Neutralizing antibody responses to SARS-CoV-2 in a COVID-19 recovered patient cohort and their implications

  1. Fan Wu
  2. Aojie Wang
  3. Mei Liu
  4. Qimin Wang
  5. Jun Chen
  6. Shuai Xia
  7. Yun Ling
  8. Yuling Zhang
  9. Jingna Xun
  10. Lu Lu
  11. Shibo Jiang
  12. Hongzhou Lu
  13. Yumei Wen
  14. Jinghe Huang

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Abstract

The COVID-19 pandemic caused by SARS-CoV-2 coronavirus threatens global public health. Currently, neutralizing antibodies (NAbs) versus this virus are expected to correlate with recovery and protection of this disease. However, the characteristics of these antibodies have not been well studied in association with the clinical manifestations in patients.

Methods

Plasma collected from 175 COVID-19 recovered patients with mild symptoms were screened using a safe and sensitive pseudotyped-lentiviral-vector-based neutralization assay. Spike-binding antibody in plasma were determined by ELISA using RBD, S1, and S2 proteins of SARS-CoV-2. The levels and the time course of SARS-CoV-2-specific NAbs and the spike-binding antibodies were monitored at the same time.

Findings

SARS-CoV-2 NAbs were unable to cross-reactive with SARS-CoV virus. SARS-CoV-2-specific NAbs were detected in patients from day 10-15 after the onset of the disease and remained thereafter. The titers of NAb among these patients correlated with the spike-binding antibodies targeting S1, RBD, and S2 regions. The titers of NAbs were variable in different patients. Elderly and middle-age patients had significantly higher plasma NAb titers (P<0.0001) and spike-binding antibodies (P=0.0003) than young patients. Notably, among these patients, there were ten patients whose NAb titers were under the detectable level of our assay (ID50: < 40); while in contrast, two patients, showed very high titers of NAb, with ID50 :15989 and 21567 respectively. The NAb titers were positive correlated with plasma CRP levels but negative correlated with the lymphocyte counts of patients at the time of admission, indicating an association between humoral response and cellular immune response.

Interpretation

The variations of SARS-CoV-2 specific NAbs in recovered COVID-19 patients may raise the concern about the role of NAbs on disease progression. The correlation of NAb titers with age, lymphocyte counts, and blood CRP levels suggested that the interplay between virus and host immune response in coronavirus infections should be further explored for the development of effective vaccine against SARS-CoV-2 virus. Furthermore, titration of NAb is helpful prior to the use of convalescent plasma for prevention or treatment.

Funding

Ministry of Science and Technology of China, National Natural Science Foundation of China, Shanghai Municipal Health Commission, and Chinese Academy of Medical Sciences

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  1. ScreenIT

    SciScore for 10.1101/2020.03.30.20047365: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: The study was conducted under a clinical protocol approved by the Investigational Review Board in the Shanghai Public Health Clinical Center (Study number: YJ-2020-S021-01).
    Consent: All participants signed an informed consent approved by the IRB.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    A 1:2500 dilution of horseradish peroxidase (HRP)-conjugated goat anti-human IgG antibody was added for 1 hour at room temperature.
    anti-human IgG
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    293T cells expressing human angiotensin converting enzyme II (ACE2) (293 T/ACE2) were obtained from the American Type Culture Collection (ATCC; Manassas, VA, USA) and were cultured in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum (FBS).
    293T
    suggested: None
    First, 293 T/ACE2 cells were seeded in a 96-well plate at a concentration of 104 cells per well and cultured for 12 hours.
    T/ACE2
    suggested: RRID:CVCL_YZ65)
    Software and Algorithms
    SentencesResources
    Statistical analysis: Statistical analyses were carried out using GraphPad Prism 7.0.
    GraphPad Prism
    suggested: (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: We detected the following sentences addressing limitations in the study:
    This study is preliminary and has several limitations. First, viral RNA was not detectable in patients’ blood. Owing to the lack of respiratory specimens, information about the kinetics of viral loads was not available. Second, patients in severe and critical condition were excluded from the study because they received passive antibody treatment before sample collection. Thus, we were not able to directly evaluate the effect of NAbs on virus clearance or disease progression of COVID-19 patients in this study. A further comprehensive study should be made to address the question. To the best of our knowledge, this is the first report about NAbs drawn from the plasma of a COVID-19 recovered patient cohort, potentially providing useful information for passive antibody therapy and vaccine development against SARS-CoV-2 virus. The highly variable levels of NAbs in the patients of COVID-19 indicated that convalescent plasma and serum from recovered donors should be titrated before use in passive antibody therapy, an easy task that can be performed using the PsV neutralization assay. Correlation of NAbs titers with the age, lymphocyte counts and blood CRP levels of patients also lays the groundwork for further study to explore the mechanism of NAbs development in COVID-19 patients.

    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 10.1101/2020.03.30.20047365v2 on medRxiv
  3. Version published to 10.1101/2020.03.30.20047365v1 on medRxiv