 2mL Collection Tubes RNase free Buffers Benefits Rapid isolation of high quality ready to use RNA No organic extraction or alcohol precipitation Consistent high yields Complete removal of contaminants and inhibitors
    https://www.bioz.com/result/dna colonic hiv rna/product/Qiagen
    Average 90 stars, based on 30806 article reviews
    Price from $9.99 to $1999.99
    dna colonic hiv rna - by Bioz Stars, 2020-04
    90/100 stars

    Images

    1) Product Images from "Initiation of antiretroviral therapy before detection of colonic infiltration by HIV reduces viral reservoirs, inflammation and immune activation"

    Article Title: Initiation of antiretroviral therapy before detection of colonic infiltration by HIV reduces viral reservoirs, inflammation and immune activation

    Journal: Journal of the International AIDS Society

    doi: 10.7448/IAS.19.1.21163

    HIV RNA in the peripheral blood and cerebrospinal fluid during acute HIV infection. HIV RNA measurements during acute HIV infection are compared between participants with detectable colonic HIV RNA and undetectable colonic HIV RNA. Statistically significant pairwise comparisons ( p
    Figure Legend Snippet: HIV RNA in the peripheral blood and cerebrospinal fluid during acute HIV infection. HIV RNA measurements during acute HIV infection are compared between participants with detectable colonic HIV RNA and undetectable colonic HIV RNA. Statistically significant pairwise comparisons ( p

    Techniques Used: Infection

    Total HIV DNA in the peripheral blood and colon before and after ART. Total HIV DNA measurements in the (a) peripheral blood and (b) colon are compared before and after 24 weeks of ART. Statistically significant pairwise comparisons ( p
    Figure Legend Snippet: Total HIV DNA in the peripheral blood and colon before and after ART. Total HIV DNA measurements in the (a) peripheral blood and (b) colon are compared before and after 24 weeks of ART. Statistically significant pairwise comparisons ( p

    Techniques Used:

    2) Product Images from "Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings"

    Article Title: Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings

    Journal: The Journal of infectious diseases

    doi: 10.1086/650388

    Integrated assays using plasma spiked with Armored RNA HIV. Armored RNA HIV was spiked into human plasma at concentrations ranging from 10,000 copies/mL to 1,000 copies/mL amd extracted with a QIAamp® Viral RNA Mini Kit according to the manufacturer’s
    Figure Legend Snippet: Integrated assays using plasma spiked with Armored RNA HIV. Armored RNA HIV was spiked into human plasma at concentrations ranging from 10,000 copies/mL to 1,000 copies/mL amd extracted with a QIAamp® Viral RNA Mini Kit according to the manufacturer’s

    Techniques Used:

    3) Product Images from "Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿"

    Article Title: Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿

    Journal: Journal of Virology

    doi: 10.1128/JVI.02672-08

    Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.
    Figure Legend Snippet: Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.

    Techniques Used: Activity Assay, Plaque Assay, Infection, Transfection, Construct, Plasmid Preparation, Over Expression, Polymerase Chain Reaction, Cell Culture, Expressing, FACS, Fluorescence, Modification

    4) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    5) Product Images from "High level of HIV-1 drug resistance mutations in patients with unsuppressed viral loads in rural northern South Africa"

    Article Title: High level of HIV-1 drug resistance mutations in patients with unsuppressed viral loads in rural northern South Africa

    Journal: AIDS Research and Therapy

    doi: 10.1186/s12981-017-0161-z

    Comparing the DRM in RNA (plasma) and DNA (peripheral blood mononuclear cells). a NRTI. b NNRTI
    Figure Legend Snippet: Comparing the DRM in RNA (plasma) and DNA (peripheral blood mononuclear cells). a NRTI. b NNRTI

    Techniques Used:

    6) Product Images from "Structural basis for human respiratory syncytial virus NS1-mediated modulation of host responses"

    Article Title: Structural basis for human respiratory syncytial virus NS1-mediated modulation of host responses

    Journal: Nature microbiology

    doi: 10.1038/nmicrobiol.2017.101

    NS1 unique regions are important for modulating host responses Human monocyte-derived dendritic cells (MDDCs) were transduced with either only empty lentiviral vectors (empty) or lentivirus vectors that express either EBOV-VP35, hRSV-NS1, hRSV-NS1-(1-118), hRSV-NS1-L132A/L133A. RNA was isolated from transduced MDDCs that were either mock-infected or infected with SeV. RNA was extracted at the indicated hours post-infection; and a, IFN-β, b, TNF-α, c, ISG54, and d, ISG56 mRNA levels were quantified RT-qPCR, normalizing their levels to that of β-actin mRNA. The graph indicates the fold-change relative to the mock-infected samples. For a–d, symbols are: ○, empty vector; ▲, hRSV NS1 1-118; ▼, hRSV NS1 L132A/L133A; ■, hRSV NS1 WT; and ●, EBOV VP35. hRSV NS1 inhibits upregulation of DC maturation markers. Transduced MDDCs were infected with SeV for 20h, harvested and stained for expression of CD40, CD80, CD83 and CD86. Fold change in mean fluorescence intensity (MFI) for the indicated proteins in transduced MDDCs where fold increase indicates comparisons of SeV-infected to uninfected MDDCs. Error bars indicate standard deviations from three independent experiments (*p
    Figure Legend Snippet: NS1 unique regions are important for modulating host responses Human monocyte-derived dendritic cells (MDDCs) were transduced with either only empty lentiviral vectors (empty) or lentivirus vectors that express either EBOV-VP35, hRSV-NS1, hRSV-NS1-(1-118), hRSV-NS1-L132A/L133A. RNA was isolated from transduced MDDCs that were either mock-infected or infected with SeV. RNA was extracted at the indicated hours post-infection; and a, IFN-β, b, TNF-α, c, ISG54, and d, ISG56 mRNA levels were quantified RT-qPCR, normalizing their levels to that of β-actin mRNA. The graph indicates the fold-change relative to the mock-infected samples. For a–d, symbols are: ○, empty vector; ▲, hRSV NS1 1-118; ▼, hRSV NS1 L132A/L133A; ■, hRSV NS1 WT; and ●, EBOV VP35. hRSV NS1 inhibits upregulation of DC maturation markers. Transduced MDDCs were infected with SeV for 20h, harvested and stained for expression of CD40, CD80, CD83 and CD86. Fold change in mean fluorescence intensity (MFI) for the indicated proteins in transduced MDDCs where fold increase indicates comparisons of SeV-infected to uninfected MDDCs. Error bars indicate standard deviations from three independent experiments (*p

    Techniques Used: Derivative Assay, Transduction, Isolation, Infection, Quantitative RT-PCR, Plasmid Preparation, Staining, Expressing, Fluorescence

    7) Product Images from "Antiviral Hammerhead Ribozymes Are Effective for Developing Transgenic Suppression of Chikungunya Virus in Aedes aegypti Mosquitoes"

    Article Title: Antiviral Hammerhead Ribozymes Are Effective for Developing Transgenic Suppression of Chikungunya Virus in Aedes aegypti Mosquitoes

    Journal: Viruses

    doi: 10.3390/v8060163

    Generalized structure of a minimal hammerhead ribozyme (hRz) and organization of the chikungunya virus (CHIKV) viral genome showing hRz target sites. ( A ) hRzs consist of helix I and III which mediate complementary base-pairing with the target, and helix II which acts as the catalytic core mediating cleavage of the target RNA; ( B ) Schematic of the CHIKV genome. nsP1–nsP4 = polyprotein encoded by the genomic RNA. Capsid, E3, E2, 6K and E1 = polyprotein encoded by the sub-genomic RNA. 9–15 = target sites for hRz#9-#15.
    Figure Legend Snippet: Generalized structure of a minimal hammerhead ribozyme (hRz) and organization of the chikungunya virus (CHIKV) viral genome showing hRz target sites. ( A ) hRzs consist of helix I and III which mediate complementary base-pairing with the target, and helix II which acts as the catalytic core mediating cleavage of the target RNA; ( B ) Schematic of the CHIKV genome. nsP1–nsP4 = polyprotein encoded by the genomic RNA. Capsid, E3, E2, 6K and E1 = polyprotein encoded by the sub-genomic RNA. 9–15 = target sites for hRz#9-#15.

    Techniques Used:

    8) Product Images from "Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿"

    Article Title: Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿

    Journal: Journal of Virology

    doi: 10.1128/JVI.02672-08

    Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.
    Figure Legend Snippet: Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.

    Techniques Used: Activity Assay, Plaque Assay, Infection, Transfection, Construct, Plasmid Preparation, Over Expression, Polymerase Chain Reaction, Cell Culture, Expressing, FACS, Fluorescence, Modification

    9) Product Images from "Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing"

    Article Title: Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing

    Journal: Genome Biology and Evolution

    doi: 10.1093/gbe/evv034

    Confirmation of contaminated columns as origin of CHIV14 DNA by qPCR. ( A ) Verification of CHIV14 genome assembled from the Illumina deep-sequencing data by overlapping PCR and inverted PCR. Five sets of overlapping primer pairs and one set of inverted primer pair were designed and used to amplify overlapping DNA fragments. (Left) Schematic diagram of the positions of primer pairs for the overlapping PCR and inverted PCR. (Right) Amplification overlapping viral DNA fragments. The numbers above indicate the primer pair used for the PCR as illustrated on the left. The numbers on the left indicate the molecular weight in base pairs. ( B ) Scatterplot showing copy number of CHIV14 per microliter of DNA extraction. DNA from patients ( n = 13), healthy controls ( n = 13), and water ( n = 31) was extracted using QIAamp mini spin columns (QIAamp Viral RNA Mini kit; Qiagen). In parallel, seven DNA extractions for each specimen type (patients, healthy individuals, and water) were performed using the UCP columns. Each dot represents one specimen. Bars show the average copy numbers of the viral genome.
    Figure Legend Snippet: Confirmation of contaminated columns as origin of CHIV14 DNA by qPCR. ( A ) Verification of CHIV14 genome assembled from the Illumina deep-sequencing data by overlapping PCR and inverted PCR. Five sets of overlapping primer pairs and one set of inverted primer pair were designed and used to amplify overlapping DNA fragments. (Left) Schematic diagram of the positions of primer pairs for the overlapping PCR and inverted PCR. (Right) Amplification overlapping viral DNA fragments. The numbers above indicate the primer pair used for the PCR as illustrated on the left. The numbers on the left indicate the molecular weight in base pairs. ( B ) Scatterplot showing copy number of CHIV14 per microliter of DNA extraction. DNA from patients ( n = 13), healthy controls ( n = 13), and water ( n = 31) was extracted using QIAamp mini spin columns (QIAamp Viral RNA Mini kit; Qiagen). In parallel, seven DNA extractions for each specimen type (patients, healthy individuals, and water) were performed using the UCP columns. Each dot represents one specimen. Bars show the average copy numbers of the viral genome.

    Techniques Used: Real-time Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction, Amplification, Molecular Weight, DNA Extraction

    10) Product Images from "Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing"

    Article Title: Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing

    Journal: Genome Biology and Evolution

    doi: 10.1093/gbe/evv034

    Confirmation of contaminated columns as origin of CHIV14 DNA by qPCR. ( A ) Verification of CHIV14 genome assembled from the Illumina deep-sequencing data by overlapping PCR and inverted PCR. Five sets of overlapping primer pairs and one set of inverted primer pair were designed and used to amplify overlapping DNA fragments. (Left) Schematic diagram of the positions of primer pairs for the overlapping PCR and inverted PCR. (Right) Amplification overlapping viral DNA fragments. The numbers above indicate the primer pair used for the PCR as illustrated on the left. The numbers on the left indicate the molecular weight in base pairs. ( B ) Scatterplot showing copy number of CHIV14 per microliter of DNA extraction. DNA from patients ( n = 13), healthy controls ( n = 13), and water ( n = 31) was extracted using QIAamp mini spin columns (QIAamp Viral RNA Mini kit; Qiagen). In parallel, seven DNA extractions for each specimen type (patients, healthy individuals, and water) were performed using the UCP columns. Each dot represents one specimen. Bars show the average copy numbers of the viral genome.
    Figure Legend Snippet: Confirmation of contaminated columns as origin of CHIV14 DNA by qPCR. ( A ) Verification of CHIV14 genome assembled from the Illumina deep-sequencing data by overlapping PCR and inverted PCR. Five sets of overlapping primer pairs and one set of inverted primer pair were designed and used to amplify overlapping DNA fragments. (Left) Schematic diagram of the positions of primer pairs for the overlapping PCR and inverted PCR. (Right) Amplification overlapping viral DNA fragments. The numbers above indicate the primer pair used for the PCR as illustrated on the left. The numbers on the left indicate the molecular weight in base pairs. ( B ) Scatterplot showing copy number of CHIV14 per microliter of DNA extraction. DNA from patients ( n = 13), healthy controls ( n = 13), and water ( n = 31) was extracted using QIAamp mini spin columns (QIAamp Viral RNA Mini kit; Qiagen). In parallel, seven DNA extractions for each specimen type (patients, healthy individuals, and water) were performed using the UCP columns. Each dot represents one specimen. Bars show the average copy numbers of the viral genome.

    Techniques Used: Real-time Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction, Amplification, Molecular Weight, DNA Extraction

    11) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    12) Product Images from "Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens"

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens

    Journal: Analytical chemistry

    doi: 10.1021/acs.analchem.5b01594

    Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).
    Figure Legend Snippet: Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).

    Techniques Used: RT Lamp Assay, Purification, Nucleic Acid Electrophoresis, Flow Cytometry, Amplification, In Situ, Positive Control, Negative Control

    13) Product Images from "Detection of Zika virus in mouse mammary gland and breast milk"

    Article Title: Detection of Zika virus in mouse mammary gland and breast milk

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0007080

    Transmission of ZIKV RNA in mouse breast milk. Postpartum AG129 dams were retro-orbitally inoculated with 1 x 10 2 FFU of ZIKV FSS13025 or 10% FBS-PBS as Mock within 24 hours of parturition. Pups were sacrificed on d1, d3, d5 and d7 after birth. (A and B) Levels of ZIKV RNA in the head and the rest of the body were measured by qRT-PCR (n = 6 mice, each day). (C and D) ZIKV RNA levels in stomach milk clots (SMC) and stomach tissues were quantified via qRT-PCR (n = 3 mice, each day). (E and F) Presence of infectious ZIKV in SMC and stomach was assessed using FFA (n = 3 mice, each day). Data represent two independent experiments.
    Figure Legend Snippet: Transmission of ZIKV RNA in mouse breast milk. Postpartum AG129 dams were retro-orbitally inoculated with 1 x 10 2 FFU of ZIKV FSS13025 or 10% FBS-PBS as Mock within 24 hours of parturition. Pups were sacrificed on d1, d3, d5 and d7 after birth. (A and B) Levels of ZIKV RNA in the head and the rest of the body were measured by qRT-PCR (n = 6 mice, each day). (C and D) ZIKV RNA levels in stomach milk clots (SMC) and stomach tissues were quantified via qRT-PCR (n = 3 mice, each day). (E and F) Presence of infectious ZIKV in SMC and stomach was assessed using FFA (n = 3 mice, each day). Data represent two independent experiments.

