Stimulation of human T cells by in vitro–differentiated DCs after engraftment of transduced CD34⁺ cells.

In this MLR T-cell proliferation assay, human PBLs (as effectors) were harvested from an unrelated (allogeneic) healthy donor and seeded (2 × 10⁵/well) in 96-well plates in triplicate. Differentiated human DCs from BM (Figure 4) or spleen (SP) of the NOD/SCID mice that received transplants of DR.GFP-, EF.GFP-, or mock-transduced CD34⁺ cells were added in a serial dilution as APC stimulators. DC and T cells were cultured together for 3 days at stimulator-effector ratios of 1:10, 1:80, and 1:640. The stimulation of lymphocyte proliferation was determined by [³H]thymidine incorporation after the 3-day culture and 18-hour pulsing. The mean incorporations (counts per minute, cpm) from 3 triplicates were calculated and plotted, in comparison to the DCs differentiated from CB CD34⁺ cells without the transduction and engraftment steps (CB CD34-DC).