Growth and functional analysis of T cells from patients with B-cell NHL after culture with γc cytokines. (A) Purified CD3⁺ T cells from tumor-containing lymph nodes of 10 patients with B-NHL were seeded at the initial concentration of 1 × 10⁶/mL and cultured for 3 weeks with autologous B-tumor cells in the presence of IL-2 at 3 ng/mL (LD–IL-2) or at 300 ng/mL (HD–IL-2). IL-15 was used at 10 ng/mL. Irradiated autologous bone marrow–derived cells (BM), autologous monocytes (MØ), or autologous monocyte-derived DCs (DCs) were also added in some cultures. Total number of viable cells was assessed by trypan blue exclusion and the number of CD3⁺ cells by immunophenotyping on days 0, 7, 14, and 21 of culture. Viable cells were always greater than 97% CD3⁺ T cells. (B) T cells isolated from tumor-containing lymph nodes were cultured for 3 weeks with HD–IL-2 alone (1), or HD–IL-2 plus IL-15 (2), or HD–IL-2 plus IL-15 and irradiated autologous bone marrow–derived cells (3). The T-cell cultures were then evaluated in a 24-hour IFN-γ–ELISPOT assay in response to no stimulus (white bars, negative control), autologous (light gray bars), or allogeneic (dark gray bars) B-cell tumor, or IL-2 at 300 ng/mL (black bars, positive control). (C) T cells were cultured with HD–IL-2 plus IL-15 and autologous bone marrow–derived cells and then tested for lysis of autologous B-cell tumor preincubated (empty bars) or not (black bars) with a mAb to a monomorphic determinant of HLA class I antigens. T cells were also tested for lysis of an allogeneic HLA-mismatched B-LCL (gray bars). (B-C) Error bars indicate SD of the mean. Statistical analysis was annotated as follows: *P < .05, **P < .01.