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BEI Resources vero cells
In vivo rescue of rZIKV from <t>Vero</t> cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by <t>immunofocus</t> assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.
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1) Product Images from "In vivo rescue of recombinant Zika virus from an infectious cDNA clone and its implications in vaccine development"

Article Title: In vivo rescue of recombinant Zika virus from an infectious cDNA clone and its implications in vaccine development

Journal: Scientific Reports

doi: 10.1038/s41598-020-57545-2

In vivo rescue of rZIKV from Vero cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by immunofocus assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.
Figure Legend Snippet: In vivo rescue of rZIKV from Vero cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by immunofocus assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.

Techniques Used: In Vivo, Transfection, BAC Assay, Mouse Assay

2) Product Images from "In vivo rescue of recombinant Zika virus from an infectious cDNA clone and its implications in vaccine development"

Article Title: In vivo rescue of recombinant Zika virus from an infectious cDNA clone and its implications in vaccine development

Journal: Scientific Reports

doi: 10.1038/s41598-020-57545-2

In vivo rescue of rZIKV from Vero cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by immunofocus assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.
Figure Legend Snippet: In vivo rescue of rZIKV from Vero cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by immunofocus assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.

Techniques Used: In Vivo, Transfection, BAC Assay, Mouse Assay

3) Product Images from "Polyamine Depletion Inhibits Bunyavirus Infection via Generation of Noninfectious Interfering Virions"

Article Title: Polyamine Depletion Inhibits Bunyavirus Infection via Generation of Noninfectious Interfering Virions

Journal: Journal of Virology

doi: 10.1128/JVI.00530-19

Polyamines enhance specific infectivity of enveloped viruses. (A to C) Huh7 cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with ZIKV (strain MR766) at an MOI of 0.1 PFU per cell. At 48 hpi, viral titers were determined via plaque assay (A) and relative viral genomes were determined by qRT-PCR (B), and the ratio of genomes to PFU was calculated (C). (D to F) Vero cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with CVB3 (Nancy strain) at an MOI of 0.1 PFU per cell. At 24 hpi, viral titers were determined via plaque assay (D) and relative viral genomes were determined by qRT-PCR (E), and the ratio of genomes to PFU was calculated (F). (G to I) Identical infection and analysis methods were performed for KEYV in Huh7 cells to determine viral titers (G), viral genomes (H), and genome-to-PFU ratio (I) at 48 hpi. (J to L) These methods were also used for VSV (strain Indiana) infected at an MOI of 0.01 PFU per cell for 24 h. Viral titers (J), genomes (K), and genome-to-PFU ratio (L) were calculated. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001 (using Student’s t test [ n ≥ 3]). Error bars represent ±1 SEM. Statistical comparisons were performed between treated and untreated conditions.
Figure Legend Snippet: Polyamines enhance specific infectivity of enveloped viruses. (A to C) Huh7 cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with ZIKV (strain MR766) at an MOI of 0.1 PFU per cell. At 48 hpi, viral titers were determined via plaque assay (A) and relative viral genomes were determined by qRT-PCR (B), and the ratio of genomes to PFU was calculated (C). (D to F) Vero cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with CVB3 (Nancy strain) at an MOI of 0.1 PFU per cell. At 24 hpi, viral titers were determined via plaque assay (D) and relative viral genomes were determined by qRT-PCR (E), and the ratio of genomes to PFU was calculated (F). (G to I) Identical infection and analysis methods were performed for KEYV in Huh7 cells to determine viral titers (G), viral genomes (H), and genome-to-PFU ratio (I) at 48 hpi. (J to L) These methods were also used for VSV (strain Indiana) infected at an MOI of 0.01 PFU per cell for 24 h. Viral titers (J), genomes (K), and genome-to-PFU ratio (L) were calculated. *, P ≤ 0.05; **, P ≤ 0.01; ***, P ≤ 0.001 (using Student’s t test [ n ≥ 3]). Error bars represent ±1 SEM. Statistical comparisons were performed between treated and untreated conditions.

