Protocol to characterize Gi/o and Gs protein-coupled receptors in transiently transfected cells using ELISA and cAMP measurements

Abstract

Summary: Activation of Gs or Gi/o protein-coupled receptors (GPCRs) leads to changes of intracellular cyclic adenosine monophosphate (cAMP) levels. This protocol describes steps for cloning HA- and FLAG-tagged GPCRs, transient transfection of CHO-K1 or HEK293-T cells, and determination of basal and ligand-induced changes in intracellular cAMP levels. We detail enzyme-linked immunosorbent assays to determine relative GPCR plasma membrane and total expression levels.For complete details on the use and execution of this protocol, please refer to Schulze et al. (2022).1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell Biology
• Cell Culture
• Cell Membrane
• Cell-based Assays
• Molecular Biology
• Signal Transduction