ABSTRACT The structure of a late endosomal compartment, which contains the bulk of the cation-independent mannose 6-phosphate receptor (MPR) in NRK cells, is documented using immunocytochemistry and cryo-sections, as well as conventional Epon sections. This compartment, which we refer to as the prelysosomal compartment (PLC), has a complex three-dimensional structure consisting of tubulo-reticular domains in continuity with vesicular parts. The latter are characterized by a high density of internal membranes, which may be either tubular or sheet-like, that label extensively for the MPR. This structural organization was also maintained after fractionation in sucrose gradients. The amount of MPR immunolabelling was then quantitated with respect to the membrane surface areas of the four compartments where it is found: namely, the plasma membrane, early endosomes, the trans Golgi network and the PLC. The results showed that in NRK cells 90% of the labelling for the receptor was found in the PLC, with the rest distributed over the other three compartments. Cytochemical studies indicated that the PLC is the first structure along the endocytic pathway that gives a significant reaction for acid phosphatase. However, the PLC is clearly distinct from the MPR-negative lysosomes, which are also acid phosphatase-positive, since the two organelles could be physically separated from each other after fractionation on Percoll gradients.