Characterization of Runx1-deficient B-cell progenitors. (A) Shown is the analysis of pre-B- and pro-B-cells (gated on CD19⁺Lin2^negIgL^neg) isolated from BM of mice with the indicated genotype and examined for expression of cKit, CD25 (IL2Rα), or for size (FSC). Gates for pro-B cells were set using cells isolated from Cd79a^hCre/hCre mice. (B) FACS analysis of sorted pro-/pre-B-cells isolated from mice with the indicated genotypes. Sorted cells were plated on OP9 cells supplemented with IL7 and analyzed after 3 days. Half of the sorted cells were stained with carboxyfluorescein diacetate succinimidyl ester (CFSE) before plating. The proliferation index (PI; based on proliferating cells only) and the division index (DI; all viable cells) were determined by calculation of the fluorescent intensity of the CFSE-stained cells. Results shown are representative of 3 independent experiments. To obtain sufficient cells for sorting, staining, and consequent culture, 3 Runx1-deficient mice were used per experiment. (C) Expression analysis of sLC and LC transcripts (λ5 encoded by Igll1 and rearranged VJ-κ transcripts, respectively) was performed on sorted pro-B/pre-B cells from mice with the given genotypes. cDNA levels were normalized to Hprt expression levels. (D) Schematic representation of the observed blocks in B-cell development in Runx1-deficient mice and the impact of Bcl2 on the cell survival of the pro-B- and pre-B-cell compartments, but their low frequency of transition to the immature stage.