    Techniques Used: Transmission Assay, Quantitative RT-PCR, Mouse Assay

    14) Product Images from "Efficacy of Human Monoclonal Antibody Monotherapy Against Bundibugyo Virus Infection in Nonhuman Primates"

    Article Title: Efficacy of Human Monoclonal Antibody Monotherapy Against Bundibugyo Virus Infection in Nonhuman Primates

    Journal: The Journal of Infectious Diseases

    doi: 10.1093/infdis/jiy295

    Therapeutic potency of monoclonal antibody (mAb) BDBV289-N to prevent death and reduce viremia. Nonhuman primates (NHPs) received 825 plaque-forming units (PFU) of Bundibugyo virus (BDBV) intramuscularly and a dose of mAb (35 mg/kg) intravenously 8 and 11 days after inoculation (data are for 6 treated animals and 1 untreated control this study and for 9 untreated historical controls). A , Kaplan-Meier survival plot. Arrows indicate the day of mAb dosing. Untreated groups represent 1 animal from this study, and 9 historical controls represent animals (cumulative 60% survival) inoculated by the same route with the same stock of virus. The proportion surviving at day 28 after infection was compared using a 2-sided exact unconditional test of homogeneity. A P value of ≤ .05 was considered significant. B , A comparison of blood viral RNA load was determined at 11 days after virus inoculation for treated or control animals, as described in panel A . The group of 6 untreated animals included 1 NHP from this study and 5 historical controls that also were assessed for viremia on day 11 after infection. Virus titers were compared using the Mann–Whitney U test. The median titer for each group is shown. C , Kinetics of the blood viral RNA load, as determined by real-time quantitative polymerase chain reaction, for 6 treated or 6 of 10 control animals, comprising 1 NHP assessed at the same time point and 9 historical controls. The dotted line indicates the limit of detection (LOD), which was 3.7 log 10 genome equivalents (GEq) per milliliter. Each measurement represented the mean value of technical duplicates. Numbers indicate individual NHPs from the respective group. Abbreviations: C, untreated, this study (gray); HC, untreated, historical control (gray curves); M, treated, this study (orange curves).
    Figure Legend Snippet: Therapeutic potency of monoclonal antibody (mAb) BDBV289-N to prevent death and reduce viremia. Nonhuman primates (NHPs) received 825 plaque-forming units (PFU) of Bundibugyo virus (BDBV) intramuscularly and a dose of mAb (35 mg/kg) intravenously 8 and 11 days after inoculation (data are for 6 treated animals and 1 untreated control this study and for 9 untreated historical controls). A , Kaplan-Meier survival plot. Arrows indicate the day of mAb dosing. Untreated groups represent 1 animal from this study, and 9 historical controls represent animals (cumulative 60% survival) inoculated by the same route with the same stock of virus. The proportion surviving at day 28 after infection was compared using a 2-sided exact unconditional test of homogeneity. A P value of ≤ .05 was considered significant. B , A comparison of blood viral RNA load was determined at 11 days after virus inoculation for treated or control animals, as described in panel A . The group of 6 untreated animals included 1 NHP from this study and 5 historical controls that also were assessed for viremia on day 11 after infection. Virus titers were compared using the Mann–Whitney U test. The median titer for each group is shown. C , Kinetics of the blood viral RNA load, as determined by real-time quantitative polymerase chain reaction, for 6 treated or 6 of 10 control animals, comprising 1 NHP assessed at the same time point and 9 historical controls. The dotted line indicates the limit of detection (LOD), which was 3.7 log 10 genome equivalents (GEq) per milliliter. Each measurement represented the mean value of technical duplicates. Numbers indicate individual NHPs from the respective group. Abbreviations: C, untreated, this study (gray); HC, untreated, historical control (gray curves); M, treated, this study (orange curves).

    Techniques Used: Infection, MANN-WHITNEY, Real-time Polymerase Chain Reaction

    15) Product Images from "Non-Catalytic Site HIV-1 Integrase Inhibitors Disrupt Core Maturation and Induce a Reverse Transcription Block in Target Cells"

    Article Title: Non-Catalytic Site HIV-1 Integrase Inhibitors Disrupt Core Maturation and Induce a Reverse Transcription Block in Target Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0074163

    NCINIs do not inhibit Gag-Pol processing, ERT, or target cell entry. ( A ) Western blot analysis of purified HIV-1 particles produced in the presence of different inhibitors at 1 µM concentration. Gag/Gag-Pol processing was assessed using anti-CA ( upper ), anti-IN ( middle ), and anti-RT antibodies ( lower ). ( B ) Equal p24 amounts of virus shown in A were analyzed for virion-associated endogenous reverse transcriptase activity. Values were averaged from duplicate measurements. ( C ) Percentage of cells positive for beta-lactamase activity following infection with Blam-Vpr complemented HIV-1 produced in the presence of DMSO or 1 µM tested compounds. Values are normalized to DMSO-treated control and represent a mean of two independent experiments performed in duplicate.
    Figure Legend Snippet: NCINIs do not inhibit Gag-Pol processing, ERT, or target cell entry. ( A ) Western blot analysis of purified HIV-1 particles produced in the presence of different inhibitors at 1 µM concentration. Gag/Gag-Pol processing was assessed using anti-CA ( upper ), anti-IN ( middle ), and anti-RT antibodies ( lower ). ( B ) Equal p24 amounts of virus shown in A were analyzed for virion-associated endogenous reverse transcriptase activity. Values were averaged from duplicate measurements. ( C ) Percentage of cells positive for beta-lactamase activity following infection with Blam-Vpr complemented HIV-1 produced in the presence of DMSO or 1 µM tested compounds. Values are normalized to DMSO-treated control and represent a mean of two independent experiments performed in duplicate.

    Techniques Used: Western Blot, Purification, Produced, Concentration Assay, Activity Assay, Infection

    16) Product Images from "APOBEC3G and APOBEC3F Act in Concert To Extinguish HIV-1 Replication"

    Article Title: APOBEC3G and APOBEC3F Act in Concert To Extinguish HIV-1 Replication

    Journal: Journal of Virology

    doi: 10.1128/JVI.03275-15

    Absolute restriction of HIV in vivo requires both APOBEC3G and APOBEC3F. (A) (Top) The plasma of BLT humanized mice infected with JRCSFvifH42/43D (blue lines) was monitored longitudinally for the presence of HIV RNA. Wild-type JRCSF RNA is presented in aggregate ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic-DNA extracts from each tissue listed from the infected humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (B) (Top) Viral RNA was present in the plasma of 4/4 mice infected with JRCSFvifW79S (red lines) at levels comparable to those of wild-type JRCSF ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic DNA extracted from each tissue listed from the infected BLT humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (C) (Top) Plasma viral RNA was not observed in 4/4 mice infected with JRCSFvifH42/43DW79S (green lines) for up to 8 weeks postexposure, in contrast to infection with wild-type JRCSF (black diamonds). (Bottom) Viral DNA was not detected in any tissue of JRCSFvifH42/43DW79S-inoculated BLT humanized mice (−). (D) Sequencing of viral DNA amplified from the tissues of mice infected with JRCSFvifH42/43D (G3 and G4) revealed heavy G-to-A mutation at GG sites (blue bar). Few G-to-A mutations were present at GA sites in viral DNA from mice infected with JRCSFvifH42/43D or JRCSFvifW79S, and no mutations were observed in WT1 JRCSF DNA. The data are presented as means ± SEM.
    Figure Legend Snippet: Absolute restriction of HIV in vivo requires both APOBEC3G and APOBEC3F. (A) (Top) The plasma of BLT humanized mice infected with JRCSFvifH42/43D (blue lines) was monitored longitudinally for the presence of HIV RNA. Wild-type JRCSF RNA is presented in aggregate ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic-DNA extracts from each tissue listed from the infected humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (B) (Top) Viral RNA was present in the plasma of 4/4 mice infected with JRCSFvifW79S (red lines) at levels comparable to those of wild-type JRCSF ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic DNA extracted from each tissue listed from the infected BLT humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (C) (Top) Plasma viral RNA was not observed in 4/4 mice infected with JRCSFvifH42/43DW79S (green lines) for up to 8 weeks postexposure, in contrast to infection with wild-type JRCSF (black diamonds). (Bottom) Viral DNA was not detected in any tissue of JRCSFvifH42/43DW79S-inoculated BLT humanized mice (−). (D) Sequencing of viral DNA amplified from the tissues of mice infected with JRCSFvifH42/43D (G3 and G4) revealed heavy G-to-A mutation at GG sites (blue bar). Few G-to-A mutations were present at GA sites in viral DNA from mice infected with JRCSFvifH42/43D or JRCSFvifW79S, and no mutations were observed in WT1 JRCSF DNA. The data are presented as means ± SEM.

    Techniques Used: In Vivo, Mouse Assay, Infection, Nested PCR, Sequencing, Amplification, Mutagenesis

    17) Product Images from "NEDD4 family ubiquitin ligases associate with LCMV Z’s PPXY domain and are required for virus budding, but not via direct ubiquitination of Z"

    Article Title: NEDD4 family ubiquitin ligases associate with LCMV Z’s PPXY domain and are required for virus budding, but not via direct ubiquitination of Z

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.1008100

    Loss of ubiquitination sites in LCMV Z has little impact on virus particle release. (A-B) Reverse genetics was used to generate recombinant (r)LCMV containing mutations at the two ubiquitination sites (K10 and K77) as well as a virus with a lysine-free (NoK) or a late domain-free (PPPY > AAAA) Z protein. The growth kinetics of these viruses were then measured by infecting A549 cells with these viruses and measuring the infectious titers by plaque assay (A) and DI particle titers by plaque interfering assay (B) at 1, 16, 24, 40, 48 and 72 hours after infection. (C) Summary of a two-way ANOVA with Holm-Sidak’s test for multiple comparisons used to compare the log-transformed mean titer values from the growth curve in (A-B). (D-E) Viral RNA was isolated from clarified cell culture media from the 72 hours post-infection time point in (A-B) and the quantities of genomic S-segment (D) or L-segment (E) vRNA were determined by quantitative real time RT-PCR. (F) The release of virus-like particles (VLPs) from HEK293T cells transfected with plasmids expressing SBP-tagged LCMV Z protein with the indicated mutations was measured. The graphs in (A-B and D-F) represent the mean values ± SEM of three independent experiments with two technical replicates each. A one-way ANOVA with Holm-Sidak’s test for multiple comparisons was used to compare the mean values to the WT control in (D-F). *p
    Figure Legend Snippet: Loss of ubiquitination sites in LCMV Z has little impact on virus particle release. (A-B) Reverse genetics was used to generate recombinant (r)LCMV containing mutations at the two ubiquitination sites (K10 and K77) as well as a virus with a lysine-free (NoK) or a late domain-free (PPPY > AAAA) Z protein. The growth kinetics of these viruses were then measured by infecting A549 cells with these viruses and measuring the infectious titers by plaque assay (A) and DI particle titers by plaque interfering assay (B) at 1, 16, 24, 40, 48 and 72 hours after infection. (C) Summary of a two-way ANOVA with Holm-Sidak’s test for multiple comparisons used to compare the log-transformed mean titer values from the growth curve in (A-B). (D-E) Viral RNA was isolated from clarified cell culture media from the 72 hours post-infection time point in (A-B) and the quantities of genomic S-segment (D) or L-segment (E) vRNA were determined by quantitative real time RT-PCR. (F) The release of virus-like particles (VLPs) from HEK293T cells transfected with plasmids expressing SBP-tagged LCMV Z protein with the indicated mutations was measured. The graphs in (A-B and D-F) represent the mean values ± SEM of three independent experiments with two technical replicates each. A one-way ANOVA with Holm-Sidak’s test for multiple comparisons was used to compare the mean values to the WT control in (D-F). *p

    Techniques Used: Recombinant, Plaque Assay, Infection, Transformation Assay, Isolation, Cell Culture, Quantitative RT-PCR, Transfection, Expressing

    18) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    19) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    20) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    21) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    22) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    23) Product Images from "Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology"

    Article Title: Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S191784

    Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.
    Figure Legend Snippet: Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Real-time Polymerase Chain Reaction

    Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.
    Figure Legend Snippet: Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Amplification, Polymerase Chain Reaction, Agarose Gel Electrophoresis, DNA Sequencing, Marker, Reverse Transcription Polymerase Chain Reaction, Negative Control

    24) Product Images from "Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿"

    Article Title: Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿

    Journal: Journal of Virology

    doi: 10.1128/JVI.02672-08

    Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.
    Figure Legend Snippet: Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.

    Techniques Used: Activity Assay, Plaque Assay, Infection, Transfection, Construct, Plasmid Preparation, Over Expression, Polymerase Chain Reaction, Cell Culture, Expressing, FACS, Fluorescence, Modification

    25) Product Images from "Evidence of Ebola Virus Replication and High Concentration in Semen of a Patient During Recovery"

    Article Title: Evidence of Ebola Virus Replication and High Concentration in Semen of a Patient During Recovery

    Journal: Clinical Infectious Diseases: An Official Publication of the Infectious Diseases Society of America

    doi: 10.1093/cid/cix518

    High viral load and identical genomes within a single patient with Ebola virus disease. A , Viral load in semen and serum. We measured viral load from extracted RNA using primers in Supplementary Table 2 (Trombley et al, 2010). Samples were quantified against a standard curve (assay limit of detection [LOD]: 1 copy/µL) to determine copies per microliter of extracted RNA. Based on the different extraction methods, we converted these values into copies per milliliter of indicated fluid (blue: serum, n = 6 replicates per sample; orange: semen, n = 3). Asterisk indicates below-assay LOD. B , Quantification of Ebola virus (EBOV) antisense (vRNA) and sense (c/mRNA) RNA in semen samples using strand-specific quantitative reverse-transcription polymerase chain reaction (RT-qPCR). We designed RT primers for a 2-step RT-qPCR to detect antisense or sense viral RNA separately. Day 180 semen sample was not tested. C , Quantification of EBOV vRNA and c/mRNA in semen samples using single-molecule tagged amplicon sequencing. We uniquely tagged and amplified RNA molecules with random barcodes in a strand-specific manner. We deduplicated reads and counted unique numbers of barcodes. Molecules at day 180 likely represent background amplification. See Supplementary Figure 1 C for quantification in peripheral blood leukocyte (PBL) samples. D , Deep sequencing metrics. We performed RNA-seq either without (unbiased) or with hybrid selection using EBOV-specific baits to reduce host RNA background. Lower viral genome coverage is observed in PBL and semen samples due to abundant cellular RNA. E , Phylogeny of the SL4 clade. We combined 1489 publicly available genomes (Diehl et al 2016) with this patient’s viral genome and generated a phylogenetic tree with 1000 bootstrap replicates. The patient sample falls within a SL4 subclade (maroon; 100% bootstrap support) of samples collected near Freetown, Sierra Leone, from late January 2015 through March 2015, representing a likely transmission network. See Supplementary Figure 2 for full tree.
    Figure Legend Snippet: High viral load and identical genomes within a single patient with Ebola virus disease. A , Viral load in semen and serum. We measured viral load from extracted RNA using primers in Supplementary Table 2 (Trombley et al, 2010). Samples were quantified against a standard curve (assay limit of detection [LOD]: 1 copy/µL) to determine copies per microliter of extracted RNA. Based on the different extraction methods, we converted these values into copies per milliliter of indicated fluid (blue: serum, n = 6 replicates per sample; orange: semen, n = 3). Asterisk indicates below-assay LOD. B , Quantification of Ebola virus (EBOV) antisense (vRNA) and sense (c/mRNA) RNA in semen samples using strand-specific quantitative reverse-transcription polymerase chain reaction (RT-qPCR). We designed RT primers for a 2-step RT-qPCR to detect antisense or sense viral RNA separately. Day 180 semen sample was not tested. C , Quantification of EBOV vRNA and c/mRNA in semen samples using single-molecule tagged amplicon sequencing. We uniquely tagged and amplified RNA molecules with random barcodes in a strand-specific manner. We deduplicated reads and counted unique numbers of barcodes. Molecules at day 180 likely represent background amplification. See Supplementary Figure 1 C for quantification in peripheral blood leukocyte (PBL) samples. D , Deep sequencing metrics. We performed RNA-seq either without (unbiased) or with hybrid selection using EBOV-specific baits to reduce host RNA background. Lower viral genome coverage is observed in PBL and semen samples due to abundant cellular RNA. E , Phylogeny of the SL4 clade. We combined 1489 publicly available genomes (Diehl et al 2016) with this patient’s viral genome and generated a phylogenetic tree with 1000 bootstrap replicates. The patient sample falls within a SL4 subclade (maroon; 100% bootstrap support) of samples collected near Freetown, Sierra Leone, from late January 2015 through March 2015, representing a likely transmission network. See Supplementary Figure 2 for full tree.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Amplification, Sequencing, RNA Sequencing Assay, Selection, Generated, Transmission Assay

    26) Product Images from "Discordant congenital Zika syndrome twins show differential in vitro viral susceptibility of neural progenitor cells"

    Article Title: Discordant congenital Zika syndrome twins show differential in vitro viral susceptibility of neural progenitor cells

    Journal: Nature Communications

    doi: 10.1038/s41467-017-02790-9

    NPC gene expression analyses by RNA-Seq in cultured cells prior to ZIKV infection (see also Supplementary Fig. 3 ). a Heatmap representation and clusterization of top 20 most significant DEGs ( p
    Figure Legend Snippet: NPC gene expression analyses by RNA-Seq in cultured cells prior to ZIKV infection (see also Supplementary Fig. 3 ). a Heatmap representation and clusterization of top 20 most significant DEGs ( p

    Techniques Used: Expressing, RNA Sequencing Assay, Cell Culture, Infection

    27) Product Images from "Enhanced pathogenicity of low-pathogenic H9N2 avian influenza virus after vaccination with infectious bronchitis live attenuated vaccine"

    Article Title: Enhanced pathogenicity of low-pathogenic H9N2 avian influenza virus after vaccination with infectious bronchitis live attenuated vaccine

    Journal: Veterinary World

    doi: 10.14202/vetworld.2018.977-985

    Column chart showing RNA copies of H9 using rRT-polymerase chain reaction in chicken groups and different time conditions. (a) Experiment 1, (b) Experiment 2.
    Figure Legend Snippet: Column chart showing RNA copies of H9 using rRT-polymerase chain reaction in chicken groups and different time conditions. (a) Experiment 1, (b) Experiment 2.

