sars cov 2 2019 ncov nucleocapsid his recombinant protein covid 19 nucleocapsid research  (Sino Biological)


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    SARS CoV 2 2019 nCoV Nucleocapsid His recombinant Protein COVID 19 Nucleocapsid Research
    Description:
    A DNA sequence encoding the SARS CoV 2 2019 nCoV Nucleocapsid Protein YP 009724397 2 335Gly Ala Met1 Ala419 was expressed with a polyhistidine tag at the C terminus
    Catalog Number:
    40588-V08B
    Price:
    None
    Category:
    recombinant protein
    Product Aliases:
    coronavirus NP Protein 2019-nCoV, coronavirus Nucleocapsid Protein 2019-nCoV, coronavirus Nucleoprotein Protein 2019-nCoV, cov np Protein 2019-nCoV, ncov NP Protein 2019-nCoV, NCP-CoV Nucleocapsid Protein 2019-nCoV, novel coronavirus NP Protein 2019-nCoV, novel coronavirus Nucleocapsid Protein 2019-nCoV, novel coronavirus Nucleoprotein Protein 2019-nCoV, np Protein 2019-nCoV, nucleocapsid Protein 2019-nCoV, Nucleoprotein Protein 2019-nCoV
    Host:
    Baculovirus-Insect Cells
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    Structured Review

    Sino Biological sars cov 2 2019 ncov nucleocapsid his recombinant protein covid 19 nucleocapsid research
    GCG suppresses <t>SARS-CoV-2</t> replication. a , b Immunofluorescence analysis of N protein in A549-hACE2-Flag cells infected with SARS-CoV-2 for 24 h ( a ). The percentage of cells with N protein foci was quantified, n = 8 biologically independent samples, 20 randomly selected views were analyzed in each sample ( b ). Scale bar, 10 μm. c 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 . d , e The inhibitory effect of GCG on the replication of SARS-CoV-2, n = 6 biologically independent samples ( d ). IC 50 was calculated, n = 5 biologically independent samples ( e ). The infection was performed after 1-h pretreatment of GCG. f , g Representative immunofluorescent images showed the inhibitory effect of GCG on SARS-CoV-2 N protein. 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 ( f ). Violin plots showing foci of cells ( n = 50 biologically independent cells) from each group, lines within the plots, with 25th, 50th, and 75th percentiles marked ( g ). h Cells were infected with SARS-CoV-2 for 1 h followed by 24-h GCG treatment, n = 3 biologically independent samples. Representative images were shown. SARS-CoV-2 was used at an MOI of 1. Hoechst (blue), nuclear staining ( a , c , f ). Error bars, mean with s.d. ( b , d , e , g , h ). Two-tailed unpaired Student’s t -test, * P
    A DNA sequence encoding the SARS CoV 2 2019 nCoV Nucleocapsid Protein YP 009724397 2 335Gly Ala Met1 Ala419 was expressed with a polyhistidine tag at the C terminus
    https://www.bioz.com/result/sars cov 2 2019 ncov nucleocapsid his recombinant protein covid 19 nucleocapsid research/product/Sino Biological
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    Images

    1) Product Images from "GCG inhibits SARS-CoV-2 replication by disrupting the liquid phase condensation of its nucleocapsid protein"

    Article Title: GCG inhibits SARS-CoV-2 replication by disrupting the liquid phase condensation of its nucleocapsid protein

    Journal: Nature Communications

    doi: 10.1038/s41467-021-22297-8

    GCG suppresses SARS-CoV-2 replication. a , b Immunofluorescence analysis of N protein in A549-hACE2-Flag cells infected with SARS-CoV-2 for 24 h ( a ). The percentage of cells with N protein foci was quantified, n = 8 biologically independent samples, 20 randomly selected views were analyzed in each sample ( b ). Scale bar, 10 μm. c 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 . d , e The inhibitory effect of GCG on the replication of SARS-CoV-2, n = 6 biologically independent samples ( d ). IC 50 was calculated, n = 5 biologically independent samples ( e ). The infection was performed after 1-h pretreatment of GCG. f , g Representative immunofluorescent images showed the inhibitory effect of GCG on SARS-CoV-2 N protein. 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 ( f ). Violin plots showing foci of cells ( n = 50 biologically independent cells) from each group, lines within the plots, with 25th, 50th, and 75th percentiles marked ( g ). h Cells were infected with SARS-CoV-2 for 1 h followed by 24-h GCG treatment, n = 3 biologically independent samples. Representative images were shown. SARS-CoV-2 was used at an MOI of 1. Hoechst (blue), nuclear staining ( a , c , f ). Error bars, mean with s.d. ( b , d , e , g , h ). Two-tailed unpaired Student’s t -test, * P
    Figure Legend Snippet: GCG suppresses SARS-CoV-2 replication. a , b Immunofluorescence analysis of N protein in A549-hACE2-Flag cells infected with SARS-CoV-2 for 24 h ( a ). The percentage of cells with N protein foci was quantified, n = 8 biologically independent samples, 20 randomly selected views were analyzed in each sample ( b ). Scale bar, 10 μm. c 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 . d , e The inhibitory effect of GCG on the replication of SARS-CoV-2, n = 6 biologically independent samples ( d ). IC 50 was calculated, n = 5 biologically independent samples ( e ). The infection was performed after 1-h pretreatment of GCG. f , g Representative immunofluorescent images showed the inhibitory effect of GCG on SARS-CoV-2 N protein. 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 ( f ). Violin plots showing foci of cells ( n = 50 biologically independent cells) from each group, lines within the plots, with 25th, 50th, and 75th percentiles marked ( g ). h Cells were infected with SARS-CoV-2 for 1 h followed by 24-h GCG treatment, n = 3 biologically independent samples. Representative images were shown. SARS-CoV-2 was used at an MOI of 1. Hoechst (blue), nuclear staining ( a , c , f ). Error bars, mean with s.d. ( b , d , e , g , h ). Two-tailed unpaired Student’s t -test, * P

    Techniques Used: Immunofluorescence, Infection, Microscopy, Staining, Two Tailed Test

    RNA triggers the LLPS of N protein. a Schematic drawing of SARS-CoV-2. b IDR scores of 29 proteins encoded by SARS-CoV-2 genome. FUS and mEGFP are positive and negative controls, respectively. IUPred2 and ANCHOR2 were used as prediction tools. c Time-lapse imaging of N-mEGFP protein (20 μM) in the presence of Cy5-labeled 60-nt vRNA (100 ng/μl), scale bar, 10 μm. d Representative fluorescent images of N-mEGFP-vRNA (60 nt) condensates fusion from a time-lapse movie, scale bar, 3 μm. e – g LLPS of N-mEGFP protein (20 μM) in the presence of indicated concentrations of 60-nt vRNA, scale bar, 10 μm ( e ). The partition coefficient of fluorescence intensity per droplet ( f ) and the partition coefficient of total fluorescence intensity in each view ( g ) were calculated. From left to right, n = 209, 1170, 1026, 1170 droplets ( f ) from 10 randomly selected views ( g ). h , i FRAP analysis of vRNA-induced liquid droplets of N-mEGFP protein, scale bar, 2 μm ( h ), and quantification of fluorescence intensity recovery of a photobleached N-mEGFP protein, n = 3 biologically independent experiments ( i ). The white dotted circle in h indicated the region of photobleaching. 20 μM N-mEGFP protein and 100 ng/μl 60-nt vRNA were used. Error bars, mean with s.d. ( f , g , i ). Two-tailed unpaired Student’s t -test ( f , g ), **** P
    Figure Legend Snippet: RNA triggers the LLPS of N protein. a Schematic drawing of SARS-CoV-2. b IDR scores of 29 proteins encoded by SARS-CoV-2 genome. FUS and mEGFP are positive and negative controls, respectively. IUPred2 and ANCHOR2 were used as prediction tools. c Time-lapse imaging of N-mEGFP protein (20 μM) in the presence of Cy5-labeled 60-nt vRNA (100 ng/μl), scale bar, 10 μm. d Representative fluorescent images of N-mEGFP-vRNA (60 nt) condensates fusion from a time-lapse movie, scale bar, 3 μm. e – g LLPS of N-mEGFP protein (20 μM) in the presence of indicated concentrations of 60-nt vRNA, scale bar, 10 μm ( e ). The partition coefficient of fluorescence intensity per droplet ( f ) and the partition coefficient of total fluorescence intensity in each view ( g ) were calculated. From left to right, n = 209, 1170, 1026, 1170 droplets ( f ) from 10 randomly selected views ( g ). h , i FRAP analysis of vRNA-induced liquid droplets of N-mEGFP protein, scale bar, 2 μm ( h ), and quantification of fluorescence intensity recovery of a photobleached N-mEGFP protein, n = 3 biologically independent experiments ( i ). The white dotted circle in h indicated the region of photobleaching. 20 μM N-mEGFP protein and 100 ng/μl 60-nt vRNA were used. Error bars, mean with s.d. ( f , g , i ). Two-tailed unpaired Student’s t -test ( f , g ), **** P

    Techniques Used: Imaging, Labeling, Fluorescence, Two Tailed Test

    NR203K/G204R gained greater ability to undergo RNA-induced LLPS. a Distribution of N gene variants among 100,849 SARS-CoV-2 genomes obtained from GISAID database. Colors indicated the nucleotide variability numbers from 100,849 genomes. The high-frequency trio-nucleotide polymorphism variant (GGG-to-AAC) is shown. b Coomassie brilliant blue-stained SDS-PAGE gel of purified variants of N-mEGFP protein. c – e LLPS of different N-mEGFP variants, in the presence of 50 ng/μl 60-nt vRNA ( c ). The partition coefficient of fluorescence intensity per droplet ( d ) and the partition coefficient of total fluorescence intensity in each view ( e ) were calculated. From left to right, n = 1232, 803, 897, 431 droplets ( d ) from 10 randomly selected views ( e ). f , g Time-lapse imaging of N R203/G204 -mEGFP and N R203K/G204R -mEGFP proteins (20 μM) in the presence of Cy5-labeled 60-nt vRNA (40 ng/μl) ( f ), and the partition coefficient ( n = 8 randomly selected views) of total fluorescence intensity in each view ( g ). Scale bars, 10 μm ( c , f ). Error bars, mean with s.d. ( d , e ) and mean with s.e.m. ( g ). Two-tailed unpaired Student’s t -test ( d , e ), **** P
    Figure Legend Snippet: NR203K/G204R gained greater ability to undergo RNA-induced LLPS. a Distribution of N gene variants among 100,849 SARS-CoV-2 genomes obtained from GISAID database. Colors indicated the nucleotide variability numbers from 100,849 genomes. The high-frequency trio-nucleotide polymorphism variant (GGG-to-AAC) is shown. b Coomassie brilliant blue-stained SDS-PAGE gel of purified variants of N-mEGFP protein. c – e LLPS of different N-mEGFP variants, in the presence of 50 ng/μl 60-nt vRNA ( c ). The partition coefficient of fluorescence intensity per droplet ( d ) and the partition coefficient of total fluorescence intensity in each view ( e ) were calculated. From left to right, n = 1232, 803, 897, 431 droplets ( d ) from 10 randomly selected views ( e ). f , g Time-lapse imaging of N R203/G204 -mEGFP and N R203K/G204R -mEGFP proteins (20 μM) in the presence of Cy5-labeled 60-nt vRNA (40 ng/μl) ( f ), and the partition coefficient ( n = 8 randomly selected views) of total fluorescence intensity in each view ( g ). Scale bars, 10 μm ( c , f ). Error bars, mean with s.d. ( d , e ) and mean with s.e.m. ( g ). Two-tailed unpaired Student’s t -test ( d , e ), **** P

    Techniques Used: Variant Assay, Staining, SDS Page, Purification, Fluorescence, Imaging, Labeling, Two Tailed Test

    2) Product Images from "SARS-CoV-2 RapidPlex: A Graphene-Based Multiplexed Telemedicine Platform for Rapid and Low-Cost COVID-19 Diagnosis and Monitoring"

    Article Title: SARS-CoV-2 RapidPlex: A Graphene-Based Multiplexed Telemedicine Platform for Rapid and Low-Cost COVID-19 Diagnosis and Monitoring

    Journal: Matter

    doi: 10.1016/j.matt.2020.09.027

    Evaluation of Analytical Sensor Performance for the Detection of Physiological Levels of Target COVID-19 Biomarkers (A) Scheme of sensor preparation for detection of SARS-CoV-2 NP and CRP based on double-sandwich and sandwich assay configurations, respectively. CAb, capture antibody; DAb, detector antibody; DAb 2 , secondary detector antibody; HRP, horseradish peroxidase. (B and C) Calibration curves constructed for NP (B) and CRP (C) detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3). (D) Scheme of sensor preparation for detection of S1-IgG and S1-IgM isotypes based on direct assay configurations. (E and F) Calibration curves constructed for S1-IgG (E) and S1-IgM (F) isotype detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3).
    Figure Legend Snippet: Evaluation of Analytical Sensor Performance for the Detection of Physiological Levels of Target COVID-19 Biomarkers (A) Scheme of sensor preparation for detection of SARS-CoV-2 NP and CRP based on double-sandwich and sandwich assay configurations, respectively. CAb, capture antibody; DAb, detector antibody; DAb 2 , secondary detector antibody; HRP, horseradish peroxidase. (B and C) Calibration curves constructed for NP (B) and CRP (C) detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3). (D) Scheme of sensor preparation for detection of S1-IgG and S1-IgM isotypes based on direct assay configurations. (E and F) Calibration curves constructed for S1-IgG (E) and S1-IgM (F) isotype detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3).

