C-Rel synergistically enhances the proliferation of normal B lymphocytes upon CD40 ligand stimulation. (A) G₀ B cells were purified from healthy donors and the purity was assessed by flow cytometry as described in “Material and methods.” Cytosolic and nuclear fractions of normal B cells were extracted with or without CD40 ligand activation for 24 hours and subjected to Western blotting. C indicates cytosolic fraction; N, nuclear fraction. (B) [³H] thymidine incorporation assay was performed after transfection of c-Rel into normal B cells. Twenty-four hours after transfection, live cells were counted and same number of live cells (20 000) were aliquoted and treated with/without CD40 ligand for 24 hours, and then pulse labeled with [³H] thymidine overnight. Cells were harvested and signals were counted in a Beckman LS3800 liquid scintillation counter. Error bars indicate SD of triplicate samples.