Techniques Used: Infection, Plaque Assay, Quantitative RT-PCR

4) Product Images from "Polyamine Depletion Inhibits Bunyavirus Infection via Generation of Noninfectious Interfering Virions"

Article Title: Polyamine Depletion Inhibits Bunyavirus Infection via Generation of Noninfectious Interfering Virions

Journal: Journal of Virology

doi: 10.1128/JVI.00530-19

Polyamines enhance specific infectivity of enveloped viruses. (A to C) Huh7 cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with ZIKV (strain MR766) at an MOI of 0.1 PFU per cell. At 48 hpi, viral titers were determined via plaque assay (A) and relative viral genomes were determined by qRT-PCR (B), and the ratio of genomes to PFU was calculated (C). (D to F) Vero cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with CVB3 (Nancy strain) at an MOI of 0.1 PFU per cell. At 24 hpi, viral titers were determined via plaque assay (D) and relative viral genomes were determined by qRT-PCR (E), and the ratio of genomes to PFU was calculated (F). (G to I) Identical infection and analysis methods were performed for KEYV in Huh7 cells to determine viral titers (G), viral genomes (H), and genome-to-PFU ratio (I) at 48 hpi. (J to L) These methods were also used for VSV (strain Indiana) infected at an MOI of 0.01 PFU per cell for 24 h. Viral titers (J), genomes (K), and genome-to-PFU ratio (L) were calculated. *, P  ≤ 0.05; **, P  ≤ 0.01; ***, P  ≤ 0.001 (using Student’s t test [ n  ≥ 3]). Error bars represent ±1 SEM. Statistical comparisons were performed between treated and untreated conditions.
Figure Legend Snippet: Polyamines enhance specific infectivity of enveloped viruses. (A to C) Huh7 cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with ZIKV (strain MR766) at an MOI of 0.1 PFU per cell. At 48 hpi, viral titers were determined via plaque assay (A) and relative viral genomes were determined by qRT-PCR (B), and the ratio of genomes to PFU was calculated (C). (D to F) Vero cells were treated with 500 μM or 1 mM DFMO for 4 days prior to infection with CVB3 (Nancy strain) at an MOI of 0.1 PFU per cell. At 24 hpi, viral titers were determined via plaque assay (D) and relative viral genomes were determined by qRT-PCR (E), and the ratio of genomes to PFU was calculated (F). (G to I) Identical infection and analysis methods were performed for KEYV in Huh7 cells to determine viral titers (G), viral genomes (H), and genome-to-PFU ratio (I) at 48 hpi. (J to L) These methods were also used for VSV (strain Indiana) infected at an MOI of 0.01 PFU per cell for 24 h. Viral titers (J), genomes (K), and genome-to-PFU ratio (L) were calculated. *, P  ≤ 0.05; **, P  ≤ 0.01; ***, P  ≤ 0.001 (using Student’s t test [ n  ≥ 3]). Error bars represent ±1 SEM. Statistical comparisons were performed between treated and untreated conditions.

Techniques Used: Infection, Plaque Assay, Quantitative RT-PCR

5) Product Images from "Establishment of a Replicon System for Bourbon Virus and Identification of Small Molecules that Efficiently Inhibit Virus Replication"

Article Title: Establishment of a Replicon System for Bourbon Virus and Identification of Small Molecules that Efficiently Inhibit Virus Replication

Journal: bioRxiv

doi: 10.1101/2020.04.24.058693

Determination of the CC 50 and EC 50 of compound #9 in cell cultures. (A B) EC 50 of compound #9. HEK293T cells (A) or Vero cells (B) were infected with BRBV at an MOI of 10, followed by application of compound #9 at various concentrations as indicated. At 3 days post treatment, RT-qPCR was performed to quantify the viral genome copy (vgc) numbers in the cell culture media. Values are the means with standard deviation and are normalized to a mock group, and were obtained from three experiments, each performed in duplicate. The EC 50 was calculated using GraphPad Prism software. (C D) CC 50 of compound #9 . A cytotoxicity assay was used to measure the viability of HEK293T cells (C) or Vero cells (D) affected by compound #9 at various concentrations as indicated. The percentage of viable cells were determined using a CytoTox-Glo cytotoxicity assay kit (Promega). The results are shown as relative values to the mock control cells. Values are the means with standard deviation obtained from three experiments performed in duplicate. The CC 50 was calculated using GraphPad Prism software.
Figure Legend Snippet: Determination of the CC 50 and EC 50 of compound #9 in cell cultures. (A B) EC 50 of compound #9. HEK293T cells (A) or Vero cells (B) were infected with BRBV at an MOI of 10, followed by application of compound #9 at various concentrations as indicated. At 3 days post treatment, RT-qPCR was performed to quantify the viral genome copy (vgc) numbers in the cell culture media. Values are the means with standard deviation and are normalized to a mock group, and were obtained from three experiments, each performed in duplicate. The EC 50 was calculated using GraphPad Prism software. (C D) CC 50 of compound #9 . A cytotoxicity assay was used to measure the viability of HEK293T cells (C) or Vero cells (D) affected by compound #9 at various concentrations as indicated. The percentage of viable cells were determined using a CytoTox-Glo cytotoxicity assay kit (Promega). The results are shown as relative values to the mock control cells. Values are the means with standard deviation obtained from three experiments performed in duplicate. The CC 50 was calculated using GraphPad Prism software.