    Techniques Used: Construct

    Application of SARS-CoV-2 RapidPlex in SARS-CoV-2 Detection in Blood and Saliva Samples from COVID-19-Positive and -Negative Subjects (A and B) Experimental readings obtained with SARS-CoV-2 RapidPlex after 10-min incubation of the sensor array with serum samples from a representative COVID-19 RT-PCR-negative (A) and -positive (B) patient. (C) Signal of individual sensor obtained after 1-min incubation with a serum sample from a COVID-19-positive patient (dark color) versus the signal obtained after 10-min incubation with a serum sample from a COVID-19-negative patient (light color). (D) Box-and-whisker plot of measured signal-to-blank ratios (S/B) for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 6) serum samples. (E) Box-and-whisker plot of measured S/B for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 3) saliva samples. (F) CRP levels in diluted serum samples plotted against given COVID-19 symptom severity, with “Healthy” referring to COVID-19-negative patient samples (n = 7). Positive COVID-19 patients were classified according to disease severity as asymptomatic (n = 2), mild (n = 5), and moderate (n = 2).
    Figure Legend Snippet: Application of SARS-CoV-2 RapidPlex in SARS-CoV-2 Detection in Blood and Saliva Samples from COVID-19-Positive and -Negative Subjects (A and B) Experimental readings obtained with SARS-CoV-2 RapidPlex after 10-min incubation of the sensor array with serum samples from a representative COVID-19 RT-PCR-negative (A) and -positive (B) patient. (C) Signal of individual sensor obtained after 1-min incubation with a serum sample from a COVID-19-positive patient (dark color) versus the signal obtained after 10-min incubation with a serum sample from a COVID-19-negative patient (light color). (D) Box-and-whisker plot of measured signal-to-blank ratios (S/B) for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 6) serum samples. (E) Box-and-whisker plot of measured S/B for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 3) saliva samples. (F) CRP levels in diluted serum samples plotted against given COVID-19 symptom severity, with “Healthy” referring to COVID-19-negative patient samples (n = 7). Positive COVID-19 patients were classified according to disease severity as asymptomatic (n = 2), mild (n = 5), and moderate (n = 2).

    Techniques Used: Incubation, Reverse Transcription Polymerase Chain Reaction, Whisker Assay

    3) Product Images from "COVID-19 Patients Upregulate Toll-like Receptor 4-mediated Inflammatory Signaling That Mimics Bacterial Sepsis"

    Article Title: COVID-19 Patients Upregulate Toll-like Receptor 4-mediated Inflammatory Signaling That Mimics Bacterial Sepsis

    Journal: Journal of Korean Medical Science

    doi: 10.3346/jkms.2020.35.e343

    COVID-19 infection boosts NF-κB signaling pathway. ( A ) The left (MILD) and right (SEVERE) sides of box represent the mean fold change in mRNA levels, compared with HC. The NF-κB signaling pathway was adopted from KEGG database (accession number: hsa04064). ( B ) The expression levels of IRF3 , TLR3 , TLR7 , TLR8 and TLR9 were represented by FPKM. Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, NF = nuclear factor, MILD = mild/moderate, SEVERE = severe/critical, HC = healthy controls, KEGG = Kyoto Encyclopedia of Genes and Genomes, FPKM = fragments per kilobase exon-model per million reads mapped, FDR = false discovery rate, TLR = toll-like receptor, IRF = interferon regulatory factor. * P
    Figure Legend Snippet: COVID-19 infection boosts NF-κB signaling pathway. ( A ) The left (MILD) and right (SEVERE) sides of box represent the mean fold change in mRNA levels, compared with HC. The NF-κB signaling pathway was adopted from KEGG database (accession number: hsa04064). ( B ) The expression levels of IRF3 , TLR3 , TLR7 , TLR8 and TLR9 were represented by FPKM. Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, NF = nuclear factor, MILD = mild/moderate, SEVERE = severe/critical, HC = healthy controls, KEGG = Kyoto Encyclopedia of Genes and Genomes, FPKM = fragments per kilobase exon-model per million reads mapped, FDR = false discovery rate, TLR = toll-like receptor, IRF = interferon regulatory factor. * P

    Techniques Used: Infection, Expressing, MANN-WHITNEY

    Transcriptome analysis reveals that immune gene expression profiles of COVID-19 patients are distinct to HC. ( A ) Schematic diagram of the immune transcriptome analysis in this study. ( B ) A result of principal component analysis of log2-transformed 579 immune gene expression levels. ( C ) The scatter plots representing 579 immune genes with the log2-transformed FPKM for COVID-19 patients compared to HC. ( D ) The ten most significantly enriched KEGG pathways of the 298 DEiGs from COVID-19 patients compared to HC. ( E ) Log2-transformed fold changes of chemokine and chemokine receptor genes from MILD (x-axis) and SEVERE (y-axis) vs. HC. ( F ) Expression levels (FPKM) of marked chemokines in (E). Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, HC = healthy controls, FPKM = fragments per kilobase exon-model per million reads mapped, KEGG = Kyoto Encyclopedia of Genes and Genomes, DEiG = differentially expressed immune gene, MILD = mild/moderate, SEVERE = severe/critical, FDR = false discovery rate, IBD = inflammatory bowel disease, TNF = tumor necrosis factor, CCL = C-C motif chemokine ligand, CXCL = C-X-C motif chemokine ligand. * P
    Figure Legend Snippet: Transcriptome analysis reveals that immune gene expression profiles of COVID-19 patients are distinct to HC. ( A ) Schematic diagram of the immune transcriptome analysis in this study. ( B ) A result of principal component analysis of log2-transformed 579 immune gene expression levels. ( C ) The scatter plots representing 579 immune genes with the log2-transformed FPKM for COVID-19 patients compared to HC. ( D ) The ten most significantly enriched KEGG pathways of the 298 DEiGs from COVID-19 patients compared to HC. ( E ) Log2-transformed fold changes of chemokine and chemokine receptor genes from MILD (x-axis) and SEVERE (y-axis) vs. HC. ( F ) Expression levels (FPKM) of marked chemokines in (E). Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, HC = healthy controls, FPKM = fragments per kilobase exon-model per million reads mapped, KEGG = Kyoto Encyclopedia of Genes and Genomes, DEiG = differentially expressed immune gene, MILD = mild/moderate, SEVERE = severe/critical, FDR = false discovery rate, IBD = inflammatory bowel disease, TNF = tumor necrosis factor, CCL = C-C motif chemokine ligand, CXCL = C-X-C motif chemokine ligand. * P

    Techniques Used: Expressing, Transformation Assay, MANN-WHITNEY

    4) Product Images from "The Characterization of Disease Severity Associated IgG Subclasses Response in COVID-19 Patients"

    Article Title: The Characterization of Disease Severity Associated IgG Subclasses Response in COVID-19 Patients

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2021.632814

    IgG1 and IgG3 were the main subclasses induced in COVID-19 patients and related with disease severity. IgG1 (A) and IgG3 (B) responses to NP, S, and RBD and neutralizing antibody (C) against SARS-CoV-2 pseudo-virus with luciferase reporter gene in the healthy controls ( n = 11) and the severe ( n = 24), moderate ( n = 54), mild ( n = 35), asymptomatic ( n = 10) patients were detected. Differences of medium values between groups were analyzed by Mann-Whitney U -test. Significant correlations among Severity, NAb and IgG subclasses (D) including anti-NP IgG1, IgG3, anti-S IgG1, IgG3, anti-RBD IgG1, IgG3 were shown. Spearman correlation coefficient was calculated. A two-tailed P value
    Figure Legend Snippet: IgG1 and IgG3 were the main subclasses induced in COVID-19 patients and related with disease severity. IgG1 (A) and IgG3 (B) responses to NP, S, and RBD and neutralizing antibody (C) against SARS-CoV-2 pseudo-virus with luciferase reporter gene in the healthy controls ( n = 11) and the severe ( n = 24), moderate ( n = 54), mild ( n = 35), asymptomatic ( n = 10) patients were detected. Differences of medium values between groups were analyzed by Mann-Whitney U -test. Significant correlations among Severity, NAb and IgG subclasses (D) including anti-NP IgG1, IgG3, anti-S IgG1, IgG3, anti-RBD IgG1, IgG3 were shown. Spearman correlation coefficient was calculated. A two-tailed P value

    Techniques Used: Luciferase, MANN-WHITNEY, Two Tailed Test

    IgA, IgG, and IgM antibodies responses in COVID-19 ranging from asymptomatic to severe patients. Serum samples collected from COVID-19 patients were used for detecting IgA, IgG, and IgM levels to NP (A) , S (B) , and RBD (C) antigens of SARS-CoV-2 via ELISA. Antibody titers of the healthy controls ( n = 11) and the severe ( n = 24), moderate ( n = 54), mild ( n = 35), asymptomatic ( n = 10) patients were shown in (A–C) . Mann-Whitney U -test was used to compare differences of medium values between groups, a two-tailed P value
    Figure Legend Snippet: IgA, IgG, and IgM antibodies responses in COVID-19 ranging from asymptomatic to severe patients. Serum samples collected from COVID-19 patients were used for detecting IgA, IgG, and IgM levels to NP (A) , S (B) , and RBD (C) antigens of SARS-CoV-2 via ELISA. Antibody titers of the healthy controls ( n = 11) and the severe ( n = 24), moderate ( n = 54), mild ( n = 35), asymptomatic ( n = 10) patients were shown in (A–C) . Mann-Whitney U -test was used to compare differences of medium values between groups, a two-tailed P value

    Techniques Used: Enzyme-linked Immunosorbent Assay, MANN-WHITNEY, Two Tailed Test

    5) Product Images from "SARS-CoV-2–Specific Antibody Detection for Seroepidemiology: A Multiplex Analysis Approach Accounting for Accurate Seroprevalence"

    Article Title: SARS-CoV-2–Specific Antibody Detection for Seroepidemiology: A Multiplex Analysis Approach Accounting for Accurate Seroprevalence

