SA anti-CD28 treatment induces accumulation of mature thymocytes in HIS (BALB-Rag/γ) mice. (A) Number of human CD45⁺ thymocytes at 3, 6, and 9 days after SA anti-CD28 mAb (mean ± SEM; □ indicates control animals; ▪, treated animals; ^*P < .05; ^**P < .01). The effect on size (forward scatter [FSC]) and granularity (side scatter [SSC]) at day 6 is shown in overlay histograms, comparing thymocytes from IgG1 control (thin line) and SA anti-CD28 treated (filled histogram) mice. (B) Surface expression of CD25, GITR, and CD45RO was compared between control (thin line) and treated HIS (BALB-Rag/γ) mice (filled histogram) in the DP, SP4, and SP8 thymocyte subsets. Values in each overlay histogram indicate the percentage of positive cells in control (top) versus treated mice (bottom). (C) The relative distribution of thymocytes between the DN (▥), DP (▪), SP4 (□) and SP8 (▤) was calculated at days 3, 6, and 9 in control and treated mice. Representative FACS profiles of CD4 and CD8 expression obtained at day 6 are shown for control (top) and treated mice (bottom). (D) Similarly, the relative distribution of thymocytes according to CD1a and CD3 expression is shown, from immature CD1a^-CD3^-(▥), CD1a⁺CD3^-(▪), and CD1a⁺CD3⁺ (□) to mature CD1a^-CD3⁺ T cells (▤). Numeric results correspond to the mean values obtained from 4 mice, and are representative of 1 experiment out of 3.