SARS-CoV-2 proteome microarray for global profiling of COVID-19 specific IgG and IgM responses

  1. He-wei Jiang
  2. Yang Li
  3. Hai-nan Zhang
  4. Wei Wang
  5. Xiao Yang
  6. Huan Qi
  7. Hua Li
  8. Dong Men
  9. Jie Zhou
  10. Sheng-ce Tao

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Abstract

We still know very little about how the human immune system responds to SARS-CoV-2. Here we construct a SARS-CoV-2 proteome microarray containing 18 out of the 28 predicted proteins and apply it to the characterization of the IgG and IgM antibodies responses in the sera from 29 convalescent patients. We find that all these patients had IgG and IgM antibodies that specifically bind SARS-CoV-2 proteins, particularly the N protein and S1 protein. Besides these proteins, significant antibody responses to ORF9b and NSP5 are also identified. We show that the S1 specific IgG signal positively correlates with age and the level of lactate dehydrogenase (LDH) and negatively correlates with lymphocyte percentage. Overall, this study presents a systemic view of the SARS-CoV-2 specific IgG and IgM responses and provides insights to aid the development of effective diagnostic, therapeutic and vaccination strategies.

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  1. Version published to 10.1038/s41467-020-17488-8
  2. ScreenIT

    SciScore for 10.1101/2020.03.20.20039495: (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 Institutional Ethics Review Committee of Foshan Fourth Hospital, Foshan, China approved this study, and written informed consent was obtained from each patient.
    Consent: Patients and samples: The Institutional Ethics Review Committee of Foshan Fourth Hospital, Foshan, China approved this study, and written informed consent was obtained from each patient.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    The purified proteins were analyzed by SDS-PAGE followed by Western blotting using an anti-His antibody (Merck millipore, USA) and Coomassie brilliant blue staining.
    anti-His
    suggested: None
    Protein microarray fabrication: The proteins, along with negative (BSA) and positive controls (anti-Human IgG and IgM antibody), were printed in quadruplicate on PATH substrate slide (Grace Bio-Labs, Oregon, USA) to generate identical arrays in a 2 × 7 subarray format using Super Marathon printer (Arrayjet, UK)
    IgM
    suggested: None
    The arrays were washed with PBST and bound autoantibodies were detected by incubating with Cy3-conjugated goat anti-human IgG and Alexa Fluor 647-conjugated donkey anti-human IgM (Jackson ImmunoResearch, PA, USA), the antibodies were diluted 1: 1,000 in PBST, and incubated at room temperature for 1 h.
    anti-human IgG
    suggested: (Bio-Rad Cat# MCA647F, RRID:AB_808612)
    anti-human IgM
    suggested: None
    Software and Algorithms
    SentencesResources
    Pearson correlation coefficient between two proteins or indicators and the corresponding p value was calculated by SPSS software under the default parameters.
    SPSS
    suggested: (SPSS, RRID:SCR_002865)
    Cluster analysis was performed by pheatmap package in R30.
    pheatmap
    suggested: (pheatmap, RRID:SCR_016418)

    Results from OddPub: Thank you for sharing your data.

    Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:
    There are some limitations of the current SARS-CoV-2 proteome microarray. Firstly, due to the difficulty of protein expression and purification, there are still 10 proteins missing3. We will try to obtain these proteins through vigorous optimization or other sources, interesting finding is anticipated in the near future for these missing proteins. Secondly, most of the proteins on the microarray are not expressed in mammalian cells, critical post-translational modifications, such as glycosylation is absent. It is known that there are 23 N-glycosylation sites on S protein, which is heavily glycosylated, and the glycosylation may play critical roles in antibody-antigen recognition5,41. We are preparing these proteins using mammalian cell systems. Once the microarray is upgraded with proteins purified from mammalian cells, PTM specific IgG/IgM response may could be elicited. Thirdly, only 29 samples at collected at a single time point were analyzed. Though there are some interesting findings, we believe some of the current conclusions could be strengthened by including more samples. Furthermore, longitudinal samples29,42 collected at different time points from the same individual after diagnosis or even after cured may enable us to reveal the dynamics of the SARS-CoV-2 specific IgG/IgM responses. The data may could be further linked to the severity of COVID-19 among different patients. The application of the SARS-CoV-2 proteome microarray is not limited to serum profiling. It co...

    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.

  3. Version published to 10.1101/2020.03.20.20039495v1 on medRxiv