    Journal: The Journal of Infectious Diseases

    doi: 10.1093/infdis/jiaa479

    Discrimination of COVID-19 patients with varying severity from a cross-sectional population panel and ILI patients. A , Individuals from the cross-sectional panel aged 3–90 years (n = 224), ILI patients with noncoronavirus (n = 75), and non-SARS-CoV-2 seasonal coronavirus-infected ILI patients (n = 109) were compared to hospitalized and nonhospitalized COVID-19 patients. Median concentration and 95% confidence intervals and statistical results (adjusted P values of Tukey multiple comparison) between the groups are shown. B , Laboratory-confirmed viral infections (see Supplementary Table 2 ) and concentration data of ILI patients are shown to confirm that the assay discriminates SARS-CoV-2–specific antibodies from antibodies induced by various laboratory-confirmed viral infections. Abbreviations: AU, arbitrary unit; COVID-19, coronavirus disease 2019; HCoV, human coronavirus; MERS-CoV, Middle East respiratory syndrome coronavirus; N, nucleoprotein; non-HCoV, noncoronavirus; RBD, receptor binding domain; RSV, respiratory syncytial virus; S1, spike protein subunit 1.
    Figure Legend Snippet: Discrimination of COVID-19 patients with varying severity from a cross-sectional population panel and ILI patients. A , Individuals from the cross-sectional panel aged 3–90 years (n = 224), ILI patients with noncoronavirus (n = 75), and non-SARS-CoV-2 seasonal coronavirus-infected ILI patients (n = 109) were compared to hospitalized and nonhospitalized COVID-19 patients. Median concentration and 95% confidence intervals and statistical results (adjusted P values of Tukey multiple comparison) between the groups are shown. B , Laboratory-confirmed viral infections (see Supplementary Table 2 ) and concentration data of ILI patients are shown to confirm that the assay discriminates SARS-CoV-2–specific antibodies from antibodies induced by various laboratory-confirmed viral infections. Abbreviations: AU, arbitrary unit; COVID-19, coronavirus disease 2019; HCoV, human coronavirus; MERS-CoV, Middle East respiratory syndrome coronavirus; N, nucleoprotein; non-HCoV, noncoronavirus; RBD, receptor binding domain; RSV, respiratory syncytial virus; S1, spike protein subunit 1.

    Techniques Used: Infection, Concentration Assay, Binding Assay

    Kinetics of antibody production after disease onset in hospitalized and nonhospitalized COVID-19 patients. Paired samples were analyzed to identify changes in IgG concentrations in hospitalized ( A ) and nonhospitalized ( B ) COVID-19 patients. C , The log-transformed concentration data of the samples shown in ( A ) and ( B ) were fitted with a 4-parameter nonlinear least squared fit. D , Of each patient with paired samples available, 1 sample was selected randomly and data were fitted to estimate the slope, R 2 of the fits, and the difference between the fitted lines determined. Abbreviations: AU, arbitrary unit; COVID-19, coronavirus disease 2019; IgG, immunoglobulin G; N, nucleoprotein; RBD, receptor binding domain; S1, spike protein subunit 1.
    Figure Legend Snippet: Kinetics of antibody production after disease onset in hospitalized and nonhospitalized COVID-19 patients. Paired samples were analyzed to identify changes in IgG concentrations in hospitalized ( A ) and nonhospitalized ( B ) COVID-19 patients. C , The log-transformed concentration data of the samples shown in ( A ) and ( B ) were fitted with a 4-parameter nonlinear least squared fit. D , Of each patient with paired samples available, 1 sample was selected randomly and data were fitted to estimate the slope, R 2 of the fits, and the difference between the fitted lines determined. Abbreviations: AU, arbitrary unit; COVID-19, coronavirus disease 2019; IgG, immunoglobulin G; N, nucleoprotein; RBD, receptor binding domain; S1, spike protein subunit 1.

    Techniques Used: Transformation Assay, Concentration Assay, Binding Assay

    6) Product Images from "SARS-CoV-2 infection induces germinal center responses with robust stimulation of CD4 T follicular helper cells in rhesus macaques"

    Article Title: SARS-CoV-2 infection induces germinal center responses with robust stimulation of CD4 T follicular helper cells in rhesus macaques

    Journal: bioRxiv

    doi: 10.1101/2020.07.07.191007

    Humoral responses to SARS-CoV-2 are dominated by IgG antibodies Concentrations of (A) IgM, (B) IgG, and (C) IgA antibodies specific for S1, S2, and N proteins were measured by BAMA or ELISA in serum of macaques infused with human COVID-19 convalescent plasma (CP; blue symbols) or naive plasma (NP; red symbols) and control non-infused animals (black symbols). The dashed line represents the median pre-infection (day 0) concentration for all animals. (D) The magnitude of the IgM, IgG and IgA antibody responses in animals that were not given human convalescent plasma was determined by dividing post-infection concentrations by those measured on day 0 in each animal. Geometric mean fold increases with SEM are shown. (E) Correlations between day 10 levels of S1-specific IgG and IgM, N-specific IgA and IgG, and pseudovirus neutralizing antibody titers and anti-RBD IgG antibodies measured by ELISA.
    Figure Legend Snippet: Humoral responses to SARS-CoV-2 are dominated by IgG antibodies Concentrations of (A) IgM, (B) IgG, and (C) IgA antibodies specific for S1, S2, and N proteins were measured by BAMA or ELISA in serum of macaques infused with human COVID-19 convalescent plasma (CP; blue symbols) or naive plasma (NP; red symbols) and control non-infused animals (black symbols). The dashed line represents the median pre-infection (day 0) concentration for all animals. (D) The magnitude of the IgM, IgG and IgA antibody responses in animals that were not given human convalescent plasma was determined by dividing post-infection concentrations by those measured on day 0 in each animal. Geometric mean fold increases with SEM are shown. (E) Correlations between day 10 levels of S1-specific IgG and IgM, N-specific IgA and IgG, and pseudovirus neutralizing antibody titers and anti-RBD IgG antibodies measured by ELISA.

    Techniques Used: Enzyme-linked Immunosorbent Assay, Infection, Concentration Assay

    SARS-CoV-2 infection leads to a rapid and transient shift in innate immune responses and increases the number CD4 T follicular helper cells in peripheral blood. (A) Experimental design. Indian-origin rhesus macaques were inoculated with SARS-CoV-2 (SARS-CoV-2/human/USA/CA-CZB-59×002/2020) via the intranasal (IN), intratracheal (IT) and ocular route. Twenty-four hours later, animals were infused with either COVID-19 convalescent human plasma (I+CP; blue symbols), or normal plasma (I+NP; red symbols) (both at 4ml/kg), and four animals did not receive any plasma (infected; black symbols). Blood was sampled over the course of infection and tissues were collected at necropsy (11-14 DPI) for immune profiling. (B) Mean viral RNA (+range) in each of the groups within nasal washes (C) Flow plot illustrating gating strategy to identify innate immune subsets in whole blood. (D ) Kinetics of innate immune responses (*p
    Figure Legend Snippet: SARS-CoV-2 infection leads to a rapid and transient shift in innate immune responses and increases the number CD4 T follicular helper cells in peripheral blood. (A) Experimental design. Indian-origin rhesus macaques were inoculated with SARS-CoV-2 (SARS-CoV-2/human/USA/CA-CZB-59×002/2020) via the intranasal (IN), intratracheal (IT) and ocular route. Twenty-four hours later, animals were infused with either COVID-19 convalescent human plasma (I+CP; blue symbols), or normal plasma (I+NP; red symbols) (both at 4ml/kg), and four animals did not receive any plasma (infected; black symbols). Blood was sampled over the course of infection and tissues were collected at necropsy (11-14 DPI) for immune profiling. (B) Mean viral RNA (+range) in each of the groups within nasal washes (C) Flow plot illustrating gating strategy to identify innate immune subsets in whole blood. (D ) Kinetics of innate immune responses (*p

    Techniques Used: Infection

    7) Product Images from "Clonal dissection of immunodominance and cross-reactivity of the CD4+ T cell response to SARS-CoV-2"

    Article Title: Clonal dissection of immunodominance and cross-reactivity of the CD4+ T cell response to SARS-CoV-2

    Journal: bioRxiv

    doi: 10.1101/2021.03.23.436642

    Sorting of T cell subsets and identification of SARS-CoV-2 Spike- and Nucleoprotein-reactive CD4 + T cells in COVID-19 and pre-pandemic samples. ( A ) Sorting strategy to isolate CD4 + total memory T cells and Tcm, Tem and cTfh subsets. ( B, C ) Characterization of antigen-specific T cells by CFSE dilution combined with CD25 and ICOS co-expression at day 7 following stimulation with Spike or Nucleoprotein in the presence of autologous monocytes. Negative controls of T cells cultured with monocytes alone are reported as dashed lines. Shown are data from patient P2 and from a pre-pandemic healthy donor sample (HD1).
    Figure Legend Snippet: Sorting of T cell subsets and identification of SARS-CoV-2 Spike- and Nucleoprotein-reactive CD4 + T cells in COVID-19 and pre-pandemic samples. ( A ) Sorting strategy to isolate CD4 + total memory T cells and Tcm, Tem and cTfh subsets. ( B, C ) Characterization of antigen-specific T cells by CFSE dilution combined with CD25 and ICOS co-expression at day 7 following stimulation with Spike or Nucleoprotein in the presence of autologous monocytes. Negative controls of T cells cultured with monocytes alone are reported as dashed lines. Shown are data from patient P2 and from a pre-pandemic healthy donor sample (HD1).

    Techniques Used: Transmission Electron Microscopy, Expressing, Cell Culture

    8) Product Images from "Evaluation of Humoral Immunity to SARS-CoV-2: Diagnostic Value of a New Multiplex Addressable Laser Bead Immunoassay"

    Article Title: Evaluation of Humoral Immunity to SARS-CoV-2: Diagnostic Value of a New Multiplex Addressable Laser Bead Immunoassay

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2020.603931

    Detection, titration, and cross-reactivity of anti–SARS-CoV-2 Spike S1, nucleocapsid N protein IgG, and anti–SARS-CoV-2 Spike S1 IgM antibodies by ALBIA-IgG-S1/N and ALBIA-IgM-S1. (A) A calibration curve was obtained after serial dilutions of the calibrator, i.e., one highly positive sample. A plateau of MFI was reached for dilutions 1:400 or lower. (B) Calculation of antibody titer by reference to the MFI value of the calibrator (gray bar) used at a 1:400 dilution in the assay and its level arbitrarily set to 100 arbitrary units (AU)/mL. The assay was first performed using a 1:100 screening dilution of the serum. In case the sample’s MFI at 1/100 dilution was higher than 70% of the calibrator’s MFI, further dilutions were performed, and the first dilution yielding an MFI inferior to 70% of calibrator MFI was retained for calculation. An example is given: at 1:100 dilution, the MFI was higher than 70% of the calibrator’s MFI (23,311 × 0.7 = 16,318), requiring a 1/800 dilution for computing the titer, i.e., 94 AU/mL anti-S1 IgG level. Specificity toward non–COVID-19 patients: (C) anti-Spike S1 and (D) anti-N IgG, IgM, and (E) anti-Spike S1 IgM antibody reactivity in patients with different conditions: PCR-confirmed infection with other CoV (17 sera from 13 patients; HKU1, n = 3; OC43, n = 11; NL63, n = 3). RA, rheumatoid arthritis; SS, Sjögren syndrome; ASS, antisynthetase syndrome; SLE, systemic lupus erythematosus.
    Figure Legend Snippet: Detection, titration, and cross-reactivity of anti–SARS-CoV-2 Spike S1, nucleocapsid N protein IgG, and anti–SARS-CoV-2 Spike S1 IgM antibodies by ALBIA-IgG-S1/N and ALBIA-IgM-S1. (A) A calibration curve was obtained after serial dilutions of the calibrator, i.e., one highly positive sample. A plateau of MFI was reached for dilutions 1:400 or lower. (B) Calculation of antibody titer by reference to the MFI value of the calibrator (gray bar) used at a 1:400 dilution in the assay and its level arbitrarily set to 100 arbitrary units (AU)/mL. The assay was first performed using a 1:100 screening dilution of the serum. In case the sample’s MFI at 1/100 dilution was higher than 70% of the calibrator’s MFI, further dilutions were performed, and the first dilution yielding an MFI inferior to 70% of calibrator MFI was retained for calculation. An example is given: at 1:100 dilution, the MFI was higher than 70% of the calibrator’s MFI (23,311 × 0.7 = 16,318), requiring a 1/800 dilution for computing the titer, i.e., 94 AU/mL anti-S1 IgG level. Specificity toward non–COVID-19 patients: (C) anti-Spike S1 and (D) anti-N IgG, IgM, and (E) anti-Spike S1 IgM antibody reactivity in patients with different conditions: PCR-confirmed infection with other CoV (17 sera from 13 patients; HKU1, n = 3; OC43, n = 11; NL63, n = 3). RA, rheumatoid arthritis; SS, Sjögren syndrome; ASS, antisynthetase syndrome; SLE, systemic lupus erythematosus.

