IkarosDN fails to antagonize the effect of IRF4 on light chain rearrangement and transcription in pre-B cells. (A) IRF4,8^−/− pre-B cells were cultivated in Opti-MEM medium containing 5 ng/mL IL-7 and infected with virus expressing IRF4, Ikaros, and Aiolos. At 24 hours after infection, the infected cells were washed and replated on top of an irradiated S17 stromal layer in the absence of IL7 for another 36 hours. The cells were then stained with antibodies against kappa and CD25 and analyzed by FACS. The expression of kappa and CD25 in the GFP^hi cells are shown. (B) IRF4,8^−/− pre-B cells were coinfected with IRF4 (YFP) and IkarosDN (GFP) as described in Figure 5. At 24 hours after infection, IL-7 was removed from the culture media. After another 72 hours, the cells were stained with anti-kappa antibody and Hoechst dye and analyzed by FACS. The infected cells were analyzed using strategies described in Figure 5. The result is a representative of 3 independent experiments. Numbers in quadrants represent percentages of gated cells.