Discriminating cross-reactivity in polyclonal IgG1 responses against SARS-CoV-2 variants of concern
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Abstract
Existing assays to measure antibody cross-reactivity against different SARS-CoV-2 spike (S) protein variants lack the discriminatory power to provide insights at the level of individual clones. Using a mass spectrometry-based approach we are able to monitor individual donors’ IgG1 clonal responses following a SARS-CoV-2 infection. We monitor the plasma clonal IgG1 profiles of 8 donors who had experienced an infection by either the wild type Wuhan Hu-1 virus or one of 3 VOCs (Alpha, Beta and Gamma). In these donors we chart the full plasma IgG1 repertoires as well as the IgG1 repertoires targeting the SARS-CoV-2 spike protein trimer VOC antigens. The plasma of each donor contains numerous anti-spike IgG1 antibodies, accounting for <0.1% up to almost 10% of all IgG1s. Some of these antibodies are VOC-specific whereas others do recognize multiple or even all VOCs. We show that in these polyclonal responses, each clone exhibits a distinct cross-reactivity and also distinct virus neutralization capacity. These observations support the need for a more personalized look at the antibody clonal responses to infectious diseases.
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SciScore for 10.1101/2022.02.24.481778: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IRB: , location AMC, in the Netherlands and approved by the local ethical committee of the AMC (NL 73281.018.20).
Consent: All individuals included in this study gave written informed consent before participating.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
Experimental Models: Cell Lines Sentences Resources All S constructs were verified by Sanger sequencing and the protein was subsequently produced in human embryonic kidney (HEK) 293F cells (Thermo Fisher Scientific) and purified as previously described (4, 8). HEKsuggested: RRID:CVCL_6642)293Fsuggested: RRID:CVCL_D615)Pse… SciScore for 10.1101/2022.02.24.481778: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Ethics IRB: , location AMC, in the Netherlands and approved by the local ethical committee of the AMC (NL 73281.018.20).
Consent: All individuals included in this study gave written informed consent before participating.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
Experimental Models: Cell Lines Sentences Resources All S constructs were verified by Sanger sequencing and the protein was subsequently produced in human embryonic kidney (HEK) 293F cells (Thermo Fisher Scientific) and purified as previously described (4, 8). HEKsuggested: RRID:CVCL_6642)293Fsuggested: RRID:CVCL_D615)Pseudo-viruses were produced by co-transfecting the SARS-CoV-2-S expression plasmid with the pHIV-1NL43 ΔEnv-NanoLuc reporter virus plasmid in HEK293T cells (ATCC, CRL-11268), as previously described (13). HEK293Tsuggested: ATCC Cat# CRL-11268, RRID:CVCL_1926)Shortly, HEK293T/ACE2 cells, kindly provided by Dr. Paul Bieniasz (13), were seeded at a density of 20,000 cells/well in a 96-well plate coated with 50 μg/mL poly-L-lysine one day prior to the start of the neutralization assay. HEK293T/ACE2suggested: NoneRecombinant DNA Sentences Resources The genes were ordered as gBlock gene fragments (Integrated DNA Technologies) and cloned Pst I/Not I in a pPPI4 expression vector containing a hexahistidine (his) tag with Gibson Assembly (Thermo Fisher Scientific). pPPI4suggested: NonePseudo-virus construction: The WT, Alpha, Beta and Gamma pseudovirus S constructs were ordered as gBlock gene fragments (Integrated DNA Technologies) and cloned using SacI and Apal in the pCR3 SARS-CoV-2-SΔ19 expression plasmid (13) using Gibson Assembly (ThermoFisher). pCR3 SARS-CoV-2-SΔ19suggested: NonePseudo-viruses were produced by co-transfecting the SARS-CoV-2-S expression plasmid with the pHIV-1NL43 ΔEnv-NanoLuc reporter virus plasmid in HEK293T cells (ATCC, CRL-11268), as previously described (13). pHIV-1NL43 ΔEnv-NanoLucsuggested: NoneSoftware and Algorithms Sentences Resources The LC was directly coupled to an Orbitrap Exploris 480 mass spectrometer with BioPharma option (Thermo Fisher Scientific, Bremen, Germany). BioPharmasuggested: (TransCelerate BioPharma, RRID:SCR_003728)Data analysis: The retention times and masses of each of the Fab molecules were retrieved from the generated RAW files using BioPharmaFinder 3.2 (Thermo Scientific). BioPharmaFindersuggested: NoneFurther data analysis was performed using in-house scripts using Python 3.8.3 (with libraries: Pandas 1.0.5, Numpy 1.18.5, Scipy 1.5.0, matplotlib 3.2.2 and seaborn 0.11.0). Pythonsuggested: (IPython, RRID:SCR_001658)Numpysuggested: (NumPy, RRID:SCR_008633)Scipysuggested: (SciPy, RRID:SCR_008058)matplotlibsuggested: (MatPlotLib, RRID:SCR_008624)The inhibitory concentration (IC50) and neutralization titers (ID50) were determined as the NAb concentration and plasmadilution at which infectivity was inhibited by 50%, respectively, using a non-linear regression curve fit (GraphPad Prism software version 8.3). 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: 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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