Organization of rhodopsin and a high molecular weight glycoprotein in rod photoreceptor disc membranes using monoclonal antibodies.
Four monoclonal antibodies obtained from the fusion of mouse myeloma cells with lymphocytes of mice immunized with bovine rod outer segment disc membranes were shown to bind to the surface of sealed discs. Radioimmune labeling of rod outer segment membrane proteins separated by sodium dodecyl sulfate gel electrophoresis indicated that two monoclonal antibodies (3D6 and 4B4) were against rhodopsin. Limited proteolysis of rod outer segment membranes with trypsin and Streptomyces griseus protease indicated that the 3D6 antibody bound to the trypsin-sensitive region close to the carboxyl-terminal end of rhodopsin. The 4B4 antibody bound at a trypsin insensitive, but S. griseus protease-sensitive internal region of rhodopsin accessible on the cytoplasmic surface of discs. Two other monoclonal antibodies (3D12 and 4B2) were found to bind to different regions of the Mr = 220,000 concanavalin A binding glycoprotein of rod outer segment disc membranes. Proteolysis studies indicated that these antibodies also bound to a Mr = 140,000 fragment which does not contain the concanavalin A binding site. Immunoferritin-labeling studies for transmission electron microscopy confirm the location of the 3D6 and 4B2 antigens on the cytoplasmic or interdisc surface of disc membranes.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D002417:Cattle; D002462:Cell Membrane / Q000648:ultrastructure; D005260:Female; D006023:Glycoproteins / Q000032:analysis*; D006825:Hybridomas / Q000276:immunology; D008214:Lymphocytes / Q000276:immunology; D008565:Membrane Proteins / Q000032:analysis*; D051379:Mice; D008807:Mice, Inbred BALB C; D008854:Microscopy, Electron; D010786:Photoreceptor Cells / Q000648:ultrastructure*; D010954:Plasmacytoma / Q000276:immunology; D011863:Radioimmunoassay; D012168:Retinal Pigments / Q000032:analysis*; D012243:Rhodopsin / Q000032:analysis*; D012374:Rod Cell Outer Segment / Q000648:ultrastructure*
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; D006023:Glycoproteins; D008565:Membrane Proteins; D012168:Retinal Pigments; D012243:Rhodopsin