False Positive Results in SARS-CoV-2 Serological Tests for Samples From Patients With Chronic Inflammatory Diseases

  1. Nastya Kharlamova
  2. Nicky Dunn
  3. Sahl K. Bedri
  4. Svante Jerling
  5. Malin Almgren
  6. Francesca Faustini
  7. Iva Gunnarsson
  8. Johan RΓΆnnelid
  9. Rille Pullerits
  10. Inger Gjertsson
  11. Karin Lundberg
  12. Anna MΓ₯nberg
  13. Elisa Pin
  14. Peter Nilsson
  15. Sophia Hober
  16. Katharina Fink
  17. Anna Fogdell-Hahn

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Abstract

Patients with chronic inflammatory diseases are often treated with immunosuppressants and therefore are of particular concern during the SARS-CoV-2 pandemic. Serological tests will improve our understanding of the infection and immunity in this population, unless they tests give false positive results. The aim of this study was to evaluate the specificity of SARS-Cov-2 serological assays using samples from patients with chronic inflammatory diseases collected prior to April 2019, thus defined as negative. Samples from patients with multiple sclerosis (MS, n=10), rheumatoid arthritis (RA, n=47) with or without rheumatoid factor (RF) and/or anti-cyclic citrullinated peptide antibodies (anti-CCP2) and systemic lupus erythematosus (SLE, n=10) with or without RF, were analyzed for SARS-CoV-2 antibodies using 17 commercially available lateral flow assays (LFA), two ELISA kits and one in-house developed IgG multiplex bead-based assay. Six LFA and the in-house validated IgG assay correctly produced negative results for all samples. However, the majority of assays (n=13), gave false positive signal for samples from patients with RA and SLE. This was most notable in samples from RF positive RA patients. No false positive samples were detected in any assay using samples from patients with MS. Poor specificity of commercial serological assays could possibly be, at least partly, due to interfering antibodies in samples from patients with chronic inflammatory diseases. For these patients, the risk of false positivity should be considered when interpreting results of the SARS-CoV-2 serological assays.

Article activity feed

  1. Version published to 10.3389/fimmu.2021.666114
  2. Rapid Reviews Infectious Diseases

    Kouji H. Harada, Zhaoqing Lyu, Mariko Harada Sassa

    Review 4: "SARS-CoV-2 serological tests can generate false positive results for samples from patients with chronic inflammatory diseases"

    This potentially reliable study suggests certain serologic assays for viral antibodies may have increased false-positives in patients with chronic inflammatory diseases. Additional independent verification in well-defined cohorts is needed.

  3. Rapid Reviews Infectious Diseases

    Review 3: "SARS-CoV-2 serological tests can generate false positive results for samples from patients with chronic inflammatory diseases"

    This potentially reliable study suggests certain serologic assays for viral antibodies may have increased false-positives in patients with chronic inflammatory diseases. Additional independent verification in well-defined cohorts is needed.

  4. Rapid Reviews Infectious Diseases

    Marcelo Fruehwirth, Robson Michael Delai

    Review 2: "SARS-CoV-2 serological tests can generate false positive results for samples from patients with chronic inflammatory diseases"

    This potentially reliable study suggests certain serologic assays for viral antibodies may have increased false-positives in patients with chronic inflammatory diseases. Additional independent verification in well-defined cohorts is needed.

  5. Rapid Reviews Infectious Diseases

    Sarah Wheeler

    Review 1: "SARS-CoV-2 serological tests can generate false positive results for samples from patients with chronic inflammatory diseases"

    This potentially reliable study suggests certain serologic assays for viral antibodies may have increased false-positives in patients with chronic inflammatory diseases. Additional independent verification in well-defined cohorts is needed.

  6. Rapid Reviews Infectious Diseases

    Strength of evidence

    Reviewers: Sarah Wheeler (University of Pittsburgh) | πŸ“’πŸ“’πŸ“’β—»οΈβ—»οΈ
    Marcelo Fruehwirth, Robson Michael Delai (Itaiguapy Foundation) | πŸ“—πŸ“—πŸ“—πŸ“—β—»οΈ
    πŸ“—πŸ“—πŸ“—πŸ“—β—»οΈ
    Kouji H. Harada, Zhaoqing Lyu, Mariko Harada Sassa (Kyoto University) | πŸ“—πŸ“—πŸ“—πŸ“—β—»οΈ

  7. ScreenIT

    SciScore for 10.1101/2020.11.13.20231076: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: Study design and ethical approval: This retrospective cohort study was approved by the Ethics Review Authority and all participants provided informed consent at the time of sample collection to participate in future ethically approved studies.
    Consent: Study design and ethical approval: This retrospective cohort study was approved by the Ethics Review Authority and all participants provided informed consent at the time of sample collection to participate in future ethically approved studies.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    MS patient samples were collected in a research laboratory doing routine testing for anti-drug antibodies (ADAs) at The Centre for Molecular Medicine, Stockholm and had been treated with interferon beta (IFNΞ²).
    anti-drug
    suggested: None
    Specificity of this assay was determined by the manufacturer using anti-influenza A, anti-influenza B, Hepatitis C virus (HCV), anti-nuclear antibodies (ANA) and respiratory syncytial virus (RSV).
    anti-influenza A
    suggested: None
    anti-influenza B
    suggested: None
    anti-nuclear
    suggested: None
    Software and Algorithms
    SentencesResources
    Statistical analyses and figures were generated using GraphPad Prism (version 8.2.1).
    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:
    There are some limitations in this study. Firstly, there are over 200 SARS-CoV-2 serology assays available on the market and with the limited set of stored samples, we could only analyse a fraction of these. For reliable use of serological assays for patients with chronic inflammatory diseases, each assay would need to be individually analysed with negative serum from that patient population, before one starts to screen that patient group. Here we can only report the specificity in relation to MS, RA and SLE. Due to the limited availability of sample material, only one test result per sample, per assay, was retrieved and it was not possible to further elucidate the molecular mechanism behind the positive signals. Secondly, the LFA’s are primary made for whole blood, to enable individual to do a rapid test with a drop a blood from the fingertip, but here we only had stored serum to use for testing. However, all of the assay also indicate that they work with serum and plasma. Given that the serum from MS patients did not give any signal in any assay, the false positive signals detected in this study is most probably not an issue of having a different matrix, but more likely the unspecific antibody contents of the serum. In conclusion, serological assays are sensitive to interfering antibodies, especially from persons with autoimmune diseases. There is a trade-off between requiring extensive screening for unspecific binding in these assays and the harm the delay the process of m...

    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.

  8. Version published to 10.1101/2020.11.13.20231076v1 on medRxiv