SARS-CoV-2 Neutralizing Antibody Evaluation

To evaluate the neutralizing activity of monoclonal antibodies, plaque reduction neutralizing tests for SARS-CoV-2 were performed as described previously^{43 (link)}. Briefly, twofold serially diluted mAbs ranging from 10³ to 1 ng/ml and an equal volume of virus (40 pfu/well) were incubated at 37 °C for 2 h. The antibody–virus mixture was inoculated into a 24-well plate seeded with VeroE6 cells (1 × 10⁵ cells/well) and incubated at 37 °C for 1 h, followed by an overlay of 1 ml of 0.5% agarose (Lonza). After 2 to 3 days of incubation, the cells were fixed with 4% paraformaldehyde and visualized plaques with crystal violet. Two independent experiments were performed in duplicate for each mAb. The data were fitted to a dose–response inhibition model, and the half-maximal inhibitory concentration (IC₅₀) of each mAb was calculated using GraphPad Prism6 software.