LC15 growth profile and IL-15 expression after withdrawal from exogenous cytokine support. PBLs were activated with OKT3 and IL-2. Cells were transduced 4 times with IL-15 retroviral supernatants on culture days 2 and 3. Control cells were not transduced or were transduced with a retroviral vector encoding irrelevant proteins. On day 7 of the culture, the PBLs were washed extensively, and 5 × 10⁶ cells from each culture were plated in fresh media, in the absence of exogenous cytokine. (A) Viable cells were enumerated every 4 to 7 days by trypan blue exclusion; concurrently, the cell culture media were refreshed by replacing half of the spent media with fresh media. Cells were maintained at a density of 1 × 10⁶ cells/mL. (B) Twenty-two other experiments were performed in which OKT3-activated PBLs were transduced with the IL-15 retroviral vector and subsequently withdrawn from IL-2. The growth of these cultures, and the growth of LC15, was plotted over the first 40 days after IL-2 withdrawal. (C) LC15 cell culture media were periodically assayed for IL-15 content by ELISA.