Ad35K++-sensitized cytolysis of lymphoma cells in vitro. (A top) Scheme of the experiment with Raji cells. Raji cells were incubated with anti-CD46 mAb, Ad35K-279, Ad35K, or Ad35K++. Eight hours later, daclizumab or rituximab were added to cells and incubated at room temperature. After 30 minutes, NHS was added, and viable cells were counted 3 hours later based on trypan blue exclusion. (Bottom) Percentage of viable cells compared with PBS-treated cells. Shown are the mean values and standard deviations; n > 6. (B-C) Studies with other lymphoma cell lines (B) or primary cells from patients with B-CLL (C). The experimental conditions were as described in panel A. Significant differences (P < .05) are marked by an asterisk. (D) Studies with normal human PBMCs. Human PBMCs (pooled from 3 healthy donors) were sorted for CD20+ cells with the use of fluorescence-activated cell sorting. CD20+ cells were cultured for 3 days. A total of 105 cells were treated with Ad35K++ (25 μg/mL), followed by rituximab (15 μg/mL) and NHS (25% final concentration) 8 hours later. Four hours after adding NHS, viable cells were counted based on trypan blue exclusion. Cell viability of PBS-treated cells was taken as 100%. There was no change in cell viability for cells incubated with Ad35K++, rituximab, or NHS alone. (E) Human PBMCs were cultured for 3 days and treated as described in panel D. Shown are the mean values and standard deviations; n = 5.