    Techniques Used: Titration, Polymerase Chain Reaction, Infection

    9) Product Images from "SARS-CoV-2 Proteome Microarray for Mapping COVID-19 Antibody Interactions at Amino Acid Resolution"

    Article Title: SARS-CoV-2 Proteome Microarray for Mapping COVID-19 Antibody Interactions at Amino Acid Resolution

    Journal: ACS Central Science

    doi: 10.1021/acscentsci.0c00742

    Landscape of the humoral antibody response to SARS-CoV-2 proteins other than Orf1ab. (a, b) The distribution of human IgM and IgG antibodies to SARS-CoV-2 individual proteins (S, E, M, N, Orf3a, Orf6, Orf7a, Orf8, and Orf10), respectively. The x -axis represents the sequence of amino acids of SARS-CoV-2 proteins from the N-terminal to C-terminal. The y -axis represents the serum samples from COVID-19 patients. The false-colored rainbow color from blue to red corresponds to the signals of antibody binding from low to high, respectively.
    Figure Legend Snippet: Landscape of the humoral antibody response to SARS-CoV-2 proteins other than Orf1ab. (a, b) The distribution of human IgM and IgG antibodies to SARS-CoV-2 individual proteins (S, E, M, N, Orf3a, Orf6, Orf7a, Orf8, and Orf10), respectively. The x -axis represents the sequence of amino acids of SARS-CoV-2 proteins from the N-terminal to C-terminal. The y -axis represents the serum samples from COVID-19 patients. The false-colored rainbow color from blue to red corresponds to the signals of antibody binding from low to high, respectively.

    Techniques Used: Sequencing, Binding Assay

    Landscape of humoral IgM antibody response to SARS-CoV-2 Orf1ab proteome. The x -axis represents the sequence of amino acids of SARS-CoV-2 nonstructural proteins (nsps) from the N-terminal to C-terminal. The y -axis represents the serum samples from COVID-19 patients. The false-colored rainbow color from blue to red corresponds to the signals of antibody binding from low to high, respectively.
    Figure Legend Snippet: Landscape of humoral IgM antibody response to SARS-CoV-2 Orf1ab proteome. The x -axis represents the sequence of amino acids of SARS-CoV-2 nonstructural proteins (nsps) from the N-terminal to C-terminal. The y -axis represents the serum samples from COVID-19 patients. The false-colored rainbow color from blue to red corresponds to the signals of antibody binding from low to high, respectively.

    Techniques Used: Sequencing, Binding Assay

    Landscape of the humoral IgG antibody response to the SARS-CoV-2 Orf1ab proteome. The x -axis represents the sequence of amino acids of the SARS-CoV-2 nonstructural proteins (nsps) from the N-terminal to C-terminal. The y -axis represents the serum samples from the COVID-19 patients. The false-colored rainbow color from blue to red corresponds to the signals of antibody binding from low to high, respectively.
    Figure Legend Snippet: Landscape of the humoral IgG antibody response to the SARS-CoV-2 Orf1ab proteome. The x -axis represents the sequence of amino acids of the SARS-CoV-2 nonstructural proteins (nsps) from the N-terminal to C-terminal. The y -axis represents the serum samples from the COVID-19 patients. The false-colored rainbow color from blue to red corresponds to the signals of antibody binding from low to high, respectively.

    Techniques Used: Sequencing, Binding Assay

    Identification of potential peptide epitopes for SARS-CoV-2 detection and neutralization. (a) Box-plot analysis of antibody responses to immunogenic epitopes of SARS-COV-2 between COVID-19 patients and control patients. The significance was performed using the Mann–Whitney U-test ( p -value
    Figure Legend Snippet: Identification of potential peptide epitopes for SARS-CoV-2 detection and neutralization. (a) Box-plot analysis of antibody responses to immunogenic epitopes of SARS-COV-2 between COVID-19 patients and control patients. The significance was performed using the Mann–Whitney U-test ( p -value

    Techniques Used: Neutralization, MANN-WHITNEY

    10) Product Images from "Novel ELISA Protocol Links Pre-Existing SARS-CoV-2 Reactive Antibodies With Endemic Coronavirus Immunity and Age and Reveals Improved Serologic Identification of Acute COVID-19 via Multi-Parameter Detection"

    Article Title: Novel ELISA Protocol Links Pre-Existing SARS-CoV-2 Reactive Antibodies With Endemic Coronavirus Immunity and Age and Reveals Improved Serologic Identification of Acute COVID-19 via Multi-Parameter Detection

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2021.614676

    Detection and quantification of SARS-CoV-2 RBD- and N- reactive antibodies in pre-pandemic samples. (A) Representative dilution curves of seven pre-pandemic samples with SARS-CoV-2 RBD-reactive antibodies (three subjects per graph). Open and solid symbols represent buffer only coat and SARS-CoV-2 RBD coat, respectively. Arbitrary Units (AU) were calculated as described in Methods and shown beneath the respective isotype graph for diluent only and SARS-CoV-2 RBD coat. AUs for SARS-CoV-2 RBD (B) and N (C) reactive IgM, IgG, and IgA in pre-pandemic samples. Open and solid symbols represent negative and positive results, respectively, as determined by Metric 1. Enumeration of the positive samples for each isotype in the pre-pandemic cohort is shown beneath each graph with percentages of total in parentheses. (D) Correlation between AUs for IgG reactive to SARS-CoV-2 RBD and N (n=32). Values were log-transformed to obtain a parametric distribution. Statistical analyses were performed using an unpaired non-parametric Mann-Whitney t-test in (B, C) and Pearson’s correlation of normally distributed AU values for (D) .
    Figure Legend Snippet: Detection and quantification of SARS-CoV-2 RBD- and N- reactive antibodies in pre-pandemic samples. (A) Representative dilution curves of seven pre-pandemic samples with SARS-CoV-2 RBD-reactive antibodies (three subjects per graph). Open and solid symbols represent buffer only coat and SARS-CoV-2 RBD coat, respectively. Arbitrary Units (AU) were calculated as described in Methods and shown beneath the respective isotype graph for diluent only and SARS-CoV-2 RBD coat. AUs for SARS-CoV-2 RBD (B) and N (C) reactive IgM, IgG, and IgA in pre-pandemic samples. Open and solid symbols represent negative and positive results, respectively, as determined by Metric 1. Enumeration of the positive samples for each isotype in the pre-pandemic cohort is shown beneath each graph with percentages of total in parentheses. (D) Correlation between AUs for IgG reactive to SARS-CoV-2 RBD and N (n=32). Values were log-transformed to obtain a parametric distribution. Statistical analyses were performed using an unpaired non-parametric Mann-Whitney t-test in (B, C) and Pearson’s correlation of normally distributed AU values for (D) .

    Techniques Used: Transformation Assay, MANN-WHITNEY

    The modified ELISA (BU ELISA) protocol exhibits low background signal at high sample concentration and use of SARS-Cov-2 RBD-recombinant antibody standard curves allows for accurate sample quantification via accounting for OD drift between experimental runs. (A) Dilution curves of buffer only coated wells from five donor samples after using an automated plate washer or the BU ELISA method of multichannel plate washing. Experiment was performed once. (B) Representative dilution curves of buffer only coated wells from 30 subjects, average and range of 1:5 sample dilution for each isotype from all subjects; IgM, IgG, and IgA were detected in individual assays. (C) Representative IgM, IgG, and IgA standard curves from 15 different experimental runs are shown. The average of all runs shown as red triangles.
    Figure Legend Snippet: The modified ELISA (BU ELISA) protocol exhibits low background signal at high sample concentration and use of SARS-Cov-2 RBD-recombinant antibody standard curves allows for accurate sample quantification via accounting for OD drift between experimental runs. (A) Dilution curves of buffer only coated wells from five donor samples after using an automated plate washer or the BU ELISA method of multichannel plate washing. Experiment was performed once. (B) Representative dilution curves of buffer only coated wells from 30 subjects, average and range of 1:5 sample dilution for each isotype from all subjects; IgM, IgG, and IgA were detected in individual assays. (C) Representative IgM, IgG, and IgA standard curves from 15 different experimental runs are shown. The average of all runs shown as red triangles.

    Techniques Used: Modification, Enzyme-linked Immunosorbent Assay, Concentration Assay, Recombinant

    SARS-CoV-2 RBD and N reactive IgG in pre-pandemic samples track with IgG recognizing analogous proteins of eCoV strains. (A) AUs of IgG reactive to RBD of NL63 and HKU1 and N of all four eCoV strains (NL63, 2293, OC43, and HKU1). (B) Correlation between SARS-CoV-2 RBD IgG levels with NL63, HKU1 RBD IgG levels in individual subjects. (C) Correlation between SARS-CoV-2 N IgG and NL63, 229E, OC43, and HKU1 N IgG levels, n=30-42. Values were log-transformed to obtain a parametric distribution. Statistical analyses were performed using Pearson’s correlation of normally distributed log transformed AU values in (B, C) and an unpaired non-parametric Mann-Whitney t-test in (A) .
    Figure Legend Snippet: SARS-CoV-2 RBD and N reactive IgG in pre-pandemic samples track with IgG recognizing analogous proteins of eCoV strains. (A) AUs of IgG reactive to RBD of NL63 and HKU1 and N of all four eCoV strains (NL63, 2293, OC43, and HKU1). (B) Correlation between SARS-CoV-2 RBD IgG levels with NL63, HKU1 RBD IgG levels in individual subjects. (C) Correlation between SARS-CoV-2 N IgG and NL63, 229E, OC43, and HKU1 N IgG levels, n=30-42. Values were log-transformed to obtain a parametric distribution. Statistical analyses were performed using Pearson’s correlation of normally distributed log transformed AU values in (B, C) and an unpaired non-parametric Mann-Whitney t-test in (A) .

    Techniques Used: Transformation Assay, MANN-WHITNEY

    Older age is associated with lower circulating antibodies reactive with SARS-CoV-2 and eCoV RBD and N antigens. Quantification of IgG reactive to RBD of NL63, HKU1, and SARS-CoV-2 and N of NL63, 229E, OC43, HKU1, and CoV-2 in pre-pandemic samples regrouped based on HIV (A) or SLE (B) disease status or age (C) . Statistical analyses were performed using an unpaired non-parametric Mann-Whitney t-test.
    Figure Legend Snippet: Older age is associated with lower circulating antibodies reactive with SARS-CoV-2 and eCoV RBD and N antigens. Quantification of IgG reactive to RBD of NL63, HKU1, and SARS-CoV-2 and N of NL63, 229E, OC43, HKU1, and CoV-2 in pre-pandemic samples regrouped based on HIV (A) or SLE (B) disease status or age (C) . Statistical analyses were performed using an unpaired non-parametric Mann-Whitney t-test.

