In vitro TcR triggering enhances B-cell help function. (A) Naive CD62L⁺CD4⁺ T cells purified from the spleen of Balb/C mice were stimulated for 48 hours with plastic-coated anti-CD3 and anti-CD28 mAbs and rested 24 hours in fresh medium. Serial dilutions of irradiated recovered Th cells were incubated with purified B cells (5 × 10⁵ cells/well) and anti-CD3 mAbs (500 ng/mL). Fresh control resting CD62L⁺CD4⁺ T cells were purified at the time of the T-B coculture test. (B) CD62L⁺CD4⁺ T cells were stimulated in neutral culture medium (Th0) or under polarizing conditions, rested, and cocultured with B cells, as in panel A. Culture supernatants were tested on day 7 for IgG₁ and IgG₂a contents. Results are expressed as mean plus or minus SD of triplicates (SD < 5% did not appear in the figure). Similar results were obtained in 2 additional experiments.