CD40L expression is diminished in CD4⁺ T-cell and lymphocyte cultures. PBMCs were plated at 1 × 10⁶ cells/mL. Autologous CD4⁺ T cells (> 95% purity) or elutriated lymphocytes were plated at the same density as that present in unfractionated PBMCs. (A) CD4⁺ T cells and PBMCs were stimulated in an identical fashion as indicated. Cells were harvested and stained for CD40L expression at the indicated times. Data are mean ± SE of 6 donors. (B) Purified CD4⁺ T cells were plated in conditioned media harvested from 6-hour-stimulated PBMCs. Cultures were left unstimulated or TCR stimulated for 6 hours. Autologous PBMCs were cultured in an identical fashion. Cells were harvested and stained for CD40L expression. Results from a single donor are shown. (C) Lymphocytes and PBMCs were stimulated in an identical fashion as indicated. Cells were harvested and stained for CD40L expression at the indicated times. Data are mean ± SE of 3 donors.