Treg depletion during SCM increases the total number of polyfunctional CD8 T cells in donor grafts. FoxP3-DTR mice were treated with diphtheria toxin (DT) or vehicle control every other day for 1 week. Mice in the mobilization group were treated with G-CSF and AMD3100. The mice were euthanized and T cells in the BM, blood, and spleen were harvested and analyzed by flow cytometry. (A) Fold changes in the number of CD8 and CD4 T cells. The fold change was calculated using the mean T-cell numbers from the non-Treg-depleted, nonmobilized group (grey triangles). (B) Representative FACS plots and frequency of memory CD8 T cells in the blood. (C) Percentage of activated DNAM-1⁺TIGIT⁺ and CD38⁺CD101⁻ CD8 T cells in the blood and spleen. (D) Frequency of IFNγ⁺TNF⁺ CD8 T cells in the blood and spleen (n = 3-8 per group from 2 independent experiments). Grey triangle: non-Treg depleted and nonmobilized, Green triangle: non-Treg depleted and mobilized, Blue square: Treg depleted and nonmobilized, Red circle: Treg depleted and mobilized. Data represents mean ± SEM. One-way ANOVA for multiple sample comparisons. ∗P < .05, ∗∗P < .01, ∗∗∗P < .001, ∗∗∗∗P < .0001.