    Techniques Used: MANN-WHITNEY

    Quantification of the relative levels of IgM, IgG, and IgA-reactive SARS-CoV-2-RBD and N antibodies from acute and convalescent SARS-CoV-2 infected subjects. (A) Arbitrary Units (AUs) of SARS-CoV-2 RBD and N reactive IgM, IgG, and IgA of acute and convalescent subjects. Open and solid symbols represent negative and positive results, respectively, as determined by our Metric 1 described in Methods. (B) Correlation between SARS-CoV-2 RBD and N IgM, IgG, and IgA log transformed AUs. Values were log-transformed to obtain a parametric distribution. (C) Quantification of SARS-CoV-2 RBD and N reactive IgM, IgG, and IgA of acute subjects regrouped based on results from Abbott’s SARS-CoV-2 IgG CMIA. Correlation between SARS-CoV-2 RBD (D) and N (E) IgM, IgG, and IgA AUs (log transformed) with the number of days post symptom (dps) onset at time of sample collection for acute subjects. Quantification of SARS-CoV-2 RBD reactive IgM and N reactive IgA (F) and RBD N reactive for IgM, IgG, and IgA (G) for pre-pandemics (n = 19) and Acutes re-classified based on Abbott test results. Light blue bars depict AU range of pre-pandemics for each respective antigen and isotype. Statistical analyses were performed using an unpaired non-parametric Mann-Whitney t-test in (A, C, F, G) and Pearson’s correlation of normally distributed log transformed AU values in (B, D, E) dps, days post symptom.
    Figure Legend Snippet: Quantification of the relative levels of IgM, IgG, and IgA-reactive SARS-CoV-2-RBD and N antibodies from acute and convalescent SARS-CoV-2 infected subjects. (A) Arbitrary Units (AUs) of SARS-CoV-2 RBD and N reactive IgM, IgG, and IgA of acute and convalescent subjects. Open and solid symbols represent negative and positive results, respectively, as determined by our Metric 1 described in Methods. (B) Correlation between SARS-CoV-2 RBD and N IgM, IgG, and IgA log transformed AUs. Values were log-transformed to obtain a parametric distribution. (C) Quantification of SARS-CoV-2 RBD and N reactive IgM, IgG, and IgA of acute subjects regrouped based on results from Abbott’s SARS-CoV-2 IgG CMIA. Correlation between SARS-CoV-2 RBD (D) and N (E) IgM, IgG, and IgA AUs (log transformed) with the number of days post symptom (dps) onset at time of sample collection for acute subjects. Quantification of SARS-CoV-2 RBD reactive IgM and N reactive IgA (F) and RBD N reactive for IgM, IgG, and IgA (G) for pre-pandemics (n = 19) and Acutes re-classified based on Abbott test results. Light blue bars depict AU range of pre-pandemics for each respective antigen and isotype. Statistical analyses were performed using an unpaired non-parametric Mann-Whitney t-test in (A, C, F, G) and Pearson’s correlation of normally distributed log transformed AU values in (B, D, E) dps, days post symptom.

    Techniques Used: Infection, Transformation Assay, MANN-WHITNEY

    11) Product Images from "A Next Generation Bivalent Human Ad5 COVID-19 Vaccine Delivering Both Spike and Nucleocapsid Antigens Elicits Th1 Dominant CD4+, CD8+ T-cell and Neutralizing Antibody Responses"

    Article Title: A Next Generation Bivalent Human Ad5 COVID-19 Vaccine Delivering Both Spike and Nucleocapsid Antigens Elicits Th1 Dominant CD4+, CD8+ T-cell and Neutralizing Antibody Responses

    Journal: bioRxiv

    doi: 10.1101/2020.07.29.227595

    cPass and Vero E6 cell SARS-CoV-2 confirm neutralization by antibodies. (a) In the cPass assay, inhibition of S RBD interaction with ACE2 was significant at both 1:20 and 1:60 dilutions of serum from hAd5 S-Fusion + N-ETSD vaccinated mice. (b) The results in the Vero E6 cell SARS-CoV-2 viral infection for mice that showed S-specific antibodies by ELISA also showed high neutralization for mice and very high neutralization for pooled sera (G4 pool, blue line) even compared to COVID-19 convalescent serum. G4 pool – mice with S-specific antibodies; M1, M2, M3, M4 – mouse ID; +C – convalescent serum; and media – media only negative control.
    Figure Legend Snippet: cPass and Vero E6 cell SARS-CoV-2 confirm neutralization by antibodies. (a) In the cPass assay, inhibition of S RBD interaction with ACE2 was significant at both 1:20 and 1:60 dilutions of serum from hAd5 S-Fusion + N-ETSD vaccinated mice. (b) The results in the Vero E6 cell SARS-CoV-2 viral infection for mice that showed S-specific antibodies by ELISA also showed high neutralization for mice and very high neutralization for pooled sera (G4 pool, blue line) even compared to COVID-19 convalescent serum. G4 pool – mice with S-specific antibodies; M1, M2, M3, M4 – mouse ID; +C – convalescent serum; and media – media only negative control.

    Techniques Used: Neutralization, Inhibition, Mouse Assay, Infection, Enzyme-linked Immunosorbent Assay, Negative Control

    12) Product Images from "Heterogeneous antibodies against SARS-CoV-2 spike receptor binding domain and nucleocapsid with implications for COVID-19 immunity"

    Article Title: Heterogeneous antibodies against SARS-CoV-2 spike receptor binding domain and nucleocapsid with implications for COVID-19 immunity

    Journal: JCI Insight

    doi: 10.1172/jci.insight.142386

    Comparison of seroconversion in patients with COVID-19 and healthy individuals. ( A ) ELISA with S-RBD protein coating and 1:100 dilution of repeated serum samples of patients with SARS-CoV-2 and healthy individuals. Absorbance normalized to the respective no antigen control for each sample at 450 nm reported. SARS-CoV-2 (blue), n = 88 (from 21 patients); HS 2017–2019 (white), n = 104; HS 2020 (white), n = 308. Arrows list consecutive serum samples evaluated for each case. Inset graphs depict the data separated based on healthy serum collected from 2017 to 2019 (left inset) and 2020 (right inset). ( B ) ELISA with N-protein coating and 1:100 dilution of the first and last serum samples of patients with SARS-CoV-2 and healthy individuals. Absorbance normalized to the respective no antigen control for each sample at 450 nm reported. SARS-CoV-2 (blue), n = 37 (from 21 patients); HS 2017–2019 (white), n = 103; HS 2020 (white), n = 308. Arrows list consecutive serum samples evaluated for each case. Inset graphs depict the data separated based on healthy serum collected from 2017 to 2019 (top inset) and 2020 (bottom inset). ( C ) Pie charts depicting percentage of samples positive for indicated antigens. SARS-CoV-2, n = 21; HS 2017–2019, n = 103; HS 2020, n = 308; non–COVID-19 samples (NCSs), n = 45; HIV, n = 7; all, n = 484.
    Figure Legend Snippet: Comparison of seroconversion in patients with COVID-19 and healthy individuals. ( A ) ELISA with S-RBD protein coating and 1:100 dilution of repeated serum samples of patients with SARS-CoV-2 and healthy individuals. Absorbance normalized to the respective no antigen control for each sample at 450 nm reported. SARS-CoV-2 (blue), n = 88 (from 21 patients); HS 2017–2019 (white), n = 104; HS 2020 (white), n = 308. Arrows list consecutive serum samples evaluated for each case. Inset graphs depict the data separated based on healthy serum collected from 2017 to 2019 (left inset) and 2020 (right inset). ( B ) ELISA with N-protein coating and 1:100 dilution of the first and last serum samples of patients with SARS-CoV-2 and healthy individuals. Absorbance normalized to the respective no antigen control for each sample at 450 nm reported. SARS-CoV-2 (blue), n = 37 (from 21 patients); HS 2017–2019 (white), n = 103; HS 2020 (white), n = 308. Arrows list consecutive serum samples evaluated for each case. Inset graphs depict the data separated based on healthy serum collected from 2017 to 2019 (top inset) and 2020 (bottom inset). ( C ) Pie charts depicting percentage of samples positive for indicated antigens. SARS-CoV-2, n = 21; HS 2017–2019, n = 103; HS 2020, n = 308; non–COVID-19 samples (NCSs), n = 45; HIV, n = 7; all, n = 484.

    Techniques Used: Enzyme-linked Immunosorbent Assay

    Detection of serum binding antibodies against SARS-CoV-2 proteins in patients with PCR-confirmed COVID-19 and healthy samples. ( A ) Timeline of COVID-19 diagnosis/ICU admittance, serum sample collection, and convalescent plasma (CP) administration. Time 0 is defined as day of COVID-19 diagnosis (PCR positive for SARS-CoV-2) and ICU admittance. Blood collections are denoted in gray and CP administration is denoted in pink. Patients were stratified based on current status (recovered, hospitalized, or deceased). Patient 29 from our cohort had symptoms but was PCR negative for SARS-CoV-2; this sample was not included in figures since there was no proof of disease. ( B ) Schematic of SARS-CoV-2 viral structure (top panel) and antigens assayed (bottom panel). S-protein, light orange; envelope protein, yellow; membrane glycoprotein, dark orange; RNA, blue; N-protein, green. Absorbance normalized to the respective no antigen control for each sample at 450 nm plotted for S-RBD (left panel), and N-protein (right panel), antigen coating with the most recent (or only) SARS-CoV-2 samples not treated with CP ( n = 21) and healthy samples collected in 2017–2019 (HS 2017–2019, n = 104 for S-RBD, n = 103 for N-protein) and 2020 (HS 2020, n = 308). Data are presented with each dot representing the mean normalized absorbance for a given serum sample; the red bar depicts the median ± interquartile range of all samples. HS, healthy sample; NC (line), negative control cutoff (see Methods). Kruskal-Wallis with Dunn’s multiple-comparisons test performed. **** P
    Figure Legend Snippet: Detection of serum binding antibodies against SARS-CoV-2 proteins in patients with PCR-confirmed COVID-19 and healthy samples. ( A ) Timeline of COVID-19 diagnosis/ICU admittance, serum sample collection, and convalescent plasma (CP) administration. Time 0 is defined as day of COVID-19 diagnosis (PCR positive for SARS-CoV-2) and ICU admittance. Blood collections are denoted in gray and CP administration is denoted in pink. Patients were stratified based on current status (recovered, hospitalized, or deceased). Patient 29 from our cohort had symptoms but was PCR negative for SARS-CoV-2; this sample was not included in figures since there was no proof of disease. ( B ) Schematic of SARS-CoV-2 viral structure (top panel) and antigens assayed (bottom panel). S-protein, light orange; envelope protein, yellow; membrane glycoprotein, dark orange; RNA, blue; N-protein, green. Absorbance normalized to the respective no antigen control for each sample at 450 nm plotted for S-RBD (left panel), and N-protein (right panel), antigen coating with the most recent (or only) SARS-CoV-2 samples not treated with CP ( n = 21) and healthy samples collected in 2017–2019 (HS 2017–2019, n = 104 for S-RBD, n = 103 for N-protein) and 2020 (HS 2020, n = 308). Data are presented with each dot representing the mean normalized absorbance for a given serum sample; the red bar depicts the median ± interquartile range of all samples. HS, healthy sample; NC (line), negative control cutoff (see Methods). Kruskal-Wallis with Dunn’s multiple-comparisons test performed. **** P

    Techniques Used: Binding Assay, Polymerase Chain Reaction, Negative Control

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    Article Snippet: Purified proteins were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis to ensure purity and appropriate molecular weights. .. SARS-CoV-2 nucleocapsid-His recombinant protein was purchased from Sino Biological (100 μg, 40588-V08B) The protein was difficult to dissolve and required treatment before S-trap preparation below with 40 μL dimethyl sulfoxide (276855—100 mL, Sigma), 126 μL 1% TFA, and 100 μL 1× S-Trap lysis buffer (5% SDS, 50 mM TEAB, pH adjusted to 7.55 using 12% phosphoric acid). ..

