The behavior of the cell surface concanavalin A (ConA) receptors of two ascites sublines of the 13762 rat mammary adenocarcinoma differing in morphology and xenotransplantability was examined using fluorescein-labeled ConA. MAT-C1 cells have a highly convoluted cell surface with branched microvilli extending from the cell body and are transplantable into mice. MAT-B1 have a more usual surface architecture and are not xenotransplantable. Rapid formation of caps is observed with the MAT-B1 cells, but redistribution is undetectable with the MAT-C1 cells. Colchicine treatment enhances redistribution and shedding of the MAT-B1 ConA receptors. Cytochalasin B (CB) causes the receptors to collect in protuberances at the cell surface. All of the receptor movements are energy- and ConA-dependent. None of these treatments causes significant receptor redistributions in the MAT-C1 cells. All of the treatments cause massive changes in the cell surface morphology of MAT-B1 cells, including disappearance of microvilli, but none of the treatments has substantial effects on the MAT-C1 morphology or microvilli structure. The possibility that immobility of the MAT-C1 receptors results from restrictions imposed by the major sialoglycoprotein was investigated by pronase treatments of the cells. Degradation of most of the sialoglycoprotein resulted in no increase in mobility of receptors on microvilli. Although the ConA receptors on MAT-C1 microvilli do not show observable mobility by fluorescence microscopy, the branched microvilli themselves can be observed to move and be shed from the cell surface of MAT-C1 cells which have been swollen in hypotonic buffers. The shed microvilli were isolated by centrifugation on a Percoll gradient. They contain actin, a polypeptide migrating on polyacrylamide gels in dodecyl sulfate (SDS) with gizzard α-actinin, 5′-nucleotidase and the major cell surface sialoglycoprotein of these ascites cells. Labeling studies with 125I and lactoperoxidase in the presence and absence of detergent indicate that they are sealed. We suggest that the lack of redistribution of the ConA receptors results from transmembrane effects of the microvillus cytoskeleton on the receptors of the stabilized MAT-C1 microvilli. The isolated microvilli should provide a useful tool for investigating molecular interactions involved in such transmembrane effects and in the organization of microvillus structure.