Rapid recovery of CD4⁺CD25⁺Foxp3⁺T_Reg cells in recipients that received T_Reg cell–depleted donor lymphocytes after transplantation. (A,B) Syngeneic transplantations were set up as described in Figure 1A. On day 0, recipients bearing A20-HA, A20-WT, or no tumors (No tumor) were transferred with either T_Reg cell–depleted (Donor T_Reg depletion) or unmanipulated (no T_Reg depletion) donor lymphocytes. Fourteen days after transplantation, mice were vaccinated, an, 7 days after vaccination, splenocytes were stained with anti-CD4, anti-CD25, and anti-Foxp3 antibodies. The percentage of CD4⁺CD25⁺Foxp3⁺T_Reg cells among total CD4⁺ T cells in A20-HA–bearing mice is indicated (A). The mean absolute number (± SD; n = 4) of CD4⁺CD25⁺Foxp3⁺ T cells per spleen in each group is indicated (B). (C,D) Syngeneic transplantations were set up as described with the addition of HA-specific CD4⁺ T cells (Thy1.1⁺Thy1.2⁺) into donor tumor-bearing mice. On day 0, recipients bearing A20-HA, A20-WT, or no tumors (No tumor) were transferred with either T_Reg cell–depleted (Donor T_Reg depletion) or unmanipulated (no T_Reg depletion) donor lymphocytes. Twenty-one days later, the percentage of HA-specific T_Reg cells (Thy1.1⁺Thy1.2⁺CD4⁺CD25⁺Foxp3⁺) among total clonotypic (Thy1.1⁺Thy1.2⁺) CD4⁺ T cells in A20-HA–bearing mice is indicated (C). The mean absolute number (± SD; n = 4) of HA-specific T_Reg cells per spleen in each group is indicated (D). Representative results of 2 independent experiments are shown.