Reduced polarization and migration of human and murine WASp-deficient B cells. (A) B cells from WAS patients, controls, or the WAS^I294T patient were stimulated with IL-4 or anti-CD40 mAbs for 20 hours. Polarized cells were defined as having a tapered cell body and uropods, whereas nonpolarized cells were spherical in shape. Background values were between 15% to 20% and subtracted from the IL-4 and anti-CD40 values. Results are presented as mean values of triplicate cultures. Error bars indicate one SD. (B) Migration of murine B cells toward CXCL13, CXCL12, CCL19, or medium containing FCS (referred to as background) was determined in a Transwell assay. B cells were purified from wild-type (WT) or WASp-deficient (WASp–) spleens. B cells (100 000) were loaded on the filter in the upper chamber. Cells were allowed to migrate for 2 to 6 hours, after which the number of cells that had migrated to the lower chamber was determined. This observation is representative of at least 3 similar experiments. (C) B cells from WAS patients, controls, or the WAS^I294T patient were allowed to migrate toward CXCL13 or medium containing FCS (referred to as background). The migrating cell population was enumerated after 2 hours as in panel A. Results are presented as mean values of triplicate wells, error bars represent one SD, and brackets indicate a significant difference of P < .01 using a 2-tailed t test. (D) F-actin content in EBV-transformed control or WAS^I294T B cells. Note the increased F-actin content in WAS^I294T B cells. Upper images show control cells, and lower images, WAS-I294T cells.