Abstract
G protein-coupled receptors (GPCRs) are the most structurally diverse family of signaling proteins and regulate a variety of cell function. For most GPCRs, the cell surface is their functional destination where they are able to respond a wide range of extracellular stimuli, leading to the activation of intracellular signal transduction cascades. Thus, the quantity of receptor expression at the cell surface is a crucial factor regulating the functionality of the receptors. Over the past decades, many methods have been developed to measure the cell surface expression of GPCRs. Here, we describe an intact live-cell radioligand binding assay to quantify the surface expression of GPCRs at the endogenous levels or after overexpression. In this assay, cell cultures will be incubated with specific cell-nonpermeable radioligands which selectively and stoichiometrically bind to individual GPCRs and the receptor numbers at the cell surface are quantified by the radioactivity of receptor-bound ligands. This method is highly specific for measuring the functional GPCRs at the surface of intact live cells and is particularly useful for endogenous, low-abundant GPCRs.