Ag uptake by human circulating DC subsets. (A-B) Uptake of soluble antigens (Ags). Human CD1c⁺ mDCs, CD16⁺ mDCs, BDCA3⁺ mDCs and pDCs were incubated with 5μM (■), 10μM (▴), or 50μM (▾) of FITC-labeled gp100_272-300 long peptide. Ag uptake was measured by flow cytometry after 0.5, 1, and 4 hours (A) and 18 hours (B). Extracellularly bound, noninternalized Ag was quenched by the addition of trypan blue. The figures show mean MFI ± SEM of 3 independent experiments performed in duplicate. (C-D) Uptake of particulate Ags. Human CD1c⁺ mDCs, CD16⁺ mDCs, BDCA3⁺ mDCs and pDCs were incubated with Atto647-encapsulating PGLA microparticles for 16 hours. Uptake was analyzed by flow cytometry (C-D) and confocal microscopy (E). (C) The graph shows the mean MFI ± SEM of 3 independent experiments. Depicted in panel E are the merged pictures of the fluorescence of Atto647-containing PGLA particles (red) and surface MHC class II staining (green), magnification 60×. (D-E) The figures show the result from 1 representative experiment.