Aortic endothelial cells obtained from a normal dog maintain a normal Weibel-Palade body distribution of VWFpp, VWF, and P-selectin while canine VWD aortic endothelial cells neither express VWF nor contain Weibel-Palade bodies. (A-I) Endothelial cells were harvested from the aortas of normal dogs, grown on gelatin-coated slides, fixed, permeabilized, immunostained, and examined by confocal microscopy as described in “Materials and methods.” Cells immunostained with a mouse monoclonal anti-PECAM (A) and a polyclonal anti-VWF (B) demonstrated a homogeneous population of endothelial cells. Staining with a mixture of several mouse monoclonal anti-VWFpp (D) and a polyclonal anti-VWF (E) demonstrated the presence of both proteins in granules where they colocalized (F, colocalization is indicated by yellow pixels). Immunostaining of the normal endothelial cells with the mouse monoclonal anti-VWF, AVW-1 (G), and rabbit anti–P-selectin (H) revealed that VWF was colocalized with P-selectin in granules (I). The aortic endothelial cells from normal dogs maintain characteristic Weibel-Palade bodies. (J-R) Endothelial cells were harvested from the aortas of type 3 VWD dogs, cultured, immunostained, and examined by confocal microscopy. Cells were dual-labeled with mouse monoclonal anti-PECAM (J) and polyclonal anti-VWF (K). The homogeneous population of endothelial cells was VWF-negative. Neither VWF nor VWFpp could be detected in these cells by immunostaining with a mix of several mouse monoclonal anti-VWFpp antibodies (M) and polyclonal anti-VWF (N). Cells were also labeled with a rabbit anti–P-selectin antibody (P) in addition to the canine-specific monoclonal VWFpp antibody (Q). Staining for P-selectin was faint and diffuse with many small granules (P). No apparent Weibel-Palade bodies were detected in the canine VWD aortic endothelial cells.