Human-IgG-Neutralizing Monoclonal Antibodies Block the SARS-CoV-2 Infection
This article has been Reviewed by the following groups
Listed in
- Evaluated articles (ScreenIT)
Abstract
Article activity feed
-
-
SciScore for 10.1101/2020.05.19.104117: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: The overall study was reviewed and approved by the SHAPHC Ethics Committee (approval no. 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 fluorescently labeled S1 bait was previously prepared by incubating 5 μg of His tag-S1 protein with Anti His tag antibody-PE for at least 1 hr at 4 °C in the dark. Anti His tag antibody-PE for at least 1suggested: NonePCR products were then purified using DNA FragSelect XP Magnetic Beads (SMART-Lifesciences) and cloned into human IgG1, lambda or kappa expression plasmids … SciScore for 10.1101/2020.05.19.104117: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: The overall study was reviewed and approved by the SHAPHC Ethics Committee (approval no. 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 fluorescently labeled S1 bait was previously prepared by incubating 5 μg of His tag-S1 protein with Anti His tag antibody-PE for at least 1 hr at 4 °C in the dark. Anti His tag antibody-PE for at least 1suggested: NonePCR products were then purified using DNA FragSelect XP Magnetic Beads (SMART-Lifesciences) and cloned into human IgG1, lambda or kappa expression plasmids for antibody expression by seamless cloning method (see below). human IgG1suggested: NoneHRP-conjugated anti-human IgG Fab antibody (Sigma) was added at the dilution of 1:10,000 in PBST containing 3% BSA (Sangon Biotech) and incubated at 37 C for 0.5 h. HRP-conjugated anti-human IgG Fab antibodysuggested: Noneanti-human IgGsuggested: NoneBriefly, 10 thousand cells in 100 μl were incubated with indicated antibodies for 30 min at room temperature, after twice washes then PE-labeled goat anti-human IgG-Fc antibody was added (1:5,000; Abcam) for 30 min, followed by flow cytometry analyses. anti-human IgG-Fcsuggested: NoneFor Figure S3, to test S protein expression on cell membrane, recombinant ACE2-Cter-6XHis labeled by rabbit anti-His-PE antibody in 1.2:1 (n:n) ratio was added to 10 thousand cells expressing S protein. anti-His-PEsuggested: (Miltenyi Biotec Cat# 130-092-691, RRID:AB_1103227)Experimental Models: Cell Lines Sentences Resources Cell lines and Viruses: Vero E6, A549-Spike (A549 expressing SARS-CoV-2 S protein), and A549-ACE2 (A549 expressing human ACE2) cell lines were supplied by Shanghai Public Health Clinical Center, Fudan University Vero E6suggested: NoneA549-Spikesuggested: NoneA549suggested: NoneA549-ACE2suggested: NoneHEK293E cells were transfected using polyethylenimine (PEI, Sigma), after 4-5 days of cell culture, antibodies purification was processed from supernatants. HEK293Esuggested: NoneFlow cytometry assays: For Figure 4, flow cytometry analyses were performed to detect the binding abilities of antibodies to Spike protein in HEK293T cells freshly expressing of SARS-CoV-2, SARS-CoV and MERS-CoV. HEK293Tsuggested: NCBI_Iran Cat# C498, RRID:CVCL_0063)Briefly, pseudovirus were generated by co-transfection of 293T cells with pNL4-3. 293Tsuggested: NoneSoftware and Algorithms Sentences Resources Sequences were analyzed using IMGT/ V-QUEST (http://www.imgt.org/IMGT_vquest) and IgBlast (IgBLAST, http://www.ncbi.nlm.nih.gov/igblast). IgBLASTsuggested: (IgBLAST, RRID:SCR_002873)Expression and purification of human monoclonal antibodies: The antibody VH/VL and constant region genes were then amplified and cloned into expression vector pcDNA3.4 using SMART Assembly Cloning Kit (SMART-Lifesciences), subsequently antibodies plasmids were amplified in competent cells (SMART-Lifesciences). SMARTsuggested: (SMART, RRID:SCR_005026)The curves and EC50 were analyzed by GraphPad Prism 8.0. GraphPad Prismsuggested: (GraphPad Prism, RRID:SCR_002798)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).
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.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: Please consider improving the rainbow (“jet”) colormap(s) used on pages 28 and 31. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.
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.
-
-
-