Production and characterization of a monoclonal antibody to dopamine D2 receptor: comparison with a polyclonal antibody to a different epitope.
A monoclonal antibody (Mab) that recognizes the rat dopamine D2 receptor (DAR) has been generated using DAR specific peptide. The Mab, IgM isotype recognizes five proteins (Mr 220, 145, 95, 66 and 47 kDa) in striatal membrane on Western blot. Preincubation of Mab with free peptide blocked the labeling of all five bands. A polyclonal antibody against peptide from a different region of the DAR, reacted with three out of five proteins (220, 66, and 47 kDa) in these membranes. The DAR antagonist NAPS-biotinyl binds to a 220 kDa protein in striatal membrane on ligand blotts; the labeling can be blocked by the addition of 2 microM sulpride. The 220 kDa Mab reactive protein was less in cerebellum and was absent in the liver. Neither the Mab nor polyclonal antibody inhibited binding of a DAR antagonist, [3H]YM09151-2, to the striatal membranes. These antibodies will enable us to study the structure/function and regulation of the synthesis of DAR protein.
Laboratory of Neurosciences, Pennington Biomedical Research Center, Baton Rouge, LA 70808.
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D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D000911:Antibodies, Monoclonal; D014150:Antipsychotic Agents / Q000378:metabolism; D001549:Benzamides / Q000378:metabolism; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D003342:Corpus Striatum / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006825:Hybridomas / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000138:chemical synthesis / Q000276:immunology; D011869:Radioligand Assay; D051381:Rats; D011954:Receptors, Dopamine / Q000032:analysis / Q000276:immunology* / Q000378:metabolism; D017448:Receptors, Dopamine D2; D014316:Tritium
D000906:Antibodies; D000911:Antibodies, Monoclonal; D014150:Antipsychotic Agents; D001549:Benzamides; D009842:Oligopeptides; D011954:Receptors, Dopamine; D017448:Receptors, Dopamine D2; D014316:Tritium; C030265:nemonapride