    Techniques Used: Polymerase Chain Reaction

    28) Product Images from "Prime-boost vaccination strategy against avian influenza and Newcastle disease viruses reduces shedding of the challenge viruses"

    Article Title: Prime-boost vaccination strategy against avian influenza and Newcastle disease viruses reduces shedding of the challenge viruses

    Journal: VirusDisease

    doi: 10.1007/s13337-018-0463-3

    qRT-PCR analysis of interferon-γ (IFN-γ) mRNA expression in chickens’ peripheral blood mononuclear cells (PBMCs) at 1 and 2 weeks post vaccination. Different letters within the same week are significantly different at p value (≤ 0.05)
    Figure Legend Snippet: qRT-PCR analysis of interferon-γ (IFN-γ) mRNA expression in chickens’ peripheral blood mononuclear cells (PBMCs) at 1 and 2 weeks post vaccination. Different letters within the same week are significantly different at p value (≤ 0.05)

    Techniques Used: Quantitative RT-PCR, Expressing

    29) Product Images from "Inhibition of autophagy limits vertical transmission of Zika virus in pregnant mice"

    Article Title: Inhibition of autophagy limits vertical transmission of Zika virus in pregnant mice

    Journal: The Journal of Experimental Medicine

    doi: 10.1084/jem.20170957

    HCQ treatment during pregnancy reduces maternal-fetal transmission of ZIKV. (A) Schematic depiction of experiment of administration of HCQ (40 mg/kg/day via intraperitoneal route) as a treatment for ZIKV infection during pregnancy. Pregnant WT female mice were treated with HCQ or PBS as a mock control from day +1 post–ZIKV infection (E10.5) to E14.5. (B) Immunohistochemical staining for autophagy protein, p62, in mouse placentas. Image represents four to six placentas from four to six independent litters of each treatment group. Bars, 100 µm. (C) ZIKV RNA from infected placentas treated with either HCQ or PBS and detected by quantitative RT-PCR (qRT-PCR); n = 14–15 placental samples from four or five litters. Bars represent medians. (D and E) Viral burden assayed by qRT-PCR from maternal serum (D) and spleen (E) shows no difference. Symbols represent individual pregnant mice ( n = 6 per group) from three independent experiments. (F) ISH images showing ZIKV infection in mock- or HCQ-treated placentas. Higher power images of area within red box are depicted in the lower panels. The images are representative of five to seven placentas from three independent dams. (G) Representative hematoxylin and eosin staining shows histopathological features of placentas from indicated treatment groups; solid lines mark the labyrinth. Blue arrows, apoptotic trophoblasts. Black arrows, increased number of nucleated fetal erythrocytes in the labyrinth. (H and I) Measurement of placental and labyrinth area; n = 6–7 samples from at least three independent dams. (J) ZIKV RNA in fetal heads measured by qRT-PCR. Mock, n = 14; HCQ, n = 13. Bars represent the median values of 13 or 14 samples from four independent experiments. (K) Fetal size was measured as crown-rump length (CRL) times occipital-frontal (OF) diameter expressed as mm 2 . Each symbol represents data from individual fetuses; n = 12–13. In all dot plot panels, *, P
    Figure Legend Snippet: HCQ treatment during pregnancy reduces maternal-fetal transmission of ZIKV. (A) Schematic depiction of experiment of administration of HCQ (40 mg/kg/day via intraperitoneal route) as a treatment for ZIKV infection during pregnancy. Pregnant WT female mice were treated with HCQ or PBS as a mock control from day +1 post–ZIKV infection (E10.5) to E14.5. (B) Immunohistochemical staining for autophagy protein, p62, in mouse placentas. Image represents four to six placentas from four to six independent litters of each treatment group. Bars, 100 µm. (C) ZIKV RNA from infected placentas treated with either HCQ or PBS and detected by quantitative RT-PCR (qRT-PCR); n = 14–15 placental samples from four or five litters. Bars represent medians. (D and E) Viral burden assayed by qRT-PCR from maternal serum (D) and spleen (E) shows no difference. Symbols represent individual pregnant mice ( n = 6 per group) from three independent experiments. (F) ISH images showing ZIKV infection in mock- or HCQ-treated placentas. Higher power images of area within red box are depicted in the lower panels. The images are representative of five to seven placentas from three independent dams. (G) Representative hematoxylin and eosin staining shows histopathological features of placentas from indicated treatment groups; solid lines mark the labyrinth. Blue arrows, apoptotic trophoblasts. Black arrows, increased number of nucleated fetal erythrocytes in the labyrinth. (H and I) Measurement of placental and labyrinth area; n = 6–7 samples from at least three independent dams. (J) ZIKV RNA in fetal heads measured by qRT-PCR. Mock, n = 14; HCQ, n = 13. Bars represent the median values of 13 or 14 samples from four independent experiments. (K) Fetal size was measured as crown-rump length (CRL) times occipital-frontal (OF) diameter expressed as mm 2 . Each symbol represents data from individual fetuses; n = 12–13. In all dot plot panels, *, P

    Techniques Used: Transmission Assay, Infection, Mouse Assay, Immunohistochemistry, Staining, Quantitative RT-PCR, In Situ Hybridization

    30) Product Images from "Comparative oncology evaluation of intravenous recombinant oncolytic Vesicular Stomatitis Virus therapy in spontaneous canine cancer"

    Article Title: Comparative oncology evaluation of intravenous recombinant oncolytic Vesicular Stomatitis Virus therapy in spontaneous canine cancer

    Journal: Molecular cancer therapeutics

    doi: 10.1158/1535-7163.MCT-17-0432

    Assessment of virus pharmacokinetics and transgene expression following two-dose intravenous VSV-IFNβ-NIS treatment in a dog with spontaneous osteosarcoma (a) RNA was isolated from whole blood, PBMCs and plasma collected immediately following VSV-IFNβ-NIS treatment to detect VSV-N gene copies by qRT-PCR. Infectious virus was detected in PBMCs and samples positive for recovery of infectious VSV are denoted using an asterisk (*). (B) Human IFNβ was monitored in plasma collected at indicated time points following administration of two IV doses of VSV-IFNβ-NIS (denoted by ▲)
    Figure Legend Snippet: Assessment of virus pharmacokinetics and transgene expression following two-dose intravenous VSV-IFNβ-NIS treatment in a dog with spontaneous osteosarcoma (a) RNA was isolated from whole blood, PBMCs and plasma collected immediately following VSV-IFNβ-NIS treatment to detect VSV-N gene copies by qRT-PCR. Infectious virus was detected in PBMCs and samples positive for recovery of infectious VSV are denoted using an asterisk (*). (B) Human IFNβ was monitored in plasma collected at indicated time points following administration of two IV doses of VSV-IFNβ-NIS (denoted by ▲)

    Techniques Used: Expressing, Isolation, Quantitative RT-PCR

    Assessment of virus pharmacokinetics, bio-distribution, viremia and antibody neutralization following intravenous VSV-IFNβ-NIS treatment Blood samples were collected immediately following intravenous VSV-IFNβ-NIS treatment in tumor bearing dogs for pharmacokinetic studies. RNA was isolated from (A) whole blood to detect VSV-N gene copies by qRT-PCR; (B) PBMCs were isolated to detect infectious virus by overlay on susceptible BHK cells to measure TCID 50 . Virus bio-distribution was assessed in (c) PBMCs or (d) plasma indicating VSV-N genes were detectable primarily in PBMCs and below limit of detection (LOD) in plasma. Whole blood and serum were collected to monitor (e) viremia and (f) anti-VSV neutralizing antibodies at indicated time points following intravenous VSV-IFNβ-NIS treatment.
    Figure Legend Snippet: Assessment of virus pharmacokinetics, bio-distribution, viremia and antibody neutralization following intravenous VSV-IFNβ-NIS treatment Blood samples were collected immediately following intravenous VSV-IFNβ-NIS treatment in tumor bearing dogs for pharmacokinetic studies. RNA was isolated from (A) whole blood to detect VSV-N gene copies by qRT-PCR; (B) PBMCs were isolated to detect infectious virus by overlay on susceptible BHK cells to measure TCID 50 . Virus bio-distribution was assessed in (c) PBMCs or (d) plasma indicating VSV-N genes were detectable primarily in PBMCs and below limit of detection (LOD) in plasma. Whole blood and serum were collected to monitor (e) viremia and (f) anti-VSV neutralizing antibodies at indicated time points following intravenous VSV-IFNβ-NIS treatment.

    Techniques Used: Neutralization, Isolation, Quantitative RT-PCR

    31) Product Images from "West Nile Virus Temperature Sensitivity and Avian Virulence Are Modulated by NS1-2B Polymorphisms"

    Article Title: West Nile Virus Temperature Sensitivity and Avian Virulence Are Modulated by NS1-2B Polymorphisms

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0004938

    Growth and temperature sensitivity of WNV NY99/KN3829 chimeras in AMCR PBMCs. (A) Growth of chimeric viruses in PBMCs at 37°C. (B) Growth of selected chimeric viruses in PBMCs at 42°C. N = 3 replicates per virus. (C) Variability among replicates in PBMC culture at 37°C. Individual replicates of PBMC cultures of KN-str/KN-NS3-4B and NY-str/KN-NS1-4B are shown. No supernatant was available for one replicate of NY-str/KN-NS1-4B at 6 dpi due to fungal contamination. Filled squares represent KN-str/KN-NS3-4B, and open diamonds represent NY-str/KN-NS1-4B. Viral RNA was extracted from 5 dpi (NY-str/KN-NS1-4B) or 6 dpi (KN-str/KN-NS3-4B) supernatants of cultures shown in black, and the NS3-249 region was sequenced. Cultures shown in grey were not sequenced.
    Figure Legend Snippet: Growth and temperature sensitivity of WNV NY99/KN3829 chimeras in AMCR PBMCs. (A) Growth of chimeric viruses in PBMCs at 37°C. (B) Growth of selected chimeric viruses in PBMCs at 42°C. N = 3 replicates per virus. (C) Variability among replicates in PBMC culture at 37°C. Individual replicates of PBMC cultures of KN-str/KN-NS3-4B and NY-str/KN-NS1-4B are shown. No supernatant was available for one replicate of NY-str/KN-NS1-4B at 6 dpi due to fungal contamination. Filled squares represent KN-str/KN-NS3-4B, and open diamonds represent NY-str/KN-NS1-4B. Viral RNA was extracted from 5 dpi (NY-str/KN-NS1-4B) or 6 dpi (KN-str/KN-NS3-4B) supernatants of cultures shown in black, and the NS3-249 region was sequenced. Cultures shown in grey were not sequenced.

    Techniques Used:

    32) Product Images from "SIV/SHIV-Zika co-infection does not alter disease pathogenesis in adult non-pregnant rhesus macaque model"

    Article Title: SIV/SHIV-Zika co-infection does not alter disease pathogenesis in adult non-pregnant rhesus macaque model

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0006811

    Viral loads of simian immunodeficiency virus, chimeric simian human immunodeficiency virus (SIV/SHIV) and Zika virus (ZIKV) in co-infected macaques. Female rhesus macaques (n = 6) chronically infected with SIVmac239 (n = 4) or SHIV3618MTF (n = 2) were also inoculated subcutaneously with 10 4 plaque forming unit (PFU) of ZIKV PRVABC59. Blood collection was performed according to the study plan on 0, 4, 7, 9, 15, 26 and 51 days post inoculation (dpi) with ZIKV for SIV co-infected individuals and on 0, 3, 5, 7, 10 and 20 dpi with ZIKV for SHIV co-infected individuals. Day 0 (D0) was the day of inoculation with ZIKV. RNA was extracted from collected plasma samples through the use of QiAmp RNA mini kit (Qiagen, Valencia, CA), and viral loads were measured using one-step real time RT-PCR detection method. Viral loads were presented in Log10 RNA copies per milliliter (ml) of plasma. A , schema of time-course sampling in study plan of SIV (n = 4) and SHIV (n = 2) co-infection with ZIKV in rhesus macaques. B , viral load status of SIV in individual (black) and the mean value (red) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. C , viral load status of ZIKV in individual (black) and the mean value (blue) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. Bars indicate standard deviation (±SD) of mean values (n = 4). D , viral load status of SHIV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation. E , viral load status of ZIKV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation.
    Figure Legend Snippet: Viral loads of simian immunodeficiency virus, chimeric simian human immunodeficiency virus (SIV/SHIV) and Zika virus (ZIKV) in co-infected macaques. Female rhesus macaques (n = 6) chronically infected with SIVmac239 (n = 4) or SHIV3618MTF (n = 2) were also inoculated subcutaneously with 10 4 plaque forming unit (PFU) of ZIKV PRVABC59. Blood collection was performed according to the study plan on 0, 4, 7, 9, 15, 26 and 51 days post inoculation (dpi) with ZIKV for SIV co-infected individuals and on 0, 3, 5, 7, 10 and 20 dpi with ZIKV for SHIV co-infected individuals. Day 0 (D0) was the day of inoculation with ZIKV. RNA was extracted from collected plasma samples through the use of QiAmp RNA mini kit (Qiagen, Valencia, CA), and viral loads were measured using one-step real time RT-PCR detection method. Viral loads were presented in Log10 RNA copies per milliliter (ml) of plasma. A , schema of time-course sampling in study plan of SIV (n = 4) and SHIV (n = 2) co-infection with ZIKV in rhesus macaques. B , viral load status of SIV in individual (black) and the mean value (red) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. C , viral load status of ZIKV in individual (black) and the mean value (blue) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. Bars indicate standard deviation (±SD) of mean values (n = 4). D , viral load status of SHIV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation. E , viral load status of ZIKV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation.

    Techniques Used: Infection, Quantitative RT-PCR, Sampling, Standard Deviation

    33) Product Images from "Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings"

    Article Title: Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings

    Journal: The Journal of infectious diseases

    doi: 10.1086/650388

    Integrated assays using plasma spiked with Armored RNA HIV. Armored RNA HIV was spiked into human plasma at concentrations ranging from 10,000 copies/mL to 1,000 copies/mL amd extracted with a QIAamp® Viral RNA Mini Kit according to the manufacturer’s
    Figure Legend Snippet: Integrated assays using plasma spiked with Armored RNA HIV. Armored RNA HIV was spiked into human plasma at concentrations ranging from 10,000 copies/mL to 1,000 copies/mL amd extracted with a QIAamp® Viral RNA Mini Kit according to the manufacturer’s

    Techniques Used:

    34) Product Images from "Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens"

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens

    Journal: Analytical chemistry

    doi: 10.1021/acs.analchem.5b01594

    Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).
    Figure Legend Snippet: Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).

    Techniques Used: RT Lamp Assay, Purification, Nucleic Acid Electrophoresis, Flow Cytometry, Amplification, In Situ, Positive Control, Negative Control

    35) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    36) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    37) Product Images from "Capturing and concentrating adenovirus using magnetic anionic nanobeads"

    Article Title: Capturing and concentrating adenovirus using magnetic anionic nanobeads

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S104926

    Detection of the DNA genome of adenovirus adsorbed onto anionic polymer-coated magnetic beads. Notes: Adenovirus AxCAwt2 (20 μL) in 500 μL PBS were mixed with poly(MVE-MA)-coated magnetic beads. After incubation, the following fractions were obtained: BD, supernatant after incubation with the beads (SP), sample before incubation with the beads (BF), and samples containing the same quantity of adenovirus as in the BD (total fraction, TL). Viral genomic DNA was subsequently extracted from the various fractions using a QIAamp Viral RNA mini kit. These experiments confirmed that adenoviral DNA could be extracted from the beads and subjected to the PCR. DNA corresponding to the adenovirus hexon gene (1597-bp) was amplified by PCR. The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The positions of the 100 bp, 500 bp, and 1,500 bp bands are indicated by arrows. Abbreviations: BD, bead fraction; PBS, phosphate-buffered solution; poly(MVE-MA), poly(methyl vinyl ether-maleic anhydrate); PCR, polymerase chain reaction.
    Figure Legend Snippet: Detection of the DNA genome of adenovirus adsorbed onto anionic polymer-coated magnetic beads. Notes: Adenovirus AxCAwt2 (20 μL) in 500 μL PBS were mixed with poly(MVE-MA)-coated magnetic beads. After incubation, the following fractions were obtained: BD, supernatant after incubation with the beads (SP), sample before incubation with the beads (BF), and samples containing the same quantity of adenovirus as in the BD (total fraction, TL). Viral genomic DNA was subsequently extracted from the various fractions using a QIAamp Viral RNA mini kit. These experiments confirmed that adenoviral DNA could be extracted from the beads and subjected to the PCR. DNA corresponding to the adenovirus hexon gene (1597-bp) was amplified by PCR. The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The positions of the 100 bp, 500 bp, and 1,500 bp bands are indicated by arrows. Abbreviations: BD, bead fraction; PBS, phosphate-buffered solution; poly(MVE-MA), poly(methyl vinyl ether-maleic anhydrate); PCR, polymerase chain reaction.