    Article Title: SARS-CoV-2 RapidPlex: A Graphene-Based Multiplexed Telemedicine Platform for Rapid and Low-Cost COVID-19 Diagnosis and Monitoring
    Article Snippet: CRP polyclonal antibody labeled with horseradish peroxidase (HRP) (PA1-28329) and 3,3′,5,5′-tetramethylbenzidine (TMB) colorimetric substrate was purchased from Invitrogen. .. Mouse NP monoclonal antibody (mAb) (40143-MM05), SARS-CoV-2 NP antigen (40588-V08B), SARS-CoV/SARS-CoV-2 nucleocapsid antibody, rabbit mAb (40143-R001), SARS-CoV NP antigen (HCoV-OC43; 40643-V07E), SARS-CoV-2 Spike S1-His recombinant protein (HPLC-verified) (40591-V08H), and SARS-CoV Spike S1 protein (S1 subunit, His tag) (40150-V08B1) were purchased from Sino Biological. ..

    other:

    Article Title: Clonal dissection of immunodominance and cross-reactivity of the CD4+ T cell response to SARS-CoV-2
    Article Snippet: 40588-V08B), SARS-CoV (S577A, Isolate Tor2) Spike protein (S1+S2 ECD, cat.no.

    Article Title: The Characterization of Disease Severity Associated IgG Subclasses Response in COVID-19 Patients
    Article Snippet: ProteinsSARS-CoV-2 Spike Protein (S1+S2) (cat# 40589-VO8B1), SARS-CoV-2 Spike RBD Protein (cat# 40592-V08B), SARS-CoV-2 Nucleocapsid Protein (cat# 40588-V08B) were purchased from Sino Biological (China).

    Multiplex Assay:

    Article Title: SARS-CoV-2–Specific Antibody Detection for Seroepidemiology: A Multiplex Analysis Approach Accounting for Accurate Seroprevalence
    Article Snippet: Assay Procedure The steps in assay validation were similar to recently developed bead-based multiplex immunoassays for CMV, EBV, and RSV, with minor modifications as described below [ , ]. .. For the multiplex bead-based immune assay the following antigens obtained from Sino Biological were used: SARS-CoV-2 monomeric spike S1 (40591-V08H), RBD (40592-V08B), and nucleoprotein (N) (40588-V08B). ..

    Lysis:

    Article Title: Development of a Parallel Reaction Monitoring Mass Spectrometry Assay for the Detection of SARS-CoV-2 Spike Glycoprotein and Nucleoprotein
    Article Snippet: Purified proteins were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis to ensure purity and appropriate molecular weights. .. SARS-CoV-2 nucleocapsid-His recombinant protein was purchased from Sino Biological (100 μg, 40588-V08B) The protein was difficult to dissolve and required treatment before S-trap preparation below with 40 μL dimethyl sulfoxide (276855—100 mL, Sigma), 126 μL 1% TFA, and 100 μL 1× S-Trap lysis buffer (5% SDS, 50 mM TEAB, pH adjusted to 7.55 using 12% phosphoric acid). ..

    High Performance Liquid Chromatography:

    Article Title: SARS-CoV-2 RapidPlex: A Graphene-Based Multiplexed Telemedicine Platform for Rapid and Low-Cost COVID-19 Diagnosis and Monitoring
    Article Snippet: CRP polyclonal antibody labeled with horseradish peroxidase (HRP) (PA1-28329) and 3,3′,5,5′-tetramethylbenzidine (TMB) colorimetric substrate was purchased from Invitrogen. .. Mouse NP monoclonal antibody (mAb) (40143-MM05), SARS-CoV-2 NP antigen (40588-V08B), SARS-CoV/SARS-CoV-2 nucleocapsid antibody, rabbit mAb (40143-R001), SARS-CoV NP antigen (HCoV-OC43; 40643-V07E), SARS-CoV-2 Spike S1-His recombinant protein (HPLC-verified) (40591-V08H), and SARS-CoV Spike S1 protein (S1 subunit, His tag) (40150-V08B1) were purchased from Sino Biological. ..

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    Sino Biological sars cov 2 2019 ncov nucleocapsid his recombinant protein covid 19 nucleocapsid research
    GCG suppresses <t>SARS-CoV-2</t> replication. a , b Immunofluorescence analysis of N protein in A549-hACE2-Flag cells infected with SARS-CoV-2 for 24 h ( a ). The percentage of cells with N protein foci was quantified, n = 8 biologically independent samples, 20 randomly selected views were analyzed in each sample ( b ). Scale bar, 10 μm. c 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 . d , e The inhibitory effect of GCG on the replication of SARS-CoV-2, n = 6 biologically independent samples ( d ). IC 50 was calculated, n = 5 biologically independent samples ( e ). The infection was performed after 1-h pretreatment of GCG. f , g Representative immunofluorescent images showed the inhibitory effect of GCG on SARS-CoV-2 N protein. 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 ( f ). Violin plots showing foci of cells ( n = 50 biologically independent cells) from each group, lines within the plots, with 25th, 50th, and 75th percentiles marked ( g ). h Cells were infected with SARS-CoV-2 for 1 h followed by 24-h GCG treatment, n = 3 biologically independent samples. Representative images were shown. SARS-CoV-2 was used at an MOI of 1. Hoechst (blue), nuclear staining ( a , c , f ). Error bars, mean with s.d. ( b , d , e , g , h ). Two-tailed unpaired Student’s t -test, * P
    Sars Cov 2 2019 Ncov Nucleocapsid His Recombinant Protein Covid 19 Nucleocapsid Research, supplied by Sino Biological, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov 2 2019 ncov nucleocapsid his recombinant protein covid 19 nucleocapsid research/product/Sino Biological
    Average 86 stars, based on 1 article reviews
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    sars cov 2 2019 ncov nucleocapsid his recombinant protein covid 19 nucleocapsid research - by Bioz Stars, 2021-05
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    Sino Biological sars cov 2 nucleocapsid his recombinant protein
    Sequence coverage and proteotypic target peptide selection for development of a PRM assay for the SARS <t>CoV-2</t> Spike protein and NP. (Top panel) Diagram of SARS CoV-2 recombinant spike glycoprotein showing the location of NTD, RBD, fusion peptide and heptad repeats 1 and 2, and the protease cleavage sites, His and Strep tags. The amino acid sequence is given below. Glycosylation sites are indicated in green. (Bottom panel) Diagram of SARS CoV-2 recombinant NP showing intrinsically disordered regions, RNA binding and dimerization regions. Phosphorylation sites (S) are indicated in yellow. Bold italics indicate sites where sequence coverage was not obtained. Peptides monitored in the spectral library are boxed, peptides selected for the final PRM assay are boxed and indicated in red text. Overall 97.1% of the spike protein and 77.2% of the NP sequence was obtained from the DDA analysis.
    Sars Cov 2 Nucleocapsid His Recombinant Protein, supplied by Sino Biological, used in various techniques. Bioz Stars score: 95/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sars cov 2 nucleocapsid his recombinant protein/product/Sino Biological
    Average 95 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    sars cov 2 nucleocapsid his recombinant protein - by Bioz Stars, 2021-05
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      Buy from Supplier

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    GCG suppresses SARS-CoV-2 replication. a , b Immunofluorescence analysis of N protein in A549-hACE2-Flag cells infected with SARS-CoV-2 for 24 h ( a ). The percentage of cells with N protein foci was quantified, n = 8 biologically independent samples, 20 randomly selected views were analyzed in each sample ( b ). Scale bar, 10 μm. c 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 . d , e The inhibitory effect of GCG on the replication of SARS-CoV-2, n = 6 biologically independent samples ( d ). IC 50 was calculated, n = 5 biologically independent samples ( e ). The infection was performed after 1-h pretreatment of GCG. f , g Representative immunofluorescent images showed the inhibitory effect of GCG on SARS-CoV-2 N protein. 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 ( f ). Violin plots showing foci of cells ( n = 50 biologically independent cells) from each group, lines within the plots, with 25th, 50th, and 75th percentiles marked ( g ). h Cells were infected with SARS-CoV-2 for 1 h followed by 24-h GCG treatment, n = 3 biologically independent samples. Representative images were shown. SARS-CoV-2 was used at an MOI of 1. Hoechst (blue), nuclear staining ( a , c , f ). Error bars, mean with s.d. ( b , d , e , g , h ). Two-tailed unpaired Student’s t -test, * P

    Journal: Nature Communications

    Article Title: GCG inhibits SARS-CoV-2 replication by disrupting the liquid phase condensation of its nucleocapsid protein

    doi: 10.1038/s41467-021-22297-8

    Figure Lengend Snippet: GCG suppresses SARS-CoV-2 replication. a , b Immunofluorescence analysis of N protein in A549-hACE2-Flag cells infected with SARS-CoV-2 for 24 h ( a ). The percentage of cells with N protein foci was quantified, n = 8 biologically independent samples, 20 randomly selected views were analyzed in each sample ( b ). Scale bar, 10 μm. c 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 . d , e The inhibitory effect of GCG on the replication of SARS-CoV-2, n = 6 biologically independent samples ( d ). IC 50 was calculated, n = 5 biologically independent samples ( e ). The infection was performed after 1-h pretreatment of GCG. f , g Representative immunofluorescent images showed the inhibitory effect of GCG on SARS-CoV-2 N protein. 3D images were obtained by Zeiss LSM 880 confocal microscope and reconstituted by Volocity 6.1.1 ( f ). Violin plots showing foci of cells ( n = 50 biologically independent cells) from each group, lines within the plots, with 25th, 50th, and 75th percentiles marked ( g ). h Cells were infected with SARS-CoV-2 for 1 h followed by 24-h GCG treatment, n = 3 biologically independent samples. Representative images were shown. SARS-CoV-2 was used at an MOI of 1. Hoechst (blue), nuclear staining ( a , c , f ). Error bars, mean with s.d. ( b , d , e , g , h ). Two-tailed unpaired Student’s t -test, * P

    Article Snippet: The recombinant N protein (40588-V08B) was from Sino-Biological.

    Techniques: Immunofluorescence, Infection, Microscopy, Staining, Two Tailed Test

    RNA triggers the LLPS of N protein. a Schematic drawing of SARS-CoV-2. b IDR scores of 29 proteins encoded by SARS-CoV-2 genome. FUS and mEGFP are positive and negative controls, respectively. IUPred2 and ANCHOR2 were used as prediction tools. c Time-lapse imaging of N-mEGFP protein (20 μM) in the presence of Cy5-labeled 60-nt vRNA (100 ng/μl), scale bar, 10 μm. d Representative fluorescent images of N-mEGFP-vRNA (60 nt) condensates fusion from a time-lapse movie, scale bar, 3 μm. e – g LLPS of N-mEGFP protein (20 μM) in the presence of indicated concentrations of 60-nt vRNA, scale bar, 10 μm ( e ). The partition coefficient of fluorescence intensity per droplet ( f ) and the partition coefficient of total fluorescence intensity in each view ( g ) were calculated. From left to right, n = 209, 1170, 1026, 1170 droplets ( f ) from 10 randomly selected views ( g ). h , i FRAP analysis of vRNA-induced liquid droplets of N-mEGFP protein, scale bar, 2 μm ( h ), and quantification of fluorescence intensity recovery of a photobleached N-mEGFP protein, n = 3 biologically independent experiments ( i ). The white dotted circle in h indicated the region of photobleaching. 20 μM N-mEGFP protein and 100 ng/μl 60-nt vRNA were used. Error bars, mean with s.d. ( f , g , i ). Two-tailed unpaired Student’s t -test ( f , g ), **** P

    Journal: Nature Communications

    Article Title: GCG inhibits SARS-CoV-2 replication by disrupting the liquid phase condensation of its nucleocapsid protein

    doi: 10.1038/s41467-021-22297-8

    Figure Lengend Snippet: RNA triggers the LLPS of N protein. a Schematic drawing of SARS-CoV-2. b IDR scores of 29 proteins encoded by SARS-CoV-2 genome. FUS and mEGFP are positive and negative controls, respectively. IUPred2 and ANCHOR2 were used as prediction tools. c Time-lapse imaging of N-mEGFP protein (20 μM) in the presence of Cy5-labeled 60-nt vRNA (100 ng/μl), scale bar, 10 μm. d Representative fluorescent images of N-mEGFP-vRNA (60 nt) condensates fusion from a time-lapse movie, scale bar, 3 μm. e – g LLPS of N-mEGFP protein (20 μM) in the presence of indicated concentrations of 60-nt vRNA, scale bar, 10 μm ( e ). The partition coefficient of fluorescence intensity per droplet ( f ) and the partition coefficient of total fluorescence intensity in each view ( g ) were calculated. From left to right, n = 209, 1170, 1026, 1170 droplets ( f ) from 10 randomly selected views ( g ). h , i FRAP analysis of vRNA-induced liquid droplets of N-mEGFP protein, scale bar, 2 μm ( h ), and quantification of fluorescence intensity recovery of a photobleached N-mEGFP protein, n = 3 biologically independent experiments ( i ). The white dotted circle in h indicated the region of photobleaching. 20 μM N-mEGFP protein and 100 ng/μl 60-nt vRNA were used. Error bars, mean with s.d. ( f , g , i ). Two-tailed unpaired Student’s t -test ( f , g ), **** P

    Article Snippet: The recombinant N protein (40588-V08B) was from Sino-Biological.