Techniques Used: Infection, Quantitative RT-PCR, Cell Culture, Standard Deviation, Software, Cytotoxicity Assay

Comparisons of BRBV-KS infection between Vero and HEK293T cells (A) BRBV growth kinetics. Cells were infected with BRBV-KS at an MOI=10 pfu/cell. Cell culture media (supernatants) were collected daily for 6 days. Viral RNA was extracted from the samples of supernatants and then titrated by a RT-qPCR assay. The data presented the means with standard deviations, which are obtained from three independent experiments, with each experiment analyzing samples in duplicate. (B) Western blotting. The M protein in BRBV-infected Vero and HEK293T cells was detected at ~30 kDa as indicated, but not in mock-infected cells. β-action was probed as a loading control.
Figure Legend Snippet: Comparisons of BRBV-KS infection between Vero and HEK293T cells (A) BRBV growth kinetics. Cells were infected with BRBV-KS at an MOI=10 pfu/cell. Cell culture media (supernatants) were collected daily for 6 days. Viral RNA was extracted from the samples of supernatants and then titrated by a RT-qPCR assay. The data presented the means with standard deviations, which are obtained from three independent experiments, with each experiment analyzing samples in duplicate. (B) Western blotting. The M protein in BRBV-infected Vero and HEK293T cells was detected at ~30 kDa as indicated, but not in mock-infected cells. β-action was probed as a loading control.

Techniques Used: Infection, Cell Culture, Quantitative RT-PCR, Western Blot

Titration of BRBV stock. (A) Plaque assay. Vero cell monolayers on a 6-well plate were infected with 200 μl of serial 10-fold diluted BRBV-KS as indicated. Plaques were visualized by staining with crystal violet after 5 days of infection on Vero cells. Arrows indicate plaques. (B) Comparison between plaque assay and RT-qPCR. The same virus stock was diluted by serial 10-fold dilution. RT-qPCR was used to quantify the viral genome copy (vgc) number. The linear regression has a R 2 of 0.9875, indicating 1 plaque forming unit (pfu) equals ~100 vgc. The data presented are averages and standard deviations from three independent experiments, with each experiment analyzing samples in duplicate.
Figure Legend Snippet: Titration of BRBV stock. (A) Plaque assay. Vero cell monolayers on a 6-well plate were infected with 200 μl of serial 10-fold diluted BRBV-KS as indicated. Plaques were visualized by staining with crystal violet after 5 days of infection on Vero cells. Arrows indicate plaques. (B) Comparison between plaque assay and RT-qPCR. The same virus stock was diluted by serial 10-fold dilution. RT-qPCR was used to quantify the viral genome copy (vgc) number. The linear regression has a R 2 of 0.9875, indicating 1 plaque forming unit (pfu) equals ~100 vgc. The data presented are averages and standard deviations from three independent experiments, with each experiment analyzing samples in duplicate.

Techniques Used: Titration, Plaque Assay, Infection, Staining, Quantitative RT-PCR

Related Articles

Amplification:

Article Title: Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity
Article Snippet: .. To better measure viral fitness via competition assay, we infected Vero cells with equal quantities of infectious virus of wildtype and 2A29K or 3C52R , and after five passages, we extracted RNA, reverse transcribed, amplified viral genomes by PCR, and Sanger sequenced the appropriate genomic locus corresponding to each protease ( ). .. After the fifth passage, we observed that the wildtype and either protease mutant virus existed as a mixed population, with peaks corresponding to the wildtype and mutant nucleotide present in the samples ( E, representative of three independent passages).

