There is an urgent need for an accurate antibody test for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this paper, we have developed 3 ELISA methods, trimer spike
IgA, trimer spike
IgG, and nucleocapsid
IgG, for detecting anti-SARS-CoV-2
antibodies. We evaluated their performance in comparison with four commercial ELISAs, EDI™ Novel Coronavirus
COVID-19 ELISA
IgG and
IgM, Euroimmun Anti-SARS-CoV-2 ELISA
IgG and
IgA, and one lateral flow assay, DPP®
COVID-19 IgM/
IgG System (Chembio). Both sensitivity and specificity were evaluated and the causes of false-positive reactions were determined. The assays were compared using 300 pre-epidemic samples and 100 PCR-confirmed
COVID-19 samples. The sensitivities and specificities of the assays were as follows: 90%/100% (in-house trimer spike
IgA), 90%/99.3% (in-house trimer spike
IgG), 89%/98.3% (in-house nucleocapsid
IgG), 73.7%/100% (EDI nucleocapsid
IgM), 84.5%/95.1% (EDI nucleocapsid
IgG), 95%/93.7% (Euroimmun S1
IgA), 82.8%/99.7% (Euroimmun S1
IgG), 82.0%/91.7% (Chembio nucleocapsid
IgM), 92%/93.3% (Chembio nucleocapsid
IgG). The presumed causes of positive signals from pre-epidemic samples in commercial and in-house assays were mixed. In some cases, positivity varied with assay repetition. In other cases, reactivity was abrogated by competitive inhibition (spiking the sample with analyte prior to performing the assay). In other cases, reactivity was consistently detected but not abrogated by analyte spiking. Overall, there was wide variability in assay performance using our samples, with in-house tests exhibiting the highest combined sensitivity and specificity. The causes of "false positivity" in pre-epidemic samples may be due to plasma
antibodies apparently reacting with the analyte, or spurious reactivity may be directed against non-specific components in the assay system. Identification of these targets will be essential to improving assay performance.