Intranasal vaccination induced cross-protective secretory IgA antibodies against SARS-CoV-2 variants with reducing the potential risk of lung eosinophilic immunopathology

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1. Version published to 10.1016/j.vaccine.2022.08.049

   Sep 1, 2022
2. 

   ScreenIT

   May 27, 2022

   SciScore for 10.1101/2022.05.24.493348: (What is this?)

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

   Table 1: Rigor

   Ethics	IACUC: All animal experiments were performed in accordance with the Guide for Animal Experiments Performed at the National Institute of Infectious Diseases (NIID) and were approved by the Animal Care and Use Committee of NIID.
   Sex as a biological variable	Immunization and sampling: Female BALB/c mice (20-24 weeks old) (Japan SLC Inc., Hamamatsu, Shizuoka, Japan) were maintained in specific pathogen-free facilities.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   The SARS-CoV-2 challenge was performed in a biosafety level 3 facility according to the Guidelines for Animal Experiments …

   SciScore for 10.1101/2022.05.24.493348: (What is this?)

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

   Table 1: Rigor

   Ethics	IACUC: All animal experiments were performed in accordance with the Guide for Animal Experiments Performed at the National Institute of Infectious Diseases (NIID) and were approved by the Animal Care and Use Committee of NIID.
   Sex as a biological variable	Immunization and sampling: Female BALB/c mice (20-24 weeks old) (Japan SLC Inc., Hamamatsu, Shizuoka, Japan) were maintained in specific pathogen-free facilities.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   The SARS-CoV-2 challenge was performed in a biosafety level 3 facility according to the Guidelines for Animal Experiments performed at NIID. 2.4. Estimation of SARS-CoV-2 S-specific antibody responses: SARS-CoV-2 S-specific antibodies were estimated using ELISA.	SARS-CoV-2 S-specific suggested: None
   IgG antibodies were detected using biotin-conjugated goat anti-mouse IgG antibody (Jackson Immunoresearch, West Grove, PA), followed by alkaline phosphatase-conjugated streptavidin (Invitrogen, CA, USA).	anti-mouse IgG suggested: None
   The S-specific IgG antibody titer was defined as the reciprocal of the highest dilution of the test sample, giving a higher absorbance than the cut-off value obtained as 2-fold mean absorbance of serial dilutions of control naive mouse serum set in each plate.	S-specific IgG suggested: None
   Quantification of S-specific IgG1 or IgG2a antibodies in the serum and IgA antibodies in nasal or lung washes was performed as previously described [23].	IgG1 suggested: None IgG2a suggested: None
   Chimeric human-mouse monoclonal IgG1, IgG2a, and IgA antibodies bearing variable regions of the S-specific human monoclonal antibody S309 [33] were used as standard antibodies for quantification.	IgA suggested: None
   Horseradish peroxidase (HRP)-conjugated polyclonal anti-mouse IgG1 antibody (Bethyl Laboratories, Montgomery, TX), anti-mouse IgG2a antibody (Bethyl Laboratories), or polyclonal anti-mouse IgA antibody (Bethyl Laboratories) were used as detection antibodies.	anti-mouse IgG1 suggested: None anti-mouse IgG2a suggested: None anti-mouse IgA suggested: None
   Briefly, in plates pre-coated with anti-mouse IFN-γ, IL-4, or IL-5 antibodies, 3 × 105 cells harvested from the spleen or cervical lymph nodes were incubated for 16 h in the presence of a peptide pool derived from the S protein of SARS-CoV-2 (a mixture of PepTivator SARS-CoV-2 Prot_S, S1, and S+; Miltenyi Biotec, Bergisch Gladbach, Germany).	anti-mouse IFN-γ suggested: None IL-4 suggested: None IL-5 suggested: None
   After washing the cells with PBS, biotin-conjugated anti-IFN-γ, IL-4, or IL-5 detection antibodies were added and incubated at RT for 2 h, followed by incubation with ALP-conjugated streptavidin at RT for 1 h.	anti-IFN-γ suggested: None
   One million cells were stained with FVD506 (Thermo Fisher Scientific) for dead cell removal and blocked with anti-mouse CD16/CD32 monoclonal antibody (BD Pharmingen, San Jose, CA).	anti-mouse CD16/CD32 suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Briefly, 50uL of QHmusX (100TCID50) and 50uL of heat-inactivated serum serially diluted by two-fold were mixed and incubated in 96-well microtiter plates for 1h at 37□, followed by the addition of 100 µL of VeroE6-TMPRSS2 cells (JCRB1819, Japanese Collection of Research Bioresources Cell Bank) [35, 36].	VeroE6-TMPRSS2 suggested: JCRB Cat# JCRB1819, RRID:CVCL_YQ49)
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Immunization and sampling: Female BALB/c mice (20-24 weeks old) (Japan SLC Inc., Hamamatsu, Shizuoka, Japan) were maintained in specific pathogen-free facilities.	BALB/c suggested: None
   Software and Algorithms
   Sentences	Resources
   Horseradish peroxidase (HRP)-conjugated polyclonal anti-mouse IgG1 antibody (Bethyl Laboratories, Montgomery, TX), anti-mouse IgG2a antibody (Bethyl Laboratories), or polyclonal anti-mouse IgA antibody (Bethyl Laboratories) were used as detection antibodies.	Bethyl Laboratories suggested: (Bethyl, RRID:SCR_013554)
   Spots formed by cytokine-secreting cells were counted and analyzed using ELISpot reader S6 Universal with ImmunoSpot 7.0 software (Cellular Technology, Ltd., Shaker Heights, OH). 2.7.	ImmunoSpot suggested: None
   Samples were analyzed with CantoII (BD Biosciences), and data were analyzed using FlowJo software version 10.8.0 (Tree Star Inc., Ashland, OR). 2.8. Quantification of SARS-CoV-2 subgenomic RNA: Total RNA was extracted from 125 µL of nasal or lung wash using ISOGEN-LS (Nippon gene, Toyko, Japan) and purified using a Maxwell RSC 48 Instrument (Promega, Madison, WI) with a Maxwell RSC miRNA Plasma and Serum Kit (Promega).	BD Biosciences suggested: (BD Biosciences, RRID:SCR_013311) FlowJo suggested: (FlowJo, RRID:SCR_008520)
   Statistical analysis: Data analysis and visualization were performed using GraphPad Prism 7.0 software (GraphPad Software Inc., San Diego, CA).	GraphPad 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: 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 10.1101/2022.05.24.493348 on bioRxiv

   May 25, 2022