Article Title: Zika viruses of African and Asian lineages cause fetal harm in a mouse model of vertical transmission
Article Snippet: .. ZIKV strain DAK AR 41524 (ZIKV-DAK; GenBank:KX601166) was originally isolated from Aedes luteocephalus mosquitoes in Senegal in 1984, with a round of amplification on Aedes pseudocutellaris cells, followed by amplification on C6/36 cells, followed by two rounds of amplification on Vero cells, was obtained from BEI Resources (Manassas, VA). ..

Isolation:

Article Title: Zika viruses of African and Asian lineages cause fetal harm in a mouse model of vertical transmission
Article Snippet: .. ZIKV strain DAK AR 41524 (ZIKV-DAK; GenBank:KX601166) was originally isolated from Aedes luteocephalus mosquitoes in Senegal in 1984, with a round of amplification on Aedes pseudocutellaris cells, followed by amplification on C6/36 cells, followed by two rounds of amplification on Vero cells, was obtained from BEI Resources (Manassas, VA). ..

Infection:

Article Title: Establishment of a Replicon System for Bourbon Virus and Identification of Small Molecules that Efficiently Inhibit Virus Replication
Article Snippet: .. BRBV-KS infection of HEK293T cells We propagated BRBV-KS, obtained from BEI Resources ( www.beiresources.org/ ), in Vero cells. ..

Article Title: Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity
Article Snippet: .. To better measure viral fitness via competition assay, we infected Vero cells with equal quantities of infectious virus of wildtype and 2A29K or 3C52R , and after five passages, we extracted RNA, reverse transcribed, amplified viral genomes by PCR, and Sanger sequenced the appropriate genomic locus corresponding to each protease ( ). .. After the fifth passage, we observed that the wildtype and either protease mutant virus existed as a mixed population, with peaks corresponding to the wildtype and mutant nucleotide present in the samples ( E, representative of three independent passages).

other:

Article Title: Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity
Article Snippet: Plaque Size Measurement To quantify the relative plaque sizes of the different CVB3 mutants, Vero cells were seeded in 10 cm dishes and grown to confluence.

Derivative Assay:

Article Title: Polyamine Depletion Inhibits Bunyavirus Infection via Generation of Noninfectious Interfering Virions
Article Snippet: .. ZIKV (MR766) was derived from the first passage of virus in Vero cells. .. CVB3 (Nancy strain) was derived from the first passage of virus in HeLa cells.

Polymerase Chain Reaction:

Article Title: Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity
Article Snippet: .. To better measure viral fitness via competition assay, we infected Vero cells with equal quantities of infectious virus of wildtype and 2A29K or 3C52R , and after five passages, we extracted RNA, reverse transcribed, amplified viral genomes by PCR, and Sanger sequenced the appropriate genomic locus corresponding to each protease ( ). .. After the fifth passage, we observed that the wildtype and either protease mutant virus existed as a mixed population, with peaks corresponding to the wildtype and mutant nucleotide present in the samples ( E, representative of three independent passages).

Competitive Binding Assay:

Article Title: Coxsackievirus B3 Responds to Polyamine Depletion via Enhancement of 2A and 3C Protease Activity
Article Snippet: .. To better measure viral fitness via competition assay, we infected Vero cells with equal quantities of infectious virus of wildtype and 2A29K or 3C52R , and after five passages, we extracted RNA, reverse transcribed, amplified viral genomes by PCR, and Sanger sequenced the appropriate genomic locus corresponding to each protease ( ). .. After the fifth passage, we observed that the wildtype and either protease mutant virus existed as a mixed population, with peaks corresponding to the wildtype and mutant nucleotide present in the samples ( E, representative of three independent passages).

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    BEI Resources vero e6 cells
    Positive control data collected on <t>Vero</t> <t>E6</t> cells of various passage ages. Twenty eight measurements of the EBOV Positive Control (PC) collected over pre-, post- and on-validation studies were performed on cell passages ranging from 30 to 41 (filled circles). Averages and standard deviations are presented for multiple results on the same passage. The solid line depicts the nominal PC target titer of 1.09 × 10 5 PFU/mL, and the dotted line denotes the lower PC cutoff limit of 34,469 PFU/mL, below which the assay fails and a repeat is required.
    Vero E6 Cells, supplied by BEI Resources, used in various techniques. Bioz Stars score: 92/100, based on 8 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vero e6 cells/product/BEI Resources
    Average 92 stars, based on 8 article reviews
    Price from $9.99 to $1999.99
    vero e6 cells - by Bioz Stars, 2020-11
    92/100 stars
      Buy from Supplier

