Figure 3. Inefficient production of CD4+ and CD8+ SP cells in B-Raf-deficient thymocytes. (A) Single-cell suspensions of thymocytes (upper panels) and spleen cells (lower panels) were isolated from age-matched B-Raf+/+ and B-Raf−/− FL chimeric mice 8 wk after transfer, stained with Ab against CD4, CD8, B220 and TCR-β chain, and analyzed by flow cytometry. Lymphoid cells were gated based on SSC and FSC. The percentages of total thymocytes and spleen cells that fall into each region are indicated. (B) Cell surface expression of CD44 on peripheral T cells isolated from spleen and lymph nodes of B-Raf +/+ (filled gray) and B-Raf−/− (bold line) FL chimeric mice. (C) Thymi were harvested 8–12 wk after transfer and the cell suspension enumerated. After calculating the percentage of lymphoid cells (~85% of total) on the basis of FSC/SSC, the numbers of the indicated population were calculated (mean ± SD). Data are based on four to five samples of thymus (*p<0.01). All data are representative of at least three B-Raf +/+ and B-Raf−/− chimeric mice in two independent experiments.