Single domain shark VNAR antibodies neutralize SARS‐CoV‐2 infection in vitro

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1. Version published to 10.1096/fj.202100986rr

   Oct 12, 2021
2. 

   ScreenIT

   Jun 10, 2021

   SciScore for 10.1101/2021.06.08.447530: (What is this?)

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

   Table 1: Rigor

   Ethics	not detected.
   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
   Anti-M13-HRP antibody (Sino Biological) was used for detection at 1:4000 dilution in blocking buffer.	Anti-M13-HRP suggested: None
   Plates were blocked for 1 hr at RT with 5% bovine serum albumin (BSA) in PBS before a 1 hr incubation with serially diluted VNAR-hFc antibodies or ACE2.	ACE2 suggested: None
   The plates were washed, and binding was detected with an anti-human IgG (Fc specific) (1:5000 dilution, Sigma-Aldrich, A0170) or anti-FLAG (1:1000 dilution, Sigma-Aldrich, A8592) …

   SciScore for 10.1101/2021.06.08.447530: (What is this?)

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

   Table 1: Rigor

   Ethics	not detected.
   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
   Anti-M13-HRP antibody (Sino Biological) was used for detection at 1:4000 dilution in blocking buffer.	Anti-M13-HRP suggested: None
   Plates were blocked for 1 hr at RT with 5% bovine serum albumin (BSA) in PBS before a 1 hr incubation with serially diluted VNAR-hFc antibodies or ACE2.	ACE2 suggested: None
   The plates were washed, and binding was detected with an anti-human IgG (Fc specific) (1:5000 dilution, Sigma-Aldrich, A0170) or anti-FLAG (1:1000 dilution, Sigma-Aldrich, A8592) HRP-conjugated antibodies for VNAR-hFc antibodies or ACE2, respectively.	anti-human IgG suggested: (Sigma-Aldrich Cat# A0170, RRID:AB_257868) anti-FLAG suggested: (Sigma-Aldrich Cat# A8592, RRID:AB_439702)
   ACE2 binding was measured with an anti-FLAG HRP conjugated antibody (Sigma-Aldrich, A8592) diluted 1:1000.	anti-FLAG HRP suggested: None A8592 suggested: (Sigma-Aldrich Cat# A8592, RRID:AB_439702)
   After blocking in 5% BSA in PBS for 1 hr, 50 nM biotinylated S1-RBD alone or premixed with 500 nM secondary (competitor) VNAR-hFc antibodies were added to the plate pairwise.	VNAR-hFc suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Expi293 cells were transiently transfected as VNAR human Fc fusions (VNAR-hFc) and produced following the manufacturer’s manual.	Expi293 suggested: RRID:CVCL_D615)
   Vero CCL81 cells were seeded at a cell density of 100,000 cells per well in 48-well plates and incubated at 37°C in serum free OptiPro SFM (Thermo Fisher Scientific) for 24 hr before infection.	Vero CCL81 suggested: None
   Recombinant DNA
   Sentences	Resources
   The ACE2 (UniProt entry: Q9BYF1) ectodomain (18-740) was synthesized with FLAG and 6xHis tags at the C-terminal end separated by a single G4S linker before cloning into the pFUSE vector.	pFUSE suggested: None
   Flow cytometry: Expi293 cells were transiently transfected with ACE2 cloned into the pCMV3-C-GFPSpark expression vector (Sino Biological).	pCMV3-C-GFPSpark suggested: None
   Software and Algorithms
   Sentences	Resources
   After gating on cells expressing ACE2-GFP fusion protein in FL1 channel, median fluorescence intensity (MFI) of cells positive for S1 binding was used to determine IC50 by 4-parametric non-linear regression analysis using GraphPad Prism 8.0.	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: 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 10.1101/2021.06.08.447530v1 on bioRxiv

   Jun 8, 2021