    94
    BEI Resources vero cells
    In vivo rescue of rZIKV from <t>Vero</t> cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by <t>immunofocus</t> assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.
    Vero Cells, supplied by BEI Resources, used in various techniques. Bioz Stars score: 94/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/vero cells/product/BEI Resources
    Average 94 stars, based on 4 article reviews
    Price from $9.99 to $1999.99
    vero cells - by Bioz Stars, 2020-11
    94/100 stars
      Buy from Supplier

    Image Search Results


    Positive control data collected on Vero E6 cells of various passage ages. Twenty eight measurements of the EBOV Positive Control (PC) collected over pre-, post- and on-validation studies were performed on cell passages ranging from 30 to 41 (filled circles). Averages and standard deviations are presented for multiple results on the same passage. The solid line depicts the nominal PC target titer of 1.09 × 10 5 PFU/mL, and the dotted line denotes the lower PC cutoff limit of 34,469 PFU/mL, below which the assay fails and a repeat is required.

    Journal: Viruses

    Article Title: Validation of the Filovirus Plaque Assay for Use in Preclinical Studies

    doi: 10.3390/v8040113

    Figure Lengend Snippet: Positive control data collected on Vero E6 cells of various passage ages. Twenty eight measurements of the EBOV Positive Control (PC) collected over pre-, post- and on-validation studies were performed on cell passages ranging from 30 to 41 (filled circles). Averages and standard deviations are presented for multiple results on the same passage. The solid line depicts the nominal PC target titer of 1.09 × 10 5 PFU/mL, and the dotted line denotes the lower PC cutoff limit of 34,469 PFU/mL, below which the assay fails and a repeat is required.

    Article Snippet: Cells The specific Vero E6 cells cultured and plated for the plaque assay (C1008) were obtained from BEI Resources at passage Number 24 (Manassas, VA, USA).

    Techniques: Positive Control

    10× images of MDM, Vero E6, and Huh-7 cells taken 10 days following exposure to select volumes of normal semen, breast milk, or cell culture media (control). Breast milk was toxic at 50 and 100 µL/T-25 flask on Vero E6 cells (no cells present after 10 days) and MDM cells, but toxicity was less on these cells than on Huh7 cells.

    Journal: Viruses

    Article Title: Ebola Virus Isolation Using Huh-7 Cells has Methodological Advantages and Similar Sensitivity to Isolation Using Other Cell Types and Suckling BALB/c Laboratory Mice

    doi: 10.3390/v11020161

    Figure Lengend Snippet: 10× images of MDM, Vero E6, and Huh-7 cells taken 10 days following exposure to select volumes of normal semen, breast milk, or cell culture media (control). Breast milk was toxic at 50 and 100 µL/T-25 flask on Vero E6 cells (no cells present after 10 days) and MDM cells, but toxicity was less on these cells than on Huh7 cells.

    Article Snippet: When target titers ranged from 1 to 10 PFU/mL, isolation success decreased to 25% (2/8) in Vero E6 cells, 50% (5/10) in Huh-7 cells, 38% (3/8) in MDM cells, and 50% (2/4) in mice.

    Techniques: Cell Culture

    Antiviral activity of GC-376 analogues. a-d Antiviral activity of GC-376 analogues against SARS-CoV-2 in the immunofluorescence assay. a GC-376 ; b UAWJ246 ; c UAWJ247 ; d UAWJ248 . Vero E6 cells in a 96-well plate were infected with SARS-CoV-2 (USA-WA1/2020 isolate) at MOI of 0.05 in the presence of the indicated concentrations of the tested compounds. At 48 hpi, the cells were fixed and stained with a rabbit monoclonal antibody against the SARS-CoV-2 NP and a secondary antibody conjugated with Alexa 488 (Green). The nuclei were counterstained with Hoechst dye (Blue). For each well, fluorescence images of approximately 10K cells were acquired and shown. The images are representatives of two repeats. e-h Antiviral activity of GC-376 analogues against SARS-CoV-2 in the plaque assay. e GC-376 ; f UAWJ246 ; g UAWJ247 ; h UAWJ248 . Vero E6 cells in 6-well plates were infected with approximately 40 PFU/well of SARS-CoV-2 (USA-WA1/2020 isolate). After 1 hour, the inoculum was removed, and the cells were overlaid with medium containing the indicated concentrations of the tested compounds and 1.2% Avicel RC-591. At 3 dpi, the overlay was removed, and the cells were stained with 0.2% crystal violet. The images are representatives of two repeats. Data fitting of the antiviral activity of GC-376 analogues against SARS-CoV-2 in the immunofluorescence assay ( i ) and the plaque assay ( j ).