    Techniques Used: Magnetic Beads, Incubation, Polymerase Chain Reaction, Amplification, DNA Sequencing, Marker

    38) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    39) Product Images from "Flavivirus NS4A-induced Autophagy Protects Cells against Death and Enhances Virus Replication *"

    Article Title: Flavivirus NS4A-induced Autophagy Protects Cells against Death and Enhances Virus Replication *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M110.192500

    Flavivirus infection leads to PI3K-dependent autophagy. A , cytoplasmic distribution of LC3-GFP observed in mock − infected HeLa cells indicates a low level of autophagy typical of healthy cells. However, following infection of HeLa cells with either Dengue-2 virus ( B ) or Modoc virus ( C ), LC3-GFP shifts to a punctate pattern, indicating up-regulation of autophagy and accumulation of LC3-GFP at autophagosomal membranes. D , mock − infected MDCK cells transiently expressing LC3-GFP display little LC3-GFP punctation, indicating a low level of autophagy. Both Dengue-2 ( E ) and Modoc virus-infected MDCK ( F ) cells display LC3-GFP punctation, indicating up-regulation of autophagy following infection. G , inhibition of PI3K by exposure to wortmannin has no effect on LC3-GFP punctation in mock − infected cells but abolishes LC3-GFP punctation induced by both Dengue-2 virus ( H ) and Modoc virus ( I ) infections, indicating that flavivirus-induced autophagy is PI3K-dependent. J , cells were scored for LC3-GFP punctation and expressed as the percent cells with punctate LC3-GFP. A limited amount of LC3-GFP punctation is seen in mock control cells, whereas infection by either Dengue-2 or Modoc virus increases punctation dramatically (to ∼70%), indicating flavivirus-induced up-regulation of autophagy. PI3K inhibition by wortmannin completely eliminates both Modoc and Dengue-2 virus-induced up-regulation of autophagy. K, boxes , mock − infected MDCK cells display double-membrane autophagosomes typical of housekeeping functions in healthy cells. Both Dengue-2 ( L, boxes ) and Modoc ( M, boxes ) virus-infected MDCK cells show an increase in double-membrane autophagosomes, demonstrating autophagy up-regulation following infection in these cells.
    Figure Legend Snippet: Flavivirus infection leads to PI3K-dependent autophagy. A , cytoplasmic distribution of LC3-GFP observed in mock − infected HeLa cells indicates a low level of autophagy typical of healthy cells. However, following infection of HeLa cells with either Dengue-2 virus ( B ) or Modoc virus ( C ), LC3-GFP shifts to a punctate pattern, indicating up-regulation of autophagy and accumulation of LC3-GFP at autophagosomal membranes. D , mock − infected MDCK cells transiently expressing LC3-GFP display little LC3-GFP punctation, indicating a low level of autophagy. Both Dengue-2 ( E ) and Modoc virus-infected MDCK ( F ) cells display LC3-GFP punctation, indicating up-regulation of autophagy following infection. G , inhibition of PI3K by exposure to wortmannin has no effect on LC3-GFP punctation in mock − infected cells but abolishes LC3-GFP punctation induced by both Dengue-2 virus ( H ) and Modoc virus ( I ) infections, indicating that flavivirus-induced autophagy is PI3K-dependent. J , cells were scored for LC3-GFP punctation and expressed as the percent cells with punctate LC3-GFP. A limited amount of LC3-GFP punctation is seen in mock control cells, whereas infection by either Dengue-2 or Modoc virus increases punctation dramatically (to ∼70%), indicating flavivirus-induced up-regulation of autophagy. PI3K inhibition by wortmannin completely eliminates both Modoc and Dengue-2 virus-induced up-regulation of autophagy. K, boxes , mock − infected MDCK cells display double-membrane autophagosomes typical of housekeeping functions in healthy cells. Both Dengue-2 ( L, boxes ) and Modoc ( M, boxes ) virus-infected MDCK cells show an increase in double-membrane autophagosomes, demonstrating autophagy up-regulation following infection in these cells.

    Techniques Used: Infection, Expressing, Inhibition

    Flavivirus NS4A-induced protection in epithelial cells is dependent upon autophagy. A , Western blotting using a specific antibody for cleaved LC3II reveals an increase in LC3II cleavage following both live Dengue-2 and Modoc infections, as well as after individual expression of NS4A from either virus. B , β-tubulin was used as a loading control. C , whole-protein lysates were collected from MDCK cells transiently expressing Dengue-2 nonstructural proteins NS1, NS2A, NS4A, and NS4B at 24 h post-transfection, as well as cells transfected with empty plasmid and cells exposed to the transfection agent as controls. All conditions stimulate a moderate amount of LC3 cleavage, likely because of stress occurring during our transfection protocol as indicated by LC3 cleavage after exposure to the transfection agent alone. NS4A expression leads to a substantial increase in LC3 cleavage compared with controls and expression of other Dengue-2 nonstructural genes, indicating NS4A expression alone is uniquely sufficient to up-regulate autophagy in MDCK cells. D , β-tubulin was used as a loading control. E , mock − infected HeLa cells stably expressing LC3-GFP exhibit little LC3-GFP punctation, whereas Dengue-2 ( F ) and Modoc ( G ) virus infections induce LC3-GFP punctation following infection, indicating autophagy up-regulation. H , expression of empty plasmid vector control in MDCK cells transiently expressing LC3-GFP does not induce LC3-GFP punctation, whereas expression of Dengue-2 NS4A ( I ) or Modoc NS4A ( J ) both induce LC3-GFP punctation, indicating autophagy up-regulation following NS4A expression.
    Figure Legend Snippet: Flavivirus NS4A-induced protection in epithelial cells is dependent upon autophagy. A , Western blotting using a specific antibody for cleaved LC3II reveals an increase in LC3II cleavage following both live Dengue-2 and Modoc infections, as well as after individual expression of NS4A from either virus. B , β-tubulin was used as a loading control. C , whole-protein lysates were collected from MDCK cells transiently expressing Dengue-2 nonstructural proteins NS1, NS2A, NS4A, and NS4B at 24 h post-transfection, as well as cells transfected with empty plasmid and cells exposed to the transfection agent as controls. All conditions stimulate a moderate amount of LC3 cleavage, likely because of stress occurring during our transfection protocol as indicated by LC3 cleavage after exposure to the transfection agent alone. NS4A expression leads to a substantial increase in LC3 cleavage compared with controls and expression of other Dengue-2 nonstructural genes, indicating NS4A expression alone is uniquely sufficient to up-regulate autophagy in MDCK cells. D , β-tubulin was used as a loading control. E , mock − infected HeLa cells stably expressing LC3-GFP exhibit little LC3-GFP punctation, whereas Dengue-2 ( F ) and Modoc ( G ) virus infections induce LC3-GFP punctation following infection, indicating autophagy up-regulation. H , expression of empty plasmid vector control in MDCK cells transiently expressing LC3-GFP does not induce LC3-GFP punctation, whereas expression of Dengue-2 NS4A ( I ) or Modoc NS4A ( J ) both induce LC3-GFP punctation, indicating autophagy up-regulation following NS4A expression.

    Techniques Used: Western Blot, Expressing, Transfection, Plasmid Preparation, Infection, Stable Transfection

    Dengue-2 virus induces LC3 cleavage. Upper panel , analysis of whole-protein lysates of Dengue-2-infected cells at 0, 3, 6, 12, 24, 36, and 48 hpi demonstrates a steady increase in LC3 cleavage to 36 hpi, indicating Dengue-2-induced autophagy in infected cells. The decline in LC3 cleavage at 48 hpi suggests complete LC3 turnover at late hpi. Lower panel , β-tubulin was used as a loading control.
    Figure Legend Snippet: Dengue-2 virus induces LC3 cleavage. Upper panel , analysis of whole-protein lysates of Dengue-2-infected cells at 0, 3, 6, 12, 24, 36, and 48 hpi demonstrates a steady increase in LC3 cleavage to 36 hpi, indicating Dengue-2-induced autophagy in infected cells. The decline in LC3 cleavage at 48 hpi suggests complete LC3 turnover at late hpi. Lower panel , β-tubulin was used as a loading control.

    Techniques Used: Infection

    Flaviviruses establish a productive infection in renal epithelial cells. Dengue-2 and Modoc virus-infected MDCK cells were fixed at 48 hpi and stained with phalloidin-TRITC ( red , actin) and antibody Di-4G2-15 ( green ) flavivirus Envelope protein. A , mock − infected cells lack positive staining for virus protein. Massive flavivirus Envelope protein accumulation in cytoplasmic vesicles within the ER-rich perinuclear region of both Dengue-2 virus ( B ) and Modoc virus ( C )- infected cells demonstrates the establishment of flavivirus translation and assembly. D , qRT-PCR of infected cell supernatant demonstrates rapid replication and release of virus.
    Figure Legend Snippet: Flaviviruses establish a productive infection in renal epithelial cells. Dengue-2 and Modoc virus-infected MDCK cells were fixed at 48 hpi and stained with phalloidin-TRITC ( red , actin) and antibody Di-4G2-15 ( green ) flavivirus Envelope protein. A , mock − infected cells lack positive staining for virus protein. Massive flavivirus Envelope protein accumulation in cytoplasmic vesicles within the ER-rich perinuclear region of both Dengue-2 virus ( B ) and Modoc virus ( C )- infected cells demonstrates the establishment of flavivirus translation and assembly. D , qRT-PCR of infected cell supernatant demonstrates rapid replication and release of virus.

    Techniques Used: Infection, Staining, Quantitative RT-PCR

    Inhibition of autophagy reduces extracellular flavivirus titer. A , plaque assay of Dengue-2 virus replication in MDCK renal epithelial cells with and without inhibitors of autophagy by wortmannin or 3MA demonstrates a role for autophagy in Dengue-2 replication. Inhibition of autophagy by either wortmannin or 3MA reduces viral titer by 27% at 96 hpi. B , plaque assay analysis of Modoc virus replication in MDCK cells with and without 3MA reveals a similar role for autophagy in Modoc replication. Inhibition of autophagy by 3MA leads to a 71% decline in Modoc virus replication in MDCK cells at 96 hpi. C , RT-PCR quantification of extracellular Modoc viral RNA at 96 hpi also demonstrates that autophagy inhibition by 3MA reduces replication. Note that CP value indicates number of rounds of replication to measurability, and thus lower initial levels result in higher CP values.
    Figure Legend Snippet: Inhibition of autophagy reduces extracellular flavivirus titer. A , plaque assay of Dengue-2 virus replication in MDCK renal epithelial cells with and without inhibitors of autophagy by wortmannin or 3MA demonstrates a role for autophagy in Dengue-2 replication. Inhibition of autophagy by either wortmannin or 3MA reduces viral titer by 27% at 96 hpi. B , plaque assay analysis of Modoc virus replication in MDCK cells with and without 3MA reveals a similar role for autophagy in Modoc replication. Inhibition of autophagy by 3MA leads to a 71% decline in Modoc virus replication in MDCK cells at 96 hpi. C , RT-PCR quantification of extracellular Modoc viral RNA at 96 hpi also demonstrates that autophagy inhibition by 3MA reduces replication. Note that CP value indicates number of rounds of replication to measurability, and thus lower initial levels result in higher CP values.

    Techniques Used: Inhibition, Plaque Assay, Reverse Transcription Polymerase Chain Reaction

    Expression of flavivirus NS4A alone leads to protection similar to that provided by infection with live flavivirus. A , individual expression of any of the 10 Dengue-2 structural and nonstructural genes does not lead to significant death in MDCK renal epithelial cells. Dengue-2 NS4A is the only gene that protects against cell killing by CPT, similar to live virus infection. NS2A and NS5 each allow slightly more death by CPT than mock − infected cells. Individual expression of empty plasmid vector, the structural protein capsid or core ( C ), premembrane ( prM ), and envelope ( E ) proteins. NS1, NS2B, NS3, or NS4B has no effect on CPT-induced death. B , infection with live Modoc virus infection or individual expression of Modoc NS4A also protects cells against CPT-induced death. Expression of NS2A, NS4A, or NS4B does not lead to significant cell killing in MDCK cells (
    Figure Legend Snippet: Expression of flavivirus NS4A alone leads to protection similar to that provided by infection with live flavivirus. A , individual expression of any of the 10 Dengue-2 structural and nonstructural genes does not lead to significant death in MDCK renal epithelial cells. Dengue-2 NS4A is the only gene that protects against cell killing by CPT, similar to live virus infection. NS2A and NS5 each allow slightly more death by CPT than mock − infected cells. Individual expression of empty plasmid vector, the structural protein capsid or core ( C ), premembrane ( prM ), and envelope ( E ) proteins. NS1, NS2B, NS3, or NS4B has no effect on CPT-induced death. B , infection with live Modoc virus infection or individual expression of Modoc NS4A also protects cells against CPT-induced death. Expression of NS2A, NS4A, or NS4B does not lead to significant cell killing in MDCK cells (

    Techniques Used: Expressing, Infection, Cycling Probe Technology, Plasmid Preparation

    Inhibition of autophagy abolishes flavivirus-induced protection against death. A , nonspecific PI3K inhibition by wortmannin eliminates protection by both Dengue-2 and Modoc virus in MDCK renal epithelial cells. B , inhibition of class III PI3K activity by 3MA also eliminates protection against death in MDCK cells, indicating class III PI3K involvement in flavivirus-induced protection. C , inhibition of autophagy by Beclin-1 knockdown eliminates protection by both Dengue-2 and Modoc virus against CPT-induced death. D , inhibition of autophagy by Atg5 knock-out also eliminates protection by both Dengue-2 and Modoc virus . E , up-regulation of autophagy by starvation prior to infection allows no significant death by CPT treatment with or without infection by either virus. F , up-regulation of autophagy signaling by mammalian target of rapamycin inactivation using rapamycin treatment allowed no significant death by high concentration of CPT with or without infection by either virus.
    Figure Legend Snippet: Inhibition of autophagy abolishes flavivirus-induced protection against death. A , nonspecific PI3K inhibition by wortmannin eliminates protection by both Dengue-2 and Modoc virus in MDCK renal epithelial cells. B , inhibition of class III PI3K activity by 3MA also eliminates protection against death in MDCK cells, indicating class III PI3K involvement in flavivirus-induced protection. C , inhibition of autophagy by Beclin-1 knockdown eliminates protection by both Dengue-2 and Modoc virus against CPT-induced death. D , inhibition of autophagy by Atg5 knock-out also eliminates protection by both Dengue-2 and Modoc virus . E , up-regulation of autophagy by starvation prior to infection allows no significant death by CPT treatment with or without infection by either virus. F , up-regulation of autophagy signaling by mammalian target of rapamycin inactivation using rapamycin treatment allowed no significant death by high concentration of CPT with or without infection by either virus.

