Proliferation of CD4⁺FoxP3⁺ Treg population after ablative conditioning. B6 mice were subjected to 9.5 Gy TBI. On the following day, mice received 5 × 10⁶ T-depleted BM cells isolated from Thy1.1 congenic B6 mice. Mice were killed 20, 35, and 56 days post-BMT and spleen and lymph nodes subjected to FAC analysis. Four days before death, mice were placed on BrdU (0.8 mg/mL) drinking water. (A) Representative dot plot showing the Thy1.1 (donor BM) staining and IgG of BrdU staining of the gated CD4⁺Foxp3⁺ cells in the spleen (left panels) and lymph nodes (right panels). (B) Percentage BrdU⁺ cells in the host (Thy1.1⁻) and donor (Thy1.1⁺) CD4⁺Foxp3⁺ and CD4⁺Foxp3⁻ compartments in the spleen (left) and lymph nodes (right). (C) Wild-type B6 mice (n = 3) were placed on BrdU drinking water for 4 days, killed, and the spleen and lymph nodes subjected to FACS analysis. Representative dot plot showing CD4 and Foxp3 expression with corresponding histograms illustrating staining by isotype control (IgG) and anti-BrdU mAb of the gated CD4⁺FoxP3⁺ and CD4⁺Foxp3⁻ populations in the spleen (left) and lymph nodes (right). Data in panel B are mean plus or minus SEM for 3 mice per group per time point.