Journal: Cell reports. Medicine

Article Title: Safety and immunogenicity of SARS-CoV-2 self-amplifying RNA vaccine expressing an anchored RBD: A randomized, observer-blind phase 1 study.

doi: 10.1016/j.xcrm.2023.101134

Figure Lengend Snippet: Figure 3. SARS-CoV-2 IgG and neutralizing antibody responses (A and B) Serum IgG titers against wild-type SARS- CoV-2 RBD protein (A) and pseudovirus neutralizing antibody titers (ID50) against SARS-CoV-2 variants (B) for non-elderly (left side) and elderly (right side) participants (n = 10 per dose group, and n = 6 for the placebo group). Participants received one injection of VLPCOV-01 (0.3, 1.0, or 3.0 mg), 30 mg BNT162b2, or placebo on day 1 (week 0). Logarithmic values are reported as geometric mean titers for serum IgG and neutralizing antibody against pseudovirus. Bars indicate 95% CIs. (C) The correlation between serum neutralizing antibody titers against pseudovirus Wuhan (wild type) and IgG antibody titers against SARS-CoV-2 RBD protein following booster vaccination. Pearson’s product-moment correlation coefficient and p value were calculated following log trans- formation of source data (r = 0.950, p < 0.001).

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse anti-human CD154-FITC (clone: TRAP1) BD Biosciences cat# 555699; RRID: AB_396049 Mouse anti-human CD3-BUV615 (clone: SP34-2) BD Biosciences cat# 751249; RRID: AB_2875266 Mouse anti-human CD4-PE-Cy5.5 (clone: S3.5) Thermo Ficher Scientific cat# MHCD0418; RRID: AB_10376013 Mouse anti-human CD8-BUV563 (clone: RPA-T8) BD Biosciences cat# 612914; RRID: AB_2870200 Mouse anti-human CD27-PE-Cy5 (clone: 1A4CD27) Beckman coulter cat# 6607107 Mouse anti-human CD45RO-BUV805 (clone: UCHL1) BD Biosciences cat# 748367; RRID: AB_2872786 Mouse anti-human IFN-g-BV786 (clone: 4S.B3) BioLegend cat# 502542; RRID: AB_2563882 Mouse anti-human TNF-BV650 (clone: MAb11) BioLegend cat# 502938; RRID: AB_2562741 Rat anti-human IL-13-BV421 (clone: JES10-5A2) BD Biosciences cat# 563580; RRID: AB_2738290 Mouse anti-human IL-21-Ax647 (clone: 3A3-N21) BD Biosciences cat# 560493; RRID: AB_1645421 Mouse anti-human IL-4-PE-Cy7 (clone:8D4-8) BD Biosciences cat# 560672; RRID: AB_1727547 Mouse anti-human IL-17A-BV605 (clone: BL168) BioLegend cat# 512326; RRID: AB_2563887 Rat anti-human IL-2-BUV737 (clone: MQ117H12) BD Biosciences cat# 612836 Mouse anti-human CD107A-BV711 (clone: H4A3) BioLegend cat# 328640; RRID: AB_2565840 Mouse anti-human MIP1b-Alexa700 (clone: D21-1351) BD Biosciences cat# 561278; RRID: AB_10612008 Anti-SARS-CoV-2 Spike RBD Neutralizing Antibody, Human IgG1 (AS35) ACROBiosystems cat# SAD-S35 Anti-SARS-CoV-2 Spike RBD Neutralizing Antibody, Human IgG2 (AS35) ACROBiosystems cat# SAD-S66 Anti-SARS-CoV-2 Spike RBD Neutralizing Antibody, Human IgG3 (AS35) ACROBiosystems cat# SAD-S67 Anti-SARS-CoV-2 Spike RBD Neutralizing Antibody, Human IgG4 (AS35) ACROBiosystems cat# SAD-S68 Mouse anti-Human IgG1 Fc Secondary Antibody, HRP Thermo Fisher Scientific cat# MH1715; RRID: AB_2539710 Mouse Anti-Human IgG2 Fc-BIOT HP6002 SouthernBiotech cat# 9070-08; RRID: AB_2796638 Mouse Anti-Human IgG3 Hinge-BIOT HP6050 SouthernBiotech cat# 9210-08; RRID: AB_2796700 Mouse Anti-Human IgG4 Fc-BIOT HP6025 SouthernBiotech cat# 9200-08; RRID: AB_2796692 Biological samples Human PBMCs This study This study Chemicals, peptides, and recombinant proteins Benzonase Nuclease, Purity >90% MERCK Millipore cat# 70746 (Continued on next page) Cell Reports Medicine 4, 101134, August 15, 2023 e1

