Binding of Rituxan and FITC-Rituxan monomers, homodimers, and trimers to various cell lines and their dissociation from cell lines.
(A) Binding of FITC-Rituxan monomers to different cell lines: Daudi (●); DHL-4 (○); Ramos (▾); Raji (▿); Namalwa (▪); and Namalwa/MDR1 (■). (B) Binding of Rituxan monomers (●), homodimers (○), and trimers (▾) to Ramos cells. In this procedure, 1 × 106 cells per milliliter were incubated with different concentrations of mAbs (0.1 to 100 μg/mL) for 30 minutes on ice; then excess mAbs were washed out, and cells were treated with FITC-GAMIg for 20 minutes on ice, washed, and analyzed in FACScan. The percentage of positive cells was determined for each mAb, and values were plotted against the concentration of mAbs. The amount of mAb needed to saturate 50% of cells was determined and used to compare the relative avidity of monomers, homodimers, and trimers. (C) Dissociation of FITC-Rituxan monomers (●), homodimers (○), and trimers (▾) from Ramos cells. Cells at 1 × 106/mL were treated with 10 μg/mL (a saturating concentration) of FITC-Rituxan monomers, homodimers, or trimers on ice for 30 minutes, then washed, and analyzed by FACScan. The percentage of positive cells was determined at this point, which was considered time 0. Cells were incubated at 37°C for 2, 4, 6, 24, 48, and 72 hours, and at each time point, samples of cells were taken, washed, and analyzed by FACScan for FITC-positive cells. The results were plotted against time. This is 1 of 2 experiments carried out.
