Abstract
A coronavirus antigen microarray (COVAM) was constructed containing 11 SARS-CoV-2, 5 SARS-1, 5 MERS, and 12 seasonal coronavirus recombinant proteins. The array is designed to measure immunoglobulin isotype and subtype levels in serum or plasma samples against each of the individual antigens printed on the array. We probed the COVAM with COVID-19 convalescent plasma (CCP) collected from 99 donors who recovered from a PCR+ confirmed SARS-CoV-2 infection. The results were analyzed using two computational approaches, a generalized linear model (glm) and random forest (RF) prediction model, to classify individual specimens as either Reactive or non-reactive against the SARS-CoV-2 antigens. A training set of 88 pre-COVID-19 specimens (PreCoV) collected in August 2019 and102 positive specimens from SARS-CoV-2 PCR+ confirmed COVID-19 cases was used for these analyses. Results compared with an FDA emergency use authorized (EUA) SARS-CoV2 S1-based total Ig chemiluminescence immunoassay (Ortho Clinical Diagnostics VITROS Anti-SARS-CoV-2 Total, CoV2T) and with a SARS-CoV-2 S1-S2 spike-based pseudovirus micro neutralization assay (SARS-CoV-2 reporter viral particle neutralization titration (RVPNT) showed high concordance between the three assays. Three CCP specimens that were negative by the VITROS CoV2T immunoassay were also negative by both COVAM and the RVPNT assay. Concordance between VITROS CoV2T and COVAM was 96%, VITROS CoV2T and RVPNT 93%, and RVPNT and COVAM 91%. The discordances were all weakly reactive samples near the cutoff threshold of the VITROS CoV2T immunoassay. The multiplex COVAM allows CCP to be grouped according to antibody reactivity patterns against 11 SARS-CoV-2 antigens. Unsupervised K-means analysis, via the gap statistics, as well as hierarchical clustering analysis revealed three main clusters with distinct reactivity intensities and patterns. These patterns were not recapitulated by adjusting the VITROS CoV2T or RVPNT assay thresholds. Plasma classified by COVAM reactivity patterns offers potential to improve CCP therapeutic efficacy CoV2T alone. The use of a SARS-CoV-2 antigen array can qualify CCP for administration as a treatment for acute COVID-19, and interrogate vaccine immunogenicity and performance in preclinical, clinical studies, and routine vaccination to identify antibody responses predictive of protection from infection and disease.
Highlights
A coronavirus antigen microarray (COVAM) was constructed containing SARS-CoV-2, 5 SARS1, 5 MERS, and seasonal coronavirus recombinant proteins
Rising within the first week after infection and about a week later for IgG with the seroconversion times around 10–14 days respectively[1,2]. It is not perfectly clear how long a protective antibody response will typically last, it has been reported that the antibody titers seem to wane down as soon as 12 weeks after symptom onset reaching with baseline levels after 52 weeks[1]
We probed COVAM with convalescent plasma (CCP) collected from 99 US PCR confirmed COVID-19 plasma donors who recovered from the infection 1–2 months prior to CCP collection
Summary
A coronavirus antigen microarray (COVAM) was constructed containing SARS-CoV-2, 5 SARS1, 5 MERS, and seasonal coronavirus recombinant proteins. Unsupervised K-means analysis, via the gap statistics, as well as hierarchical clustering analysis revealed three main clusters with distinct reactivity intensities and patterns These patterns were not recapitulated by adjusting the VITROS CoV2T or RVPNT assay thresholds. The use of a SARS-CoV-2 antigen array can qualify CCP for administration as a treatment for acute COVID19, and interrogate vaccine immunogenicity and performance in preclinical, clinical studies, and routine vaccination to identify antibody responses predictive of protection from infection and disease. Following exposure and recovery from SARS-CoV-2 infection, convalescent patients develop antigen specific adaptive T- and B-cell immune responses including binding and neutralizing antibodies (Ab). The receptor binding domain (RBD) is a critical domain of the S1 subunit, responsible for binding to the host cell ACE receptor and is the main target for neutralizing antibodies[4] These immune responses may prevent reinfection or blunt the clinical consequences of future infectious exposures to the virus. As the use of CCP evolved, it became apparent that not all CCP contained effective neutralizing efficacy, and anti-SARS CoV2 activity correlated with clinical outcomes[5,6]
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