Rabbit antiserum against purified Electrophorus electricus acetylcholine receptor is studied using an immunoprecipitin assay to measure either antibody titer or concentration of toxin-binding sites in solubilized receptor preparations. This antiserum, unlike control serum, blocks the electrophysiological response of the electroplax to carbamylcholine. Toxin (a-neurotoxin, Naja naja) and several cholinergic ligands produce partial inhibition of the reaction of antiserum with purified acetylcholine receptor. Evidence is presented that some of the toxin-binding sites on receptor, purified by affinity chromatography on toxin-agarose conjugates, are occluded by toxin. In addition, evidence is presented that antireceptor antiserum will cause precipitation of more toxinbinding sites present in an initial extract than in purified receptor preparations.Over the past few years, considerable effort has been directed towards understanding the molecular biology of the acetylcholine receptor (AChR), a large portion of which has been concerned with its purification (1-8). Implicit in these efforts is the assumption that knowledge of the structural properties of purified receptor will provide insight into its function in situ. In order to generate a probe of receptor structure and function which could be used to compare receptor purified and in situ, we stimulated anti-AChR antibody production in rabbits. As we have reported (9), the injection of rabbits with purified receptor resulted in the formation of antireceptor antibodies and subsequently the appearance of flaccid paralysis leading to death. In this paper, we present properties of one antiserum thus obtained and use this antiserum and antitoxin antiserum to study purified eel acetylcholine receptor. MATERIALS AND METHODSPreparation of Neurotoxin. Toxin was purified from the venom of the Indian Cobra N. Naja Naja, as previously described (10). Purified toxin was iodinated with Na 1251 (New England Nuclear Corp., carrier-free) to specific activities of 30-40 Ci/mmol (11).Preparation of Acetylcholine Receptor. Acetylcholine receptor was purified from the electric eel, Electrophorus electricus, by affinity chromatography on columns of toxin-agarose conjugates (6). Protein concentrations determined by the Lowry reaction (12) were corrected to dry-weight protein by stanAbbreviations: GAR, goat anti-rabbit immunoglobulin G antiserum; AChR, acetylcholine receptor; Toxin, a-neurotoxin,
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