    Techniques: Imaging, Labeling, Fluorescence, Two Tailed Test

    NR203K/G204R gained greater ability to undergo RNA-induced LLPS. a Distribution of N gene variants among 100,849 SARS-CoV-2 genomes obtained from GISAID database. Colors indicated the nucleotide variability numbers from 100,849 genomes. The high-frequency trio-nucleotide polymorphism variant (GGG-to-AAC) is shown. b Coomassie brilliant blue-stained SDS-PAGE gel of purified variants of N-mEGFP protein. c – e LLPS of different N-mEGFP variants, in the presence of 50 ng/μl 60-nt vRNA ( c ). The partition coefficient of fluorescence intensity per droplet ( d ) and the partition coefficient of total fluorescence intensity in each view ( e ) were calculated. From left to right, n = 1232, 803, 897, 431 droplets ( d ) from 10 randomly selected views ( e ). f , g Time-lapse imaging of N R203/G204 -mEGFP and N R203K/G204R -mEGFP proteins (20 μM) in the presence of Cy5-labeled 60-nt vRNA (40 ng/μl) ( f ), and the partition coefficient ( n = 8 randomly selected views) of total fluorescence intensity in each view ( g ). Scale bars, 10 μm ( c , f ). Error bars, mean with s.d. ( d , e ) and mean with s.e.m. ( g ). Two-tailed unpaired Student’s t -test ( d , e ), **** P

    Journal: Nature Communications

    Article Title: GCG inhibits SARS-CoV-2 replication by disrupting the liquid phase condensation of its nucleocapsid protein

    doi: 10.1038/s41467-021-22297-8

    Figure Lengend Snippet: NR203K/G204R gained greater ability to undergo RNA-induced LLPS. a Distribution of N gene variants among 100,849 SARS-CoV-2 genomes obtained from GISAID database. Colors indicated the nucleotide variability numbers from 100,849 genomes. The high-frequency trio-nucleotide polymorphism variant (GGG-to-AAC) is shown. b Coomassie brilliant blue-stained SDS-PAGE gel of purified variants of N-mEGFP protein. c – e LLPS of different N-mEGFP variants, in the presence of 50 ng/μl 60-nt vRNA ( c ). The partition coefficient of fluorescence intensity per droplet ( d ) and the partition coefficient of total fluorescence intensity in each view ( e ) were calculated. From left to right, n = 1232, 803, 897, 431 droplets ( d ) from 10 randomly selected views ( e ). f , g Time-lapse imaging of N R203/G204 -mEGFP and N R203K/G204R -mEGFP proteins (20 μM) in the presence of Cy5-labeled 60-nt vRNA (40 ng/μl) ( f ), and the partition coefficient ( n = 8 randomly selected views) of total fluorescence intensity in each view ( g ). Scale bars, 10 μm ( c , f ). Error bars, mean with s.d. ( d , e ) and mean with s.e.m. ( g ). Two-tailed unpaired Student’s t -test ( d , e ), **** P

    Article Snippet: The recombinant N protein (40588-V08B) was from Sino-Biological.

    Techniques: Variant Assay, Staining, SDS Page, Purification, Fluorescence, Imaging, Labeling, Two Tailed Test

    Evaluation of Analytical Sensor Performance for the Detection of Physiological Levels of Target COVID-19 Biomarkers (A) Scheme of sensor preparation for detection of SARS-CoV-2 NP and CRP based on double-sandwich and sandwich assay configurations, respectively. CAb, capture antibody; DAb, detector antibody; DAb 2 , secondary detector antibody; HRP, horseradish peroxidase. (B and C) Calibration curves constructed for NP (B) and CRP (C) detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3). (D) Scheme of sensor preparation for detection of S1-IgG and S1-IgM isotypes based on direct assay configurations. (E and F) Calibration curves constructed for S1-IgG (E) and S1-IgM (F) isotype detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3).

    Journal: Matter

    Article Title: SARS-CoV-2 RapidPlex: A Graphene-Based Multiplexed Telemedicine Platform for Rapid and Low-Cost COVID-19 Diagnosis and Monitoring

    doi: 10.1016/j.matt.2020.09.027

    Figure Lengend Snippet: Evaluation of Analytical Sensor Performance for the Detection of Physiological Levels of Target COVID-19 Biomarkers (A) Scheme of sensor preparation for detection of SARS-CoV-2 NP and CRP based on double-sandwich and sandwich assay configurations, respectively. CAb, capture antibody; DAb, detector antibody; DAb 2 , secondary detector antibody; HRP, horseradish peroxidase. (B and C) Calibration curves constructed for NP (B) and CRP (C) detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3). (D) Scheme of sensor preparation for detection of S1-IgG and S1-IgM isotypes based on direct assay configurations. (E and F) Calibration curves constructed for S1-IgG (E) and S1-IgM (F) isotype detection in PBS (pH 7.4) supplemented with 1.0% BSA. Data are presented as mean ± SD (n = 3).

    Article Snippet: Mouse NP monoclonal antibody (mAb) (40143-MM05), SARS-CoV-2 NP antigen (40588-V08B), SARS-CoV/SARS-CoV-2 nucleocapsid antibody, rabbit mAb (40143-R001), SARS-CoV NP antigen (HCoV-OC43; 40643-V07E), SARS-CoV-2 Spike S1-His recombinant protein (HPLC-verified) (40591-V08H), and SARS-CoV Spike S1 protein (S1 subunit, His tag) (40150-V08B1) were purchased from Sino Biological.

    Techniques: Construct

    Application of SARS-CoV-2 RapidPlex in SARS-CoV-2 Detection in Blood and Saliva Samples from COVID-19-Positive and -Negative Subjects (A and B) Experimental readings obtained with SARS-CoV-2 RapidPlex after 10-min incubation of the sensor array with serum samples from a representative COVID-19 RT-PCR-negative (A) and -positive (B) patient. (C) Signal of individual sensor obtained after 1-min incubation with a serum sample from a COVID-19-positive patient (dark color) versus the signal obtained after 10-min incubation with a serum sample from a COVID-19-negative patient (light color). (D) Box-and-whisker plot of measured signal-to-blank ratios (S/B) for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 6) serum samples. (E) Box-and-whisker plot of measured S/B for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 3) saliva samples. (F) CRP levels in diluted serum samples plotted against given COVID-19 symptom severity, with “Healthy” referring to COVID-19-negative patient samples (n = 7). Positive COVID-19 patients were classified according to disease severity as asymptomatic (n = 2), mild (n = 5), and moderate (n = 2).

    Journal: Matter

    Article Title: SARS-CoV-2 RapidPlex: A Graphene-Based Multiplexed Telemedicine Platform for Rapid and Low-Cost COVID-19 Diagnosis and Monitoring

    doi: 10.1016/j.matt.2020.09.027

    Figure Lengend Snippet: Application of SARS-CoV-2 RapidPlex in SARS-CoV-2 Detection in Blood and Saliva Samples from COVID-19-Positive and -Negative Subjects (A and B) Experimental readings obtained with SARS-CoV-2 RapidPlex after 10-min incubation of the sensor array with serum samples from a representative COVID-19 RT-PCR-negative (A) and -positive (B) patient. (C) Signal of individual sensor obtained after 1-min incubation with a serum sample from a COVID-19-positive patient (dark color) versus the signal obtained after 10-min incubation with a serum sample from a COVID-19-negative patient (light color). (D) Box-and-whisker plot of measured signal-to-blank ratios (S/B) for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 6) serum samples. (E) Box-and-whisker plot of measured S/B for NP, S1-IgG, S1-IgM, and CRP in RT-PCR-confirmed COVID-19-positive (n = 5) and -negative (n = 3) saliva samples. (F) CRP levels in diluted serum samples plotted against given COVID-19 symptom severity, with “Healthy” referring to COVID-19-negative patient samples (n = 7). Positive COVID-19 patients were classified according to disease severity as asymptomatic (n = 2), mild (n = 5), and moderate (n = 2).

    Article Snippet: Mouse NP monoclonal antibody (mAb) (40143-MM05), SARS-CoV-2 NP antigen (40588-V08B), SARS-CoV/SARS-CoV-2 nucleocapsid antibody, rabbit mAb (40143-R001), SARS-CoV NP antigen (HCoV-OC43; 40643-V07E), SARS-CoV-2 Spike S1-His recombinant protein (HPLC-verified) (40591-V08H), and SARS-CoV Spike S1 protein (S1 subunit, His tag) (40150-V08B1) were purchased from Sino Biological.

    Techniques: Incubation, Reverse Transcription Polymerase Chain Reaction, Whisker Assay

    COVID-19 infection boosts NF-κB signaling pathway. ( A ) The left (MILD) and right (SEVERE) sides of box represent the mean fold change in mRNA levels, compared with HC. The NF-κB signaling pathway was adopted from KEGG database (accession number: hsa04064). ( B ) The expression levels of IRF3 , TLR3 , TLR7 , TLR8 and TLR9 were represented by FPKM. Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, NF = nuclear factor, MILD = mild/moderate, SEVERE = severe/critical, HC = healthy controls, KEGG = Kyoto Encyclopedia of Genes and Genomes, FPKM = fragments per kilobase exon-model per million reads mapped, FDR = false discovery rate, TLR = toll-like receptor, IRF = interferon regulatory factor. * P

    Journal: Journal of Korean Medical Science

    Article Title: COVID-19 Patients Upregulate Toll-like Receptor 4-mediated Inflammatory Signaling That Mimics Bacterial Sepsis

    doi: 10.3346/jkms.2020.35.e343

    Figure Lengend Snippet: COVID-19 infection boosts NF-κB signaling pathway. ( A ) The left (MILD) and right (SEVERE) sides of box represent the mean fold change in mRNA levels, compared with HC. The NF-κB signaling pathway was adopted from KEGG database (accession number: hsa04064). ( B ) The expression levels of IRF3 , TLR3 , TLR7 , TLR8 and TLR9 were represented by FPKM. Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, NF = nuclear factor, MILD = mild/moderate, SEVERE = severe/critical, HC = healthy controls, KEGG = Kyoto Encyclopedia of Genes and Genomes, FPKM = fragments per kilobase exon-model per million reads mapped, FDR = false discovery rate, TLR = toll-like receptor, IRF = interferon regulatory factor. * P

    Article Snippet: SARS-CoV-2 (2019-nCoV) NC-His recombinant protein (cat. No. 40588-V08B), Spike S1-His recombinant protein (cat. No. 40591-V08H), Spike S2 extracellular domain (ECD)-His recombinant protein (cat. No. 40590-V08B), and Spike RBD-His recombinant protein (cat. No. 40592-V08H) were purchased from Sino Biological, Beijing, China.

