Recognition by a mouse monoclonal antibody of a glycoprotein antigen of rat brain which is expressed intracellularly by neurons.
We describe a previously uncharacterised glycoprotein antigen of rat brain. The antigen was localised by immunofluorescence on 10 micron cryostat tissue sections, and was found to be present intracellularly in neurons. No other cell types or structures within the brain were stained. The antigen is recognised by a mouse monoclonal antibody called NGP41. The antibody was produced after immunising a mouse with glycoproteins purified by lentil lectin affinity chromatography of solubilised rat brain membranes. Spleen cells from the immunised mouse were fused with the myeloma P3X63Ag8. The antigen is expressed by neurons in all brain regions, and also in the dorsal root ganglion neurons of the peripheral nervous system. In all brain regions, the large projection neurons are the most intensely stained by immunofluorescence, but some small neurons also express the antigen. Although dendrites were not stained, sections of sciatic nerve were stained by NGP41, suggesting that the antigen is expressed by axonal processes. The cell bodies of neurons in the inferior olive were stained by NGP41, but their terminals on Purkinje cell dendrites in the cerebellar cortex were not stained, suggesting that the antigen is absent or expressed below the limit of detection in terminals. Both crude brain membranes and a lentil lectin affinity purified brain glycoprotein fraction absorbed the antibody, suggesting that the antigen is a membrane bound glycoprotein. In immunoblotting experiments, the antigen was detected in homogenates of brain and spinal cord membranes, where it appeared as a triplet of bands with molecular weights of 41K, 38K and 36K. Antigen was not detected by immunoblotting in homogenates of six different tissues of non-nervous origin. The antigen was enriched in glycoprotein fractions from adult and juvenile cerebellum as assessed by immunoblotting. Adult brain glycoprotein preparations had a triplet structure similar to that in the homogenates, although most of the antigenic activity of the juvenile preparation was found in a position corresponding to the upper two bands of the triplet.
D000818:Animals; D000911:Antibodies, Monoclonal; D000941:Antigens / Q000032:analysis; D001369:Axons / Q000378:metabolism; D002531:Cerebellum / Q000276:immunology* / Q000378:metabolism; D000939:Epitopes / Q000032:analysis; D005455:Fluorescent Antibody Technique; D006023:Glycoproteins / Q000276:immunology*; D008565:Membrane Proteins / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009411:Nerve Endings / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000276:immunology*; D051381:Rats / Q000276:immunology
D000911:Antibodies, Monoclonal; D000941:Antigens; D000939:Epitopes; D006023:Glycoproteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins