Loss of Bmf enhances the survival of premalignant Eμ-myc B lymphocytes. For analysis of preleukemic mice, absence of transplantable tumor cells was confirmed by injecting 2 × 10⁶ spleen cells into wt C57BL/6 recipients followed for at least 2 months. (A) Cell number and B-cell subset composition determined by cell counting and flow cytometric analysis of bone marrow (2 femora), spleen, and blood from 4-week-old mice of the indicated genotypes. Data represent means ± SD from 3 to 4 mice per genotype. *P < .05, ** P < .01, ***P < .001. Total B cells (CD19⁺), Pro/pre-B (CD19⁺IgM⁻CD43⁻), T1 (IgM^highCD21⁺), T2 (IgM^highCD21⁺CD23⁺), and mature (IgM⁺D⁺) B cells. (B) Pre-B cells (CD19⁺IgM⁻CD43⁻) sorted from bone marrow and immature (IgM^highIgD^low) as well as mature B cells (IgD^high) sorted from spleens from 4-week-old mice of the indicated genotypes, were cultured up to 48 hours ex vivo. Percentages of surviving cells were determined by annexin-V/propidium iodide staining. Data represent means ± SEM from 3 to 4 independent experiments for each genotype. *P < .05, **P < .01, ***P < .001 compared with Eμ-myc, #P < .05, ###P < .001 compared with wt. (C) Freshly isolated splenocytes from 4-week-old mice of the indicated genotypes were immediately stained with anti-CD19-phycoerythrin together with fluorescein isothiocyanate -annexin-V plus 7-AAD and analyzed by flow cytometry. Percentages of apoptotic cells in the CD19⁺ gate were determined. Data represent means ± SD from 3 independent experiments for each genotype. *P < .05, **P < .01. (D) At 4 hours after in vivo labeling, the percentage of BrdU⁺ CD19⁺IgM⁻ pro/pre-B cells in the bone marrow and of mature BrdU⁺CD19⁺IgM⁺ B cells in the spleen was evaluated by combined cell-surface and intracellular antigen staining. Data represent means ± SD from 2 experiments for each genotype.