Tissue factor (TF) is the most important trigger of blood coagulation in vascular pathology. Rabbit TF, with or without (C) its C-terminal intracellular tail, has been conjugated to the green fluorescent protein (GFP) to study the subcellular localization and other functions of TF. TF-GFP and TFC-GFP are associated with Na₂CO₃-resistant buoyant fractions in HEK 293 cells (lipid rafts); there is no morphological difference in the surface distribution of these or other GFP-labeled membrane proteins present in or excluded from rafts (confocal microscopy, HEK 293 cells). Endogenous TF expressed by rabbit aortic smooth muscle cells (SMCs) is also raft-associated. Membranes from HEK 293 cells expressing recombinant TF-GFP or the wild type TF were equipotent to clot human plasma; however TFC-GFP is ~20-fold more active (per membrane weight). Immunoblot confirms that the deletion mutant is more abundantly expressed and confocal microscopy shows that it has preferential membrane localization, whereas TF-GFP is mainly intracellular (nuclear lining and multiple granules). With a similar half-life (< 4 h), the two constructions differ by their intracellular retention, lower for TFC-GFP. In serum-starved SMCs, the expression of endogenous TF is upregulated by interleukin-1 and/or FBS treatments (immunoblot, immunofluorescence, clotting assay). However, TF secretion or surface expression is not regulated by stimuli of physiological intensity (such as stimulation of the co-expressed kinin B₁ receptors), although a calcium ionophore is highly active in this respect. TF is a raft-associated molecule whose surface expression (secretion) is apparently retarded or impaired by structural determinant(s) located in its C-terminal tail.