B7 costimulation, Batf3 and perforin-1 were required for optimal tumor-reactive CD8⁺ T-lymphocyte expansion induced by vaccination with gp96-Ig-secreting tumor cells following syngeneic HSCT. BM transplants were performed as in Figure 1 using mice deficient in (A-B) CD80 and CD86 (B7KO), (C) Batf3 (Batf3KO), or (D) perforin-1 (PrfKO) as donors and/or recipients with (A) coinfusion or (B-D) delayed infusion of (A-B,D) 1.0 × 10⁶ or (C) 0.5 × 10⁶ tumor-reactive CD8⁺ T cells (OT-I), and the peritoneal cavity was analyzed 5 days following intraperitoneal vaccination with tumor cells secreting gp96-Ig. (A) Donor and recipient APC contributed to expansion of tumor-reactive CD8⁺ T cells, that were cotransplanted with the BM, following vaccination with gp96-Ig-secreting tumor cells; n = 3 to 9 from a pool of 3 experiments. (B) Only donor APC contributed to expansion of tumor-reactive CD8⁺ T lymphocytes, that were infused 5 days after HSCT, induced by vaccination with tumor cells secreting gp96-Ig; n = 2 to 4 from a pool of 2 experiments. (C) Donor cross-presenting CD11c⁺ CD8α⁺ dendritic cells were required for optimal expansion of tumor-reactive CD8⁺ T cells following vaccination with gp96-Ig-secreting tumor cells; n = 5. (D) Perforin-1 could be supplied by either donor or recipient cells for optimal expansion of tumor-reactive CD8⁺ T lymphocytes following vaccination with tumor cells secreting gp96-Ig; n = 2 to 4 from a pool of 2 experiments.