title: Development of a high-throughput calcium flux assay for identification of all ligand types including positive, negative, and silent allosteric modulators for G protein-coupled receptors.
abstract: In recent years, the increased use of cell-based functional assays for G protein-coupled receptors in high-throughput screening has enabled the design of robust assays to identify allosteric modulators (AMs) in addition to the more traditional orthosteric agonists and antagonists. In this article, the authors describe a screening format able to identify all ligand types using a triple-add assay that measures changes in cytosolic calcium concentration with three separate additions and reads in the same assay plate. This triple-add assay captures more small molecule ligand types than previously described assay formats without a significant increase in screening cost. Finally, the customizability of the triple-add assay to suit the needs of various AM screening programs is demonstrated.
authors: Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, Connecticut, USA.
authors: Devin J
authors: DJ
authors: Noblin
authors: Robert L
authors: RL
authors: Bertekap
authors: Neil T
authors: NT
authors: Burford
authors: Adam
authors: A
authors: Hendricson
authors: Litao
authors: L
authors: Zhang
authors: Ronald
authors: R
authors: Knox
authors: Martyn
authors: M
authors: Banks
authors: Jonathan
authors: J
authors: O'Connell
authors: Andrew
authors: A
authors: Alt
mesh_terms: D000494:Allosteric Regulation / Q000502:physiology; D000818:Animals; D016466:CHO Cells; D020013:Calcium Signaling / Q000502:physiology*; D006224:Cricetinae; D003412:Cricetulus; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D008024:Ligands; D011485:Protein Binding / Q000502:physiology; D043562:Receptors, G-Protein-Coupled / Q000502:physiology*
chemical_list: D008024:Ligands; D043562:Receptors, G-Protein-Coupled