ELISA Assay for Nipah mAb Binding

ELISA protocol was adapted from Quinlan et al.^{52 (link)}. Briefly, 100 μL of the wild-type GP in PBS was adsorbed to clear 96-well Maxisorp plates (Nunc) at a concentration of 1 μg/mL and left at 4 °C overnight. Plates were washed with PBS with 0.05% Tween 20 (PBST) and incubated with 100 μL of 2% BSA in PBST for 1 h at room temperature. Plates were washed with PBST then threefold serially diluted of anti-Nipah mAb were added to wells followed by incubation for 2 h. After washing with PBST, 100 μL of secondary antibody, goat anti-human IgG H&L conjugated with HRP (1:10,000, Abcam) were incubated for 1 h. Finally, plates were washed, and incubated with 100 μL of TMB substrate (KPL) for 5 min and quenched with 100 μL of 1 N H₂SO₄. Plates were read using a Tecan Infinite N200 Pro plate reader at 450 nm.