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. 2021 Jan 4;12(1):6.
doi: 10.1038/s41467-020-20095-2.

Analysis of SARS-CoV-2 antibodies in COVID-19 convalescent blood using a coronavirus antigen microarray

Rafael R de Assis  1 , Aarti Jain  1 , Rie Nakajima  1 , Algis Jasinskas  1 , Jiin Felgner  1 , Joshua M Obiero  1 , Philip J Norris  2   3 , Mars Stone  2   3 , Graham Simmons  2   3 , Anil Bagri  4 , Johannes Irsch  4 , Martin Schreiber  5 , Andreas Buser  6 , Andreas Holbro  5 , Manuel Battegay  6 , Philip Hosimer  7 , Charles Noesen  7 , Oluwasanmi Adenaiye  8 , Sheldon Tai  8 , Filbert Hong  8 , Donald K Milton  8 , D Huw Davies  1 , Paul Contestable  7 , Laurence M Corash  4 , Michael P Busch  2   3 , Philip L Felgner  1 , Saahir Khan  9
Affiliations

Analysis of SARS-CoV-2 antibodies in COVID-19 convalescent blood using a coronavirus antigen microarray

Rafael R de Assis et al. Nat Commun. .

Abstract

The current practice for diagnosis of COVID-19, based on SARS-CoV-2 PCR testing of pharyngeal or respiratory specimens in a symptomatic patient at high epidemiologic risk, likely underestimates the true prevalence of infection. Serologic methods can more accurately estimate the disease burden by detecting infections missed by the limited testing performed to date. Here, we describe the validation of a coronavirus antigen microarray containing immunologically significant antigens from SARS-CoV-2, in addition to SARS-CoV, MERS-CoV, common human coronavirus strains, and other common respiratory viruses. A comparison of antibody profiles detected on the array from control sera collected prior to the SARS-CoV-2 pandemic versus convalescent blood specimens from virologically confirmed COVID-19 cases demonstrates near complete discrimination of these two groups, with improved performance from use of antigen combinations that include both spike protein and nucleoprotein. This array can be used as a diagnostic tool, as an epidemiologic tool to more accurately estimate the disease burden of COVID-19, and as a research tool to correlate antibody responses with clinical outcomes.

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Conflict of interest statement

The coronavirus antigen microarray is the intellectual property of the Regents of the University of California that is licensed for commercialization to Nanommune Inc. (Irvine, CA), a private company for which P. Felgner is the largest shareholder and several co-authors (R. de Assis, A. Jain, R. Nakajima, A. Jasinskas, J. Obiero, Jiin Felgner, H. Davies, and S. Khan) also own shares. Nanommune Inc. has a business partnership with Sino Biological Inc. (Beijing, China) which expressed and purified the antigens used in this study. The convalescent plasma used in this study was collected for clinical use by independent blood centers using licensed plasma or platelet processing systems manufactured by Cerus Corporation, for which multiple authors (L. Corash, A. Bagri, and Johannes Irsch) are shareholders and employees. Convalescent sera were also provided by Ortho Clinical Diagnostics, which is using these specimens to validate a clinical diagnostic test, and P. Hosimer, C. Noesen, and P. Contestable are shareholders and employees of this company. M. Battegay, A. Buser, A. Holbro, Philip J. Norris, Mars Stone, Graham Simmons, Martin Schreiber, Oluwasanmi Adenaiye, Sheldon Tai, Filbert Hong, Donald K. Milton, Michael P. Busch, and the Prometheus-UMD consortium investigators have no conflicts of interest to disclose.

Figures

Fig. 1
Fig. 1. Heatmap for coronavirus antigen microarray.
The heatmap shows IgG (top) and IgM (bottom) reactivity measured as mean fluorescence intensity across four replicates, against each antigen organized into rows color coded by virus, for blood specimens organized into columns classified as positive (convalescent from PCR-positive individuals) or negative (prior to pandemic from naive individuals). Reactivity is represented by color (white = low, black = mid, red = high). Source data are provided as a Source data file.
Fig. 2
Fig. 2. Normalized antibody reactivity of positive and negative sera on coronavirus antigen microarray.
The plot shows IgG (a) and IgM (b) reactivity against each antigen measured as mean fluorescence intensity (MFI). The boxes represent the first quartile, median and third quartile and the whiskers extend 1.5 times the interquartile range (IQR). Each point represents the distribution of the reactivity for either convalescent blood specimens from PCR-positive individuals (positive, red, n = 100) and sera from naive individuals prior to pandemic (negative, blue, n = 88). Below the plot, the heatmap shows average reactivity for each group (white = low, black = mid, red = high). The antigen labels are color coded for respiratory virus group. Source data are provided as a Source data file.
Fig. 3
Fig. 3. ROC curves for high-performing combination of antigens.
ROC curves showing sensitivity versus specificity for discrimination of positive and negative blood specimens were derived for each combination of the high performing antigens for both IgM (a) and IgG (b) and compared to no discrimination (ROC AUC = 0.5, gray line). Source data are provided as a Source data File.
Fig. 4
Fig. 4. Sensitivity of CoVAM as a function of time since symptom onset.
The sensitivity in detecting SARS-CoV-2 antibody responses in PCR-positive individuals is shown as a function of time since symptom onset for IgM, IgG, and either IgM or IgG. Source data are provided as a Source data file.
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