Phage ELISA Using 384-Well Plate

Phage ELISA was performed as described previously except that the 384-well plate was used instead of the 96-well plate [18 (link)]. Briefly, the wells of the 384-well ELISA plate (Nunc) were coated with neutravidin (Thermo Fisher) for 1 hr at room temperature. The wells were washed with PBS-T (PBS containing 0.1% Tween 20) and blocked with PBS containing 0.5% (w/v) BSA (Gemini Bio) for 1hr. After removing the blocking buffer, biotinylated antigens were added to each well and washed three times with PBS-T. The 5-fold dilution of the cell culture supernatants containing phage were added to the wells and incubated for 30 min. After washing the wells with PBS-T three times, anti-M13HRP (Sino Biological) was added to the wells. SIGMAFAST™ OPD (Sigma) was used as a substrate and 2 M HCl was used as a quenching solution. The absorbance at 490 nm was measured using a BioTek Epoch 2 plate reader (BioTek).