Figure 2. Ikaros-deficient pre-B cells grow only on stroma.

a, Flow cytometric analysis of sorted large pre-B cells (CD19⁺CD43⁺BP1⁺) cultured for 7 days stromal-free with limiting serum and IL-7. Differentiation of WT and IkE5^Δ/Δ large pre-B cells is monitored by stage-specific markers. Arrows indicate the direction of pre-B cell differentiation as depicted in Supplementary Fig. 1a. b, Growth of WT and IkE5^Δ/Δ large pre-B cells in high, low, and no (5, 0.05, and 0 ng/ml, respectively) IL-7 concentrations under stromal-free conditions (left) or with OP9 BM stroma (right). The mean absolute number of cells obtained in stromal-free (n=5) and stromal-containing (n=4) cultures with replicates for each experiment is shown in a line graph ± s.d. Asterisks denote significant differences between WT and mutant cells (*P < 0.05, **P < 0.01, two-tailed Student's t-test). c, Mean percentage ± s.d. of apoptotic (AnnexinV⁺) WT and IkE5^Δ/Δ large pre-B cells in stromal-free cultures as in Fig. 2b, left panel. d, Cell cycle stage distribution (mean percentage ± s.d. of cells in S+G2+M) of WT and IkE5^Δ/Δ large pre-B stromal cultures as in Fig. 2b, right panel. Asterisks in c and d denote significant differences between WT and mutant cells (*P < 0.05, **P < 0.01, two-tailed Student's t-test). e, Cell cycle kinetics of WT and IkE5^Δ/Δ large pre-B cells grown on stroma as measured by BrdU pulse-chase. The mean fluorescence intensity (MFI) of BrdU staining is shown at 45 min of pulse and after 48 h of chase.