Evaluation of 6 Commercial SARS-CoV-2 Serology Assays Detecting Different Antibodies for Clinical Testing and Serosurveillance

  1. Suellen Nicholson
  2. Theo Karapanagiotidis
  3. Arseniy Khvorov
  4. Celia Douros
  5. Francesca Mordant
  6. Katherine Bond
  7. Julian Druce
  8. Deborah A Williamson
  9. Damian Purcell
  10. Sharon R Lewin
  11. Sheena Sullivan
  12. Kanta Subbarao
  13. Mike Catton

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Abstract

Background

Serological testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) complements nucleic acid tests for patient diagnosis and enables monitoring of population susceptibility to inform the coronavirus disease 2019 (COVID-19) pandemic response. It is important to understand the reliability of assays with different antigen or antibody targets to detect humoral immunity after SARS-CoV-2 infection and to understand how antibody (Ab) binding assays compare to those detecting neutralizing antibody (nAb), particularly as we move into the era of vaccines.

Methods

We evaluated the performance of 6 commercially available enzyme-linked immunosorbent assays (ELISAs), including a surrogate virus neutralization test (sVNT), for detection of SARS-CoV-2 immunoglobulins (IgA, IgM, IgG), total or nAb. A result subset was compared with a cell culture–based microneutralization (MN) assay. We tested sera from patients with prior reverse transcription polymerase chain reaction–confirmed SARS-CoV-2 infection, prepandemic sera, and potential cross-reactive sera from patients with other non-COVID-19 acute infections.

Results

For sera collected >14 days post–symptom onset, the assay achieving the highest sensitivity was the Wantai total Ab at 100% (95% CI, 94.6%–100%), followed by 93.1% for Euroimmun NCP-IgG, 93.1% for GenScript sVNT, 90.3% for Euroimmun S1-IgG, 88.9% for Euroimmun S1-IgA, and 83.3% for Wantai IgM. Specificity for the best-performing assay was 99.5% for the Wantai total Ab, and for the lowest-performing assay it was 97.1% for sVNT (as per the Instructions for Use [IFU]). The Wantai Total Ab had the best agreement with MN at 98% followed by Euroimmun S1-IgA, Euro NCP-IgG, and sVNT (as per IFU) with 97%, 97% and 95%, respectively; Wantai IgM had the poorest agreement at 93%.

Conclusions

Performance characteristics of the SARS-CoV-2 serology assays detecting different antibody types are consistent with those found in previously published reports. Evaluation of the surrogate virus neutralization test in comparison to the Ab binding assays and a cell culture–based neutralization assay showed good result correlation between all assays. However, correlation between the cell-based neutralization test and some assays detecting Ab’s not specifically involved in neutralization was higher than with the sVNT. This study demonstrates the reliability of different assays to detect the humoral immune response following SARS-CoV-2 infection, which can be used to optimize serological test algorithms for assessing antibody responses post–SARS-CoV-2 infection or vaccination.

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  1. Version published to 10.1093/ofid/ofab239
  2. ScreenIT

    SciScore for 10.1101/2021.01.21.21250249: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: Ethics: Project ethical approval for RMH specimens was obtained from Melbourne Health Human Research Ethics Committee (RMH HREC QA2020052).
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    SARS-CoV-2 binding antibodies are detected using enzyme-labelled anti-human-IgG or anti-human-IgA conjugates and a colourmetric substrate and are read spectrophotometrically.
    anti-human-IgG
    suggested: None
    anti-human-IgA
    suggested: None
    Patient antibody to SARS-CoV-2 that binds antigen coated on the plate, is bound by horseradish peroxidase (HRP) antigen conjugate forming an antigen-antibody-antigen-HRP complex detectable by colourmetric substrate and read spectrophotometrically.
    antigen-antibody-antigen-HRP
    suggested: None
    Wantai SARS-CoV-2 IgM: This is a capture ELISA for detection of IgM-class antibodies to SARS-CoV-2 virus.
    IgM-class
    suggested: None
    Anti-µ chain antibodies on the plate capture patient IgM antibodies; detection is by recombinant SARS-CoV-2antigen-HRP-conjugate followed by a colourmetric substrate read spectrophotometrically.
    Anti-µ chain
    suggested: None
    2 Surrogate Virus Neutralization Test (sVNT): (GenScript USA, Inc. New Jersey, USA) This is a species and isotype independent blocking ELISA which mimics virus neutralisation, detecting circulating neutralising SARS-CoV-2 antibodies that block the interaction between the viral spike RBD and host angiotensin converting enzyme 2 (ACE2) cell surface receptor.
    ACE2
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Briefly, the ability of serial 2-fold dilutions of sera to neutralize the infectivity of 100 median tissue culture infectious doses of SARS-CoV-2 was assessed by inhibition of viral cytopathic effect in Vero cells.
    Vero
    suggested: CLS Cat# 605372/p622_VERO, RRID:CVCL_0059)
    Software and Algorithms
    SentencesResources
    The 95% confidence intervals were generated using the exact binomial Clopper-Pearson method with PropCIs R package [17].
    PropCIs
    suggested: None

    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:

    The Wantai SARS-CoV-2 total Ab assay was alone among those tested in our hands in achieving sensitivity and specificity very close to that reported by the manufacturer (94.5%; and 100% respectively); one limitation of this assay is the requirement for 100ul of serum versus 10ul for the other assays. In low prevalence populations and with low test PPV’s, serological testing algorithms should use highly sensitive assays for screening such as Wantai Total Ab, followed by supplemental or confirmatory testing with highly specific assays such as sVNT or MN to ensure reliable results. An important component of this study is the comparison of sensitivities of the ELISAs and MN, which should be considered in the context of discordant screening and supplemental results in testing algorithms. Limitations of our study include the smaller number of convalescent patient RT-PCR-POS confirmed SARS-CoV-2 samples collected > 14 days post-symptom onset used for the Euroimmun and sVNT evaluations; Panel A, 49% (72/147) of samples compared to Panel B, 69% (66/96) of samples used for Wantai Total Ab and IgM assays. A rise in index value/inhibition for all assays was observed over time (Figure 2) however these measurements are semi-quantitative, indicating antibody amount present for comparison between assays, not antibody titre which can only be obtained by sample titration to ‘end-point.’ In conclusion, our study revealed different performance characteristics between six commercially available EL...

    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 scite Reference Check: We found no unreliable references.

    About SciScore

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  3. Version published to 10.1101/2021.01.21.21250249v1 on medRxiv