Mechanism of a COVID-19 nanoparticle vaccine candidate that elicits a broadly neutralizing antibody response to SARS-CoV-2 variants

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   ScreenIT

   Mar 29, 2021

   SciScore for 10.1101/2021.03.26.437274: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	IACUC: Briefly, Institutional Animal Care and Use Committee (IACUC) guidelines were followed for all of the animal studies.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   All of the vaccine antigens were transiently expressed in ExpiCHO cells and purified by a CR3022 antibody column and size-exclusion chromatography (SEC) on a Superose 6 10/300 GL column as described previously (41).	CR3022 suggested: (Imported from the IEDB Cat# CR3022, RRID:AB_2848080)
   Lymph node tissue sections were stained with human anti-spike …

   SciScore for 10.1101/2021.03.26.437274: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	IACUC: Briefly, Institutional Animal Care and Use Committee (IACUC) guidelines were followed for all of the animal studies.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   All of the vaccine antigens were transiently expressed in ExpiCHO cells and purified by a CR3022 antibody column and size-exclusion chromatography (SEC) on a Superose 6 10/300 GL column as described previously (41).	CR3022 suggested: (Imported from the IEDB Cat# CR3022, RRID:AB_2848080)
   Lymph node tissue sections were stained with human anti-spike antibody P2B-2F6 (45) (1:50) and biotinylated goat anti-human secondary antibody (Abcam, catalog no. ab7152, 1:300), followed by streptavidin-horseradish peroxidase reagent (Vectastain Elite ABC-HRP Kit, Vector, catalog no. PK-6100) then DAB (ImmPACT DAB, Vector, catalog no. SK-4105) to study the distribution and retention of the S2GΔHR2 spike alone and S2GΔHR2-presenting E2p and I3-01v9 SApNPs.	anti-spike suggested: None anti-human secondary antibody suggested: (Thermo Fisher Scientific Cat# MA1-82289, RRID:AB_933977)
   For immunofluorescent staining, tissue sections were stained for FDCs using anti-CD21 antibody (Abcam, catalog no. ab75985, 1:1800) followed by anti-rabbit secondary antibody conjugated with Alexa Fluor 555 (Thermo Fisher, catalog no. A21428; 1:200), for B cells using anti-B220 antibody (eBioscience, catalog no. 14-0452-82, 1:100) followed by anti-rat secondary antibody conjugated with Alexa Fluor 674 (Thermo Fisher, catalog no. A21247; 1:200), and for subcapsular sinus macrophages using anti-sialoadhesin (	anti-CD21 suggested: (Abcam Cat# ab75985, RRID:AB_1523292) anti-rabbit suggested: (GenWay Biotech Inc. Cat# GWB-674F40, RRID:AB_10283848) anti-B220 suggested: (Antibodies-Online Cat# ABIN334910, RRID:AB_10767277) anti-sialoadhesin suggested: None
   CD169) antibody (Abcam, catalog no. ab53443, 1:600) followed by anti-rat secondary antibody conjugated with Alexa Fluor 488 (Abcam, catalog no. ab150165; 1:200).	CD169 suggested: None anti-rat suggested: (Abcam Cat# ab150157, RRID:AB_2722511)
   Germinal center B cells were labeled using rat anti-GL7 antibody (FITC; BioLegend, catalog no. 144604, 1:250).	anti-GL7 suggested: None
   T Follicular helper cells were labeled using CD4 antibody (Biolegend, catalog no. 100402, 1:100) followed by anti-rat secondary antibody conjugated with Alexa Fluor 488 (Abcam, catalog no. ab150165; 1:1000) and Bcl6 antibody (Abcam, catalog no. ab220092, 1:300) followed by anti-rabbit secondary antibody conjugated with Alexa Fluor 555 (Thermo Fisher, catalog no. A21428; 1:1000).	CD4 suggested: (BioLegend Cat# 344617, RRID:AB_10559751) Bcl6 suggested: None
   The nonspecific binding of Fc receptors was blocked using anti-CD16/32 antibody (BioLegend, catalog no. 101302) on ice for 30 min.	anti-CD16/32 suggested: None
   FITC anti-mouse CD3 antibody (BioLegend, catalog no. 100204),	anti-mouse CD3 suggested: None
   Alexa Fluor 700 anti-mouse CD4 antibody (BioLegend, catalog no. 100536), PE anti-mouse/human GL7 antibody (BioLegend, catalog no. 144608)	anti-mouse CD4 suggested: None anti-mouse/human GL7 suggested: None
   Brilliant Violet 605 anti-mouse CD95 (Fas) antibody (BioLegend, catalog no. 152612)	anti-mouse CD95 suggested: None Fas suggested: None
   Brilliant Violet 421 anti-mouse CD185 (CXCR5) antibody (BioLegend, catalog no. 145511), and PE/Cyanine7 anti-mouse CD279 (PD-1) antibody (BioLegend, catalog no. 135216) were then mixed with the cells and placed on ice for 30 min.	anti-mouse CD185 suggested: None CXCR5 suggested: None anti-mouse CD279 suggested: None PD-1 suggested: None
   P vaccine-induced neutralizing antibody responses against SARS-CoV-2 variants of concern (VOCs). fig. S2.	S2 suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Briefly, SARS-CoV-2-pps were generated by the co-transfection of HEK293T cells with the HIV-1 pNL4-3.lucR-E-plasmid (obtained from the National Institutes of Health AIDS reagent program; https://www.aidsreagent.org/) and the expression plasmid encoding the S gene of various SARS-CoV-2 strains, including three variants: B.1.1.7, B. 1.351, and P.1	HEK293T suggested: None
   The HEK293T-hACE2 cell line (catalog no. NR-52511) and pcDNA3.1(-) vector containing the S gene of the SARS-CoV-2 isolate Wuhan-Hu-1 (catalog no. NR52420) were requested from the BEI Resources (https://www.beiresources.org/) on September 23, 2020 and used in the pseudovirus neutralization assays (42).	HEK293T-hACE2 suggested: None
   Experimental Models: Organisms/Strains
   Sentences	Resources
   BALB/c mice (6 weeks old) were purchased from The Jackson Laboratory and kept in ventilated cages in environmentally controlled rooms at The Scripps Research Institute.	BALB/c suggested: RRID:IMSR_ORNL:BALB/cRl)
   Software and Algorithms
   Sentences	Resources
   The HEK293T-hACE2 cell line (catalog no. NR-52511) and pcDNA3.1(-) vector containing the S gene of the SARS-CoV-2 isolate Wuhan-Hu-1 (catalog no. NR52420) were requested from the BEI Resources (https://www.beiresources.org/) on September 23, 2020 and used in the pseudovirus neutralization assays (42).	https://www.beiresources.org/ suggested: (BEI Resource Repository, RRID:SCR_013698)
   The bright-field images of stained S2GΔHR2 spike and S2GΔHR2-presenting SApNPs in lymph node follicles and fluorescent images of GCs were quantified using ImageJ software (National Institutes of Health) (96)	ImageJ suggested: (ImageJ, RRID:SCR_003070)
   The data were further processed using FlowJo 10 software.	FlowJo suggested: (FlowJo, RRID:SCR_008520)
   All of the statistical analyses were performed and graphs were generated using GraphPad Prism 6.01 software.	GraphPad Prism suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

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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: Please consider improving the rainbow (“jet”) colormap(s) used on page 33. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

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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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   Version published to 10.1101/2021.03.26.437274 on bioRxiv

   Mar 27, 2021