Cross-presentation of FcγR-targeted antigen by human BDCA-3⁺ DCs requires antigen processing in both the endosomal pathway and by the proteasome. Human BDCA-3⁺ DCs were allowed to process 3 μg pp65-IC in the absence (A) or presence of proteolysis inhibitors indicated (O/N, 37°C; B-G). Next, DCs were washed and cocultured with A2/NLVPMVATVCD8⁺ T cells (4 hours in the presence of Golgi-stop), and IFN-γ production by T cells was measured as a read-out for cross-presentation. Shown are representative plots (top) and summarizing graphs for each inhibitor (n = 4-5). (B) MG132 proteasome inhibitor, 50μM. (C) Lactacysin proteasome inhibitor, 100μM. (D) Epoxomicin proteasome inhibitor, 10μM. (E) chloroquine endosomal acidification inhibitor, 50μM. (F) Leupeptin lysosomal cysteine protease inhibitor, 15μM. (G) Primaquine recycling endosome inhibitor, 50μM.