title: Miniaturization of whole live cell-based GPCR assays using microdispensing and detection systems.
abstract: Cell-based beta-lactamase reporter gene assays designed to measure the functional responses of G-protein-coupled receptors (GPCRs) were miniaturized to less than 2 microL total assay volume in a 3456-well microplate. Studies were done to evaluate both receptor agonists and antagonists. The pharmacology of agonists and antagonists for target GPCRs originally developed in a 96-well format was recapitulated in a 3456-well microplate format without compromising data quality or EC(50)/IC(50) precision. These assays were employed in high-throughput screening campaigns, allowing the testing of more than 150,000 compounds in 8 h. The instrumentation used and practical aspects of the assay development are discussed.
authors.affiliation: Merck Research Laboratories, Department of Automated Biotechnology, North Wales, PA 19454, USA. Oleg_Kornienko@merck.com
authors.forename: Oleg
authors.initials: O
authors.lastname: Kornienko
authors.forename: Raul
authors.initials: R
authors.lastname: Lacson
authors.forename: Priya
authors.initials: P
authors.lastname: Kunapuli
authors.forename: Jonathan
authors.initials: J
authors.lastname: Schneeweis
authors.forename: Ira
authors.initials: I
authors.lastname: Hoffman
authors.forename: Todd
authors.initials: T
authors.lastname: Smith
authors.forename: Melissa
authors.initials: M
authors.lastname: Alberts
authors.forename: James
authors.initials: J
authors.lastname: Inglese
authors.forename: Berta
authors.initials: B
authors.lastname: Strulovici
mesh_terms: D000818:Animals; D001681:Biological Assay / Q000295:instrumentation* / Q000379:methods*; D020650:Combinatorial Chemistry Techniques; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods*; D017930:Genes, Reporter; D006801:Humans; D020128:Inhibitory Concentration 50; D036103:Nanotechnology / Q000379:methods; D043562:Receptors, G-Protein-Coupled / Q000819:agonists* / Q000037:antagonists & inhibitors*; D018028:Receptors, Neurotensin / Q000187:drug effects / Q000235:genetics; D018045:Receptors, Oxytocin / Q000819:agonists / Q000037:antagonists & inhibitors / Q000235:genetics; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D043562:Receptors, G-Protein-Coupled; D018028:Receptors, Neurotensin; D018045:Receptors, Oxytocin; D001618:beta-Lactamases