Effect of RTX on BCR expression and proteasome-mediated degradation. (A-B) BCR expression at the cell surface was evaluated by flow cytometry on fixed cells pretreated with RTX (10 μg/mL, 16 hours) and then stained with Cy5-conjugated anti-IgM antibody (RL, 1G11, 2B11) or Cy5-conjugated anti-IgG antibody (DOHH2). (Aii) Results are expressed as percentage of untreated cells and are the mean of 3 independent experiments ± SD. P < .005 compared with untreated cells. (C) Total BCR expression level was analyzed by flow cytometry with fixed and permeabilized FL cells (Ci) or by Western blot (Cii). (Ai,B,C1) Untreated and treated cells are represented by gray and black histograms, respectively. Histograms are representative of at least 3 independent experiments. (D) RL cells were cotreated with MG132 (10μM) and RTX (10 μg/mL) for 16 hours. BCR expression was analyzed as described in panel A. Results are expressed in percentage of untreated cells and are the mean of 3 independent experiments ± SD. *P < .002 compared with untreated cells. (E) RL cells were cotreated with MG132 (10μM) and RTX (10 μg/mL) for 16 hours. BCR was then stimulated with anti-IgM antibody (10 μg/mL, 5 minutes) and Syk phosphorylation (Y525) was evaluated by flow cytometry. Results are expressed in MFI and are the mean of 3 independent experiments ± SD. *P < .002 compared with BCR-stimulated cells.