A multiplex calcium assay for identification of GPCR agonists and antagonists.

Methods Development

Abstract

Activation of G(q) protein-coupled receptors can be monitored by measuring the increase in intracellular calcium with fluorescent dyes. Recent advances in fluorescent kinetic plate readers and liquid-handling technology have made it possible to follow these transient changes in intracellular calcium in a 1,536-well plate format for high-throughput screening (HTS). Here, we have applied the latest generation of fluorescence kinetic plate readers to multiplex the agonist and antagonist screens of a G protein-coupled receptor (GPCR). This multiplexed assay format provides an efficient and cost-effective method for HTS of G(q)-coupled GPCR targets.

Authors

Liu, Ke; Southall, Noel; Titus, Steve A; Inglese, James; Eskay, Robert L; Shinn, Paul; Austin, Christopher; Heilig, Markus A; Zheng, Wei;

Keywords

  • Animals
  • CHO Cells
  • Calcium/ analysis
  • Calcium/ metabolism
  • Calibration
  • Cricetinae
  • Cricetulus
  • Cyclic AMP/ metabolism
  • Dose-Response Relationship, Drug
  • Drug Evaluation, Preclinical
  • Fluorescent Dyes
  • Indicators and Reagents
  • Kinetics
  • Muscarinic Agonists/ pharmacology
  • Muscarinic Antagonists/ pharmacology
  • Radioligand Assay
  • Receptor, Muscarinic M1/ drug effects
  • Receptors, G-Protein-Coupled/ agonists
  • Receptors, G-Protein-Coupled/ antagonists & inhibitors

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