Epitope Mapping of SARS-CoV-2 Spike Protein Reveals Distinct Antibody Binding Activity of Vaccinated and Infected Individuals

  1. Nathaniel Felbinger
  2. David Trudil
  3. Lawrence Loomis
  4. Richard Ascione
  5. Gregory Siragusa
  6. Seiji Haba
  7. Shruti Rastogi
  8. Aidan Mucci
  9. Mark Claycomb
  10. Sebastian Snowberger
  11. Brian Luke
  12. Stephen Francesconi
  13. Shirley Tsang

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Abstract

Previous studies have attempted to characterize the antibody response of individuals to the SARS-CoV-2 virus on a linear peptide level by utilizing peptide microarrays. These studies have helped to identify epitopes that have potential to be used for diagnostic tests to identify infected individuals, however, the immunological responses of individuals who have received the currently available Moderna mRNA-1273 or Pfizer BNT162b2 mRNA vaccines have not been characterized. We aimed to identify linear peptides of the SARS-CoV-2 spike protein that elicited high IgG or IgA binding activity and to compare the immunoreactivity of infected individuals to those who received both doses of either vaccines by utilizing peptide microarrays. Our results revealed peptide epitopes of significant IgG binding among recently infected individuals. Some of these peptides are located near functional domains implicated in the high infectivity of SARS-CoV-2. Vaccinated individuals lacked these distinct markers despite overall binding activity being similar.

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  1. Version published to 10.1101/2022.04.13.487697v3 on bioRxiv
  2. Version published to 10.1101/2022.04.13.487697v2 on bioRxiv
  3. ScreenIT

    SciScore for 10.1101/2022.04.13.487697: (What is this?)

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

    Table 1: Rigor

    Ethicsnot detected.
    Sex as a biological variablenot detected.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    After blocking, slides were again aspirated and probed with a mixture of fluorescent secondary antibodies that would be used for peptide binding detection consisting of Rabbit Anti-Human IgG DyLight 800 (Rockland Immunochemicals)
    Anti-Human IgG
    suggested: None
    Peptides that had significantly different fluorescent intensity values were screened out, and the remaining peptides were recognized as potential epitopes (5, 13) 1.4 ELISA Testing: In-house ELISA tests were utilized to detect levels of IgA and IgG antibodies against SARS-CoV-2 receptor-binding domain (RBD) antigen.
    SARS-CoV-2 receptor-binding domain (RBD) antigen.
    suggested: None
    Secondary detection antibody solutions of HRP conjugated polyclonal mouse anti-human IgG antibodies (SouthernBiotech) diluted 1:8000 in 1% milk PBST and HRP conjugated monoclonal mouse anti-human IgA1/IgA2 antibodies (SouthernBiotech) diluted 1:4000 in 1% milk PBST were used to detect IgG and IgA respectively.
    anti-human IgA1/IgA2
    suggested: (Thermo Fisher Scientific Cat# SA5-10222, RRID:AB_2665329)

    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:
    Using a SARS-CoV-2 peptide microarray to perform IgA epitope profiling of sera may be difficult due to the limitations previously described without significant purification and concentration of IgA from samples. However, the potential for studying IgA epitopes of SARS-CoV-2 may still be viable with the use of saliva, a bodily fluid with a high content of IgA. Previous work using the peptide microarray to study the epitopes of other viral targets have shown the IgG antibody profiles of blood and saliva were similar (45). Salivary IgA has shown potential as a reliable biomarker for SARS-CoV-2 infection and a correlation between symptom severity and levels of salivary IgA have been previously demonstrated (46). Developing a method to treat microarrays with saliva may help to study IgA binding activity to SARS-CoV-2 epitopes. Previous attempts to directly treat microarrays with saliva are limited, but we have managed to obtain reproducible results screening processed saliva at low dilutions. Our ongoing study of IgA and IgG binding of saliva samples from previously infected individuals has demonstrated antibody binding to some epitopes identified in this report. These results are preliminary, with much further analysis and validation required to draw any conclusions. Yet the screening of saliva samples may be helpful with efforts to perform IgA epitope profiling of the SARS-CoV-2 proteome and lead to development of saliva-based diagnostics, immunity determinations, and therapeuti...

    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.
    • No funding statement was detected.
    • No protocol registration statement was detected.

    Results from scite Reference Check: We found no unreliable references.

    About SciScore

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  4. Version published to 10.1101/2022.04.13.487697v1 on bioRxiv