Effect of in vivo FLT3 ligand administration on B lymphopoiesis. FL^−/− mice (27-33 weeks old) were treated with FL (0.4 μg or 10 μg per mouse) or PBS (control) every second day for 2 weeks, at which time mice were killed and BM cells counted and analyzed for presence of distinct stages of B-cell development in the BM. (A-C) FACS profiles from representative mice treated with PBS (control) or 10 μg FL. Numbers indicate percentage of total BM cells within the indicated gates or quadrants. (D) Total number of B-cell progenitor subsets in the BM (per 2 tibiae and 2 femora) of mice treated with PBS (control), 0.4 μg, or 10 μg FL (based on the gated cell populations indicated in A-C). Mean (SEM) numbers from 3 mice per group. (E) PB B220⁺IgM⁺ cell analysis of mice treated with PBS (control) or 10 μg FL. Numbers in FACS plots indicate frequency of B220⁺IgM⁺ B cells among total blood cells. Bar graphs show mean (SEM) values from 3 mice per group. (F) Lethally irradiated adult WT and FL^−/− recipients were transplanted with 10⁶ WT and FL^−/− BM (test) cells, in competition with 10⁶ WT cells as indicated. Donor- and competitor-derived blood cells could be separated based on expression of different CD45 isoforms (Materials and methods). Mice were analyzed for PB B cell (B220⁺) reconstitution at 3 and 16 weeks post-transplantation. Mean (SEM) values from 14 recipient mice in each group, from 2 independent experiments. (G) BM (test) cells from adult FL^−/− (CD45.2) mice were transplanted into lethally irradiated primary (1⁰) FL^−/− (CD45.1) recipients. Four weeks after transplantation, mice were killed and ½ femur equivalent (FE) of BM cells from pooled 1⁰ recipients were transplanted into 3 different groups of lethally irradiated secondary (2⁰) recipients as indicated: either FL^−/− CD45.1 mice without competitor, WT CD45.1 mice without competitor, or FL^−/− CD45.1 mice along with 10⁶ WT CD45.1 competitor BM cells. PB was analyzed at 6 weeks post-transplantation for donor-derived B-cell reconstitution. Bar graph shows mean (SEM) total donor-derived B220⁺IgM⁺ cells per mL blood from 5-7 secondary recipient mice per group.