Development of an Automated Chemiluminescence Assay System for Quantitative Measurement of Multiple Anti-SARS-CoV-2 Antibodies

  1. Sousuke Kubo
  2. Norihisa Ohtake
  3. Kei Miyakawa
  4. Sundararaj Stanleyraj Jeremiah
  5. Yutaro Yamaoka
  6. Kota Murohashi
  7. Eri Hagiwara
  8. Takahiro Mihara
  9. Atsushi Goto
  10. Etsuko Yamazaki
  11. Takashi Ogura
  12. Takeshi Kaneko
  13. Takeharu Yamanaka
  14. Akihide Ryo

Reviewed by

Read the full article See related articles

Listed in

Log in to save this article

Abstract

Serological tests for COVID-19 have been instrumental in studying the epidemiology of the disease. However, the performance of the currently available tests is plagued by the problem of variability. We have developed a high-throughput serological test capable of simultaneously detecting total immunoglobulins (Ig) and immunoglobulin G (IgG) against nucleocapsid protein (NP) and spike protein (SP) and report its performance in detecting COVID-19 in clinical samples.

Methods

We designed and prepared reagents for measuring NP-IgG, NP-Total Ig, SP-IgG, and SP-Total Ig (using N-terminally truncated NP (ΔN-NP) or receptor-binding domain (RBD) antigen) dedicated automated chemiluminescent enzyme immunoassay analyzer AIA-CL1200. After determining the basal thresholds based on 17 sera obtained from confirmed COVID-19 patients and 600 negative sera, the clinical validity of the assay was evaluated using independent 202 positive samples and 1,000 negative samples from healthy donors.

Results

All of the four test parameters showed 100% specificity individually (1,000/1,000; 95%CI, 99.63–100). The sensitivity of the assay increased proportionally to the elapsed time from symptoms onset, and all the tests achieved 100% sensitivity (153/153; 95%CI, 97.63–100) after 13 days from symptoms onset. NP-Total Ig was the earliest to attain maximal sensitivity among the other antibodies tested.

Conclusion

Our newly developed serological testing exhibited 100% sensitivity and specificity after 13 days from symptoms onset. Hence, it could be used as a reliable method for accurate detection of COVID-19 patients and to evaluate seroprevalence and possibly for surrogate assessment of herd immunity.

Article activity feed

  2. Version published to 10.3389/fmicb.2020.628281
  3. Rapid Reviews Infectious Diseases

    Massimo Pieri

    Review 2: "Development of an automated chemiluminescence assay system for quantitative measurement of multiple anti-SARS-CoV-2 antibodies"

    The authors develop a new chemiluminescence assay system for measurement of SARS-CoV-2 antibodies. However, reviewers suggest that the study is potentially informative, though the lack of analytical validation in the study limits the generalizability of the findings.

  4. Rapid Reviews Infectious Diseases

    Kiang-Teck Yeo, Clarence W Chan

    Review 1: "Development of an automated chemiluminescence assay system for quantitative measurement of multiple anti-SARS-CoV-2 antibodies"

    The authors develop a new chemiluminescence assay system for measurement of SARS-CoV-2 antibodies. However, reviewers suggest that the study is potentially informative, though the lack of analytical validation in the study limits the generalizability of the findings.

  5. ScreenIT

    SciScore for 10.1101/2020.11.04.20225805: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementConsent: All healthy donors provided written informed consent for use of their clinical and biological data for the purpose of the scientific research.
    IRB: The study was performed according to protocols approved by our institutional review board.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Development of an antibody detection reagent for AIA-CL analyzers: We developed four types of serological test reagents (TOSOH NP-IgG, NP-Total Ig, SP-IgG, SP-Total Ig test) to detect anti–SARS-CoV-2 antibodies on the AIA-CL.
    NP-Total Ig, SP-IgG
    suggested: None
    anti–SARS-CoV-2
    suggested: None
    Patient cohorts: For initial performance evaluation, 600 samples of healthy donors collected prior to the spread of COVID-19 infection (purchased from TRINA BIOREACTIVES AG, Zurich, Swiss) and 17 samples from COVID-19 antibody–positive patients (according to the Abbott Architect SARS-CoV-2 IgG antibody test) were purchased from Biomex GmbH (Heidelberg, German).
    SARS-CoV-2 IgG
    suggested: None
    Software and Algorithms
    SentencesResources
    Patient cohorts: For initial performance evaluation, 600 samples of healthy donors collected prior to the spread of COVID-19 infection (purchased from TRINA BIOREACTIVES AG, Zurich, Swiss) and 17 samples from COVID-19 antibody–positive patients (according to the Abbott Architect SARS-CoV-2 IgG antibody test) were purchased from Biomex GmbH (Heidelberg, German).
    Abbott Architect
    suggested: (Abbott ARCHITECT i1000sr System, RRID:SCR_019328)
    All statistical analyses were performed by using GraphPad Prism 6 software.
    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:
    It should be noted that there were some limitations of this study. First, most cases in this study were of high severity. It is known that COVID-19 antibody titers tend to be lower in less severe cases (29, 30). Although there was no obvious difference in the antibody titers between moderate, severe and critical cases in this study, it was insufficient to evaluate the different in antibody titers due to the severity of COVID-19 because the number of samples in moderate cases were smaller than that of severe or critical cases. Therefore, there is the possibility that the cutoff value set in this assay may not be appropriate for detecting the antibody titers in lower severity cases, such as mild or asymptomatic patients. Second, the newly developed reagents can only measure IgG and total Ig, but not other immunoglobulin levels, such as IgM, IgA. Also, this study showed that NP-Total Ig was sustained but NP-IgG decreased after 30 days since symptoms onset. The reason for this reduction in IgG titers is unclear and the other immunoglobulin which elevated to sustain the total Ig levels also remains to be identified with further investigations. In conclusion, the automated TOSOH AIA-CL assay that can simultaneously quantify anti-SARS-CoV-2 NP-IgG, NP-Total Ig, SP-IgG and SP-Total Ig which was developed in this study, performed to be a highly accurate and specific serodiagnostic test for COVID-19. This test could play an important role in accurate detection of COVID-19 patients, eva...

    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: Please consider improving the rainbow (“jet”) colormap(s) used on page 35. At least one figure is not accessible to readers with colorblindness and/or is not true to the data, i.e. not perceptually uniform.

    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.

  6. Version published to 10.1101/2020.11.04.20225805v1 on medRxiv