Techniques: Injection

Journal: bioRxiv

Article Title: Combinatorial approach with mass spectrometry and lectin microarray dissected glycoproteomic features of virion-derived spike protein of SARS-CoV-2

doi: 10.1101/2021.04.10.439300

Figure Lengend Snippet: Amino acid sequences of S proteins from seven viruses belonging to subgenus Sarbecovirus were aligned with MUSCLE and N-!P-(S|T) sequons are highlighted. Sequons conserved across all the seven viruses are colored yellow. Sequons observed in SARS-CoV-2 but not conserved across all the members are colored green. Sequons not observed in SARS-CoV-2 are colored gray. ACE2 binding sites are indicated in red boxes. S1/S2 and S2’ proteolytic sites are shown with open red arrowheads.

Article Snippet: Rabbit monoclonal anti-SARS-CoV-2 spike S1 antibody, 007 (Cat# 40150-R007), and mouse monoclonal anti-SARS-CoV-2 spike neutralizing antibody, 43 (Cat# 40591-V05H1), were purchased from Sino Biological (Beijing, China).

Techniques: Binding Assay

Journal: Emerging Microbes & Infections

Article Title: Establishment of replication-competent vesicular stomatitis virus-based recombinant viruses suitable for SARS-CoV-2 entry and neutralization assays

doi: 10.1080/22221751.2020.1830715

Figure Lengend Snippet: Characterization of rVSV-eGFP-SARS-CoV-2. (A) Schematic diagrams showing genome organization in rVSV vectors. The original VSV G gene was replaced by SARS-CoV-2 S gene to generate rVSV-eGFP-SARS-CoV-2. eGFP was inserted into the first position of the genome. (B) Lysate of rVSV-eGFP-SARS-CoV-2 producing cells (Lane 1) and purified rVSV-eGFP-SARS-CoV-2 (Lane 2) were analysed by western blot using an antibody recognizing the RBD domain of the S protein. (C) Images of mock- (upper panels) or rVSV-eGFP-SARS-CoV-2-infected (lower panels) Vero cells stained with Hoechst to label the nuclei (blue). The GFP signal is shown in green. The overlay of blue and green is shown to visualize syncytia. (D) The titers of neutralizing antibodies (NAbs) in three serum samples from COVID-19 convalescent patients were determined against rVSV-eGFP-SARS-CoV-2 by a focus reduction neutralization test. rVSV-eGFP-EBOV (Ebola virus) were used as a control virus. (E) Growth kinetics of rVSV-eGFP-SARS-CoV-2 in Vero cells (MOI=0.0l). Viral titers were measured by a focus-forming assay and expressed as focus forming units per ml (FFU/ml). Error bars in (D-E) indicate standard deviation of the mean (n=3). (F) Plaque morphology of rVSVs in Vero cells. The plaques of rVSV-eGFP-EBOV (upper panel) or rVSV-eGFP-SARS-CoV-2 (middle panel) at 72 h post-infection indicated by yellow arrowhead were visualized under microscope. Lower panel: Mock-infected cells (control). The data represent three independent experiments.

Article Snippet: Rabbit anti-SARS-CoV-2 S-RBD polyclonal antibody (Cat. 40150-RP01) and SARS-CoV-2 Spike RBD domain (Cat: 40592-V08B-B) were purchased from Sino Biological Inc. (Beijing, China).

Techniques: Purification, Western Blot, Infection, Staining, Neutralization, Focus Forming Assay, Standard Deviation, Microscopy

Journal: Emerging Microbes & Infections

Article Title: Establishment of replication-competent vesicular stomatitis virus-based recombinant viruses suitable for SARS-CoV-2 entry and neutralization assays

doi: 10.1080/22221751.2020.1830715

Figure Lengend Snippet: Characteristics of rVSV-eGFP-SARS-CoV-2 entry. (A) Infectivity of rVSV-eGFP-SARS-CoV-2 and rVSV-eGFP-G on various cell types. Infectivity was quantified by counting GFP positive cells in the above nine cell lines. The infectious titers were expressed as log 10 FFU/ml. The limit of detection was 10 FFU/ml. Titers below 10 FFU/ml (★) are indicated. (B) Inhibition of rVSV-eGFP-SARS-CoV-2 infection by NH 4 Cl. Vero cells were infected with rVSV-eGFP-SARS-CoV-2 or rVSV-eGFP-EBOV at MOI of 0.01 after treatment with different concentrations of NH 4 Cl. The results are expressed as the percentage of the control (without NH 4 Cl treatment). (C) Effect of SARS-CoV-2 RBD fragment on the infectivity of rVSV-eGFP-SARS-CoV-2. All infection experiments were performed in triplicate. Error bars in (A–C) indicate standard deviation of the mean (n=3).

