Antitumor activity after CD4⁺Trp1⁺ T cell transfer is specific to B16/BL6 melanoma and does not prevent growth of an unrelated tumor. (a–e) Mice were challenged with B16/BL6 melanoma and EL-4 in opposing flanks (a and b) or coinjected in the same flank (c–e). 10 d after tumor challenge, all mice received 5 Gy of RT and anti–CTLA-4 mAb. Half of the mice were also treated with 50,000 CD4⁺Trp1⁺ T cells and all mice were monitored for tumor growth (a–c). (d) Representative images of mice challenged with B16/BL6 and EL-4 in the same site shows the growth of a pigmented tumor in the absence of CD4⁺Trp1⁺ transfer and growth of a nonpigmented tumor after administration of 50,000 tumor-reactive CD4⁺Trp1⁺ T cells Data are representative of three independent experiments (n = 5 mice per group). To verify that B16/BL6 melanoma is rejected in the coinjection setting, mice were challenged with EL-4 and B16/BL6-luciferase in the same site, and light emission was determined over time after tumor challenge and triple therapy. (e) Representative images of luciferase signal disappearing from the tumor after therapy. (f) Quantification of luciferase signal and tumor growth. Data are representative of two independent experiments (n = 5 mice per group). Error bars represent means ± SD.