Assessment of CDC and ADCC. (A) Daudi cells were treated with hL243γ4P or control mAbs, as indicated, at the concentrations shown in the presence of human complement. Cell viability was measured using resazurin and reported as percentage of viable population relative to cells treated with complement only (no mAb). (B) 51Cr-labeled NHL cell lines were incubated with anti-B-cell mAbs in the presence of human complement. Following a 3-hour incubation at 37°C, supernatants were collected and counted. Percentage of specific lysis of 3 cell lines is shown. (C) Calcein-AM cytotoxicity release assay for measurement of ADCC. Labeled NHL cell lines were incubated with anti-B-cell mAbs in the presence of human mononuclear cells. Following a 4-hour incubation at 37°C, supernatants were harvested and transferred to new plates. Samples were measured using a Spectromax Gemini dual-scanning microplate spectrofluorimeter (Molecular Devices, Sunnyvale, CA); excitation filter, 485 nm; bandpass filter, 530 nm. Percentage of specific lysis of 3 cell lines is shown. ▪, with PBMCs; □, without PBMCs. Error bars represent SD.
