Neutralizing Antibodies Isolated by a site-directed Screening have Potent Protection on SARS-CoV-2 Infection

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1. 

   ScreenIT

   Jun 6, 2020

   SciScore for 10.1101/2020.05.03.074914: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	not detected.
   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
   The phage library (Tomlison I library and Domain antibody library) and the coated wells were blocked with PBST with 5% milk at room temperature for 1 h.	Domain suggested: None
   After blocking, biotin-labeled ACE2, blocked phage or the indicated antibodies were added to the wells and incubated at 37 ° C for 0.5 h.	ACE2 suggested: None
   Streptavidin-HRP (Thermo, Pudong New Area, Shanghai), a rabbit anti-M13 HRP antibody (for phage) (GE Healthcare, Milwaukee, WI), …

   SciScore for 10.1101/2020.05.03.074914: (What is this?)

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

   Table 1: Rigor

   Institutional Review Board Statement	not detected.
   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
   The phage library (Tomlison I library and Domain antibody library) and the coated wells were blocked with PBST with 5% milk at room temperature for 1 h.	Domain suggested: None
   After blocking, biotin-labeled ACE2, blocked phage or the indicated antibodies were added to the wells and incubated at 37 ° C for 0.5 h.	ACE2 suggested: None
   Streptavidin-HRP (Thermo, Pudong New Area, Shanghai), a rabbit anti-M13 HRP antibody (for phage) (GE Healthcare, Milwaukee, WI), or a goat anti-human Fcγ HRP antibody (Jackson ImmunoResearch, West Grove, PA) was added.	anti-M13 HRP suggested: None
   For capture ELISA, an anti-his antibody (5 μg/ml) was coated on an ELISA plate at 4 °C overnight.	anti-his suggested: None
   After washing, a goat anti-human Fcγ HRP antibody (Jackson ImmunoResearch, West Grove, PA) was added and incubated at 37 °C for 0.5 h.	anti-human Fcγ HRP suggested: None
   After washing, the cell suspension was labeled with goat anti-human PE antibody (Thermo, Pudong New Area, Shanghai) and incubated for 1 h on ice.	anti-human PE suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Cell binding and antibody blocking assays: For the cell binding assay, cell suspension of SARS-CoV-2-spike-CHO cells was incubated with the indicated antibody (5 μg/ml) for 1 h on ice and then incubated with goat anti-human PE antibody (Thermo, Pudong New Area, Shanghai) for 1 h on ice.	SARS-CoV-2-spike-CHO suggested: None
   For the antibody blocking assay, antibodies were preincubated with 2.5 μg/ml SARS-CoV-1-RBD-hFc or SARS-CoV-2-RBD-hFc at different concentrations for 1 h on ice, and then, the mixture was incubated with ACE2-CHO cells for 1 h on ice.	ACE2-CHO suggested: None
   Pseudovirus neutralization assay: To generate SARS-CoV-2 pseudovirus, we replaced the coding sequence of VSV-G protein with the sequence of SARS-CoV-2 spike in lentiviral packaging system19,20 and then co-transfect HEK293T cells with the pLVX-EGFP-Luciferase reporter gene.	HEK293T suggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)
   Viruses were amplified in Vero E6 cells and made as working stocks at 105 pfu/ml.	Vero E6 suggested: None
   Software and Algorithms
   Sentences	Resources
   The remodeled ectodomain trimers of SARS-CoV-2 spike (open state) and SARS-CoV-2 spike (closed state) were established by replacing the partially determined RBD of the ectodomain trimers of SARS-CoV-2 spike (open state) (PDB ID: 6VYB) and SARS-CoV-2 spike (closed state) (PDB ID: 6VXX) with the completely determined SARS-CoV-2-RBD (PDB ID: 6M0J) by using PyMOL, Discovery Studio and SWISS MODEL.	PyMOL suggested: (PyMOL, RRID:SCR_000305)
   All statistical analyses were conducted 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 found bar graphs of continuous data. We recommend replacing bar graphs with more informative graphics, as many different datasets can lead to the same bar graph. The actual data may suggest different conclusions from the summary statistics. For more information, please see Weissgerber et al (2015).

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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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   Read the original source
2. 

   Version published to 10.1101/2020.05.03.074914v2 on bioRxiv

   May 4, 2020
3. 

   Version published to 10.1101/2020.05.03.074914v1 on bioRxiv

   May 3, 2020