Figure 7

PsL-EGFmAb signals to DCs through DC-SIGN. A-C, We generated human monocyte-derived DCs by incubating MACS-isolated CD14⁺ monocytes from healthy donor blood with GM-CSF and IL-4 for 5 days (imDCs). Flow cytometry detected DC-SIGN expression on imDCs using fluorescence-labeled DC-SIGN mAb (A) and PsL-EGFmAb (B) respectively. C, imDCs pre-incubated with anti-DC-SIGN goat antiserum were also included to prevent binding of PsL-EGFmAb to DCs before staining with PsL-EGFmAb. D, Western blotting analysis of DC-SIGN in different samples: imDC lysates immunoprecipitated using PsL-EGFmAb-coated beads (left), imDC lysates without immunoprecipitation (middle), imDC lysates immunoprecipitated using mouse IgG-coated beads (right). Experiments were performed in triplicate and the data from one representative experiment is shown. E, Western blotting analysis of DC-SIGN from different samples: The lysates of imDCs with DC-SIGN siRNA were immunoprecipitated using PsL-EGFmAb-coated beads (left), the lysates of imDCs with DC-SIGN siRNA without immunoprecipitation (middle), the lysates of imDCs with DC-SIGN siRNA immunoprecipitated using mouse IgG-coated beads (right).