Development of long-term, multilineage mixed chimerism. (A) A total of 15 × 106 male 45.2 BM cells were transferred into congenic female 45.1 mice untreated (▴, n = 3) or conditioned with either 5 weekly intravenous injections of 2 × 105 to 5 × 105 DBY-Tregs (•, n = 5) or a single injection of 1 × 105 DBY-Tregs (○, n = 5). As a positive control, female 45.2 BM cells were transferred into congenic female 45.1 mice (X, n = 3). Donor chimerism expressed as a percentage of CD45.2+ cells was analyzed in PBMCs at various time points after BMT (A). Results represent the mean per group ± SEM. (B-C) Mice chimerized for more than 300 days (5 DBY-Treg injections) were killed, and cells from various organs were analyzed by FACS. (B) Splenocytes were stained with CD45.2-biotin/APC-streptavidin, PE-conjugated anti-CD8, anti-CD4, anti-B220, anti-CD11c, and anti–7-AAD. (C) Thymocytes were stained with CD4, CD3, CD45.2, and CD8 (no gate) or with CD3, CD45.2, and CD11c (FACS gated on CD3– 7-AAD– is shown).
