Inhibition of donor antihost alloresponses by BYK-1 treatment. (A-C) BDF1 recipient mice were injected intravenously with 5 × 10⁷ donor B6 spleen cells. The recipient mice were treated intraperitoneally with 200 μg of BYK-1 (■) or control Ig (□) on days 0, 3, and 6. (A) On day 9, recipient spleen cells were harvested and assessed for CTL activity against P815 (H-2^d) and EL4 (H-2^b) cells by a standard 4-hours ⁵¹Cr releasing assay. (B) On day 9, recipient spleen cells were stained with anti-H-2K^d mAb, together with either anti-CD4 or anti-CD8 mAb, and analyzed by flow cytometry. Percentages of donor CD4⁺ or CD8⁺ T cells (upper left quadrant) in the recipient spleen are shown. The experiment was repeated more than 3 times, and representative data are shown. P < .05 (control Ig vs BYK-1). (C) On days 9 and 12, absolute numbers of donor CD4⁺ and CD8⁺ T cells in the recipient spleen were assessed. Each column represents average plus or minus SD of donor T-cell number after the treatment with control Ig (white bar) or BYK-1 (black bar). (D) BDF1 recipient mice were exposed to irradiation (9 Gy) and subsequently injected intravenously with 5 × 10⁶ donor B6 spleen cells. The recipient mice were injected intraperitoneally with 200 μg BYK-1 (black bar) or control Ig (white bar) on days 0 and 3. On day 6, mesenteric lymph node cells harvested from the recipient mice were standardized for numbers of CD4⁺ T cells (5 × 10⁴ per well) and stimulated in vitro with 5 μg/mL immobilized anti-CD3 mAb and 2 μg/mL anti-CD28 mAb. After 24 hours, the culture supernatants were harvested, and the levels of interferon-γ (left panel) and IL-4 (right panel) were measured by enzyme-linked immunosorbent assay. Each column represents average plus or minus SD. P < .05 (control Ig vs BYK-1). Data are representative of 3 independent experiments.