AAV ACTOne Gs-GPCR Assay Kit-SCTR

The Secretin Receptor (SCTR) is a G protein-coupled receptor (GPCR) that binds to secretin, a peptide hormone involved in regulating water and bicarbonate secretion in the pancreas and bile ducts. SCTR is primarily expressed in the pancreas, liver, stomach, and intestines, where it plays a crucial role in digestion and maintaining pH balance. Activation of this receptor leads to increased cyclic AMP (cAMP) levels, stimulating the secretion of bicarbonate-rich fluids, which neutralizes stomach acid and aids in digestion.
This kit uses AAV vectors with a CMV promoter to co-express the SCTR and cyclic nucleotide-gated (CNG) channel, allowing researchers to conduct high-throughput screening and functional analysis of potential SCTR-targeting compounds. The kit provides a sensitive and reliable method for evaluating the pharmacological properties of SCTR drugs, such as agonists and antagonists, in a live-cell environment.

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Background

ACTOne™ is the only high-throughput GPCR screening technology that can directly measure the intracellular changes of the secondary messenger cyclic AMP (cAMP) in living cells, in real-time. It uses a proprietary modified cyclic nucleotide-gated (CNG) channel, which is co-localized with adenylate cyclase at the plasma membrane, as a biosensor of cAMP activity. The CNG channel opens when the cAMP level near the plasma membrane increases, resulting in ion flux and cell membrane depolarization. The influx of cations through the CNG channel can be quantified using fluorescent ion indicators or membrane potential (MP) dyes. It provides information on real time intracellular cAMP changes and is highly sensitive. By combining kinetic and endpoint readouts, we are able to capture and analyze transient responses from endogenous GPCRs and weak responses caused by weak Gs or Gi coupled GPCR activities. Using ACTOne, we are able to detect the subcellular cAMP concentration changes directly caused by GPCR activation. Real-time kinetic readouts minimize artifacts, and provide greater content and more statistically relevant data. The intensity of signal increase caused by GPCR activation is directly related to the receptor number on cell surface. Using ACTOne assay, we were able to detect activities of some endogenous Gs coupled receptors in HEK293 cells that have not been reported in literature. In addition, we have also detected weak Gs coupled activity of a GPCR that was widely considered to be only linked to Gq coupled pathway. The ACTOne assay also provides a useful tool for GPCR de-orphanization.