    Techniques: Infection, Expressing, MANN-WHITNEY

    Transcriptome analysis reveals that immune gene expression profiles of COVID-19 patients are distinct to HC. ( A ) Schematic diagram of the immune transcriptome analysis in this study. ( B ) A result of principal component analysis of log2-transformed 579 immune gene expression levels. ( C ) The scatter plots representing 579 immune genes with the log2-transformed FPKM for COVID-19 patients compared to HC. ( D ) The ten most significantly enriched KEGG pathways of the 298 DEiGs from COVID-19 patients compared to HC. ( E ) Log2-transformed fold changes of chemokine and chemokine receptor genes from MILD (x-axis) and SEVERE (y-axis) vs. HC. ( F ) Expression levels (FPKM) of marked chemokines in (E). Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, HC = healthy controls, FPKM = fragments per kilobase exon-model per million reads mapped, KEGG = Kyoto Encyclopedia of Genes and Genomes, DEiG = differentially expressed immune gene, MILD = mild/moderate, SEVERE = severe/critical, FDR = false discovery rate, IBD = inflammatory bowel disease, TNF = tumor necrosis factor, CCL = C-C motif chemokine ligand, CXCL = C-X-C motif chemokine ligand. * P

    Journal: Journal of Korean Medical Science

    Article Title: COVID-19 Patients Upregulate Toll-like Receptor 4-mediated Inflammatory Signaling That Mimics Bacterial Sepsis

    doi: 10.3346/jkms.2020.35.e343

    Figure Lengend Snippet: Transcriptome analysis reveals that immune gene expression profiles of COVID-19 patients are distinct to HC. ( A ) Schematic diagram of the immune transcriptome analysis in this study. ( B ) A result of principal component analysis of log2-transformed 579 immune gene expression levels. ( C ) The scatter plots representing 579 immune genes with the log2-transformed FPKM for COVID-19 patients compared to HC. ( D ) The ten most significantly enriched KEGG pathways of the 298 DEiGs from COVID-19 patients compared to HC. ( E ) Log2-transformed fold changes of chemokine and chemokine receptor genes from MILD (x-axis) and SEVERE (y-axis) vs. HC. ( F ) Expression levels (FPKM) of marked chemokines in (E). Error bar indicates standard error of mean. P values were calculated using Mann-Whitney U test and adjusted P values (FDR) were shown. COVID-19 = coronavirus disease 2019, HC = healthy controls, FPKM = fragments per kilobase exon-model per million reads mapped, KEGG = Kyoto Encyclopedia of Genes and Genomes, DEiG = differentially expressed immune gene, MILD = mild/moderate, SEVERE = severe/critical, FDR = false discovery rate, IBD = inflammatory bowel disease, TNF = tumor necrosis factor, CCL = C-C motif chemokine ligand, CXCL = C-X-C motif chemokine ligand. * P

    Article Snippet: SARS-CoV-2 (2019-nCoV) NC-His recombinant protein (cat. No. 40588-V08B), Spike S1-His recombinant protein (cat. No. 40591-V08H), Spike S2 extracellular domain (ECD)-His recombinant protein (cat. No. 40590-V08B), and Spike RBD-His recombinant protein (cat. No. 40592-V08H) were purchased from Sino Biological, Beijing, China.

    Techniques: Expressing, Transformation Assay, MANN-WHITNEY

    Sequence coverage and proteotypic target peptide selection for development of a PRM assay for the SARS CoV-2 Spike protein and NP. (Top panel) Diagram of SARS CoV-2 recombinant spike glycoprotein showing the location of NTD, RBD, fusion peptide and heptad repeats 1 and 2, and the protease cleavage sites, His and Strep tags. The amino acid sequence is given below. Glycosylation sites are indicated in green. (Bottom panel) Diagram of SARS CoV-2 recombinant NP showing intrinsically disordered regions, RNA binding and dimerization regions. Phosphorylation sites (S) are indicated in yellow. Bold italics indicate sites where sequence coverage was not obtained. Peptides monitored in the spectral library are boxed, peptides selected for the final PRM assay are boxed and indicated in red text. Overall 97.1% of the spike protein and 77.2% of the NP sequence was obtained from the DDA analysis.

    Journal: Analytical Chemistry

    Article Title: Development of a Parallel Reaction Monitoring Mass Spectrometry Assay for the Detection of SARS-CoV-2 Spike Glycoprotein and Nucleoprotein

    doi: 10.1021/acs.analchem.0c02288

    Figure Lengend Snippet: Sequence coverage and proteotypic target peptide selection for development of a PRM assay for the SARS CoV-2 Spike protein and NP. (Top panel) Diagram of SARS CoV-2 recombinant spike glycoprotein showing the location of NTD, RBD, fusion peptide and heptad repeats 1 and 2, and the protease cleavage sites, His and Strep tags. The amino acid sequence is given below. Glycosylation sites are indicated in green. (Bottom panel) Diagram of SARS CoV-2 recombinant NP showing intrinsically disordered regions, RNA binding and dimerization regions. Phosphorylation sites (S) are indicated in yellow. Bold italics indicate sites where sequence coverage was not obtained. Peptides monitored in the spectral library are boxed, peptides selected for the final PRM assay are boxed and indicated in red text. Overall 97.1% of the spike protein and 77.2% of the NP sequence was obtained from the DDA analysis.

    Article Snippet: SARS-CoV-2 nucleocapsid-His recombinant protein was purchased from Sino Biological (100 μg, 40588-V08B) The protein was difficult to dissolve and required treatment before S-trap preparation below with 40 μL dimethyl sulfoxide (276855—100 mL, Sigma), 126 μL 1% TFA, and 100 μL 1× S-Trap lysis buffer (5% SDS, 50 mM TEAB, pH adjusted to 7.55 using 12% phosphoric acid).

    Techniques: Sequencing, Selection, Recombinant, RNA Binding Assay

    Chromatograms and calibration curves for two best target peptides used in the PRM assay for SARS CoV-2 spike protein and nuceloprotein. The summed area under curve values for the top four transitions of each peptide were taken to generate calibration curves for quantitation. The right panels display chromatograms obtained for each of transitions shown in different colors for (A) DQVILLNK (NP) and (B) FQTLLALHR (S). Three technical replicates were run on two separate days. The chromatograms on the left of each panel show a low and high standard from the SARS CoV-2 S and NP in a mucin background. Calibration curves were constructed from the PRM data (top right) and zoomed in (bottom right) displaying mean values at the low end of the curve to show the LOD (left dotted line) and LOQ (right dotted line).

    Journal: Analytical Chemistry

    Article Title: Development of a Parallel Reaction Monitoring Mass Spectrometry Assay for the Detection of SARS-CoV-2 Spike Glycoprotein and Nucleoprotein

    doi: 10.1021/acs.analchem.0c02288

    Figure Lengend Snippet: Chromatograms and calibration curves for two best target peptides used in the PRM assay for SARS CoV-2 spike protein and nuceloprotein. The summed area under curve values for the top four transitions of each peptide were taken to generate calibration curves for quantitation. The right panels display chromatograms obtained for each of transitions shown in different colors for (A) DQVILLNK (NP) and (B) FQTLLALHR (S). Three technical replicates were run on two separate days. The chromatograms on the left of each panel show a low and high standard from the SARS CoV-2 S and NP in a mucin background. Calibration curves were constructed from the PRM data (top right) and zoomed in (bottom right) displaying mean values at the low end of the curve to show the LOD (left dotted line) and LOQ (right dotted line).

    Article Snippet: SARS-CoV-2 nucleocapsid-His recombinant protein was purchased from Sino Biological (100 μg, 40588-V08B) The protein was difficult to dissolve and required treatment before S-trap preparation below with 40 μL dimethyl sulfoxide (276855—100 mL, Sigma), 126 μL 1% TFA, and 100 μL 1× S-Trap lysis buffer (5% SDS, 50 mM TEAB, pH adjusted to 7.55 using 12% phosphoric acid).

    Techniques: Quantitation Assay, Construct

    PRM assay results of mock (SARS-CoV-2 spiked) samples. Three biological replicates processed on different days and averaged from three technical replicates from each mock sample were evaluated using the calibration curves for the two best performing peptides (A) DQVILLNK and (B) FQTLLALHR. The samples represent the spiked-in amounts; low (3.125 μL) and high (12.5 μL) of inactivated SARS-CoV-2 virions into in vitro derived mucus. Tables below display the average calculated amol amounts obtained on each day along with the interday mean and % CV. The dotted line indicates the calculated LOD and the dashed line indicated the LOQ determined from the calibration curves generated for each peptide.

    Journal: Analytical Chemistry

    Article Title: Development of a Parallel Reaction Monitoring Mass Spectrometry Assay for the Detection of SARS-CoV-2 Spike Glycoprotein and Nucleoprotein

    doi: 10.1021/acs.analchem.0c02288

    Figure Lengend Snippet: PRM assay results of mock (SARS-CoV-2 spiked) samples. Three biological replicates processed on different days and averaged from three technical replicates from each mock sample were evaluated using the calibration curves for the two best performing peptides (A) DQVILLNK and (B) FQTLLALHR. The samples represent the spiked-in amounts; low (3.125 μL) and high (12.5 μL) of inactivated SARS-CoV-2 virions into in vitro derived mucus. Tables below display the average calculated amol amounts obtained on each day along with the interday mean and % CV. The dotted line indicates the calculated LOD and the dashed line indicated the LOQ determined from the calibration curves generated for each peptide.

    Article Snippet: SARS-CoV-2 nucleocapsid-His recombinant protein was purchased from Sino Biological (100 μg, 40588-V08B) The protein was difficult to dissolve and required treatment before S-trap preparation below with 40 μL dimethyl sulfoxide (276855—100 mL, Sigma), 126 μL 1% TFA, and 100 μL 1× S-Trap lysis buffer (5% SDS, 50 mM TEAB, pH adjusted to 7.55 using 12% phosphoric acid).

    Techniques: In Vitro, Derivative Assay, Generated

    Schematic of the workflow used to develop a PRM assay for the detection and quantitation of SARS-CoV-2 spike and NP (A) PRM assay development was performed using recombinant SARS CoV-2 spike protein and NP. Proteotypic target peptides/transitions were selected to generate a spectral library in Skyline. (B) PRM assay was then used to quantitate the SARS-CoV-2 protein levels in a mock sample that was created by adding an inactivated virus sample to in vitro derived mucus.

    Journal: Analytical Chemistry

    Article Title: Development of a Parallel Reaction Monitoring Mass Spectrometry Assay for the Detection of SARS-CoV-2 Spike Glycoprotein and Nucleoprotein

    doi: 10.1021/acs.analchem.0c02288

    Figure Lengend Snippet: Schematic of the workflow used to develop a PRM assay for the detection and quantitation of SARS-CoV-2 spike and NP (A) PRM assay development was performed using recombinant SARS CoV-2 spike protein and NP. Proteotypic target peptides/transitions were selected to generate a spectral library in Skyline. (B) PRM assay was then used to quantitate the SARS-CoV-2 protein levels in a mock sample that was created by adding an inactivated virus sample to in vitro derived mucus.

    Article Snippet: SARS-CoV-2 nucleocapsid-His recombinant protein was purchased from Sino Biological (100 μg, 40588-V08B) The protein was difficult to dissolve and required treatment before S-trap preparation below with 40 μL dimethyl sulfoxide (276855—100 mL, Sigma), 126 μL 1% TFA, and 100 μL 1× S-Trap lysis buffer (5% SDS, 50 mM TEAB, pH adjusted to 7.55 using 12% phosphoric acid).

    Techniques: Quantitation Assay, Recombinant, In Vitro, Derivative Assay