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Multiplexing of G protein-coupled receptors (GPCRs) in microarrays promises to increase the efficiency, reduce the costs, and improve the quality of high-throughput assays. However, this technology is still nascent and has not yet achieved the status of "high throughput" or laid claim to handling a large set of receptors. In addition, the technology has been demonstrated only when using fluorescent ligands to detect binding, limiting its application to a subset of GPCRs. To expand the impact of multiplexing on this receptor class, we have developed a radiometric approach to the microarray assay. In these studies, we considered two receptors in the alpha-adrenergic receptor family, alpha2A and alpha2C, and the 125I-labeled agonist clonidine. We demonstrate that microarrays of these receptors can be readily detected (signal/noise ratio approximately 160) using a Typhoon 9210 PhosphorImager. In addition, biochemical characterization shows that ligand-binding profiles and selectivity are preserved with the selective antagonists BRL44408 and ARC239. Importantly, these microarrays use approximately 200- to 400-fold less membrane preparation required by conventional assay methods and allow two or more receptors to be assayed in an area equivalent to a standard well of a microtiter plate. The impact of this approach on screening in drug discovery is discussed.

Membrane-bound proteins represent the single most important class of drug targets. This article discusses the issues surrounding fabrication of membrane–protein microarrays by conventional robotic pin printing techniques. Ligand binding selectivity and specificity to G protein-coupled receptor (GPCR) microarrays are presented. The potential applications of these arrays for drug screening are discussed.

Conventional assay methods for discovering and profiling drug-target interactions are typically developed on a target-bytarget basis and hence can be cumbersome to enable and orchestrate. Herein the authors report a solid-state ligand-binding assay that operates in a multiplexed mode to report compound activity against a micorarray-configured panel of G-proteincoupled receptor (GPCR) targets. The pharmacological fidelity of the system is high, and its miniaturized “plug-and-play” format provides improved efficiency both in terms of execution time and reagent consumption. Taken together, these features make the system ideally suited to explore the structure-activity relationship of compounds across a broad region of target class space.(Journal of Biolmolecular Screening 2006:435-438)

The increased number of drug targets and compounds demands novel high-throughput screening technologies that could be used for parallel analysis of many genes and proteins. Protein microarrays are evolving promising technologies for the parallel analysis of many proteins with respect to their abundance, location, modifications, and interactions with other biological and chemical molecules. This chapter specifically describes the fabrication of G protein-coupled receptor (GPCR) microarrays, a unique subset of protein microarrays, using contact-pin printing technology. The bioassays and potential applications of GPCR microarrays for the determination of compound affinities and potencies are also included.

This paper describes G-protein-coupled receptor (GPCR) microarrays on porous glass substrates and functional assays based on the binding of a europium-labeled GTP analogue. The porous glass slides were made by casting a glass frit on impermeable glass slides and then coating with gamma-aminopropyl silane (GAPS). The emitted fluorescence was captured on an imager with a time-gated intensified CCD detector. Microarrays of the neurotensin receptor 1, the cholinergic receptor muscarinic 2, the opioid receptor mu, and the cannabinoid receptor 1 were fabricated by pin printing. The selective agonism of each of the receptors was observed. The screening of potential antagonists was demonstrated using a cocktail of agonists. The amount of activation observed was sufficient to permit determinations of EC50 and IC50. Such microarrays could potentially streamline drug discovery by helping integrate primary screening with selectivity and safety screening without compromising the essential functional information obtainable from cellular assays.