title: Development of a homogeneous high throughput fluorescence polarization assay for G protein-coupled receptor binding.
abstract: Traditional methods that follow receptor ligand interactions are competitive assays in which the test compound displaces a radiolabeled molecule. These assays require either a time-consuming step for separation of free ligands from bound ligands or immobilization of receptors and the scintillant on a solid-phase support. In this report, we describe the development of a homogeneous binding assay for a G protein-coupled receptor in the fluorescence polarization format. This homogeneous fluorescence polarization binding assay format is superior to the traditional binding methods because no radioisotope, separation step, or solid-phase support is required. The elimination of the separation step enhances detection of low-affinity ligands and enables a real-time, continuous readout of the binding activity in a high throughput 384-well microplate format.
authors.affiliation: Discovery Technologies, Pharmacia & Upjohn, Kalamazoo, MI 49001, USA. paul.h.lee@am.pnu.com
authors.forename: P H
authors.initials: PH
authors.lastname: Lee
authors.forename: D J
authors.initials: DJ
authors.lastname: Bevis
mesh_terms: D000345:Affinity Labels; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004353:Drug Evaluation, Preclinical; D005454:Fluorescence Polarization / Q000379:methods*; D005456:Fluorescent Dyes; D008024:Ligands; D011485:Protein Binding; D044105:Receptor, Melanocortin, Type 4; D018043:Receptors, Corticotropin / Q000378:metabolism*; D000521:alpha-MSH / Q000031:analogs & derivatives* / Q000378:metabolism
chemical_list: D000345:Affinity Labels; D005456:Fluorescent Dyes; D008024:Ligands; D044105:Receptor, Melanocortin, Type 4; D018043:Receptors, Corticotropin; D000521:alpha-MSH; C027756:MSH, 4-Nle-7-Phe-alpha-