M25_Fab, but not M25, further improves the in vivo mitogenic effect of IL-7-Fc on CD8⁺ T cells. (A) B6.CD45.2⁺ host mice received 3.5 × 10⁶ purified and CFSE-labeled CD45.1⁺CD8⁺ T cells intravenously on day 0 and intraperitoneal injections of PBS, IL-7-Fc (equivalent of 1.5 μg rhIL-7), or IL-7-Fc/M25 (equivalent of 1.5 μg rhIL-7/7.5 μg of M25) on days 2, 4, and 6. On day 8, donor cell proliferation in host LN and spleen was determined by flow cytometry. Shown are CFSE histograms of CD45.1⁺TCRβ⁺CD8⁺ lymphocytes representative of at least 3 experiments with 2 separately analyzed hosts per group. (B) CFSE-labeled B6.CD90.1⁺ LN cells (5 × 10⁶) were adoptively transferred to B6.CD90.2⁺ hosts on day 0. Intraperitoneal injections administered to hosts on days 1, 3, and 5 consisted of PBS, rhIL-7/M25 (1.5 μg/7.5 μg), IL-7-Fc (equivalent of 1.5 μg rhIL-7), IL-7-Fc/M25 (equivalent of 1.5 μg rhIL-7/7.5 μg of M25), or IL-7-Fc/M25_Fab (equivalent of 1.5 μg rhIL-7/equivalent of 7.5 μg M25). Day 7 CFSE profiles of CD90.1⁺TCRβ⁺CD8⁺ LN cells are depicted. Data are representative of at least 3 experiments with 1 to 4 separately analyzed mice per group. (C) Combined data from 2 identical experiments as described in (B), numbers of CD90.1⁺TCRβ⁺CD8⁺ cells recovered for LNs and spleens of mice treated with PBS (solid circles), IL-7-Fc (solid squares), or IL-7-Fc/M25_Fab (solid triangles). Data are representative of 3 similar experiments. *P < .05. (D) B6.CD90.2⁺ hosts received 3 × 10⁶ CFSE-labeled purified CD90.1⁺CD8⁺ T cells intravenously on day 0 and intraperitoneal injections on days 1, 3, and 5 of IL-7-Fc (IL-7 equivalents as indicated) alone or with M25_Fab (M25 equivalents of either 3.75 μg, left; or 7.5 μg, right). Donor cell proliferation in host LNs and spleens was analyzed on day 7; shown are CFSE histograms gated on CD90.1⁺CD8⁺ lymphocytes that are representative of at least 2 experiments with 2 separately analyzed hosts per condition.