    Techniques Used: Inhibition, Activity Assay, Cycling Probe Technology, Knock-Out, Infection, Concentration Assay

    40) Product Images from "In Vitro Selection of Highly Darunavir-Resistant and Replication-Competent HIV-1 Variants by Using a Mixture of Clinical HIV-1 Isolates Resistant to Multiple Conventional Protease Inhibitors ▿ Selection of Highly Darunavir-Resistant and Replication-Competent HIV-1 Variants by Using a Mixture of Clinical HIV-1 Isolates Resistant to Multiple Conventional Protease Inhibitors ▿ †"

    Article Title: In Vitro Selection of Highly Darunavir-Resistant and Replication-Competent HIV-1 Variants by Using a Mixture of Clinical HIV-1 Isolates Resistant to Multiple Conventional Protease Inhibitors ▿ Selection of Highly Darunavir-Resistant and Replication-Competent HIV-1 Variants by Using a Mixture of Clinical HIV-1 Isolates Resistant to Multiple Conventional Protease Inhibitors ▿ †

    Journal: Journal of Virology

    doi: 10.1128/JVI.00967-10

    Sequence analysis of the protease-encoding regions in the mixture of 8 HIV MDR isolates. Viral RNA was purified from each indicated supernatant using the QIAamp viral RNA minikit (Qiagen Inc., Valencia, CA), and RT-PCR was carried out using the Superscript First-Strand synthesis system for RT-PCR (Invitrogen, Carlsbad, CA) according to the manufacturer's instructions. The amino acid sequences of protease deduced from nucleotide sequences of the protease-encoding region of HIV-1 clones determined are shown. The fraction of clones examined is indicated on the right. The amino acid sequence of protease of a wild-type pNL4-3 clone is shown as a reference. Identity with this sequence at individual amino acid positions is indicated (dots).
    Figure Legend Snippet: Sequence analysis of the protease-encoding regions in the mixture of 8 HIV MDR isolates. Viral RNA was purified from each indicated supernatant using the QIAamp viral RNA minikit (Qiagen Inc., Valencia, CA), and RT-PCR was carried out using the Superscript First-Strand synthesis system for RT-PCR (Invitrogen, Carlsbad, CA) according to the manufacturer's instructions. The amino acid sequences of protease deduced from nucleotide sequences of the protease-encoding region of HIV-1 clones determined are shown. The fraction of clones examined is indicated on the right. The amino acid sequence of protease of a wild-type pNL4-3 clone is shown as a reference. Identity with this sequence at individual amino acid positions is indicated (dots).

    Techniques Used: Sequencing, Purification, Reverse Transcription Polymerase Chain Reaction, Clone Assay

    41) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    42) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    43) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    44) Product Images from "Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology"

    Article Title: Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S191784

    Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.
    Figure Legend Snippet: Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Real-time Polymerase Chain Reaction

    Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.
    Figure Legend Snippet: Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Amplification, Polymerase Chain Reaction, Agarose Gel Electrophoresis, DNA Sequencing, Marker, Reverse Transcription Polymerase Chain Reaction, Negative Control

    45) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    46) Product Images from "Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing"

    Article Title: Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing

    Journal: Genome Biology and Evolution

    doi: 10.1093/gbe/evv034

    Confirmation of contaminated columns as origin of CHIV14 DNA by qPCR. ( A ) Verification of CHIV14 genome assembled from the Illumina deep-sequencing data by overlapping PCR and inverted PCR. Five sets of overlapping primer pairs and one set of inverted primer pair were designed and used to amplify overlapping DNA fragments. (Left) Schematic diagram of the positions of primer pairs for the overlapping PCR and inverted PCR. (Right) Amplification overlapping viral DNA fragments. The numbers above indicate the primer pair used for the PCR as illustrated on the left. The numbers on the left indicate the molecular weight in base pairs. ( B ) Scatterplot showing copy number of CHIV14 per microliter of DNA extraction. DNA from patients ( n = 13), healthy controls ( n = 13), and water ( n = 31) was extracted using QIAamp mini spin columns (QIAamp Viral RNA Mini kit; Qiagen). In parallel, seven DNA extractions for each specimen type (patients, healthy individuals, and water) were performed using the UCP columns. Each dot represents one specimen. Bars show the average copy numbers of the viral genome.
    Figure Legend Snippet: Confirmation of contaminated columns as origin of CHIV14 DNA by qPCR. ( A ) Verification of CHIV14 genome assembled from the Illumina deep-sequencing data by overlapping PCR and inverted PCR. Five sets of overlapping primer pairs and one set of inverted primer pair were designed and used to amplify overlapping DNA fragments. (Left) Schematic diagram of the positions of primer pairs for the overlapping PCR and inverted PCR. (Right) Amplification overlapping viral DNA fragments. The numbers above indicate the primer pair used for the PCR as illustrated on the left. The numbers on the left indicate the molecular weight in base pairs. ( B ) Scatterplot showing copy number of CHIV14 per microliter of DNA extraction. DNA from patients ( n = 13), healthy controls ( n = 13), and water ( n = 31) was extracted using QIAamp mini spin columns (QIAamp Viral RNA Mini kit; Qiagen). In parallel, seven DNA extractions for each specimen type (patients, healthy individuals, and water) were performed using the UCP columns. Each dot represents one specimen. Bars show the average copy numbers of the viral genome.

    Techniques Used: Real-time Polymerase Chain Reaction, Sequencing, Polymerase Chain Reaction, Amplification, Molecular Weight, DNA Extraction

    47) Product Images from "Evolution of Neuroadaptation in the Periphery and Purifying Selection in the Brain Contribute to Compartmentalization of Simian Immunodeficiency Virus (SIV) in the Brains of Rhesus Macaques with SIV-Associated Encephalitis"

    Article Title: Evolution of Neuroadaptation in the Periphery and Purifying Selection in the Brain Contribute to Compartmentalization of Simian Immunodeficiency Virus (SIV) in the Brains of Rhesus Macaques with SIV-Associated Encephalitis

    Journal: Journal of Virology

    doi: 10.1128/JVI.00137-16

    Viral genomic RNA and DNA levels in the brain readily distinguish macaques based on SIVE status.
    Figure Legend Snippet: Viral genomic RNA and DNA levels in the brain readily distinguish macaques based on SIVE status.

    Techniques Used:

    48) Product Images from "Sand Fly–Associated Phlebovirus with Evidence of Neutralizing Antibodies in Humans, Kenya"

    Article Title: Sand Fly–Associated Phlebovirus with Evidence of Neutralizing Antibodies in Humans, Kenya

    Journal: Emerging Infectious Diseases

    doi: 10.3201/eid2504.180750

    Phylogenetic relationship of novel sand fly–associated phlebovirus Ntepes virus from Kenya (red bold text) in relation to other selected members of the Phlebovirus genus. A) RNA-dependent RNA polymerase; B) nucleocapsid protein; C) glycoprotein Gn; D) glycoprotein Gc. The phylogenetic trees were inferred based on complete large, medium, and small protein sequences, applying maximum likelihood analysis in PhyML version 3.0 ( http://www.atgc-montpellier.fr/phyml/versions.php ) using the LG substitution model. Statistical support of the tree topology was evaluated by bootstrap resampling of the sequences 1,000 times. Sequences are identified by virus name and branch colors. Bootstrap values > 70 are indicated at the nodes. Scale bar represents numbers of substitutions per site.
    Figure Legend Snippet: Phylogenetic relationship of novel sand fly–associated phlebovirus Ntepes virus from Kenya (red bold text) in relation to other selected members of the Phlebovirus genus. A) RNA-dependent RNA polymerase; B) nucleocapsid protein; C) glycoprotein Gn; D) glycoprotein Gc. The phylogenetic trees were inferred based on complete large, medium, and small protein sequences, applying maximum likelihood analysis in PhyML version 3.0 ( http://www.atgc-montpellier.fr/phyml/versions.php ) using the LG substitution model. Statistical support of the tree topology was evaluated by bootstrap resampling of the sequences 1,000 times. Sequences are identified by virus name and branch colors. Bootstrap values > 70 are indicated at the nodes. Scale bar represents numbers of substitutions per site.

    Techniques Used:

    49) Product Images from "Flavivirus NS4A-induced Autophagy Protects Cells against Death and Enhances Virus Replication *"

    Article Title: Flavivirus NS4A-induced Autophagy Protects Cells against Death and Enhances Virus Replication *

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M110.192500

    Inhibition of autophagy reduces extracellular flavivirus titer. A , plaque assay of Dengue-2 virus replication in MDCK renal epithelial cells with and without inhibitors of autophagy by wortmannin or 3MA demonstrates a role for autophagy in Dengue-2 replication. Inhibition of autophagy by either wortmannin or 3MA reduces viral titer by 27% at 96 hpi. B , plaque assay analysis of Modoc virus replication in MDCK cells with and without 3MA reveals a similar role for autophagy in Modoc replication. Inhibition of autophagy by 3MA leads to a 71% decline in Modoc virus replication in MDCK cells at 96 hpi. C , RT-PCR quantification of extracellular Modoc viral RNA at 96 hpi also demonstrates that autophagy inhibition by 3MA reduces replication. Note that CP value indicates number of rounds of replication to measurability, and thus lower initial levels result in higher CP values.
    Figure Legend Snippet: Inhibition of autophagy reduces extracellular flavivirus titer. A , plaque assay of Dengue-2 virus replication in MDCK renal epithelial cells with and without inhibitors of autophagy by wortmannin or 3MA demonstrates a role for autophagy in Dengue-2 replication. Inhibition of autophagy by either wortmannin or 3MA reduces viral titer by 27% at 96 hpi. B , plaque assay analysis of Modoc virus replication in MDCK cells with and without 3MA reveals a similar role for autophagy in Modoc replication. Inhibition of autophagy by 3MA leads to a 71% decline in Modoc virus replication in MDCK cells at 96 hpi. C , RT-PCR quantification of extracellular Modoc viral RNA at 96 hpi also demonstrates that autophagy inhibition by 3MA reduces replication. Note that CP value indicates number of rounds of replication to measurability, and thus lower initial levels result in higher CP values.

    Techniques Used: Inhibition, Plaque Assay, Reverse Transcription Polymerase Chain Reaction

    50) Product Images from "APOBEC3G and APOBEC3F Act in Concert To Extinguish HIV-1 Replication"

    Article Title: APOBEC3G and APOBEC3F Act in Concert To Extinguish HIV-1 Replication

    Journal: Journal of Virology

    doi: 10.1128/JVI.03275-15

    Absolute restriction of HIV in vivo requires both APOBEC3G and APOBEC3F. (A) (Top) The plasma of BLT humanized mice infected with JRCSFvifH42/43D (blue lines) was monitored longitudinally for the presence of HIV RNA. Wild-type JRCSF RNA is presented in aggregate ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic-DNA extracts from each tissue listed from the infected humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (B) (Top) Viral RNA was present in the plasma of 4/4 mice infected with JRCSFvifW79S (red lines) at levels comparable to those of wild-type JRCSF ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic DNA extracted from each tissue listed from the infected BLT humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (C) (Top) Plasma viral RNA was not observed in 4/4 mice infected with JRCSFvifH42/43DW79S (green lines) for up to 8 weeks postexposure, in contrast to infection with wild-type JRCSF (black diamonds). (Bottom) Viral DNA was not detected in any tissue of JRCSFvifH42/43DW79S-inoculated BLT humanized mice (−). (D) Sequencing of viral DNA amplified from the tissues of mice infected with JRCSFvifH42/43D (G3 and G4) revealed heavy G-to-A mutation at GG sites (blue bar). Few G-to-A mutations were present at GA sites in viral DNA from mice infected with JRCSFvifH42/43D or JRCSFvifW79S, and no mutations were observed in WT1 JRCSF DNA. The data are presented as means ± SEM.
    Figure Legend Snippet: Absolute restriction of HIV in vivo requires both APOBEC3G and APOBEC3F. (A) (Top) The plasma of BLT humanized mice infected with JRCSFvifH42/43D (blue lines) was monitored longitudinally for the presence of HIV RNA. Wild-type JRCSF RNA is presented in aggregate ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic-DNA extracts from each tissue listed from the infected humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (B) (Top) Viral RNA was present in the plasma of 4/4 mice infected with JRCSFvifW79S (red lines) at levels comparable to those of wild-type JRCSF ( n = 7) (black diamonds). (Bottom) Nested PCR to detect HIV DNA was performed on genomic DNA extracted from each tissue listed from the infected BLT humanized mice. +, tissues positive for HIV DNA; −, tissues where HIV DNA was not detected. (C) (Top) Plasma viral RNA was not observed in 4/4 mice infected with JRCSFvifH42/43DW79S (green lines) for up to 8 weeks postexposure, in contrast to infection with wild-type JRCSF (black diamonds). (Bottom) Viral DNA was not detected in any tissue of JRCSFvifH42/43DW79S-inoculated BLT humanized mice (−). (D) Sequencing of viral DNA amplified from the tissues of mice infected with JRCSFvifH42/43D (G3 and G4) revealed heavy G-to-A mutation at GG sites (blue bar). Few G-to-A mutations were present at GA sites in viral DNA from mice infected with JRCSFvifH42/43D or JRCSFvifW79S, and no mutations were observed in WT1 JRCSF DNA. The data are presented as means ± SEM.

    Techniques Used: In Vivo, Mouse Assay, Infection, Nested PCR, Sequencing, Amplification, Mutagenesis

    51) Product Images from "SIV/SHIV-Zika co-infection does not alter disease pathogenesis in adult non-pregnant rhesus macaque model"

    Article Title: SIV/SHIV-Zika co-infection does not alter disease pathogenesis in adult non-pregnant rhesus macaque model

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0006811

    Viral loads of simian immunodeficiency virus, chimeric simian human immunodeficiency virus (SIV/SHIV) and Zika virus (ZIKV) in co-infected macaques. Female rhesus macaques (n = 6) chronically infected with SIVmac239 (n = 4) or SHIV3618MTF (n = 2) were also inoculated subcutaneously with 10 4 plaque forming unit (PFU) of ZIKV PRVABC59. Blood collection was performed according to the study plan on 0, 4, 7, 9, 15, 26 and 51 days post inoculation (dpi) with ZIKV for SIV co-infected individuals and on 0, 3, 5, 7, 10 and 20 dpi with ZIKV for SHIV co-infected individuals. Day 0 (D0) was the day of inoculation with ZIKV. RNA was extracted from collected plasma samples through the use of QiAmp RNA mini kit (Qiagen, Valencia, CA), and viral loads were measured using one-step real time RT-PCR detection method. Viral loads were presented in Log10 RNA copies per milliliter (ml) of plasma. A , schema of time-course sampling in study plan of SIV (n = 4) and SHIV (n = 2) co-infection with ZIKV in rhesus macaques. B , viral load status of SIV in individual (black) and the mean value (red) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. C , viral load status of ZIKV in individual (black) and the mean value (blue) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. Bars indicate standard deviation (±SD) of mean values (n = 4). D , viral load status of SHIV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation. E , viral load status of ZIKV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation.
    Figure Legend Snippet: Viral loads of simian immunodeficiency virus, chimeric simian human immunodeficiency virus (SIV/SHIV) and Zika virus (ZIKV) in co-infected macaques. Female rhesus macaques (n = 6) chronically infected with SIVmac239 (n = 4) or SHIV3618MTF (n = 2) were also inoculated subcutaneously with 10 4 plaque forming unit (PFU) of ZIKV PRVABC59. Blood collection was performed according to the study plan on 0, 4, 7, 9, 15, 26 and 51 days post inoculation (dpi) with ZIKV for SIV co-infected individuals and on 0, 3, 5, 7, 10 and 20 dpi with ZIKV for SHIV co-infected individuals. Day 0 (D0) was the day of inoculation with ZIKV. RNA was extracted from collected plasma samples through the use of QiAmp RNA mini kit (Qiagen, Valencia, CA), and viral loads were measured using one-step real time RT-PCR detection method. Viral loads were presented in Log10 RNA copies per milliliter (ml) of plasma. A , schema of time-course sampling in study plan of SIV (n = 4) and SHIV (n = 2) co-infection with ZIKV in rhesus macaques. B , viral load status of SIV in individual (black) and the mean value (red) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. C , viral load status of ZIKV in individual (black) and the mean value (blue) of all SIV-ZIKV co-infected animals (n = 4) in days post ZIKV inoculation. Bars indicate standard deviation (±SD) of mean values (n = 4). D , viral load status of SHIV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation. E , viral load status of ZIKV in individual SHIV-ZIKV co-infected animals (n = 2) in days post ZIKV inoculation.

