Antigen expression of TADCs cultured under different conditions. The urothelial carcinoma cell lines J82 and UMUC3 and the melanoma cell lines MelIm and Mel108 were used for MCTS generation. After 4 to 5 days, half of the medium was replaced by a monocyte suspension in IL-4 and GM-CSF (A) or in IL-4, GM-CSF, and TGF beta 1 (B). In some experiments, LPS was added during the last 72 hours to induce DC maturation (B). After 5 to 7 days in coculture, MCTSs were dissociated and stained with mouse anti-human CD45 antibody in combination with antibodies against the indicated antigens. The same staining procedure was performed with control DCs. For flow cytometry analysis, CD45⁺ cells were gated. Panel A shows the mean and SEM of the median fluorescence intensity (isotype subtracted) of at least 3 experiments (CD1a expression: control DCs versus J82 TADCs [P < .001], control DCs vs UMUC3 TADCs/Mel108 TADCs/MelIm TADCs [P < .05], unpaired t test). Panel B represents 1 representative experiment of 3. Quadrant statistics are shown for the top right quadrant (CD45⁺ cells were gated and represent 100%).