Antibody evolution to SARS-CoV-2 after single-dose Ad26.COV2.S vaccine in humans

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1. Version published to 10.1084/jem.20220732

   Jul 1, 2022
2. 

   ScreenIT

   Apr 4, 2022

   SciScore for 10.1101/2022.03.31.486548: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Ethics	Consent: All participants provided written informed consent before participation in the study and the study was conducted in accordance with Good Clinical Practice. IRB: The study was performed in compliance with all relevant ethical regulations and the protocol (DRO-1006) for studies with human participants was approved by the Institutional Review Board of the Rockefeller University.
   Sex as a biological variable	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Plates were washed 6 times with washing buffer and then incubated with anti-human IgG, IgM or IgA secondary antibody …

   SciScore for 10.1101/2022.03.31.486548: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Ethics	Consent: All participants provided written informed consent before participation in the study and the study was conducted in accordance with Good Clinical Practice. IRB: The study was performed in compliance with all relevant ethical regulations and the protocol (DRO-1006) for studies with human participants was approved by the Institutional Review Board of the Rockefeller University.
   Sex as a biological variable	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Plates were washed 6 times with washing buffer and then incubated with anti-human IgG, IgM or IgA secondary antibody conjugated to horseradish peroxidase (HRP) (Jackson ImmunoResearch 109-036-088, 109-035-129, and Sigma A0295) in blocking buffer at a 1:5,000 dilution (IgM and IgG) or 1:3,000 dilution (IgA).	anti-human IgG suggested: (Jackson ImmunoResearch Labs Cat# 109-036-088, RRID:AB_2337594) IgA suggested: None
   The half-maximal neutralization titers for plasma (NT50) or half-maximal and 90% inhibitory concentrations for monoclonal antibodies (IC50 and IC90) were determined using four-parameter nonlinear regression (least squares regression method without weighting; constraints: top=1, bottom=0) (GraphPad Prism).	IC90 suggested: None
   The enriched B cells were incubated in Flourescence-Activated Cell-sorting (FACS) buffer (1× PBS, 2% FCS, 1 mM ethylenediaminetetraacetic acid (EDTA)) with the following anti-human antibodies (all at 1:200 dilution): anti-CD20-PECy7 (BD Biosciences, 335793), anti-CD3-APC-eFluro780 (Invitrogen, 47-0037-41), anti-CD8-APC-eFluor780 (Invitrogen, 47-0086-42), anti-CD16-APC-eFluor780 (Invitrogen, 47-0168-41), anti-CD14-APC-eFluor780 (Invitrogen, 47-0149-42), as well as Zombie NIR (BioLegend, 423105) and fluorophore-labeled Wuhan-Hu-1 RBD, NTD, and ovalbumin (Ova) for 30 min on ice.	anti-CD20-PECy7 suggested: None anti-CD3-APC-eFluro780 suggested: None anti-CD8-APC-eFluor780 suggested: None anti-CD16-APC-eFluor780 suggested: None anti-CD14-APC-eFluor780 suggested: None
   For B cell phenotype analysis, in addition to above antibodies, B cells were also stained with following anti-human antibodies (all at 1:200 dilution): anti-IgD-BV650 (BD, 740594), anti-CD27-BV786 (BD biosciences, 563327), anti-CD19-BV605 (Biolegend, 302244), anti-CD71-PerCP-Cy5.5 (Biolegend, 334114), anti-IgG-PECF594 (BD, 562538), anti-IgM-AF700 (Biolegend, 314538), anti-IgA-Viogreen (Miltenyi Biotec, 130-113-481)	anti-human suggested: (Abnova Cat# H00007360-A01, RRID:AB_563327) anti-IgD-BV650 suggested: None anti-CD27-BV786 suggested: None anti-CD19-BV605 suggested: None anti-CD71-PerCP-Cy5.5 suggested: None anti-IgM-AF700 ( Biolegend , 314538) , anti-IgA-Viogreen ( Miltenyi Biotec , 130-113-481) suggested: None
   The frequency distributions of human V genes in anti-SARS-CoV-2 antibodies from this study was compared to 131,284,220 IgH and IgL sequences generated by (50) and downloaded from cAb-Rep (51), a database of human shared BCR clonotypes available at https://cab-rep.c2b2.columbia.edu/.	anti-SARS-CoV-2 suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Briefly, 293T (CRL-11268) cells were obtained from ATCC, and the cells were transfected with pNL4-3 DEnv-nanoluc and pSARS-CoV-2-SΔ19.	293T suggested: ATCC Cat# CRL-11268, RRID:CVCL_1926)
   Recombinant DNA
   Sentences	Resources
   Briefly, 293T (CRL-11268) cells were obtained from ATCC, and the cells were transfected with pNL4-3 DEnv-nanoluc and pSARS-CoV-2-SΔ19.	pNL4-3 suggested: None pSARS-CoV-2-SΔ19 suggested: None
   Software and Algorithms
   Sentences	Resources
   The average of its signal was used for normalization of all the other values on the same plate with Excel software before calculating the area under the curve using Prism V9.1 (GraphPad).	Excel suggested: None Prism suggested: (PRISM, RRID:SCR_005375) GraphPad suggested: (GraphPad Prism, RRID:SCR_002798)
   For monoclonal antibodies, the ELISA half-maximal concentration (EC50) was determined using four-parameter nonlinear regression (GraphPad Prism V9.1).	GraphPad Prism suggested: (GraphPad Prism, RRID:SCR_002798)
   3,000 units/ml RNasin Ribonuclease Inhibitors (Promega, N2615)) per well using a FACS Aria III and FACSDiva software (Becton Dickinson) for acquisition and FlowJo for analysis.	FACSDiva suggested: (BD FACSDiva Software, RRID:SCR_001456) FlowJo suggested: (FlowJo, RRID:SCR_008520)
   For B cell phenotype analysis, in addition to above antibodies, B cells were also stained with following anti-human antibodies (all at 1:200 dilution): anti-IgD-BV650 (BD, 740594), anti-CD27-BV786 (BD biosciences, 563327), anti-CD19-BV605 (Biolegend, 302244), anti-CD71-PerCP-Cy5.5 (Biolegend, 334114), anti-IgG-PECF594 (BD, 562538), anti-IgM-AF700 (Biolegend, 314538), anti-IgA-Viogreen (Miltenyi Biotec, 130-113-481)	Biolegend , 314538) , anti-IgA-Viogreen ( Miltenyi Biotec suggested: None Miltenyi suggested: (Miltenyi Biotec, RRID:SCR_008984)
   Sequence analysis was performed using MacVector.	MacVector suggested: (MacVector, RRID:SCR_015700)
   : Figures arranged in Adobe Illustrator 2022.	Adobe Illustrator suggested: (Adobe Illustrator, RRID:SCR_010279)

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   Results from OddPub: Thank you for sharing your code and data.

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   Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

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   Results from JetFighter: We did not find any issues relating to colormaps.

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   Results from rtransparent:

   • Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
   • Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
   • No protocol registration statement was detected.

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   Results from scite Reference Check: We found no unreliable references.

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3. 

   Version published to 10.1101/2022.03.31.486548v1 on bioRxiv

   Apr 1, 2022