CD25 depletion before cell culture preserves the allogeneic responses of cultured murine and human T cells. (A) Induction of GVHD in mice: mT cells were initially depleted of CD25⁺ cells before culture and 6 days later 10⁷ (▪, n = 5) and 3 × 10⁷ (□, n = 4) day 6 cultured cells were injected into B6D2F1 irradiated mice. Controls were mice receiving 10⁷ CD25-depleted noncultured cells (▵, n = 8), 10⁷ nondepleted day 6 cultured cells (▴, n = 13), and noninjected mice (X, n = 11). (B) Proliferative response of hT cells to EBV B cells: total hT cells (▪) and initially CD25-depleted hT cells (□) were plated either directly (day 0) or after culture with CD3 + IL-2 (day 6) in the presence of allogeneic irradiated EBV B cells (T cells/EBV B cells ratio: 5:1). The proliferative response was measured by ³H-Tdr incorporation. Results presented as mean cpm ± SEM of triplicate cultures and are from 1 representative experiment of 3 using cells from 3 different donors.