The tissue localization of antihemophilic factor (AHF, Factor VIII) has been determined by immunofluorescent studies using monospecific rabbit antibody to human AHF. Specific staining demonstrating AHF antigens has been identified in endothelial cells of a wide range of human tissues. The staining pattern was observed in endothelial cells of arteries, capillaries, and veins as well as the cells lining hepatic and splenic sinusoids. Specific fluorescence was limited to these endothelial cells in sections of kidney, liver, spleen, lymph node, cardiac and smooth muscle, thyroid, umbilical cord, and skin. Absorption studies established that the staining was specific for cells in which there were proteins that had AHF antigens. The demonstration of fluorescence within the cytoplasm of endothelial cells suggests that these cells synthesize proteins that have AHF antigens.