Btk^−/−PLCγ2^−/− mice have reduced peripheral B-cell populations compared with single mutants. (A) Splenocytes from mice of various genotypes were analyzed via fluorescence-activated cell sorter (FACS) using fluorochrome-conjugated anti-B220 and anti-IgM antibodies. Numbers indicate percentage of cells in lymphocyte gate. Figures shown were representative of at least 4 independent experiments. (B) Enumeration of splenic B cells (mean ± SEM) in various mice. Number of cells is calculated based on total cell count and fraction of B220⁺IgM⁺ cells present in the tissues as revealed by FACS staining. Wild-type (WT), n = 6; Btk^−/−, n = 6; PLCγ2^−/−, n = 5; and Btk^−/−PLCγ2^−/− (DKO), n = 4.