Deep mutational scanning identifies SARS-CoV-2 Nucleocapsid escape mutations of currently available rapid antigen tests
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SciScore for 10.1101/2022.05.19.492641: (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 All testing was conducted in a blinded manner and results were unblinded after completion of all testing. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Cells were then washed 4 times with binding buffer followed by staining with appropriate labelled anti-Myc and PE-labelled secondary antibodies (Supplementary Table 1, antibodies and detection reagents). PE-labelled secondary antibodies ( Supplementary Table 1 ,suggested: NoneCells were washed three times and then stained with appropriate host-specific anti-IgG and anti-Myc antibodies. anti-IgGsug…SciScore for 10.1101/2022.05.19.492641: (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 All testing was conducted in a blinded manner and results were unblinded after completion of all testing. Power Analysis not detected. Cell Line Authentication not detected. Table 2: Resources
Antibodies Sentences Resources Cells were then washed 4 times with binding buffer followed by staining with appropriate labelled anti-Myc and PE-labelled secondary antibodies (Supplementary Table 1, antibodies and detection reagents). PE-labelled secondary antibodies ( Supplementary Table 1 ,suggested: NoneCells were washed three times and then stained with appropriate host-specific anti-IgG and anti-Myc antibodies. anti-IgGsuggested: NoneUsing the anti-Myc signal to account for differences in expression, diagonal gates were drawn on anti-Myc vs antibody signal plots to select the cells with the lowest 10-15% antibody signal (Supplementary Figure 2F). anti-Mycsuggested: NoneExperimental Models: Cell Lines Sentences Resources Cell culture: All surface-display experiments were performed using “Viral Production Cells” from the ThermoFisher LV-MAX Lentiviral Production system, a derivative of the HEK 293F cell line. HEK 293Fsuggested: RRID:CVCL_6642)For validation of individual mutations, HEK293 cells were transfected with Wuhan or mutants plasmids using the protocol optimized in the LV-MAX lentiviral production system (no packaging vectors were supplied in this case). HEK293suggested: CLS Cat# 300192/p777_HEK293, RRID:CVCL_0045)Recombinant DNA Sentences Resources This sequence was cloned into pLVX-IRES-ZsGreen1 (TakaraBio) at the EcoRI and NotI sites using Gibson assembly (NEB 2x Gibson Assembly Master Mix). pLVX-IRES-ZsGreen1suggested: NoneThe resulting construct was packaged into a lentivirus using the packaging vectors psPAX2 and pMD2G and the LV-MAX lentiviral production system (ThermoFisher). pMD2Gsuggested: NoneThe resulting DNA was assmbled into pLVX-IRES-ZsGreen at the EcoRI and NotI sites using Gibson assembly (NEB). pLVX-IRES-ZsGreensuggested: NoneThe resulting replicate libraries were packaged into a lentiviral library using the packaging vectors psPAX2 and pMD2G and the LV-MAX lentiviral production system (ThermoFisher). psPAX2suggested: RRID:Addgene_12260)Software and Algorithms Sentences Resources Sequencing data analysis and calculation of escape scores: Sequences were analyzed using custom scripts in Python and R. Pythonsuggested: (IPython, RRID:SCR_001658)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: We detected the following sentences addressing limitations in the study:We overcome these limitations by employing mammalian surface-display to directly probe antibody recognition of the full-length antigen and using a mutational library to generate binding measurements for all possible antigen mutations. It is important to note that our approach does not directly determine an antibody’s epitope – the area of physical contact between antibody and antigen. Most escape mutations will be at the binding interface and reduce affinity through direct mechanisms, especially in the case of linear epitopes. Some mutations, however, will reduce binding indirectly through allostery. This is especially important for antibodies recognizing three-dimensional epitopes since mutations far away from the binding site may reduce the stability of the protein or affect local structure at the epitope allosterically. Thus, while escape mutations mapped onto the structure of the antigen identify localized surface patches, they represent a combination of the epitope and additional sites that are important for maintaining the conformational integrity and accessibility of the epitope. This approach thus goes beyond the notion of an epitope and, instead provides a mutational profile reminiscent of a fingerprint of the antibody on the antigen. These fingerprints are highly valuable in the evaluation of antibodies and other detection reagents such as nanobodies or DNA aptamers used in a diagnostic test regarding their ability to detect variants of a rapidly mutating viral anti...
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: We did not find any issues relating to colormaps.
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.
Results from scite Reference Check: We found no unreliable references.
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