Evaluating opioid-mediated adenylyl cyclase inhibition in live cells using a BRET-based assay

Preeti Manandhar, Shivani Sachdev, Marina Santiago*

*Corresponding author for this work

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Quantitative measurement of receptor signaling by different ligands is important for understanding the mechanism of drug action and screening of drugs. Here, we describe a simple and cost-effective method of measuring adenylyl cyclase inhibition, one of the hallmarks of opioid receptor activation. The assay is based on bioluminescence resonance energy transfer (BRET) that involves transfection of a biosensor in human embryonic kidney (HEK)-293 cells stably transfected with μ-opioid receptor (μ receptor), enabling real-time measurement of cAMP levels.

Original languageEnglish
Title of host publicationOpioid receptors
Subtitle of host publicationmethods and protocols
EditorsSanti M. Spampinato
PublisherHumana Press Inc.
Chapter11
Pages117-125
Number of pages9
Edition2nd
ISBN (Electronic)9781071608845
ISBN (Print)9781071608838
DOIs
Publication statusPublished - 2021

Publication series

NameMethods in Molecular Biology
Volume2201
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • Adenylyl cyclase
  • BRET assay
  • cAMP
  • Forskolin
  • HEK-293 cells
  • μ-Opioid receptor

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