Figure 5 | Scientific Reports

Figure 5

From: Heat-inactivated Factor B inhibits alternative pathway fluid-phase activation and convertase formation on endothelial cell-secreted ultra-large von Willebrand factor strings

Figure 5

C3, FB and C5, even in the presence of HFB, bind to GMVEC-secreted ULVWF strings. GMVECs were stimulated with 100 µM histamine containing 20 µg/ml HFB for 2 min, followed by addition of rabbit anti-VWF + chicken anti-rabbit AF-488 (green) for 15 min (also in the presence of HFB). Cells were then p-formaldehyde-fixed and stained with: goat anti-C3 (a–c); goat anti-FB (d–f); or goat anti-C5 (g–i); and secondary donkey anti-goat IgG AF-647 (red). Cell nuclei were detected with DAPI (blue). Graphs show the fluorescent intensities (y-axis) measured along the ULVWF string (x-axis, length in pixels, 1 pixel = 0.114 µm) from merged images of detected VWF with detected C3, FB, or C5 (with HFB present during incubation). The yellow arrows in the merged images indicate the start and end points of the strings. The white arrows in the merged images of complement proteins (647 nm, red) with DAPI detection indicate high 647-intensity values (in order to assist visualization of the measured line). Images were obtained at 60X and were selected from over 130 images.

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