SARS-CoV-2 Spike Protein Antibody Assay

Serum samples were analyzed as previously described [57 (link)]. In brief, maxisorp plates (Nunc) were coated with 50 ng spike protein (generated in-house) per well. Plates were incubated overnight at 4°C. Plates were blocked with casein in phosphate buffered saline (PBS) (ThermoFisher) for 1 hour at room temperature. Serum was diluted 2-fold in blocking buffer and samples (duplicate) were incubated for 1 hour at room temperature. Secondary goat anti-hamster IgG Fc (horseradish peroxidase (HRP)-conjugated, Abcam) spike-specific antibodies were used for detection and visualized with KPL TMB 2-component peroxidase substrate kit (SeraCare, 5120–0047). The reaction was stopped with KPL stop solution (Seracare) and plates were read at 450 nm. The threshold for positivity was calculated as the average plus 3 × the standard deviation of negative control hamster sera.