Platelets take up plasma proteins into their alpha granules. Platelet activation releases the alpha granule contents. The goal of this study was to test the hypothesis that human platelets also contain some coagulation factor IX in their alpha granules. Platelets were examined by immunofluorescence microscopy. Activated and unactivated platelets were quick frozen on to slides and dehydrated in situ to preserve optimal morphology. The slides were stained by indirect immunofluorescence for factors V and IX. Unactivated platelets showed coarsely granular staining for factor V and finely granular staining for factor IX. Staining for factor V was diffuse after activation, while staining for factor IX disappeared. Thus, the results support the conclusion that platelets contain factor IX which can be released upon activation. Immunoelectron microscopic studies were conducted to more precisely localize the site of the platelet factor IX. As expected, factor V was primarily localized in the alpha granules of unactivated platelets. By contrast, factor IX was observed both in alpha granules and diffusely in the platelet cytoplasm and membrane-bounded vesicles. After activation, staining for both factors V and IX was primarily in the open canalicular system. The physiological importance of this small amount of factor IX is unknown. However, it may be significant since only a few percent of normal IX levels are required to support haemostasis in the absence of trauma, and platelet factor IX could be released at sites of active coagulation.