    Techniques Used: Infection, Quantitative RT-PCR, Sampling, Standard Deviation

    52) Product Images from "Description and characterization of a novel live-attenuated tri-segmented Machupo virus in Guinea pigs"

    Article Title: Description and characterization of a novel live-attenuated tri-segmented Machupo virus in Guinea pigs

    Journal: Virology Journal

    doi: 10.1186/s12985-018-1009-4

    IFN-β production by A549 cells infected either with MACV wt, r3MACV or SeV. A549 cells were infected for one hour with the MACV wt, r3MACV or SeV viruses at a MOI of 3, then the inoculum was removed and fresh culture medium was added. At indicated time points, a virus titers in the supernatants were determined by virus TCID 50 , as described in Fig. 2 ; b production of IFN-β was detected in supernatant solutions using the VeriKine Human IFN BETA ELISA Kit (PBL assay sciences); and c intracellular RNA was extracted using the QIAmp Viral RNA Mini kit (Qiagen). RT-qPCR were performed using the QuantiTect Probe RT-PCR kit (Qiagen) in a 30 μl final volume with 5 μl of purified RNA and primers and probe at a final concentration of 400 nM and 200 nM, respectively. The assay was carried out using a CFX96 model (Bio-Rad) with a cycling profile of 50 °C for 30 min, 95 °C for 15 min, and 40 cycles at 94 °C for 15 s followed by 60°c for 1 min. The primers and probes (available on request) were designed using the Eurofins Genomics’ online tool ( www.eurofinsgenomics.eu ). Probes were 5′- and 3′-labelled with the fluorescent reporter dye 6-carboxyfluorescein (FAM) and the Black Hole Quencher (BHQ-1), respectively. All procedures were carried out following manufacturers’ instructions. This experiment was carried out in triplicate. Asterisks denote significant differences (P
    Figure Legend Snippet: IFN-β production by A549 cells infected either with MACV wt, r3MACV or SeV. A549 cells were infected for one hour with the MACV wt, r3MACV or SeV viruses at a MOI of 3, then the inoculum was removed and fresh culture medium was added. At indicated time points, a virus titers in the supernatants were determined by virus TCID 50 , as described in Fig. 2 ; b production of IFN-β was detected in supernatant solutions using the VeriKine Human IFN BETA ELISA Kit (PBL assay sciences); and c intracellular RNA was extracted using the QIAmp Viral RNA Mini kit (Qiagen). RT-qPCR were performed using the QuantiTect Probe RT-PCR kit (Qiagen) in a 30 μl final volume with 5 μl of purified RNA and primers and probe at a final concentration of 400 nM and 200 nM, respectively. The assay was carried out using a CFX96 model (Bio-Rad) with a cycling profile of 50 °C for 30 min, 95 °C for 15 min, and 40 cycles at 94 °C for 15 s followed by 60°c for 1 min. The primers and probes (available on request) were designed using the Eurofins Genomics’ online tool ( www.eurofinsgenomics.eu ). Probes were 5′- and 3′-labelled with the fluorescent reporter dye 6-carboxyfluorescein (FAM) and the Black Hole Quencher (BHQ-1), respectively. All procedures were carried out following manufacturers’ instructions. This experiment was carried out in triplicate. Asterisks denote significant differences (P

    Techniques Used: Infection, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Reverse Transcription Polymerase Chain Reaction, Purification, Concentration Assay

    53) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

    54) Product Images from "Structural basis of Zika virus specific antibody protection"

    Article Title: Structural basis of Zika virus specific antibody protection

    Journal: Cell

    doi: 10.1016/j.cell.2016.07.020

    In vivo protection of anti-ZIKV mAbs Four to five week-old C57BL/6 mice were passively transferred 2 mg of anti-Ifnar1 mAb and 250 μg of the indicated mAbs (CHK-166, ZV-54, or ZV-57) via an intraperitoneal injection one day before subcutaneous inoculation with 10 5 FFU of ZIKV Dakar 41519. A . On day 3 after infection, serum was collected for analysis of viral RNA by qRT-PCR. B. Daily weights were measured. For A and B, statistical significance was analyzed by a one-way ANOVA with a Dunnett’s multiple comparisons test (**, P
    Figure Legend Snippet: In vivo protection of anti-ZIKV mAbs Four to five week-old C57BL/6 mice were passively transferred 2 mg of anti-Ifnar1 mAb and 250 μg of the indicated mAbs (CHK-166, ZV-54, or ZV-57) via an intraperitoneal injection one day before subcutaneous inoculation with 10 5 FFU of ZIKV Dakar 41519. A . On day 3 after infection, serum was collected for analysis of viral RNA by qRT-PCR. B. Daily weights were measured. For A and B, statistical significance was analyzed by a one-way ANOVA with a Dunnett’s multiple comparisons test (**, P

    Techniques Used: In Vivo, Mouse Assay, Injection, Infection, Quantitative RT-PCR

    55) Product Images from "Quantifying Mixed Populations of Drug-Resistant Human Immunodeficiency Virus Type 1"

    Article Title: Quantifying Mixed Populations of Drug-Resistant Human Immunodeficiency Virus Type 1

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.49.8.3334-3340.2005

    M184 viral RNA analysis. A series of mixtures of M184V and wild-type HIV-1 viral RNA containing 100% M184M or 1, 10, 40, 60, 90, 99, or 100% M184V were assayed using the M184 RTx system and ABI 7900. Real-time PCR fluorescence in relative fluorescence units (RFU) versus PCR cycles (left), negative first derivative of melt fluorescence with respect to temperature (−dRFU/dTemp) versus temperature in °C (middle), and standard curve with linear regression (right) for M184M channel (A) and M184V channel (B) are shown. Samples containing either 1% M184M or 1% M184V (asterisk) could be distinguished from those lacking either target. Note that late nonspecific amplification in the no-target control (NTC) reaction of the M184V channel is distinguished by the melt T m .
    Figure Legend Snippet: M184 viral RNA analysis. A series of mixtures of M184V and wild-type HIV-1 viral RNA containing 100% M184M or 1, 10, 40, 60, 90, 99, or 100% M184V were assayed using the M184 RTx system and ABI 7900. Real-time PCR fluorescence in relative fluorescence units (RFU) versus PCR cycles (left), negative first derivative of melt fluorescence with respect to temperature (−dRFU/dTemp) versus temperature in °C (middle), and standard curve with linear regression (right) for M184M channel (A) and M184V channel (B) are shown. Samples containing either 1% M184M or 1% M184V (asterisk) could be distinguished from those lacking either target. Note that late nonspecific amplification in the no-target control (NTC) reaction of the M184V channel is distinguished by the melt T m .

    Techniques Used: Real-time Polymerase Chain Reaction, Fluorescence, Polymerase Chain Reaction, Amplification

    56) Product Images from "Inhibition of Dengue Virus Entry and Multiplication into Monocytes Using RNA Interference"

    Article Title: Inhibition of Dengue Virus Entry and Multiplication into Monocytes Using RNA Interference

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0001410

    Extracellular dengue virus RNA load. DENV RNA in the culture supernatant of transfected and non-transfected monocytes was quantified by RT-qPCR. Result shows marked reduction in viral RNA (51.4%, 63.7%, 52.2%, and 63.0%) for CD-14 associated molecule, CLTC, DNM2, and combined silenced monocytes, respectively, when compared with non-transfected monocytes, which defined as 100% (viral RNA copy number is 1.06×10 4 /µl). This result is statistically significant (One-way ANOVA with Dunnett's post-test, P
    Figure Legend Snippet: Extracellular dengue virus RNA load. DENV RNA in the culture supernatant of transfected and non-transfected monocytes was quantified by RT-qPCR. Result shows marked reduction in viral RNA (51.4%, 63.7%, 52.2%, and 63.0%) for CD-14 associated molecule, CLTC, DNM2, and combined silenced monocytes, respectively, when compared with non-transfected monocytes, which defined as 100% (viral RNA copy number is 1.06×10 4 /µl). This result is statistically significant (One-way ANOVA with Dunnett's post-test, P

    Techniques Used: Transfection, Quantitative RT-PCR

    57) Product Images from "Inhibition of Dengue Virus Entry and Multiplication into Monocytes Using RNA Interference"

    Article Title: Inhibition of Dengue Virus Entry and Multiplication into Monocytes Using RNA Interference

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0001410

    Extracellular dengue virus RNA load. DENV RNA in the culture supernatant of transfected and non-transfected monocytes was quantified by RT-qPCR. Result shows marked reduction in viral RNA (51.4%, 63.7%, 52.2%, and 63.0%) for CD-14 associated molecule, CLTC, DNM2, and combined silenced monocytes, respectively, when compared with non-transfected monocytes, which defined as 100% (viral RNA copy number is 1.06×10 4 /µl). This result is statistically significant (One-way ANOVA with Dunnett's post-test, P
    Figure Legend Snippet: Extracellular dengue virus RNA load. DENV RNA in the culture supernatant of transfected and non-transfected monocytes was quantified by RT-qPCR. Result shows marked reduction in viral RNA (51.4%, 63.7%, 52.2%, and 63.0%) for CD-14 associated molecule, CLTC, DNM2, and combined silenced monocytes, respectively, when compared with non-transfected monocytes, which defined as 100% (viral RNA copy number is 1.06×10 4 /µl). This result is statistically significant (One-way ANOVA with Dunnett's post-test, P

    Techniques Used: Transfection, Quantitative RT-PCR

    58) Product Images from "Initiation of antiretroviral therapy before detection of colonic infiltration by HIV reduces viral reservoirs, inflammation and immune activation"

    Article Title: Initiation of antiretroviral therapy before detection of colonic infiltration by HIV reduces viral reservoirs, inflammation and immune activation

    Journal: Journal of the International AIDS Society

    doi: 10.7448/IAS.19.1.21163

    HIV RNA in the peripheral blood and cerebrospinal fluid during acute HIV infection. HIV RNA measurements during acute HIV infection are compared between participants with detectable colonic HIV RNA and undetectable colonic HIV RNA. Statistically significant pairwise comparisons ( p
    Figure Legend Snippet: HIV RNA in the peripheral blood and cerebrospinal fluid during acute HIV infection. HIV RNA measurements during acute HIV infection are compared between participants with detectable colonic HIV RNA and undetectable colonic HIV RNA. Statistically significant pairwise comparisons ( p

    Techniques Used: Infection

    59) Product Images from "Evolution of Neuroadaptation in the Periphery and Purifying Selection in the Brain Contribute to Compartmentalization of Simian Immunodeficiency Virus (SIV) in the Brains of Rhesus Macaques with SIV-Associated Encephalitis"

    Article Title: Evolution of Neuroadaptation in the Periphery and Purifying Selection in the Brain Contribute to Compartmentalization of Simian Immunodeficiency Virus (SIV) in the Brains of Rhesus Macaques with SIV-Associated Encephalitis

    Journal: Journal of Virology

    doi: 10.1128/JVI.00137-16

    Viral genomic RNA and DNA levels in the brain readily distinguish macaques based on SIVE status.
    Figure Legend Snippet: Viral genomic RNA and DNA levels in the brain readily distinguish macaques based on SIVE status.

    Techniques Used:

    60) Product Images from "Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens"

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens

    Journal: Analytical chemistry

    doi: 10.1021/acs.analchem.5b01594

    Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).
    Figure Legend Snippet: Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).

    Techniques Used: RT Lamp Assay, Purification, Nucleic Acid Electrophoresis, Flow Cytometry, Amplification, In Situ, Positive Control, Negative Control

    61) Product Images from "Comparison of Matrix-Based and Filter Paper-Based Systems for Transport of Plasma for HIV-1 RNA Quantification and Amplicon Preparation for Genotyping"

    Article Title: Comparison of Matrix-Based and Filter Paper-Based Systems for Transport of Plasma for HIV-1 RNA Quantification and Amplicon Preparation for Genotyping

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.00541-16

    Comparison of HIV-1 RNA quantification and drug resistance genotyping following preservation of plasma samples in ViveST tubes (extracted by QIAamp or MiniMag) versus on RNA Sound cards (extracted by QIAamp) versus frozen (extracted by QIAamp). Aliquots
    Figure Legend Snippet: Comparison of HIV-1 RNA quantification and drug resistance genotyping following preservation of plasma samples in ViveST tubes (extracted by QIAamp or MiniMag) versus on RNA Sound cards (extracted by QIAamp) versus frozen (extracted by QIAamp). Aliquots

    Techniques Used: Preserving

    62) Product Images from "Development of a Low Bias Method for Characterizing Viral Populations Using Next Generation Sequencing Technology"

    Article Title: Development of a Low Bias Method for Characterizing Viral Populations Using Next Generation Sequencing Technology

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0013564

    Flowchart of experimental methodology. Cell culture supernatants containing viral particles was collected from HIV infected cells. The RNA was extracted using the QIAamp Viral RNA Mini Kit and coverted into large quantities of single stranded DNA using the WT-Ovation Pico RNA Amplification System. The complementary strand for the ssDNA was then synthesized using the WT-Ovation Exon Module. The final step in the process involved using the Genomic DNA Sample Prep Kit to produce an Illumina library.
    Figure Legend Snippet: Flowchart of experimental methodology. Cell culture supernatants containing viral particles was collected from HIV infected cells. The RNA was extracted using the QIAamp Viral RNA Mini Kit and coverted into large quantities of single stranded DNA using the WT-Ovation Pico RNA Amplification System. The complementary strand for the ssDNA was then synthesized using the WT-Ovation Exon Module. The final step in the process involved using the Genomic DNA Sample Prep Kit to produce an Illumina library.

    Techniques Used: Cell Culture, Infection, Amplification, Synthesized, Sample Prep

    63) Product Images from "Capture of dengue viruses using antibody-integrated graphite-encapsulated magnetic beads produced using gas plasma technology"

    Article Title: Capture of dengue viruses using antibody-integrated graphite-encapsulated magnetic beads produced using gas plasma technology

    Journal: Molecular Medicine Reports

    doi: 10.3892/mmr.2016.5330

    Detection of genomic RNA in DENVs adsorbed onto antibody-integrated magnetic beads. Culture media containing uninfected (Mock) C6/36 cells or C6/36 cells infected with DENV1-4 were mixed with antibody-integrated magnetic beads and separated into BD and SP fractions. Viral RNA was extracted from each fraction using a QIAamp Viral RNA Mini kit and analyzed using RT-PCR analysis. The PCR primer pairs specifically annealed to a common 511 bp fragment among the DENV1-4 serotypes 1–4. Each PCR product was analyzed using agarose gel electrophoresis, and the amplified products were then sequenced to confirm they corresponded to the gene of the DENVs. A 100 bp ladder marker (M) is shown. Numbers indicated on the left are molecular size (bp). DENV, dengue virus; BD, bead fraction; SP, supernatant fraction; RT-PCR, reverse transcription-polymerase chain reaction.
    Figure Legend Snippet: Detection of genomic RNA in DENVs adsorbed onto antibody-integrated magnetic beads. Culture media containing uninfected (Mock) C6/36 cells or C6/36 cells infected with DENV1-4 were mixed with antibody-integrated magnetic beads and separated into BD and SP fractions. Viral RNA was extracted from each fraction using a QIAamp Viral RNA Mini kit and analyzed using RT-PCR analysis. The PCR primer pairs specifically annealed to a common 511 bp fragment among the DENV1-4 serotypes 1–4. Each PCR product was analyzed using agarose gel electrophoresis, and the amplified products were then sequenced to confirm they corresponded to the gene of the DENVs. A 100 bp ladder marker (M) is shown. Numbers indicated on the left are molecular size (bp). DENV, dengue virus; BD, bead fraction; SP, supernatant fraction; RT-PCR, reverse transcription-polymerase chain reaction.