Article Snippet: Rabbit anti-SARS-CoV-2 S-RBD polyclonal antibody (Cat. 40150-RP01) and SARS-CoV-2 Spike RBD domain (Cat: 40592-V08B-B) were purchased from Sino Biological Inc. (Beijing, China).

Techniques: Infection, Inhibition, Standard Deviation

Journal: Emerging Microbes & Infections

Article Title: Establishment of replication-competent vesicular stomatitis virus-based recombinant viruses suitable for SARS-CoV-2 entry and neutralization assays

doi: 10.1080/22221751.2020.1830715

Figure Lengend Snippet: Comparison of rVSV-eGFP-SARS-CoV-2-based neutralization assay with pseudotype and live viruses. (A) IC 50 values for 8 human mAbs against rVSV-eGFP-SARS-CoV-2 and VSV pseudotype. NAb titers of the mAbs against rVSV-eGFP-SARS-CoV-2 was evaluated by a focus reduction neutralization test. NAb titers against pseudovirus was evaluated by pseudovirus based neutralization assay (PBNA). IC 50 was calculated by the Reed-Muench method. (B) Correlation of IC 50 values in (A) between pseudovirus- and rVSV-based assays. (C) IC 50 values for six serum samples from COVID-19 convalescent patients against rVSV-eGFP-SARS-CoV-2 and SARS-CoV-2. NAb titers of the serum samples against SARS-CoV-2 was determined by plaque reduction neutralization test, and IC 50 was calculated by the Reed-Muench method. (D) Correlation of IC 50 values in COVID-19 patients measured by rVSV-eGFP-SARS-CoV-2 and SARS-CoV-2.

Article Snippet: Rabbit anti-SARS-CoV-2 S-RBD polyclonal antibody (Cat. 40150-RP01) and SARS-CoV-2 Spike RBD domain (Cat: 40592-V08B-B) were purchased from Sino Biological Inc. (Beijing, China).

Techniques: Neutralization, Endpoint Dilution Assay, Plaque Reduction Neutralization Test

Journal: Emerging Microbes & Infections

Article Title: Establishment of replication-competent vesicular stomatitis virus-based recombinant viruses suitable for SARS-CoV-2 entry and neutralization assays

doi: 10.1080/22221751.2020.1830715

Figure Lengend Snippet: Validation of rVSV-eGFP-SARS-CoV-2-based neutralization assay. (A) Representative inhibition curves of two positive and two negative human serum samples. (B) The limit of detection (LOD) of the assay. A subset of negative serum samples from 20 mice and 4 humans was tested against rVSV-eGFP-SARS-CoV-2 using the focus reduction neutralization test. (C) Reproducibility of the neutralization assay. Pooled positive serum samples (n=2) were run six times per experiment in total of three experiments to measure reproducibility. Individual IC 50 values were plotted to evaluate the intra- and inter-assay variability. Error bars in (A–C) indicate standard deviation of the mean.

Article Snippet: Rabbit anti-SARS-CoV-2 S-RBD polyclonal antibody (Cat. 40150-RP01) and SARS-CoV-2 Spike RBD domain (Cat: 40592-V08B-B) were purchased from Sino Biological Inc. (Beijing, China).

Techniques: Neutralization, Inhibition, Inter Assay, Standard Deviation

Journal: Microbiology Spectrum

Article Title: Comprehensive Comparison of Seven SARS-CoV-2-Specific Surrogate Virus Neutralization and Anti-Spike IgG Antibody Assays Using a Live-Virus Neutralization Assay as a Reference

doi: 10.1128/spectrum.02314-22

Figure Lengend Snippet: Correlation analyses of sVNTs and nAb titers measured by in-house live-virus NT. Spearman correlation coefficients ( r ) and coefficients of determination ( R 2 ) are indicated. (A) cPass (GenScript); (B) SARS-CoV-2-NeutraLISA (Euroimmun); (C) ACE2-RBD neutralization assay (DiaPro); (D) TECO SARS-CoV-2 neutralization antibody assay (TECOmedical). Symbols represent samples from different cohorts, as indicated at the top. The dotted lines indicate the cutoffs of the specific assays.

Article Snippet: The four antibody assays with the highest AUC included three sVNTs: the SARS-CoV-2-NeutraLISA (Euroimmun) with an AUC of 0.92, cPass (GenScript) with an AUC of 0.92, and the TECO SARS-CoV-2 neutralization antibody assay (TECOmedical) with an AUC of 0.90. shows the specific cutoffs calculated using ROC analyses for the commercial antibody assays that would be optimal to detect a nAb titer of ≥80 with a specificity of >90%, >95%, and >99%.

