Mass-Screening of Curarimimetic Neurotoxin Antagonists

Abstract : A solid phase assay for ligands of curarimimetic toxin binding sites was developed. Its principle is the autoradiographic detection of the displacement of 125 I-alphabungarotoxin from Torpedo californica acetylcholine receptor. To permit analysis of large numbers of data, receptor was immobilized to multiwell titer plates. Several parameters were investigated to optimize the procedure, among them type of plate and plastic; nature of detergent used for receptor solubilization; and treatment to increase receptor adsorption to the wells. The final protocol involves cholate as a receptor extractant and flexible PVC plates coated with polyethyleneimine for maximal receptor adsorption. An autoradiographic procedure utilizing a special exposure cassette was devised. The possibility of an ELISA version of the assay was explored but a major goal, receptor adsorption in amounts sufficient for colorimetric detection, has not been reached. Keywords: Immunoassay; Curarimimetic toxin antidotes: Solid-phase ligand binding assay; Microwell titer plate technology; Autoradiography.