RADIOLIGAND BINDING METHODS - PRACTICAL GUIDE AND TIPS

Radioligand binding assays are a relatively simple but extremely power ful tool for studying receptors. They allow an analysis of the interac tions of hormones, neurotransmitters, growth factors, and related drug s with the receptors, studies of receptor interactions with second mes senger systems, and characterization of regulatory changes in receptor number, subcellular distribution, and physiological function. As a re sult, these assays are widely used (and often misused) by investigator s in a variety of disciplines, including pharmacology, physiology, bio chemistry, immunology, and cell biology. This article presents a broad overview of the radioligand binding assay technique, primarily for th e investigator who has limited experience with this technique. Practic al guidelines for setting up a new assay are presented, including the receptor preparation to be used, choice of appropriate radioligand, op timizing assay conditions, and appropriate methods for data analysis. Tips for avoiding some of the common pitfalls in application of these assays are also included. The primary focus is on radioligand binding assays of membrane-bound receptors studied in membrane preparations. H owever, similar assay techniques can be used to study receptors on int act cells. The unique advantages and disadvantages of these intact cel l binding assays are also discussed. In particular, the occurrence of regulatory changes in receptors during the course of intact cell bindi ng assays is considered, with approaches for circumventing these compl ications and for using intact cell assays to advantage in studying the se regulatory changes.