Specificity of denileukin diftitox for deletion of Treg cells versus activated T cells. A single intraperitoneal injection of 0.75 μg denileukin diftitox was administered to C57BL/6 mice (n ≥ 10/group). Flow cytometry analysis was performed on tissues on days 1, 3, 5, and 10 after injection. (A) Spleen, bone marrow, and peripheral blood cells from individual mice were stained for CD4⁺CD25⁺Foxp3⁺ Treg cells. Individual values and means for percentage of Treg cells are represented as percentage relative to naive mice; actual values for percentage Treg cells of total CD4⁺ T cells in the naive group averaged 8.9% for spleen (range, 5.6%-12%), 8.4% for bone marrow (range, 3.7%-11.1%), and 5.0% for peripheral blood (range, 2.2%-7.8%). Statistical evaluation was performed by one-way ANOVA using GraphPad Prism (GraphPad Software, San Diego, CA). (B) Spleen cells from individual mice were stained for CD4⁺CD25⁺Foxp3⁻ and CD8⁺CD25⁺ activated T cells. Mean values (± SD) are shown on the graph as percentage relative to naive mice. Actual values for percentage activated CD4⁺ of total CD4⁺ T cells and activated CD8⁺ of total CD8⁺ T cells averaged 1.4% (range, 0.6%-2.9%) and 0.9% (range, 0.2%-1.6%), respectively, for the naive group.