Techniques: Neutralization

Journal: Microbiology Spectrum

Article Title: Comprehensive Comparison of Seven SARS-CoV-2-Specific Surrogate Virus Neutralization and Anti-Spike IgG Antibody Assays Using a Live-Virus Neutralization Assay as a Reference

doi: 10.1128/spectrum.02314-22

Figure Lengend Snippet: Correlation analyses of the IgG antibody binding tests and nAb titers measured with an in-house live-virus NT. Spearman correlation coefficients ( r ) and coefficients of determination ( R 2 ) are indicated. (A) Anti-SARS-CoV-2 QuantiVac ELISA (Euroimmun); (B) Liaison SARS-CoV-2 TrimericS IgG CLIA (DiaSorin); (C and D) SARS-CoV-2 ViraChip IgG microarray (Viramed Biotech) RBD (C) and S1-directed IgG (D). Symbols represent samples from different cohorts, as indicated at the top. The dotted lines indicate the cutoffs of the specific assays.

Article Snippet: The four antibody assays with the highest AUC included three sVNTs: the SARS-CoV-2-NeutraLISA (Euroimmun) with an AUC of 0.92, cPass (GenScript) with an AUC of 0.92, and the TECO SARS-CoV-2 neutralization antibody assay (TECOmedical) with an AUC of 0.90. shows the specific cutoffs calculated using ROC analyses for the commercial antibody assays that would be optimal to detect a nAb titer of ≥80 with a specificity of >90%, >95%, and >99%.

Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Microarray

Journal: Microbiology Spectrum

Article Title: Comprehensive Comparison of Seven SARS-CoV-2-Specific Surrogate Virus Neutralization and Anti-Spike IgG Antibody Assays Using a Live-Virus Neutralization Assay as a Reference

doi: 10.1128/spectrum.02314-22

Figure Lengend Snippet: Longitudinal changes in neutralizing antibodies measured by live-virus NT and sVNTs. Samples from 34 convalescent individuals for whom the first sample was taken <90 days postonset (dpo) and an additional sample ≥90 days postonset were tested with an NT and all sVNTs. (A) cPass (GenScript); (B) SARS-CoV-2-NeutraLISA (Euroimmun); (C) ACE2-RBD neutralization assay (DiaPro); (D) TECO SARS-CoV-2 neutralization antibody assay (TECOmedical). The dotted lines indicate the cutoffs of the specific assays. Connected lines show medians. Comparison of paired values was performed by the Wilcoxon test. Gray brackets show the results for NT values, and black brackets show those for the respective commercial antibody assay. ***, P ≤ 0.001; ****, P ≤ 0.0001; ns, not significant. IH, inhibition.

Article Snippet: The four antibody assays with the highest AUC included three sVNTs: the SARS-CoV-2-NeutraLISA (Euroimmun) with an AUC of 0.92, cPass (GenScript) with an AUC of 0.92, and the TECO SARS-CoV-2 neutralization antibody assay (TECOmedical) with an AUC of 0.90. shows the specific cutoffs calculated using ROC analyses for the commercial antibody assays that would be optimal to detect a nAb titer of ≥80 with a specificity of >90%, >95%, and >99%.

Techniques: Neutralization, Inhibition

Journal: Microbiology Spectrum

Article Title: Comprehensive Comparison of Seven SARS-CoV-2-Specific Surrogate Virus Neutralization and Anti-Spike IgG Antibody Assays Using a Live-Virus Neutralization Assay as a Reference

doi: 10.1128/spectrum.02314-22

Figure Lengend Snippet: Rise in nAb values between pre- and post-COVID-19 vaccination samples measured by live-virus NT and sVNTs for 20 convalescent individuals. (A) cPass (GenScript); (B) SARS-CoV-2-NeutraLISA (Euroimmun); (C) ACE2-RBD neutralization assay (DiaPro); (D) TECO SARS-CoV-2 neutralization antibody assay (TECOmedical). The dotted lines indicate the cutoffs of the specific assays. Connected lines show medians. Comparison of paired values was performed by the Wilcoxon test. Gray brackets show the results for NT values, and black brackets show those for the respective commercial antibody assay. ****, P ≤ 0.0001.

Article Snippet: The four antibody assays with the highest AUC included three sVNTs: the SARS-CoV-2-NeutraLISA (Euroimmun) with an AUC of 0.92, cPass (GenScript) with an AUC of 0.92, and the TECO SARS-CoV-2 neutralization antibody assay (TECOmedical) with an AUC of 0.90. shows the specific cutoffs calculated using ROC analyses for the commercial antibody assays that would be optimal to detect a nAb titer of ≥80 with a specificity of >90%, >95%, and >99%.

Techniques: Neutralization