Enzyme-linked immunosorbent assay (ELISA)
ELISA was employed to quantify human IgG1 anti-SARS-CoV-2 antibody levels in the circulation after intranasal application of NAS in rats. In brief, rat serum samples diluted in 3% BSA in PBS buffer at a 1:10 dilution were added (100 µl/well) to an ELISA plate coated with 100 ng/well of Delta-variant RBD proteins (40592-V08H90, Sino Biological). Human IgG1 anti-SARS-CoV-2 antibodies were detected with goat anti-human IgG Fcγ-HRP antibody (109-005-098, Jackson Immuno Research) diluted 1:2,000 in 3% BSA in PBS buffer. The SIGMA