Technologies & Platforms / Enzyme Fragment Complementation Technology / PathHunter EFC Cell-Based Assay Platform / Protein:Protein Interactions / GPCRs β-Arrestin

PathHunter® β-Arrestin GPCR Assay Platform

PathHunter β-Arrestin GPCR assays are whole cell, functional assays that directly measure GPCR activity by detecting the interaction of β-Arrestin with the activated GPCR. Because Arrestin recruitment is independent of G-protein signaling, these assays offer a powerful and universal screening and profiling platform that can be used for virtually any Gi-, Gs, or Gq-coupled receptor.


Technology Principle

Pathhunter GPCR Arrestin
 
In this system, the GPCR is fused in frame with the small enzyme fragment ProLink™ and co-expressed in cells stably expressing a fusion protein of β-Arrestin and the larger, N-terminal deletion mutant of β-gal (called enzyme acceptor or EA). Activation of the GPCR stimulates binding of β-Arrestin to the ProLink-tagged GPCR and forces complementation of the two enzyme fragments, resulting in the formation of an active β-gal enzyme. This interaction leads to an increase in enzyme activity that can be measured using chemiluminescent PathHunter® Detection Reagents.

Analysis of ADRB2 & CRTH2 Agonists & Partial Agonists


Full & Partial ADRB2 Agonists

       

CRTH2 Agonists
Adrenergic beta 2 receptor (ADRB2)   Prostaglandin D2 Receptor
Uncover pharmacologies for all classes of compounds. Depicted are dose curves identifying a full agonist (isoproterenol), partial agonist (clenbuterol) and antagonist (propranolol) for the Adrenergic beta 2 receptor (ADRB2) [click graph to enlarge].
  
Prostaglandin D2 Receptor, PGD2 (CRTH2) dose curves depict accurate rank order potencies of compounds. Excellent assay windows combined with chemiluminescent detection makes screening fast and simple [click graph to enlarge].

Features & Benefits

  • Validated with over 180 GPCRs – compatible with all GPCR classes (A and B)
  • Universal format – independent of G-protein coupling, ideal for orphan GPCR screening, 
  • Specific signal – tagged receptor eliminates background from endogenous receptors
  • Stable cell lines - increased reliability and reproducibility
  • Multiplexing capabilities - additionally run second messenger assays using the same cell line 
  • Stoichiometric signal generation - accurate compound efficacy measurements
  • Standardized assay conditions - efficient comparison studies across receptors or receptor families (regardless of G-protein coupling)
  • Screening-friendly format – superior assay windows and robust performance