Uptake of Bcl-2-ASO.
(A) Kinetics of Bcl-2-ASO uptake in ARH-77 myeloma cells. ARH-77 cells (high Bcl-2) were seeded at 5 × 105/mL and placed in a 24-well plate in a total volume of 1 mL FCS medium. FITC–Bcl-2-ASO was added at the indicated concentrations. FITC–Bcl-2-ASO was replenished every other day. Uptake was determined by flow cytometry using FACSCalibur, and results were analyzed by the CellQuest software; 10 000 cells were analyzed. Uptake at 4, 8, 24, 48, 72, and 96 hours is shown at 0 μg/mL, 1 μg/mL, 2.5 μg/mL, 5 μg/mL, and 10 μg/mL Bcl-2-ASO. The raw histograms are depicted for the various combinations. Note maximal uptake of FITC–Bcl-2-ASO at 5 to 10 μg/mL at 72 to 96 hours of treatment. Shaded histograms show cells with no FITC–Bcl-2-ASO. (B) Kinetics of Bcl-2-ASO uptake in ARP-1 myeloma cells. Experiment was performed as outlined in panel A except that ARP-1 cells (low Bcl-2) were used. Results are depicted as change in mean fluorescence intensity (MFI) as a function of treatment duration. Uptake was time and dose dependent, with a clear increase in the MFI between 72 and 96 hours, especially at the higher doses of FITC–Bcl-2-ASO. Note the similar kinetics of uptake for both cell lines presented in both panels. Bars are ± SD of at least 3 experiments.