    Journal: bioRxiv

    Article Title: Structure and inhibition of the SARS-CoV-2 main protease reveals strategy for developing dual inhibitors against Mpro and cathepsin L

    doi: 10.1101/2020.07.27.223727

    Figure Lengend Snippet: Antiviral activity of GC-376 analogues. a-d Antiviral activity of GC-376 analogues against SARS-CoV-2 in the immunofluorescence assay. a GC-376 ; b UAWJ246 ; c UAWJ247 ; d UAWJ248 . Vero E6 cells in a 96-well plate were infected with SARS-CoV-2 (USA-WA1/2020 isolate) at MOI of 0.05 in the presence of the indicated concentrations of the tested compounds. At 48 hpi, the cells were fixed and stained with a rabbit monoclonal antibody against the SARS-CoV-2 NP and a secondary antibody conjugated with Alexa 488 (Green). The nuclei were counterstained with Hoechst dye (Blue). For each well, fluorescence images of approximately 10K cells were acquired and shown. The images are representatives of two repeats. e-h Antiviral activity of GC-376 analogues against SARS-CoV-2 in the plaque assay. e GC-376 ; f UAWJ246 ; g UAWJ247 ; h UAWJ248 . Vero E6 cells in 6-well plates were infected with approximately 40 PFU/well of SARS-CoV-2 (USA-WA1/2020 isolate). After 1 hour, the inoculum was removed, and the cells were overlaid with medium containing the indicated concentrations of the tested compounds and 1.2% Avicel RC-591. At 3 dpi, the overlay was removed, and the cells were stained with 0.2% crystal violet. The images are representatives of two repeats. Data fitting of the antiviral activity of GC-376 analogues against SARS-CoV-2 in the immunofluorescence assay ( i ) and the plaque assay ( j ).

    Article Snippet: SARS-CoV-2, isolate USA-WA1/2020 (NR-52281), was obtained through BEI Resources and propagated once on VERO E6 cells before it was used for this study.

    Techniques: Activity Assay, Immunofluorescence, Infection, Staining, Fluorescence, Plaque Assay

    In vivo rescue of rZIKV from Vero cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by immunofocus assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.

    Journal: Scientific Reports

    Article Title: In vivo rescue of recombinant Zika virus from an infectious cDNA clone and its implications in vaccine development

    doi: 10.1038/s41598-020-57545-2

    Figure Lengend Snippet: In vivo rescue of rZIKV from Vero cells transfected with the BAC cDNA clone. (A) Schematic representation of rZIKV in vivo rescue approach using transfected Vero cells. ( B–D) Four-to-five-week-old IFNAR−/− A129 male (N = 3) and female (N = 3) mice were inoculated SC in the footpad (top), IM in the quadriceps muscle (middle) or IP in the peritoneal cavity (bottom) with 1 × 10 6 Vero cells/mouse transfected with pBAC-ZIKV. Body weight ( B ) and survival ( C ) were evaluated during 18 dpi. Mice that lost more than 20% of their initial body weight or presented hind limb paralysis were humanely euthanized. Error bars represent standard deviations (SD) of the mean for each group of mice. Mice were bled at 3 and 6 dpi and viral titers in sera were determined by immunofocus assay (FFU/ml) ( D ). Symbols represent data from individual mice and bars the geometric means of viral titers. *Virus not detected in one mouse; , virus not detected in two mice; ND, not detected. Dotted black lines indicate the limit of detection, LoD (200 FFU/ml). No statistically significant differences between the SC, IM and IP routes of inoculation of transfected cells were observed.

    Article Snippet: Virus stocks were propagated in Vero cells and titrated by immunofocus assay in Vero cells using the pan-flavivirus envelope (E) protein monoclonal antibody (mAb) 4G2 from BEI Resources (NR-50327) .

    Techniques: In Vivo, Transfection, BAC Assay, Mouse Assay