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. 2021 May 28;16(5):e0252621.
doi: 10.1371/journal.pone.0252621. eCollection 2021.

Validation of a multiplex flow immunoassay for detection of IgG antibodies against SARS-CoV-2 in dried blood spots

Coleman T Turgeon  1 , Karen A Sanders  1 , Piero Rinaldo  1 , Dane Granger  2 , Heather Hilgart  2 , Dietrich Matern  1 , Elitza S Theel  2
Affiliations

Validation of a multiplex flow immunoassay for detection of IgG antibodies against SARS-CoV-2 in dried blood spots

Coleman T Turgeon et al. PLoS One. .

Abstract

Background: Dried blood spots (DBS) are an established specimen type for clinical testing given their low cost, ease of collection and storage, and convenient shipping capabilities through the postal system. These attributes are complementary to the expansion of SARS-CoV-2 serologic testing, which may be used to inform community seroprevalence rates.

Methods: The Luminex xMAP SARS-CoV-2 Multi-Antigen assay utilizes magnetic beads labeled with three viral antigens (nucleocapsid [NC], receptor binding domain [RBD], spike S1 subunit) to detect anti-viral IgG-class antibodies, and has Food and Drug Administration (FDA) Emergency Use Authorization (EUA) for use in serum and plasma. This assay was modified for use with DBS and validated against paired sera tested by one of two reference assays: the Roche Diagnostics Elecsys anti-SARS-CoV-2 ECLIA or the Euroimmun anti-SARS-CoV-2 IgG ELISA.

Results: 159 paired DBS and serum specimens analyzed using the modified Luminex xMAP assay on DBS and the reference methods on serum showed an overall concordance of 96.9% (154/159). Use of multivariate pattern recognition software (CLIR) for post-analytical interpretation of the Luminex xMAP DBS assay results, instead of manufacturer provided interpretive thresholds, increased overall qualitative result concordance to 99.4% (158/159) between the modified Luminex xMAP DBS and reference results.

Conclusions: Use of DBS for detection of antibodies against SARS-CoV-2 provides comparable results to those obtained using serum. DBS concordance was improved with multivariate pattern recognition software (CLIR). We demonstrate that DBS are a reliable specimen type for SARS-CoV-2 antibody detection using the modified Luminex xMAP assay.

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Conflict of interest statement

I have read the journal’s policy and among the authors of this manuscript Dr. Theel has the following broad interests that could be perceived as conflicts of interest: - Consulting fees from Roche Diagnostics and Accelerate Diagnostics - Member of the College of American Pathologists’ Microbiology Resource Committee (travel compensation) - Editor, Clinical Microbiology Newsletter This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Figures

Fig 1
Fig 1. Result comparison of 107 paired DBS and serum samples analyzed for IgG against the SARS-CoV-2 nucleocapsid (NC, panel A), spike glycoprotein S1 subunit (S1, panel B) and receptor binding domain (RBD) antigens (panel C) using the Luminex xMAP assay.
The samples represented by the X and the diamond indicate two cases with discordant result interpretations.
Fig 2
Fig 2. CLIR plot by multiple conditions of 159 DBS samples with serum SARS-CoV-2 antibody results.
Abbreviations: NC, nucleocapsid protein; RBD, receptor binding domain; S1, spike glycoprotein S1 subunit; BGKR, background; C, control. Luminex (Lu)-IgG TP, ranges for true positive cases (N = 49; blue); Lu-IgG FP, false positive cases (N = 3; orange); Lu-IgG FN, false negative cases (N = 2; purple); and Lu-IgG TN, true negative cases (N = 105; red). Upper whisker end: 99th percentile; top of box: 90th percentile; line in box: median; bottom of box: 10th percentile; lower whisker end: 1st percentile. Black diamonds indicate the manufacturer provided cutoff (700 MFI) for IgG against SARS-CoV-2 NC, RBD and S1 antigens, respectively.
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