Effect of B cell-targeted expression of PKCβII on B cell populations in the peritoneum of Eµ-PKCβIItg and wt mice. Single cell suspensions prepared from peritoneal wash of Eµ-PKCβIItg and wt mice were stained with antibodies to B220, IgM, IgD, CD43, CD24, and CD5 and analyzed by flow cytometry. Quantitative comparisons were then made of the percentage B220+ cells within the following gates (A) IgD+ IgMdim, (B) IgDdim IgM+, and (C) IgDdim IgM+ CD43+ CD24+ (B-1 cells) for wt and Eµ-PKCβIItg mice as defined by the strategy illustrated in Supplementary Figure 3. Black dots in these graphs refer to wt and homozygous Eµ-PKCβIItg progeny derived from mating heterozygous Eµ-PKCβIItg mice. Red dots refer to wt mice with similar genetic backgrounds (C57BL/6) that are alike in terms of age to the transgenic mice. (D) Comparison of IgM levels in serum derived from Rag2 deficient, wt, and heterozygous and homozygous Eµ-PKCβIItg mice. Statistical analysis for parts (A) (**P = 0.0040), (B) (*P = 0.042), and (C) (***P = 0.0010) was performed using a Mann–Whitney U test, for part (D) analysis was done using a one-way ANOVA and Tukey’s multiple comparisons test (P = 0.040).