We have previously reported that ectopically expressed factor VIII (FVIII) is stored within platelets and is released upon platelet activation. Studies by others in various cell lines have suggested that having von Willebrand factor (VWF) coexpression is necessary for FVIII granular storage and for its secretion. We tested the importance of VWF coexpression for ectopic storage of FVIII in platelets and for its bioavailability. Transgenic mice expressing platelet-specific FVIII were crossed onto a VWF^-/- background. Antigenic levels of platelet FVIII in these mice were nearly unchanged whether VWF was present or not. Whole-blood clotting times and FeCl₃ carotid artery injury correction demonstrated that platelet FVIII demonstrably improved the bleeding diathesis in FVIII^null mice independent of the platelets' VWF status. Immunogold electron microscopy demonstrated that platelet FVIII is stored in platelet α-granules independent of the presence of VWF. It appears that FVIII's interaction with VWF and its intracellular transportation, storage, and secretion differ greatly depending on the cell type. The molecular basis for these differences now needs to be elucidated. (Blood. 2005;105:4674-4676)