A potent SARS-CoV-2 antibody neutralizes Omicron variant by disassembling the spike trimer

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   ScreenIT

   Mar 25, 2022

   SciScore for 10.1101/2022.03.21.485243: (What is this?)

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

   Table 1: Rigor

   NIH rigor criteria are not applicable to paper type.

   Table 2: Resources

   Experimental Models: Cell Lines
   Sentences	Resources
   Expression and Purification of recombinant antibodies: A pair of plasmids separately expressing the heavy- and the light-chain of antibodies were transiently co-transfected into HEK293F cells.	HEK293F suggested: RRID:CVCL_6642)
   Serial 1/3 dilutions of antibodies were incubated with pseudoviruses at 37°C for 1 hour, and then the mixtures were added in ACE2 expressed Huh-7 cells (104 per well in 96-well plates).	Huh-7 suggested: None
   Recombinant DNA
   Sentences	Resources
   Production of recombinant D614G and variants spike ectodomain: The gene encoding SARS-CoV-2 S ectodomain (residues 1-1208, …

   SciScore for 10.1101/2022.03.21.485243: (What is this?)

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

   Table 1: Rigor

   NIH rigor criteria are not applicable to paper type.

   Table 2: Resources

   Experimental Models: Cell Lines
   Sentences	Resources
   Expression and Purification of recombinant antibodies: A pair of plasmids separately expressing the heavy- and the light-chain of antibodies were transiently co-transfected into HEK293F cells.	HEK293F suggested: RRID:CVCL_6642)
   Serial 1/3 dilutions of antibodies were incubated with pseudoviruses at 37°C for 1 hour, and then the mixtures were added in ACE2 expressed Huh-7 cells (104 per well in 96-well plates).	Huh-7 suggested: None
   Recombinant DNA
   Sentences	Resources
   Production of recombinant D614G and variants spike ectodomain: The gene encoding SARS-CoV-2 S ectodomain (residues 1-1208, Gene Bank: MN908947) was synthesized (GeneScript) and cloned into mammalian expression constructs pcDNA-3.1, proline substitutions at residues 986 and 987, “GSAS” substitution at furin cleavage site (residues 682-685), a T4 fibritin trimerization motif, an HRV3C protease cleavage site, a TwinStrepTag, and an 8XHisTag at C-terminal were introduced simultaneously by MultiS one step cloning kit (Vazyme).	pcDNA-3.1 suggested: None
   Using the SARS-CoV-2 WT plasmid as the template, mutations such as D614G, B.1.17 (Del 69H70V, N501Y, P681H, S982A, Del 145Y, A570D, T716I, D1118H), B.1.351 (K417N, E484K, N501Y) and P1 (K417T, E484K, N501Y) was introduced by Multi Site-Directed Mutagenesis Kit (Yeasen).	SARS-CoV-2 WT suggested: None
   The expression plasmid of Omicron S with HexaPro mutations(26), “GSAS” substitution at furin cleavage site (residues 682-285), a T4 fibritin trimerization motif, a TwinStrep Tag, and a C-terminal 8 × His Tag was constructed into pcDNA3.1 vector by MultiS one step cloning kit (Vazyme).	pcDNA3.1 suggested: RRID:Addgene_79663)
   Briefly, whole spike glycoprotein sequences of SARS-CoV, wild type or variants of SARS-CoV-2 were inserted into the vector of pcDNA3.1+ and severally co-transfected into 293 T cells (ATCC, Manassas, VA, USA) with a defective HIV-1 genome that encodes luciferase reporter.	pcDNA3.1+ suggested: RRID:Addgene_117272)
   Software and Algorithms
   Sentences	Resources
   A nonlinear regression analysis was performed on the resulting curves using Prism (GraphPad) to calculate half-maximal inhibitory concentration (IC50) values.	Prism suggested: (PRISM, RRID:SCR_005375) GraphPad suggested: (GraphPad Prism, RRID:SCR_002798)
   Automated data acquisition was carried out with SerialEM software(27).	SerialEM suggested: (SerialEM, RRID:SCR_017293)
   Cryo-EM image processing: All the data processing was carried out using either modules on, or through, RELION v3.0 and cryoSPARC(28).	RELION suggested: (RELION, RRID:SCR_016274)
   For S-15 complex (Fig.S10), a total of 6,943 movie stacks was binned 2 × 2, dose weighted, and motion corrected using MotionCor2(29).	MotionCor2 suggested: (MotionCor2, RRID:SCR_016499)
   After blob-picking in cryoSPARC and 2D classification, trimer and monomer particles were observed.	cryoSPARC suggested: (cryoSPARC, RRID:SCR_016501)
   For model building of S-15 and S-60, the apo-S trimer model and the antibody (15 and 60) model generated by swiss-model were fitted into the map using UCSF Chimera(31) and followed by manually adjustment in COOT(33), as well as real space refinement in Phenix.	Phenix suggested: (Phenix, RRID:SCR_014224)
   Model validation was performed using MolProbity.	MolProbity suggested: (MolProbity, RRID:SCR_014226)

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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: 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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   Version published to 10.1101/2022.03.21.485243v1 on bioRxiv

   Mar 22, 2022