    Techniques Used: Magnetic Beads, Infection, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Amplification, Marker

    Concentrated genomic RNA of DENV following adsorption onto antibody-integrated magnetic beads. The culture medium (10 µ l) of the C6/36 cells infected with DENV1 was diluted with 1 ml phosphate-buffered saline, mixed with antibody-integrated magnetic beads and subjected to magnetic separation. Viral RNA was extracted from the samples before and after magnetic separation using a QIAamp Viral RNA Mini kit. Reverse transcription was then performed, and the resultant cDNAs were analyzed by polymerase chain reaction using forward and reverse primers for the DENV genomic RNA. The sample obtained following magnetic separation was the same as that referred to as the bead fraction. * P
    Figure Legend Snippet: Concentrated genomic RNA of DENV following adsorption onto antibody-integrated magnetic beads. The culture medium (10 µ l) of the C6/36 cells infected with DENV1 was diluted with 1 ml phosphate-buffered saline, mixed with antibody-integrated magnetic beads and subjected to magnetic separation. Viral RNA was extracted from the samples before and after magnetic separation using a QIAamp Viral RNA Mini kit. Reverse transcription was then performed, and the resultant cDNAs were analyzed by polymerase chain reaction using forward and reverse primers for the DENV genomic RNA. The sample obtained following magnetic separation was the same as that referred to as the bead fraction. * P

    Techniques Used: Adsorption, Magnetic Beads, Infection, Polymerase Chain Reaction

    64) Product Images from "A Single Mutation in Chikungunya Virus Affects Vector Specificity and Epidemic Potential"

    Article Title: A Single Mutation in Chikungunya Virus Affects Vector Specificity and Epidemic Potential

    Journal: PLoS Pathogens

    doi: 10.1371/journal.ppat.0030201

    Effect of E1-A226V Mutation on CHIKV Transmission by Ae. albopictus and Ae. aegypti Mosquitoes (A) Six 2- to 3-day-old suckling mice (Swiss Webster) were subcutaneously infected with a 20-μl mixture of ≈ 25 pfu LR-Apa-226V and ≈ 25 pfu of LR-226A viruses. (B and C) Ae. aegypti and Ae. albopictus mosquitoes were presented with a blood meal containing 10 7 pfu/ml of LR-Apa-226V and 10 7 pfu/ml of LR-226A viruses. At 13 dpi, ten to 15 mosquitoes were placed in separate paper cartons and starved for 24 h. The next day, the mosquitoes in each carton were presented with a 2- to 3-day-old suckling mouse (Swiss Webster). Mice were returned to their cage and sacrificed on day 3 post-exposure. Blood from each individual mouse (≈ 50 μl) was collected and immediately mixed with 450 μl of TRIzol reagent for RNA extraction. BM and inoc. - initial ratio of LR-ApaI-226V and LR-226A in blood meal samples and inoculum for subcutaneous infection. 1–6 ratio of LR-ApaI-226V and LR-226A RNA in six individual mice.
    Figure Legend Snippet: Effect of E1-A226V Mutation on CHIKV Transmission by Ae. albopictus and Ae. aegypti Mosquitoes (A) Six 2- to 3-day-old suckling mice (Swiss Webster) were subcutaneously infected with a 20-μl mixture of ≈ 25 pfu LR-Apa-226V and ≈ 25 pfu of LR-226A viruses. (B and C) Ae. aegypti and Ae. albopictus mosquitoes were presented with a blood meal containing 10 7 pfu/ml of LR-Apa-226V and 10 7 pfu/ml of LR-226A viruses. At 13 dpi, ten to 15 mosquitoes were placed in separate paper cartons and starved for 24 h. The next day, the mosquitoes in each carton were presented with a 2- to 3-day-old suckling mouse (Swiss Webster). Mice were returned to their cage and sacrificed on day 3 post-exposure. Blood from each individual mouse (≈ 50 μl) was collected and immediately mixed with 450 μl of TRIzol reagent for RNA extraction. BM and inoc. - initial ratio of LR-ApaI-226V and LR-226A in blood meal samples and inoculum for subcutaneous infection. 1–6 ratio of LR-ApaI-226V and LR-226A RNA in six individual mice.

    Techniques Used: Mutagenesis, Transmission Assay, Mouse Assay, Infection, RNA Extraction

    65) Product Images from "Regulation of Viral RNA Synthesis by the V Protein of Parainfluenza Virus 5"

    Article Title: Regulation of Viral RNA Synthesis by the V Protein of Parainfluenza Virus 5

    Journal: Journal of Virology

    doi: 10.1128/JVI.01832-15

    Effects of V mutants on viral RNA synthesis. Truncated V proteins inhibited PIV5 RNA replication. BSR T7 cells were transfected as described in Materials and Methods. Encapsidated PIV5 minigenome RNA was purified with NP antibody, and a primer that binds to antigenomic RNA in the trailer region was added to the reverse transcription reaction mixture. Then, the same amount of RNA from the reverse transcription reaction was added to the PCR mixture as the template, and a pair of primers that anneal to the C terminus of Renilla luciferase and the PIV5 trailer region, respectively, was also added to the PCR mixture. The expected PCR products are indicated by MG and were approximately 250 bp. DNA ladder, a 100-bp DNA size marker (the bands of the ladder at 200 bp are indicated). (A) Detection of minigenome RNA using RT-PCR. Purified RNA without RT was used in the PCR mixture to demonstrate the lack of plasmid DNA contamination, and the corresponding RNA with RT was used in the PCR mixture. All samples were amplified for 30 cycles. (B) Detection of minigenome RNA using RT-PCR and different numbers of PCR cycles. Purified RNAs were reverse transcribed and then amplified for 15, 20, 25, and 30 cycles. No MG, cells transfected with all minigenome components except the plasmid carrying the minigenome; Pos Control, a minigenome system without plasmid carrying V or the V mutants; instead, the same expression plasmid containing enhanced GFP was used to maintain the total amount of transfected plasmid DNA constant. This gel is representative of those from three experiments. (C) Analysis of PCR products. The intensity of the PCR products was quantified using ImageQuant software. The intensity of cells transfected with all minigenome components except the plasmid carrying the minigenome was set equal to 0 in PCRs with 25 and 30 cycles; the intensity of the positive control was set to 100 in PCRs with 25 and 30 cycles; the relative percentage of the V, V120, or V21-222 PCR band intensity is the ratio of the V, V120, or V21-222 band intensity to that of the positive control in 25 and 30 cycles. The average was calculated from three independent experiments. Error bars represent standards deviation of the means. *, P
    Figure Legend Snippet: Effects of V mutants on viral RNA synthesis. Truncated V proteins inhibited PIV5 RNA replication. BSR T7 cells were transfected as described in Materials and Methods. Encapsidated PIV5 minigenome RNA was purified with NP antibody, and a primer that binds to antigenomic RNA in the trailer region was added to the reverse transcription reaction mixture. Then, the same amount of RNA from the reverse transcription reaction was added to the PCR mixture as the template, and a pair of primers that anneal to the C terminus of Renilla luciferase and the PIV5 trailer region, respectively, was also added to the PCR mixture. The expected PCR products are indicated by MG and were approximately 250 bp. DNA ladder, a 100-bp DNA size marker (the bands of the ladder at 200 bp are indicated). (A) Detection of minigenome RNA using RT-PCR. Purified RNA without RT was used in the PCR mixture to demonstrate the lack of plasmid DNA contamination, and the corresponding RNA with RT was used in the PCR mixture. All samples were amplified for 30 cycles. (B) Detection of minigenome RNA using RT-PCR and different numbers of PCR cycles. Purified RNAs were reverse transcribed and then amplified for 15, 20, 25, and 30 cycles. No MG, cells transfected with all minigenome components except the plasmid carrying the minigenome; Pos Control, a minigenome system without plasmid carrying V or the V mutants; instead, the same expression plasmid containing enhanced GFP was used to maintain the total amount of transfected plasmid DNA constant. This gel is representative of those from three experiments. (C) Analysis of PCR products. The intensity of the PCR products was quantified using ImageQuant software. The intensity of cells transfected with all minigenome components except the plasmid carrying the minigenome was set equal to 0 in PCRs with 25 and 30 cycles; the intensity of the positive control was set to 100 in PCRs with 25 and 30 cycles; the relative percentage of the V, V120, or V21-222 PCR band intensity is the ratio of the V, V120, or V21-222 band intensity to that of the positive control in 25 and 30 cycles. The average was calculated from three independent experiments. Error bars represent standards deviation of the means. *, P

    Techniques Used: Transfection, Purification, Polymerase Chain Reaction, Luciferase, Marker, Reverse Transcription Polymerase Chain Reaction, Plasmid Preparation, Amplification, Expressing, Software, Positive Control

    66) Product Images from "Coordinated Implementation of Chikungunya Virus Reverse Transcription-PCR"

    Article Title: Coordinated Implementation of Chikungunya Virus Reverse Transcription-PCR

    Journal: Emerging Infectious Diseases

    doi: 10.3201/eid1503.081104

    Probit analysis of laboratories with a positive result (y axes) for chikungunya virus in relation to viral RNA concentration in positive samples (x axes). A) Laboratories using in-house reverse transcription–PCRs (RT-PCRs) (n = 18) had a 50% certainty of having a positive result at 10,000 RNA copies/mL (95% confidence interval [CI] 3,162–19,952). B) Laboratories using a preformulated RT-PCR (n = 13) had a 50% certainty of having a positive result at 1,288 RNA copies/mL (95% CI 416–2,344). Data points represent individual samples in the test panel. Thick line is the regression line calculated on the basis of a probit model (dose-response curve), and thin lines are 95% CIs. Data fit into the model with p
    Figure Legend Snippet: Probit analysis of laboratories with a positive result (y axes) for chikungunya virus in relation to viral RNA concentration in positive samples (x axes). A) Laboratories using in-house reverse transcription–PCRs (RT-PCRs) (n = 18) had a 50% certainty of having a positive result at 10,000 RNA copies/mL (95% confidence interval [CI] 3,162–19,952). B) Laboratories using a preformulated RT-PCR (n = 13) had a 50% certainty of having a positive result at 1,288 RNA copies/mL (95% CI 416–2,344). Data points represent individual samples in the test panel. Thick line is the regression line calculated on the basis of a probit model (dose-response curve), and thin lines are 95% CIs. Data fit into the model with p

    Techniques Used: Concentration Assay, Reverse Transcription Polymerase Chain Reaction

    67) Product Images from "Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿"

    Article Title: Lack of Bax Prevents Influenza A Virus-Induced Apoptosis and Causes Diminished Viral Replication ▿

    Journal: Journal of Virology

    doi: 10.1128/JVI.02672-08

    Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.
    Figure Legend Snippet: Influenza A virus replication is dependent upon opposite virus-induced effects on Bax and Bak activity that are unlikely to be interferon related. (A) Influenza A virus replication was analyzed by plaque assay. Virus replication is severely attenuated in Bax KO cells, resulting in a 2-log decrease in PFU/ml compared to the WT. Bak KO cells allow a maximum replication similar to that of the WT, while Bax/Bak DKO cells show a slight elevation of infectious titers during infection. These results indicate that Bax is proviral during infection, while Bak is dispensable for replication. (B) Bax was transiently expressed in all cell types by Lipofectamine 2000 transfection of a C2-Bax-GFP construct prior to infection, and supernatant samples were collected for plaque assay at 48 hpi. Baseline virus replication in each cell type was evaluated using empty C2-GFP plasmid transfection. Bax reconstitution in Bax KO cells resulted in a fivefold increase in infectious titers compared to the control ( P = 0.0007). A minimal effect on the virus titer was seen after Bax overexpression by transient transfection in WT cells compared to empty plasmid controls. (C) Influenza A virus replication was assessed by reverse transcription-PCR. Serial dilutions of stock virus at known concentrations were also analyzed to generate a standard curve to which experimental samples were compared, thus calculating the approximate number of influenza A virus particles/ml in each sample. By 24 hpi, Bax KO, Bak KO, and Bax/Bak DKO cells all showed significantly higher levels of influenza A virus RNA released into the cell culture supernatant than did WT cells. (D) Interferon activity was assessed by infecting mock- and influenza A virus-infected cells with interferon-sensitive, GFP-linked NDV and quantifying the mean GFP expression levels of 10,000 events per condition by FACS analysis. Each assay was run in triplicate, and data are expressed as the ratio of the numbers of influenza A virus-infected to mock-infected cells per cell type. After influenza A virus infection, Bak KO cells exhibited a slight decrease in ratio compared to the WT, representing a 30% increase in interferon activity ( P = 0.002). Bax KO and Bax/Bak DKO cells both showed similar fluorescence changes compared to the WT after infection. Due the high degree of similarity between cell types, these results suggest that the interferon response in infected cells is modulated by viral replication in the presence of Bak and is only slightly modified by Bax activity during influenza A virus infection. As an elevated interferon response typically leads to a reduced virus replication capacity, these results also suggest that it is unlikely that the observed trends in infectious virus titer are due to virus-induced interferon signaling.

    Techniques Used: Activity Assay, Plaque Assay, Infection, Transfection, Construct, Plasmid Preparation, Over Expression, Polymerase Chain Reaction, Cell Culture, Expressing, FACS, Fluorescence, Modification

    68) Product Images from "Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR"

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Figure Legend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Techniques Used: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).
    Figure Legend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Techniques Used: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    69) Product Images from "Capture of dengue viruses using antibody-integrated graphite-encapsulated magnetic beads produced using gas plasma technology"

    Article Title: Capture of dengue viruses using antibody-integrated graphite-encapsulated magnetic beads produced using gas plasma technology

    Journal: Molecular Medicine Reports

    doi: 10.3892/mmr.2016.5330

    Detection of genomic RNA in DENVs adsorbed onto antibody-integrated magnetic beads. Culture media containing uninfected (Mock) C6/36 cells or C6/36 cells infected with DENV1-4 were mixed with antibody-integrated magnetic beads and separated into BD and SP fractions. Viral RNA was extracted from each fraction using a QIAamp Viral RNA Mini kit and analyzed using RT-PCR analysis. The PCR primer pairs specifically annealed to a common 511 bp fragment among the DENV1-4 serotypes 1–4. Each PCR product was analyzed using agarose gel electrophoresis, and the amplified products were then sequenced to confirm they corresponded to the gene of the DENVs. A 100 bp ladder marker (M) is shown. Numbers indicated on the left are molecular size (bp). DENV, dengue virus; BD, bead fraction; SP, supernatant fraction; RT-PCR, reverse transcription-polymerase chain reaction.
    Figure Legend Snippet: Detection of genomic RNA in DENVs adsorbed onto antibody-integrated magnetic beads. Culture media containing uninfected (Mock) C6/36 cells or C6/36 cells infected with DENV1-4 were mixed with antibody-integrated magnetic beads and separated into BD and SP fractions. Viral RNA was extracted from each fraction using a QIAamp Viral RNA Mini kit and analyzed using RT-PCR analysis. The PCR primer pairs specifically annealed to a common 511 bp fragment among the DENV1-4 serotypes 1–4. Each PCR product was analyzed using agarose gel electrophoresis, and the amplified products were then sequenced to confirm they corresponded to the gene of the DENVs. A 100 bp ladder marker (M) is shown. Numbers indicated on the left are molecular size (bp). DENV, dengue virus; BD, bead fraction; SP, supernatant fraction; RT-PCR, reverse transcription-polymerase chain reaction.

    Techniques Used: Magnetic Beads, Infection, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Amplification, Marker

    Concentrated genomic RNA of DENV following adsorption onto antibody-integrated magnetic beads. The culture medium (10 µ l) of the C6/36 cells infected with DENV1 was diluted with 1 ml phosphate-buffered saline, mixed with antibody-integrated magnetic beads and subjected to magnetic separation. Viral RNA was extracted from the samples before and after magnetic separation using a QIAamp Viral RNA Mini kit. Reverse transcription was then performed, and the resultant cDNAs were analyzed by polymerase chain reaction using forward and reverse primers for the DENV genomic RNA. The sample obtained following magnetic separation was the same as that referred to as the bead fraction. * P
    Figure Legend Snippet: Concentrated genomic RNA of DENV following adsorption onto antibody-integrated magnetic beads. The culture medium (10 µ l) of the C6/36 cells infected with DENV1 was diluted with 1 ml phosphate-buffered saline, mixed with antibody-integrated magnetic beads and subjected to magnetic separation. Viral RNA was extracted from the samples before and after magnetic separation using a QIAamp Viral RNA Mini kit. Reverse transcription was then performed, and the resultant cDNAs were analyzed by polymerase chain reaction using forward and reverse primers for the DENV genomic RNA. The sample obtained following magnetic separation was the same as that referred to as the bead fraction. * P

    Techniques Used: Adsorption, Magnetic Beads, Infection, Polymerase Chain Reaction

    70) Product Images from "Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology"

    Article Title: Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S191784

    Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.
    Figure Legend Snippet: Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Real-time Polymerase Chain Reaction

    Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.
    Figure Legend Snippet: Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Amplification, Polymerase Chain Reaction, Agarose Gel Electrophoresis, DNA Sequencing, Marker, Reverse Transcription Polymerase Chain Reaction, Negative Control

    71) Product Images from "Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology"

    Article Title: Efficient recovery and enrichment of infectious rotavirus using separation with antibody-integrated graphite-encapsulated magnetic nanobeads produced by argon/ammonia gas plasma technology

    Journal: International Journal of Nanomedicine

    doi: 10.2147/IJN.S191784

    Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.
    Figure Legend Snippet: Quantitative analysis of the viral gene of rotavirus adsorbed onto the antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to RT-reaction. The resultant cDNA was analyzed by real-time PCR using primers for the rotavirus VP7 gene as described in Materials and methods. The value of the TL sample was taken as 100%. Abbreviations: MNBs, magnetic nanobeads; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Real-time Polymerase Chain Reaction

    Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.
    Figure Legend Snippet: Detection of viral RNA of rotavirus adsorbed onto antibody-integrated MNBs. Notes: Rotavirus-infected cell lysate (10 µL) was diluted with PBS (500 µL) and then incubated with antibody-integrated magnetic beads. After incubation, the following fractions were obtained: 1) diluted rotavirus sample before incubation with the beads (BF), 2) bead fraction after incubation with anti-rotavirus antibody-integrated MNBs (RV-BD), 3) bead fraction after incubation with anti-dengue virus antibody-integrated MNBs (DV-BD), 4) supernatant fraction after incubation with the anti-rotavirus antibody-integrated MNBs (RV-SP), 5) supernatant fraction after incubation with the anti-dengue virus antibody-integrated MNBs (DV-SP), and 6) total sample containing the same quantity of rotavirus as in 10 µL of rotavirus-infected cell lysate (total fraction, TL). Viral genomic RNA was subsequently extracted from the above fractions using a QIAamp Viral RNA mini kit and subjected to a RT-reaction. Rotavirus viral protein 7 (VP7) gene (552 bp) in the cDNA was amplified by PCR as described in Materials and methods. PCR products were analyzed by agarose gel electrophoresis (1.2% gel). The identity of the amplified products was confirmed by DNA sequencing. The left-hand lane is size marker (M), which includes DNA of 100, 200, 300, 400, 500, 600, 700, 800, 900, 1,000, 1,200, and 1,500 bp. The position of the 552 bp band for VP7 is indicated by an arrow. The NC comprised a water sample (no rotavirus) that was subjected to RT-PCR. Abbreviations: MNBs, magnetic nanobeads; NC, negative control; RT, reverse transcription.

