Effector functions. ADCC was evaluated with Raji (A) or Daudi (B) cells. Cells were incubated with the indicated mAb or mAb-IFNα (10 replicates/treatment) at 33nM in the presence of freshly isolated PBMCs) for 4 hours before quantification of cell lysis with CytoTox-One (Promega). Effector/target ratio = 50:1. Error bars indicate SD. The hMN-14 (anti-CEACAM5 mAb) and 14-2b-2b (mAb-IFNα of hMN-14) were used as a nontargeting mAb and mAb-IFNα, respectively. (C) CDC. Daudi cells were incubated with serial dilutions of C2-2b-2b, hL243γ4p, hL243 IgG1, or veltuzumab in the presence of human complement (1/20 final dilution; Quidel Corp) for 2 hours at 37°C and 5% CO₂. Viable cells were then quantified with the Vybrant Cell Metabolic Assay Resazurin kit (Invitrogen). Controls included cells treated with 0.25% Triton X-100 (100% lysis) and cells treated with complement alone (background). The percentage of complement control (number of viable cells in the test sample compared with cells treated with complement only) was plotted against the log of the molar concentration. Error bars indicate SDs.