Supplementary Material for: Quantitative Measurement of IgG to Severe Acute Respiratory Syndrome Coronavirus-2 Proteins Using ImmunoCAP

<b><i>Background:</i></b> Detailed understanding of the immune response to severe acute respiratory syndrome coronavirus (SARS-CoV)-2, the cause of coronavirus disease 2019 (CO­VID-19) has been hampered by a lack of quantitative antibody assays. <b><i>Objective:</i></b> The objective was to develop a quantitative assay for IgG to SARS-CoV-2 proteins that could be implemented in clinical and research laboratories. <b><i>Methods:</i></b> The biotin-streptavidin technique was used to conjugate SARS-CoV-2 spike receptor-binding domain (RBD) or nucleocapsid protein to the solid phase of the ImmunoCAP. Plasma and serum samples from patients hospitalized with COVID-19 (<i>n</i> = 60) and samples from donors banked before the emergence of COVID-19 (<i>n</i> = 109) were used in the assay. SARS-CoV-2 IgG levels were followed longitudinally in a subset of samples and were related to total IgG and IgG to reference antigens using an ImmunoCAP 250 platform. <b><i>Results:</i></b> At a cutoff of 2.5 μg/mL, the assay demonstrated sensitivity and specificity exceeding 95% for IgG to both SARS-CoV-2 proteins. Among 36 patients evaluated in a post-hospital follow-up clinic, median levels of IgG to spike-RBD and nucleocapsid were 34.7 μg/mL (IQR 18–52) and 24.5 μg/mL (IQR 9–59), respectively. Among 17 patients with longitudinal samples, there was a wide variation in the magnitude of IgG responses, but generally the response to spike-RBD and to nucleocapsid occurred in parallel, with peak levels approaching 100 μg/mL, or 1% of total IgG. <b><i>Conclusions:</i></b> We have described a quantitative assay to measure IgG to SARS-CoV-2 that could be used in clinical and research laboratories and implemented at scale. The assay can easily be adapted to measure IgG to mutated COVID-19 proteins, has good performance characteristics, and has a readout in standardized units.