Assessment of serological assays for identifying high titer convalescent plasma

  1. Christopher W. Farnsworth
  2. James B. Case
  3. Karl Hock
  4. Rita E. Chen
  5. Jane A. O'Halloran
  6. Rachel Presti
  7. Charles W. Goss
  8. Adriana M. Rauseo
  9. Ali Ellebedy
  10. Elitza S. Theel
  11. Michael S. Diamond
  12. Jeffrey P. Henderson

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Abstract

Background

The COVID‐19 pandemic has been accompanied by the largest mobilization of therapeutic convalescent plasma (CCP) in over a century. Initial identification of high titer units was based on dose–response data using the Ortho VITROS IgG assay. The proliferation of severe acute respiratory syndrome coronavirus 2 serological assays and non‐uniform application has led to uncertainty about their interrelationships. The purpose of this study was to establish correlations and analogous cutoffs between multiple serological assays.

Methods

We compared the Ortho, Abbott, Roche, an anti‐spike (S) ELISA, and a virus neutralization assay. Relationships relative to FDA‐approved cutoffs under the CCP emergency use authorization were identified in convalescent plasma from a cohort of 79 donors from April 2020.

Results

Relative to the neutralization assay, the spearman r value of the Ortho Clinical, Abbott, Roche, anti‐S ELISA assays was 0.65, 0.59, 0.45, and 0.76, respectively. The best correlative index for establishing high‐titer units was 3.87 signal‐to‐cutoff (S/C) for the Abbott, 13.82 cutoff index for the Roche, 1:1412 for the anti‐S ELISA, 1:219 by the neutralization assay, and 15.9 S/C by the Ortho Clinical assay. The overall agreement using derived cutoffs compared to a neutralizing titer of 1:250 was 78.5% for Abbott, 74.7% for Roche, 83.5% for the anti‐S ELISA, and 78.5% for Ortho Clinical.

DISCUSSION

Assays based on antibodies against the nucleoprotein were positively associated with neutralizing titers and the Ortho assay, although their ability to distinguish FDA high‐titer specimens was imperfect. The resulting relationships help reconcile results from the large body of serological data generated during the COVID‐19 pandemic.

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  1. Version published to 10.1111/trf.16580
  2. ScreenIT

    SciScore for 10.1101/2021.03.26.21254427: (What is this?)

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

    Table 1: Rigor

    Institutional Review Board StatementIRB: Human subjects: This study was approved by the Washington University Institutional Review Board.
    Randomizationnot detected.
    Blindingnot detected.
    Power Analysisnot detected.
    Sex as a biological variablenot detected.
    Cell Line Authenticationnot detected.

    Table 2: Resources

    Antibodies
    SentencesResources
    Three commercial serological assays and an anti-S ELISA granted EUA at Mt. Sinai Hospital, but used on a research basis for this study, were used to directly measure antibody levels in serum specimens.
    anti-S
    suggested: None
    Plates were washed and incubated with 1 µg/mL anti-S antibody (CR3022) and HRP conjugated goat anti-human IgG.
    CR3022
    suggested: None
    anti-human IgG
    suggested: None
    Experimental Models: Cell Lines
    SentencesResources
    Virus was propagated in Vero E6 cells in Dulbecco’s Modified Eagle Medium (DMEM, Corning) that was supplemented with 10% FBS, glucose, L-glutamine, and sodium pyruvate.
    Vero E6
    suggested: None
    Software and Algorithms
    SentencesResources
    The Abbott SARS-CoV-2 IgG assay was performed on an Abbott Architect i2000 and detects antibodies to the nucleocapsid protein.
    Abbott Architect
    suggested: (Abbott ARCHITECT i1000sr System, RRID:SCR_019328)
    The positive cutoffs for each assay are 1.0 (S/C), 1.4 (S/C), and 1.0 (COI) for the Ortho Clinical, the Abbott, and the Roche assays, respectively.
    Abbott
    suggested: (Abbott, RRID:SCR_010477)

    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 several limitations associated with this study. Among the greatest limitations is the lack of standardization between assays, even among the same manufacturers. This was previously noted with the neutralization assay, though the same is true among commercial assays. Since several of the assays have been designated as qualitative (i.e. the Roche, Abbott, and Ortho Clinical assays), there is limited evidence that semi-quantitative results are comparable between different instruments by the same manufacturer above the cutoff. For example, since there is no material to verify linearity at higher concentrations, a result of 15 S/C at one institution using the Ortho Clinical assay may vary from the Ortho Clinical assay at another institution. This may underlie the differences between the established cutoff and FDA cutoff for the Roche assay. In general, this problem will continue to plague the field until quantitative assays are universally adopted and standardized to SARS-CoV-2 antibody reference material, such as that recently released by the World Health Organization (27). This is further complicated by unclear direction as to how to report a qualitative assay result as quantitative under an EUA, which does not permit modification of the manufacturer’s Instructions for Use. Another limitation of the current study is that a limited number of assays were evaluated, limiting the generalizability of results. It is also important to note that these specimens were obtained e...

    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 10.1101/2021.03.26.21254427v1 on medRxiv