Apoptotic effect of mAbs on CD74⁺ cells. Induction of apoptosis was evaluated by flow cytometry determination of hypodiploid DNA on the cell line panel. Cells were cultured with the mAbs without a second antibody, with GAM IgG, or with GAH IgG, followed by DNA staining with propidium iodide. (A) Representative histograms are shown for SU-DHL-6 cells. Hypodiploid DNA peaks corresponding to apoptotic nuclei were quantified in region M1. Dashed line indicates no second antibody; solid gray line, GAM IgG; solid black line, GAH IgG. (B) Percentage apoptosis (hypodiploid DNA) induced by cross-linked mAbs is shown for a panel of cell lines. (C) Caspase-3 and -8 activities in Raji cells. Cells were cultured in the presence of hLL1 (5 μg/mL) or a GAH IgG Fcγ–specific antibody cross-linking agent (CL) (20 μg/mL), with both antibodies (LL1 [5 μg/mL] + CL [20 μg/mL]) or no antibody added (Untreated) for 24 hours. Intracellular levels of caspase-3 and -8 were measured by using ApoAlert Caspase assay kits (Clontech).