Systematic evaluation of IgG responses to SARS-CoV-2 spike protein-derived peptides for monitoring COVID-19 patients

  1. Yang Li
  2. Dan-yun Lai
  3. Qing Lei
  4. Zhao-wei Xu
  5. Feng Wang
  6. Hongyan Hou
  7. Lingyun Chen
  8. Jiaoxiang Wu
  9. Yan Ren
  10. Ming-liang Ma
  11. Bo Zhang
  12. Hong Chen
  13. Caizheng Yu
  14. Jun-biao Xue
  15. Yun-xiao Zheng
  16. Xue-ning Wang
  17. He-wei Jiang
  18. Hai-nan Zhang
  19. Huan Qi
  20. Shu-juan Guo
  21. Yandi Zhang
  22. Xiaosong Lin
  23. Zongjie Yao
  24. Pengfei Pang
  25. Dawei Shi
  26. Wei Wang
  27. Xiao Yang
  28. Jie Zhou
  29. Huiming Sheng
  30. Ziyong Sun
  31. Hong Shan
  32. Xionglin Fan
  33. Sheng-ce Tao

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Abstract

Serological tests play an essential role in monitoring and combating the COVID-19 pandemic. Recombinant spike protein (S protein), especially the S1 protein, is one of the major reagents used for serological tests. However, the high cost of S protein production and possible cross-reactivity with other human coronaviruses pose unavoidable challenges. By taking advantage of a peptide microarray with full spike protein coverage, we analyzed 2,434 sera from 858 COVID-19 patients, 63 asymptomatic patients and 610 controls collected from multiple clinical centers. Based on the results, we identified several S protein-derived 12-mer peptides that have high diagnostic performance. In particular, for monitoring the IgG response, one peptide (aa 1148–1159 or S2–78) exhibited a sensitivity (95.5%, 95% CI 93.7–96.9%) and specificity (96.7%, 95% CI 94.8–98.0%) comparable to those of the S1 protein for the detection of both symptomatic and asymptomatic COVID-19 cases. Furthermore, the diagnostic performance of the S2–78 (aa 1148–1159) IgG was successfully validated by ELISA in an independent sample cohort. A panel of four peptides, S1–93 (aa 553–564), S1–97 (aa 577–588), S1–101 (aa 601–612) and S1–105 (aa 625–636), that likely will avoid potential cross-reactivity with sera from patients infected by other coronaviruses was constructed. The peptides identified in this study may be applied independently or in combination with the S1 protein for accurate, affordable, and accessible COVID-19 diagnosis.

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  1. ScreenIT

    SciScore for 10.1101/2020.09.01.20186387: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: Patients and samples: The study was approved by the Ethical Committee of Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China (ITJ-C20200128), Institutional Ethics Review Committee of Foshan Fourth Hospital, Foshan, China (202005) and the Ethical Committee of The Fifth Affiliated Hospital of Sun Yat-sen University, Zhuhai, China (K14-2).
    Consent: Written informed consent was obtained from all participants enrolled in this
    RandomizationA few conjugates were randomly selected for examination by SDS-PAGE.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Peptide microarray fabrication: The peptide-BSA conjugates as well as S1 protein, RBD protein and N protein of SARS-CoV-2, along with the negative (BSA) and positive controls (anti-Human IgG and IgM antibody), were printed in triplicate on PATH substrate slide (Grace Bio-Labs, Oregon, USA) to generate identical arrays in a 1 x 7 (for the peptide microarray with three concentrations) or 2 x 7 subarray format (for the peptide microarray with one concentration) using Super Marathon printer (Arrayjet, UK).
    IgM
    suggested: None
    The arrays were washed with 1×PBST and bound antibodies were detected by incubating with Cy3-conjugated goat anti-human IgG and Alexa Fluor 647-conjugated donkey anti-human IgM (Jackson ImmunoResearch, PA, USA), which were diluted for 1: 1,000 in 1×PBST.
    anti-human IgG
    suggested: (Bio-Rad Cat# MCA647F, RRID:AB_808612)
    anti-human IgM
    suggested: None
    After six washes with PBST, the secondary antibody, i.e., anti-human IgG-peroxidase (Sangon Biotech, Shanghai, China) was diluted at 1:10000 and incubated at 37 ° C for 1 h, followed by eight washes.
    anti-human IgG-peroxidase
    suggested: None
    Software and Algorithms
    SentencesResources
    Graphpad 6.0 was used to generate ROC plots and calculated AUC values.
    Graphpad
    suggested: (GraphPad, RRID:SCR_000306)

    Results from OddPub: Thank you for sharing your data.

    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.

    About SciScore

    SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

  2. Version published to 10.1038/s41423-020-00612-5
  3. Version published to 10.1101/2020.09.01.20186387v1 on medRxiv