SARS-CoV-2 Spike Protein IgG ELISA

Serum samples were analyzed as previously described (Yinda et al., 2021 (link)). In brief, maxisorp plates (Nunc) were coated with 50 ng spike protein (generated in-house, purified recombinant) per well. Plates were incubated overnight at 4 °C. Plates were blocked with casein in phosphate buffered saline (PBS) (Thermo Fisher) for 1 hr at room temperature (RT). Serum was diluted twofold in blocking buffer and samples (duplicate) were incubated for 1 hr at RT. Secondary goat anti-hamster IgG Fc (horseradish peroxidase (HRP)-conjugated, Cat.No. 5220–0371 Lot. 10492253, Seracare) antibodies were used for detection and KPL TMB 2-component peroxidase substrate kit (SeraCare, Cat.No. 5120–0047) was used for visualization. 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 x the standard deviation of negative control hamster sera.