(A) Killing of cells was tested using normal cynomolgus monkey serum, heat-inactivated serum (HI serum), RPMI-1640 (no serum), or RPMI-1640 +complement, each with or without 30 µg/ml alemtuzumab. Killing is shown as the percentage of cells that became 7-AAD-positive. Normal serum and RPMI-1640 + complement were associated with increased killing of cells incubated with alemtuzumab. Heat-inactivated serum and RPMI-1640 were not associated with increased killing. (B, C) Cells were incubated with alemtuzumab at the concentrations shown to determine apoptosis and killing of cells. Fresh T cells showed increasing 7-AAD (B) and Annexin-V (C) staining as the concentration of alemtuzumab increased. Expanded Teff had a higher baseline level of Annexin-V and 7-AAD staining, but did not show any increase when alemtuzumab was added. Data are representative of 4 separate experiments.