23] Ligand-binding assays in Xenopus oocytes

Abstract

Publisher Summary This chapter discusses the applications of ligand-binding assays to ion channels and receptors expressed in Xenopus oocytes and presents some general considerations for ligand-binding assays using oocytes. Ligand-binding assays are performed by incubating intact cells, cell extracts, or purified membranes with a radiolabeled ligand for a period of time that is sufficient for equilibrium to be attained. The receptor–ligand complex is separated from the unbound ligand and the amount bound is quantified by liquid scintillation or γ counting. The chapter describes two different protocols for binding assays; cell surface receptor binding of [ 125 I]SARILE to angiotensin II receptors and 125 I-labeled α-Bungarotoxin binding to acetylcholine receptors in oocyte extracts, which is used to determine the total number of acetylcholine receptors expressed in oocytes after the injection of in vitro transcripts.