Figure 5.

Anti-IgM response depends on Btk dosage. (A) [³H]thymidine incorporation. Purified B220⁺ cells from spleen were incubated in triplicate for 48 hr with medium alone, 2 μg/ml anti-IgM, 20 μg/ml anti-IgM, or 10 ng/ml PMA and 1 μM ionomycin, then labeled overnight with [³H]thymidine. Counts per minute (cpm) incorporated in untreated or anti-IgM treated cells were normalized to the PMA and ionomycin response, which was similar for all cell types. The number of mice tested per group is: medium alone, 9; 2 μg/ml anti-IgM, 3; 20 μg/ml anti-IgM, 9; PMA and ionomycin, 9. Data are plotted as mean ± SEM. xid^1xtg is significantly different from both wild type (wt) and xid^2xtg (P < 0.005). (B) BrdUrd incorporation. Total splenocytes were incubated for 48 hr with medium alone, 2 μg/ml or 20 μg/ml anti-IgM, and labeled with BrdUrd for the final 24 hr. Samples were stained with anti-B220 PE and anti-BrdUrd FITC. The percentage of B220⁺ cells that incorporated BrdUrd is presented as mean ± SEM. The number of mice tested per group is: xid, 7; xid^1xtg, 7; xid^2xtg, 7; wt, 10; wt^1xtg, 5; wt^2xtg, 5. xid^1xtg is significantly different from both wt (P < 0.005) and xid^2xtg (P < 0.05). For wt vs. wt^2xtg, 0.05 < P < 0.1.