We have used confocal immunofluorescence microscopy and immunogold electron microscopy to localize tumor necrosis factor (TNF) receptors in cultured human umbilical vein endothelial cells. The 75-kd receptor is largely expressed on the cell surface and undergoes endocytosis and transport to lycosomes through coated pits and coated vesicles. TNF may accelerate receptor clustering and internalization. The 55-kd receptor is expressed at much lower levels on the cell surface than the 75-kd receptor but is more abundant overall. It is predominantly localized to the Golgi apparatus and, to a lesser extent, cytoplasmic vacuoles. The cellular distribution of the 55-kd receptor is largely unaffected by TNF. These results have implications for TNF-mediated endothelial cell activation and for interactions between 75- and 55-kd TNF receptors.