Generation of mice devoid of WASP and N-WASP in B cells. (A) Schematic of the PCR strategy used to monitor conditional targeting of N-WASP. Displayed is a portion of the WT N-WASP allele, the conditionally targeted allele with exon 2 flanked by loxP sites (L2L), and the conditionally targeted allele after Cre-mediated excision in B cells (L). LoxP sites are denoted by black arrowheads. Labeled arrows denote primers used in the PCR strategy to simultaneously detect L2L and L using primers a, b, and c. (B) PCR analysis of N-WASP deletion in splenic B cells from WT, WKO, cNWKO, and cDKO mice. Note that the WKO mouse in this experiment has the N-WASP L2L allele but lacks expression of CD19-Cre for deletion of N-WASP L2L, and therefore expresses N-WASP. (C) N-WASP protein detection by Western blotting in splenic B cells from WT, WKO, cNWKO, and cDKO mice using an Ab for N-WASP (top panel). IgH was used as loading control (bottom panel). The weak expression of N-WASP seen in cells from cNWKO and cDKO mice may reflect the incomplete deletion of N-WASP by CD19-Cre or the contribution of other hematopoietic cells left after B-cell purification (90%-95% B-cell purity). The asterisk denotes a band present in cNWKO B cells that may represent WASP, because the anti–N-WASP Ab shows some cross-reactivity with WASP.