Quantifying NXT-2 Antibody Titers

Microtiter plates (Immunolon 4HBX; Thermo Fisher Scientific) were coated with purified NXT-2 at 5 µg/mL in PBS. Heat-inactivated plasma samples were diluted 1:100 in blocking buffer (PBS with 5% nonfat milk) and 1:2 serial dilutions were made to determine endpoint titers, as previously described (30 (link), 33 (link)). Goat antimouse and goat antimonkey immunoglobulin conjugated horseradish peroxidase (Southern Biotech, Accurate Chemical) were used for detection, and plates were developed with TMB (Fisher Scientific). Naive (uninfected, NXT-2-negative by antibody titer) mouse and NHP plasma samples were used as negative controls.