Figure 3. Expression and characterization of the antigen and verification of binding to rhesus macaque DCs.

293 T cells were transiently transfected with grading doses (from 3 to 0,003 µg) of scISO-p27 or scDEC-p27 expressing plasmids (pV-scISO-p27 or pV-scDEC-p27). Additionally, 293 T cells were transfected with 3 µg of a GFP-expressing plasmid as negative control. Supernatants were harvested 48 h after transfection and secreted fusion proteins were detected by Western Blot analysis to confirm comparable expression levels (A). Monocyte-derived rhesus macaque DCs were incubated with supernatants of transfected 293 T cells, and bound fusion proteins were visualized by using an Alexa647-labeled α-OLLAS antibody. Subsequent FACS-analyses are shown for scDEC-p27 (filled grey histogram) and scISO-p27 (open black histogram) for immature and mature DCs (B).