The endoplasmic reticulum was initially thought to be devoid of cholesterol dependent DRMs because of its low cholesterol content. Several recent studies have however reported their existence. Due to the increase in cholesterol and sphingolipids along the secretory pathway, raft-like domains are thought to become more abundant in the Golgi and more specifically the trans-Golgi network. Raft-like domains are also present in the endocytic pathway, as highlighted by studies on the trafficking of GPI-anchored proteins, flotillins, toxins and viruses. Occurrence of rafts in the endocytic pathway is probably the combined result of de novo assembly and engulfment from the plasma membrane. Endocytosis of raft-like domains can indeed occur both via clathrin-dependent and independent-pathways. Having previously documented the occurrence of DRMs in late endosomes, we have characterized these raft-like domains in more detail using morphological approaches and subcellular fractionation followed by sub-organellar fractionation. We show that limiting and internal membranes of this multivesicular compartment both contain raft-like membranes but that these domains differ in their physico-chemical properties and protein composition. As shown in Fig. 1, Tubeimoside-I whereas GPI-anchored proteins are abundant in the DRM fraction of untreated late endosomes, treatment of the purified organelle with either saponin or filipin prior to Triton X100 solubilization, led to the redistribution of these proteins to the high density detergent soluble fractions on these Optiprep gradients. It has long been known that late endosomes have a complex morphology with tubular and vesicular regions, which in turn can be multivesicular or multilamellar. These morphological distinct areas, which by themselves Orotic acid (6-Carboxyuracil) define different membrane domains, are likely to be further divided into macro or microdomains. Consistently, Rab9 and Rab7, two late endosomal Rab proteins, occupy distinct domains within late endosomal membranes. Here we have studied the existence of lipid raftlike domains in late endosomes.