β₂GPI association with RAGE on the iDC surface. (A) Scanning confocal microscopic analysis of β₂GPI association with RAGE on iDC surface. (A) Cells were fixed in 4% formaldehyde in PBS and then labeled with a polyclonal anti-β₂GPI Ab and with a monoclonal anti-RAGE Ab, followed by Texas Red-conjugated anti–goat or FITC-conjugated anti–mouse Ab. Panel i shows β₂GPI-RAGE association in untreated (control) cells. One representative cell is shown. Panel ii shows β₂GPI-RAGE association in cells treated with M-β₂GPI. One representative cell is shown. Panel iii shows β₂GPI-RAGE association in cells treated with G-β₂GPI. One representative cell is shown. Panel iv shows β₂GPI-RAGE association in cells treated with G-β₂GPI. A group of cells is shown. (B) Coimmunoprecipitation analysis of β₂GPI association with RAGE on the iDC surface. Cells were incubated with M-β₂GPI or G-β₂GPI and then immunoprecipitated with anti-RAGE mAb. The immunoprecipitates were analyzed by Western blotting using an anti β₂GPI polyclonal Ab. Lane a shows the reactivity of anti-β₂GPI with RAGE immunoprecipitate from untreated cells. Lane b shows the reactivity of anti-β₂GPI with RAGE immunoprecipitate from M-β₂GPI–stimulated cells. Lane c shows the reactivity of anti-β₂GPI with RAGE immunoprecipitate from G-β₂GPI–stimulated cells. Lane d shows the reactivity of anti-β₂GPI with immunoprecipitate with IgG with irrelevant specificity from untreated cells.