The levels of pMst1 were increased in WASP KO B cells after sAg stimulation. WT, WASP KO, Dock8 KO, and WT B cells treated with latrunculin B (Lat)or cytochalasin D (CD) were incubated with AF546-mB-Fab′-anti-Ig without (−) or with streptavidin (sAg) at 4°C, washed, and warmed to 37°C for varying lengths of time (n = 4). (A-D) After fixation and permeabilization, the cells were stained for pMst1 and analyzed by using confocal microscopy. (E) The Pearson’s correlation coefficients between BCR and pMst1 staining in sAg-stimulated cells were determined by using NIS-Elements AR 3.2 software. (F) Western blot analysis of the Mst1 in WT and WASP KO B cells. (G) Flow cytometry analysis of pMst1 in WT, WASP KO, Dock8 KO, and WT B cells treated with latrunculin B or cytochalasin D stimulated with sAg. (H) Coimmunoprecipitation was performed with B cells from C57BL/6 WT mice. The splenic B cells were stimulated with sAg, and the cell lysates of these cells were incubated with control immunoglobulin G (IgG; lane 1, serving as negative control) or anti-Mst1 (lanes 2 and 3); the immunoprecipitated (IP) protein complex and the cell lysates were then examined by using western blot. Shown are the results from 3 independent experiments and representative images at indicated times and the average values (± standard deviation) of ∼50 cells from 3 independent experiments. Scale bars, 2.5 μm. One-way analysis of variance with the Tukey test was used to perform multiple group comparisons; *P < .01.