Immunogenicity of SARS-CoV-2 trimetric spike protein associated to Poly(I:C) plus Alum

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   ScreenIT

   Oct 8, 2021

   SciScore for 10.1101/2021.10.05.461434: (What is this?)

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

   Table 1: Rigor

   Ethics	IRB: The experiments were carried out in accordance with the Ethics Committee on the Use of Animals of the Health Sciences Center of the Federal University of Rio de Janeiro (Comitê de Ética no Uso de Animais do Centro de Ciências da Saúde da Universidade Federal do Rio de Janeiro), under the protocol number:
   Sex as a biological variable	Animals: Female BALB/c mice, 6–8 weeks old (n = 5 per group), were obtained from the breeding facility of UFRJ.
   Randomization	not detected.
   Blinding	The reader was blind in respect to the source of the supernatant.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   This protein was used in this work to …

   SciScore for 10.1101/2021.10.05.461434: (What is this?)

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

   Table 1: Rigor

   Ethics	IRB: The experiments were carried out in accordance with the Ethics Committee on the Use of Animals of the Health Sciences Center of the Federal University of Rio de Janeiro (Comitê de Ética no Uso de Animais do Centro de Ciências da Saúde da Universidade Federal do Rio de Janeiro), under the protocol number:
   Sex as a biological variable	Animals: Female BALB/c mice, 6–8 weeks old (n = 5 per group), were obtained from the breeding facility of UFRJ.
   Randomization	not detected.
   Blinding	The reader was blind in respect to the source of the supernatant.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   This protein was used in this work to immunize mice and as ELISA antigen to detect anti-SARS-CoV-2 antibodies in samples from immunized animals.	anti-SARS-CoV-2 suggested: None
   Antigen-specific antibody responses: Antigen-specific IgG (Cat. No. 1015-05, Southern Biotech, AL, USA), IgA (Cat. No.1040-05, Southern Biotech, AL, USA), IgG1 (Cat. No. 1071-01, Southern Biotech, AL, USA) and IgG2a (Cat. No.1101-01, Southern Biotech, AL, USA) levels were determined by enzyme linked immunosorbent assay (ELISA) using recombinant SARS-CoV-2 S Ptn as the capture antigen.	Antigen-specific IgG suggested: None IgA suggested: (SouthernBiotech Cat# 1040-05, RRID:AB_2714213) IgG1 suggested: (SouthernBiotech Cat# 1071-01, RRID:AB_2794423) IgG2a suggested: (SouthernBiotech Cat# 1101-01, RRID:AB_2794586)
   After this incubation the ELISA plate was washed 5 times with a washing solution consisting of PBS + 0.05% Tween 20 and then the anti-mouse IgG, IgG1, IgG2a, and IgA-HRP detection antibodies (Southern Biotech, AL, USA) were added for another hour.	anti-mouse IgG suggested: None IgG1, IgG2a suggested: None
   The following antibodies were used: B220 (anti-B220-PerCP; eBiosciences), GL7 (anti-GL7-PE; BioLegend), CD38 (anti-CD38-PeCy7; BioLegend), RBD-conjugated with Alexa 488, S Ptn-conjugated with Alexa 647, TRCβ (anti-TRCβ-Pacific Blue; BioLegend), CD4 (anti-CD4-FITC; BioLegend), CD8 (anti-CD8-PeCy7; BioLegend), and IFN-γ (anti-IFN-γ-APC; BioLegend).	anti-B220-PerCP suggested: None anti-GL7-PE suggested: None CD38 suggested: None anti-CD38-PeCy7 suggested: None anti-TRCβ-Pacific suggested: None CD4 suggested: None anti-CD4-FITC suggested: (Sigma-Aldrich Cat# SAB4700062, RRID:AB_10898908) CD8 suggested: None anti-CD8-PeCy7 suggested: None IFN-γ suggested: (BioLegend Cat# 430801, RRID:AB_2893366) anti-IFN-γ-APC suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   SARS-CoV-2 was prepared in Vero E6 cells.	Vero E6 suggested: RRID:CVCL_XD71)
   Then, the samples were placed into 96-well plates with monolayers of Vero cells (2×104 cell/well) with supernatants for 1 h at 37°C.	Vero suggested: RRID:CVCL_ZW93)
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Animals: Female BALB/c mice, 6–8 weeks old (n = 5 per group), were obtained from the breeding facility of UFRJ.	BALB/c suggested: RRID:IMSR_ORNL:BALB/cRl)
   Recombinant DNA
   Sentences	Resources
   RBD Sequence: RBD (receptor-binding domain): cloning: synthetic gene S1-seq263-685 CoVID-2019_HKU-SZ-005b_2020 (MN975262) (length: 1302 bp) cloned into pET21a(+) using cloning sites NdeI and SalI purchased from Genscript Inc. (USA).	pET21a(+ suggested: RRID:Addgene_12649)
   Expression of recombinant RBD in E. coli: Chemically competent E. coli BL21(λDE3) (Novagen, USA) were transformed with 0.2 ng of the pET21a-plasmid, and positive clones were selected in an LB-agar medium containing 100 μg/mL ampicillin at 37 °C overnight.	pET21a-plasmid suggested: None
   Software and Algorithms
   Sentences	Resources
   The data analyzes were performed using the FlowJo® software vX.0.7.	FlowJo® suggested: (FlowJo, RRID:SCR_008520)
   All samples were analyzed on a Becton-Dickinson LSR-II/Fortessa flow cytometer (BD Biosciences, San Diego, CA, USA) and analyzed by using FlowJo software (Tree Star Inc.).	FlowJo suggested: (FlowJo, RRID:SCR_008520)
   Data analysis was performed using GraphPad Prism® 8.0 software.	GraphPad suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: Thank you for sharing your 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: We found the following clinical trial numbers in your paper:
   

   Identifier	Status	Title
   NCT03721718	Completed	Evaluate the Safety, Tolerability and Immunogenicity Study o…
   NCT04672291	Recruiting	Nasal Poly-ICLC (Hiltonol®) in Healthy COVID-19 Vaccinated A…

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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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   Version published to 10.1101/2021.10.05.461434 on bioRxiv

   Oct 6, 2021