Expression of langerin and CD1a by monocytes and DCs. (A) Sorted cells cultured for 3 days in conditions as indicated showing expression of CD1a and extracellular langerin. The experiment was repeated 5 times except for the panels with X-Vivo, which were repeated 3 times. Four subsets of cells were collected from 1 donor, different in each experiment. (B) Time course of expression of CD1a and langerin double-positive cells showing a peak at 3 days and gradual decline in the percentage of positive cells up to 7 days of culture. Mean ± SEM of 5 experiments with different donors. (C) Recovery of viable cells under each condition at 7 days of culture, estimated by the total number of 4,6 diamidino-2-phenylindole (DAPI)–negative cells recorded when the culture was analyzed and run to dryness on the cytometer. Ten-thousand cells were added to each well as counted by the sorter but typically resulted in 6000 to 8000 viable cells at the start of the culture. There were no statistically significant differences between each condition for a given subset of cells. (D) Upper plot: percentage of langerin⁺ cells derived from GM-CSF+TGFβ (open bars), GM-CSF+BMP7 (gray bars), and GM-CSF+TGFβ and BMP7 (black bars) after 3 days of culture. Lower plot: percentage of langerin^high cells derived from GM-CSF+TGFβ (open bars), GM-CSF+BMP7 (gray bars), and GM-CSF+TGFβ and BMP7 (black bars) after 3 days of culture. Mean ± SEM of 5 experiments with different donors. Gating of langerin⁺ and langerin^high cells is illustrated using CD1c⁺ DCs incubated with GM-CSF and BMP7 as an example. There were no statistically significant differences between each condition for a given subset of cells. *P < .01 compared with corresponding CD14⁺ monocyte culture. Differences in langerin induction between monocyte subsets and pDCs for a given culture condition were not significant.