Antibodies against lysosomal membranes reveal a 100,000-mol-wt protein that cross-reacts with purified H+,K+ ATPase from gastric mucosa.
Specific antibodies against lysosomal membranes were prepared by using techniques previously described (Louvard, D., H. Reggio, and G. Warren, 1982, J. Cell Biol., 92:92-107) for obtaining organelle-specific antibodies. The purified antibodies stained an acidic vacuolar compartment as shown by double-labeling experiments with acridine orange and indirect immunofluorescence. Characterization of the antibodies by immunoreplica methods revealed one major protein of approximately 100,000 mol wt. The antibodies cross-reacted with purified H+,K+ ATPase from pig gastric mucosa, the enzyme responsible for HCl secretion, but not with ATPases transporting other ions. They may therefore recognize a component of the proton pump involved in the acidification of lysosomes. As was expected, secondary lysosomes contained immunoreactive antigen, as determined by the fine-structural localization of reaction product for peroxidase or immunogold probes in several cell types. The antigen was also found in vacuoles containing phagocytosed bacteria in macrophages so it is present in at least some of the compartments of an endocytic pathway. In liver, the antigen was present in small amounts on the plasma membrane and in large amounts in some coated vesicles (near the sinusoidal surface of hepatocytes), putative endosomes, two cisternae on the cis side of the Golgi complex, adjacent vesicles and vacuoles, and pericanalicular dense bodies. In summary, the antigen seems to be present in those compartments that have recently been demonstrated to be acidified by an ATP-driven pump.
D000251:Adenosine Triphosphatases / Q000032:analysis*; D000818:Animals; D000906:Antibodies; D000936:Antigen-Antibody Complex; D002458:Cell Fractionation; D002460:Cell Line; D000939:Epitopes / Q000032:analysis; D005753:Gastric Mucosa / Q000201:enzymology / Q000648:ultrastructure*; D017506:H(+)-K(+)-Exchanging ATPase; D007124:Immunoenzyme Techniques; D007425:Intracellular Membranes / Q000201:enzymology / Q000648:ultrastructure*; D007668:Kidney; D008247:Lysosomes / Q000201:enzymology / Q000648:ultrastructure*; D008565:Membrane Proteins / Q000032:analysis*; D008871:Microvilli / Q000648:ultrastructure; D008970:Molecular Weight; D051381:Rats
D000906:Antibodies; D000936:Antigen-Antibody Complex; D000939:Epitopes; D008565:Membrane Proteins; D000251:Adenosine Triphosphatases; D017506:H(+)-K(+)-Exchanging ATPase