    Techniques Used: Infection, Incubation, Magnetic Beads, Amplification, Polymerase Chain Reaction, Agarose Gel Electrophoresis, DNA Sequencing, Marker, Reverse Transcription Polymerase Chain Reaction, Negative Control

    72) Product Images from "Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿"

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.02255-08

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)
    Figure Legend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Techniques Used: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of
    Figure Legend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Techniques Used: Sample Prep

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    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer. .. The target region on the HA gene, from nucleotide 351 to 1735, was PCR amplified with the Phusion High-Fidelity PCR kit (New England BioLabs), purified via a QIAquick Gel Extraction kit (Qiagen), and cloned into the pGEM-T Easy vector (Promega).

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: Paragraph title: Comparison of different RNA extraction methods for RNA quantification and PCR amplification of HIV-1 for a panel of patient samples. ... However, we kept the QIAamp viral RNA mini kit (Qiagen) because the Biocentric viral load kit recommends this extraction method.

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR
    Article Snippet: .. One hundred ninety-two specimens tested DNA- or RNA-positive for the respective pathogen (for HBV, 100 serum samples; for CMV, 8 urine samples, 52 serum samples, and 15 plasma samples; for HGV, 17 serum samples) by virus-specific quantitative PCR (HBV [Cobas Amplicor; Roche Diagnostics, Basel, Switzerland] and CMV) ( ) or qualitative PCR (HGV) in our routine diagnostic laboratory following extraction with the QIAamp DNA Blood Mini kit or the QIAamp Viral RNA Mini kit (QIAGEN, Hilden, Germany). .. Extraction capacity was evaluated for HBV with a dilution series of the EUROHEP standard (4 × 105 to 1 × 101 copies per ml of serum), for CMV with a dilution series of the standard plasmid pCR-gpB diluted in serum (2 × 105 to 2 × 102 copies per ml of serum), and for HGV with a dilution series of viral RNA from serum samples from a highly viremic patient (dilutions of 1−6 to 10−6 ).

    Viral-load Assay:

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: The QIAamp viral RNA minikit (Qiagen, Courtaboeuf, France) combines a silica gel-based membrane with the speed of microspin centrifugation. .. The Nuclisens manual extraction kit (bioMérieux, Craponne, France) is based on the use of silica particles as described by Boom et al. ( ); this technology is used for the Nuclisens EasyQ HIV-1 commercial viral load assay.

    Magnetic Beads:

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR
    Article Snippet: A recent development is the coverage of magnetic beads with nucleic acid-binding matrices, which promises a high potential for automation ( ). .. One hundred ninety-two specimens tested DNA- or RNA-positive for the respective pathogen (for HBV, 100 serum samples; for CMV, 8 urine samples, 52 serum samples, and 15 plasma samples; for HGV, 17 serum samples) by virus-specific quantitative PCR (HBV [Cobas Amplicor; Roche Diagnostics, Basel, Switzerland] and CMV) ( ) or qualitative PCR (HGV) in our routine diagnostic laboratory following extraction with the QIAamp DNA Blood Mini kit or the QIAamp Viral RNA Mini kit (QIAGEN, Hilden, Germany).

    Isolation:

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens
    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer. .. Plasmids were isolated with a Plasmid Midi Prep Kit (Qiagen), blunt-cut linearized, and served as in vitro transcription DNA templates using a Ribomax Transcription kit (Promega).

    RNA Extraction:

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: Paragraph title: Viral RNA extraction. ... The QIAamp viral RNA minikit (Qiagen, Courtaboeuf, France) combines a silica gel-based membrane with the speed of microspin centrifugation.

    Purification:

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens
    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer. .. The target region on the HA gene, from nucleotide 351 to 1735, was PCR amplified with the Phusion High-Fidelity PCR kit (New England BioLabs), purified via a QIAquick Gel Extraction kit (Qiagen), and cloned into the pGEM-T Easy vector (Promega).

    Sequencing:

    Article Title: Multiple Layers of Chimerism in a Single-Stranded DNA Virus Discovered by Deep Sequencing
    Article Snippet: Paragraph title: CHIV14 Discovered by Deep Sequencing Is Traced Back to Spin-Column Contamination ... Definitive confirmation of the origin of CHIV14 was obtained by in-depth analyses of water that was passed through contaminated spin columns ( B ): The test for the presence of CHIV14 by PCR was positive for the non-A–E hepatitis sera, healthy sera control, and mock (water) extractions obtained using QIAamp Viral RNA Mini kit (Qiagen), but was negative for samples extracted using UCP spin columns (Qiagen).

    Gel Extraction:

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens
    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer. .. The target region on the HA gene, from nucleotide 351 to 1735, was PCR amplified with the Phusion High-Fidelity PCR kit (New England BioLabs), purified via a QIAquick Gel Extraction kit (Qiagen), and cloned into the pGEM-T Easy vector (Promega).

    Plasmid Preparation:

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens
    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer. .. The target region on the HA gene, from nucleotide 351 to 1735, was PCR amplified with the Phusion High-Fidelity PCR kit (New England BioLabs), purified via a QIAquick Gel Extraction kit (Qiagen), and cloned into the pGEM-T Easy vector (Promega).

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR
    Article Snippet: One hundred ninety-two specimens tested DNA- or RNA-positive for the respective pathogen (for HBV, 100 serum samples; for CMV, 8 urine samples, 52 serum samples, and 15 plasma samples; for HGV, 17 serum samples) by virus-specific quantitative PCR (HBV [Cobas Amplicor; Roche Diagnostics, Basel, Switzerland] and CMV) ( ) or qualitative PCR (HGV) in our routine diagnostic laboratory following extraction with the QIAamp DNA Blood Mini kit or the QIAamp Viral RNA Mini kit (QIAGEN, Hilden, Germany). .. Extraction capacity was evaluated for HBV with a dilution series of the EUROHEP standard (4 × 105 to 1 × 101 copies per ml of serum), for CMV with a dilution series of the standard plasmid pCR-gpB diluted in serum (2 × 105 to 2 × 102 copies per ml of serum), and for HGV with a dilution series of viral RNA from serum samples from a highly viremic patient (dilutions of 1−6 to 10−6 ).

    Negative Control:

    Article Title: Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings
    Article Snippet: A negative control consisted of 140 μL of HIV-negative, pooled unspiked plasma. .. Note: according to the QIAamp® Viral RNA Mini Kit Handbook, this Kit can also be performed using the QIAvac® 24 Plus (Qiagen, Valencia, CA) if a centrifuge is not available.

    Sample Prep:

    Article Title: Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings
    Article Snippet: Paragraph title: Sample preparation ... Note: according to the QIAamp® Viral RNA Mini Kit Handbook, this Kit can also be performed using the QIAvac® 24 Plus (Qiagen, Valencia, CA) if a centrifuge is not available.

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: The QIAamp viral RNA minikit (Qiagen, Courtaboeuf, France) combines a silica gel-based membrane with the speed of microspin centrifugation. .. The Abbott sample preparation system (Abbott Molecular, Rungis, France) is an iron particle-based method used for the Abbott RealTime HIV-1 commercial assay for viral load determination.

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: In order to confirm to what extent HIV-1 RNA was correctly recovered and quantified from DPS after extractions with the Abbott sample preparation system and the Nuclisens manual extraction kit (bioMérieux), we decided to extend our evaluation to real patient samples and used unlinked leftover samples from 47 patients followed at an HIV clinic of the University Hospital of Montpellier. .. However, we kept the QIAamp viral RNA mini kit (Qiagen) because the Biocentric viral load kit recommends this extraction method.

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: .. Elution from spots was performed by cutting spots into two to four pieces that were subsequently incubated with the kit lysis buffer, using 2 ml for QIAamp viral RNA minikit (Qiagen) and High Pure viral nucleic acid kit (Roche) extractions, 3 ml for Abbott sample preparation system extraction, and 9 ml for Nuclisens manual extraction kit (bioMérieux) extraction. .. After 2 h of incubation at room temperature under gentle agitation, the supernatants were clarified by centrifugation at 1,500 × g for 2 min. RNAs were then extracted according to the instructions of the corresponding extraction method and eluted in 60 μl of elution buffer.

    In Vitro:

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens
    Article Snippet: Paragraph title: H1N1 in Vitro Transcribed RNA Standards ... Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer.

    Ethanol Precipitation:

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens
    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer. .. The RNA transcripts were then purified via DNase digestion, acid phenol-chloroform extraction and ethanol precipitation.

    Lysis:

    Article Title: Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings
    Article Snippet: Extractions were performed by adding 560 μL of Viral Lysis Buffer (Qiagen, Valencia CA) with carrier RNA in a 1.5-mL microcentrifuge tube, adding 140 μL of human plasma, and incubated at room temperature for 10 min. After the incubation 560 μL of 100% ethanol was added to the mixture, and the samples were applied to QIAamp spin columns. .. Note: according to the QIAamp® Viral RNA Mini Kit Handbook, this Kit can also be performed using the QIAvac® 24 Plus (Qiagen, Valencia, CA) if a centrifuge is not available.

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿
    Article Snippet: .. Elution from spots was performed by cutting spots into two to four pieces that were subsequently incubated with the kit lysis buffer, using 2 ml for QIAamp viral RNA minikit (Qiagen) and High Pure viral nucleic acid kit (Roche) extractions, 3 ml for Abbott sample preparation system extraction, and 9 ml for Nuclisens manual extraction kit (bioMérieux) extraction. .. After 2 h of incubation at room temperature under gentle agitation, the supernatants were clarified by centrifugation at 1,500 × g for 2 min. RNAs were then extracted according to the instructions of the corresponding extraction method and eluted in 60 μl of elution buffer.

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    Qiagen qiaamp viral rna mini kit
    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic <t>DNA/RNA</t> kit and the <t>QIAamp</t> DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .
    Qiaamp Viral Rna Mini Kit, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 1490 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Journal: Journal of Clinical Microbiology

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Figure Lengend Snippet: Amplification of a serial dilution of the EUROHEP standard with HBV-specific primers in a nested PCR following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. (A) Chemagen extracts. (B) QIAGEN extracts. Lane M shows a 100-bp ladder, and lane C shows results for the negative control. Lanes 1 through 10 show results for different viral loads expressed in numbers of copies per ml: 1, 4 × 10 5 ; 2, 4 × 10 4 ; 3, 4 × 10 3 ; 4, 4 × 10 2 ; 5, 2 × 10 2 ; 6, 1 × 10 2 ; 7, 8 × 10 1 ; 8, 4 × 10 1 ; 9, 2 × 10 1 ; 10, 1 × 10 1 .

    Article Snippet: Identical results were obtained after extraction of the identical specimens with the QIAamp Viral RNA Mini kit.

    Techniques: Amplification, Serial Dilution, Nested PCR, Negative Control

    Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Journal: Journal of Clinical Microbiology

    Article Title: Efficient Extraction of Viral DNA and Viral RNA by the Chemagic Viral DNA/RNA Kit Allows Sensitive Detection of Cytomegalovirus, Hepatitis B Virus, and Hepatitis G Virus by PCR

    doi: 10.1128/JCM.41.11.5273-5276.2003

    Figure Lengend Snippet: Amplification of a serial dilution of the standard plasmid pCR-gpB by real-time PCR with CMV-specific primers (amplicon, 254 bp) on the LightCycler instrument (software, version 3.5; analysis method, fit points with manual noise band adjustment, hybridization probe format) following extraction with the Chemagic DNA/RNA kit and the QIAamp DNA Blood Mini kit. pCR-gpB concentrations (from left to right) were 2 × 10 5 , 2 × 10 4 , 2 × 10 3 , and 2 × 10 2 copies per ml of serum. Continuous line, Chemagen extracts; broken line, QIAGEN extracts. Cycle number, cycle number of the amplification reaction; F2/F1, quotient of fluorescence channel F2 (hybridization probe) and fluorescence channel F1 (fluorescein).

    Article Snippet: Identical results were obtained after extraction of the identical specimens with the QIAamp Viral RNA Mini kit.

    Techniques: Amplification, Serial Dilution, Plasmid Preparation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Software, Hybridization, Fluorescence

    Integrated assays using plasma spiked with Armored RNA HIV. Armored RNA HIV was spiked into human plasma at concentrations ranging from 10,000 copies/mL to 1,000 copies/mL amd extracted with a QIAamp® Viral RNA Mini Kit according to the manufacturer’s

    Journal: The Journal of infectious diseases

    Article Title: Nucleic acid assay system for tier II labs and moderately complex clinics to detect HIV in low-resource settings

    doi: 10.1086/650388

    Figure Lengend Snippet: Integrated assays using plasma spiked with Armored RNA HIV. Armored RNA HIV was spiked into human plasma at concentrations ranging from 10,000 copies/mL to 1,000 copies/mL amd extracted with a QIAamp® Viral RNA Mini Kit according to the manufacturer’s

    Article Snippet: Note: according to the QIAamp® Viral RNA Mini Kit Handbook, this Kit can also be performed using the QIAvac® 24 Plus (Qiagen, Valencia, CA) if a centrifuge is not available.

    Techniques:

    Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Journal: Journal of Clinical Microbiology

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    doi: 10.1128/JCM.02255-08

    Figure Lengend Snippet: Bland-Altman analysis of HIV-1 viral loads in patient samples ( n = 47) of plasma versus DPS after RNA extraction with a QIAamp viral RNA mini kit (Qiagen) (A), the Abbott sample preparation system (B), and a Nuclisens manual extraction kit (bioMérieux)

    Article Snippet: The QIAamp viral RNA minikit (Qiagen, Courtaboeuf, France) combines a silica gel-based membrane with the speed of microspin centrifugation.

    Techniques: RNA Extraction, Sample Prep

    HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Journal: Journal of Clinical Microbiology

    Article Title: Evaluation of Different RNA Extraction Methods and Storage Conditions of Dried Plasma or Blood Spots for Human Immunodeficiency Virus Type 1 RNA Quantification and PCR Amplification for Drug Resistance Testing ▿

    doi: 10.1128/JCM.02255-08

    Figure Lengend Snippet: HIV-1 viral loads in patient samples ( n = 47) of plasma and DPS after RNA extractions with a QIAamp viral RNA mini kit (Qiagen), the Abbott sample preparation system, or a Nuclisens manual extraction kit (bioMérieux). (A) Comparison of

    Article Snippet: The QIAamp viral RNA minikit (Qiagen, Courtaboeuf, France) combines a silica gel-based membrane with the speed of microspin centrifugation.

    Techniques: Sample Prep

    Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).

    Journal: Analytical chemistry

    Article Title: Paper-Based RNA Extraction, in Situ Isothermal Amplification, and Lateral Flow Detection for Low-Cost, Rapid Diagnosis of Influenza A (H1N1) from Clinical Specimens

    doi: 10.1021/acs.analchem.5b01594

    Figure Lengend Snippet: Clinical nasopharyngeal specimens. (a) Paper extractions and QIAamp kit extractions of clinical specimens A–L. (b) RT-LAMP assay performed in solution with Qiagen-extracted purified RNA from clinical specimens A–L, and gel electrophoresis of products. (c) Lateral flow detection of amplified products; test line intensities plotted as a percentage of control line intensities. (d) Paper extraction of clinical specimens A–L followed by in situ RT-LAMP and lateral flow detection. + = positive control (10 9 cp/mL RNA standard); − = negative control (no RNA).

    Article Snippet: Genomic RNA from the patient sample was extracted via the QIAamp Viral RNA Mini Kit (Qiagen) and reverse-transcribed with the Superscript III First-Strand Synthesis Kit (Invitrogen) with a gene-specific reverse primer.

    Techniques: RT Lamp Assay, Purification, Nucleic Acid Electrophoresis, Flow Cytometry, Amplification, In Situ, Positive Control, Negative Control