LC/NK cultures supplemented with allogeneic CD4⁺ T cells, IL-2, or IL-12 are able to activate NK cells. Size (forward scatter [FSC]), granularity (side scatter [SSC]), and surface expression of the CD56 and CD16 epitopes were quantified by flow cytometry. FSC and SSC were measured on purified NK cells stimulated for 7 days by Langerhans cells (LC) at an NK-cell-DC ratio of 10:1. Cultures were supplemented with irradiated allogeneic CD4⁺ T cells at an NK/CD4⁺ T-cell ratio of 1:1 (LC + CD4), 80 IU/mL rIL-2 (LC + IL-2), 500 pg/mL IL-12 (LC + IL-12), or nothing (LC). CD56 and CD16 expression was quantified on gated CD3^- lymphocytes, which lacked autofluorescence in channel FL-3 and were located in the FSC/SSC lymphocyte gate (dotted line). Percentages indicate NK-cell blasts, CD16^-CD56^bright, CD16⁺CD56^bright, and CD16⁺CD56^dim cells. LCs maintained NK-cell viability very well (bottom left panel) despite the absence of activation and blast transformation without supplements. Note that most of the peripheral blood NK cells are CD56^dim but not CD56^- directly after isolation from PBMCs or after coculture with LCs. The data are representative of 3 independent experiments.