Specializing in both standard and custom in vitro functional assays, we are focused on providing the right assays for assessing a GPCR therapeutic's potential efficacy and liabilities to help identify candidate compounds for preclinical and clinical development. Combined with our radioligand binding assays, these functional assays can provide an integrated assessment of a drug candidate's in vitro pharmacology profile.

Below is a complete list of available functional assays.  For functional autoradiographic assays please contact us.

96-well drug plate setup diagram for functional binding assays.  The functional binding plate is designed to contain two buffer controls (Column 1 and 11), one agonist control (Column 2), one antagonist control (Column 12) and 16 compounds (Columns 2 - 10) at a single concentration. Results are normalized and transformed to percentage values. For agonists, the reference agonist activity at 10 μM is set as 100% and basal activity with buffer as 0%. For antagonists, the basal activity with buffer only is set as 100% inhibition and the activity of the EC80 of the reference agonist as 0% inhibition. For allosteric potentiators, the activity of the EC20 of a reference agonist is set as 0% potentiation. Potential agonist hits are subjected to full concentration-response studies to determine their efficacy and potency. Potential antagonist hits are subjected to full concentration-response studies to determine IC50 values against EC50 to EC80 concentrations of a reference agonist.
96-well drug plate setup diagram for functional binding assays. The functional binding plate is designed to contain two buffer controls (Column 1 and 11), one agonist control (Column 2), one antagonist control (Column 12) and 16 compounds (Columns 2 - 10) at a single concentration. Results are normalized and transformed to percentage values. For agonists, the reference agonist activity at 10 μM is set as 100% and basal activity with buffer as 0%. For antagonists, the basal activity with buffer only is set as 100% inhibition and the activity of the EC80 of the reference agonist as 0% inhibition. For allosteric potentiators, the activity of the EC20 of a reference agonist is set as 0% potentiation. Potential agonist hits are subjected to full concentration-response studies to determine their efficacy and potency. Potential antagonist hits are subjected to full concentration-response studies to determine IC50 values against EC50 to EC80 concentrations of a reference agonist.
Ref#ReceptorAgonistAntagonistAssay TypeSource
FBA004Adenosine A1CPADPCPX[35S]-GTPγS
FBA018Adenosine A3IB-MECAMRS 1220[35S]-GTPγS
FBA005Adrenergic a2AClonodineYohimbine[35S]-GTPγS
FBA019Adrenergic a2BUK 14304Yohimbine[35S]-GTPγS
FBA020Adrenergic a2CUK 14304Yohimbine[35S]-GTPγS
FBA006Cannabinoid CB1CP 55940AM 251[35S]-GTPγS
FBA007Cannabinoid CB2CP 55940AM 630[35S]-GTPγS
FBA021Chemokine CCR5MIP-1βVicriviroc[35S]-GTPγS
FBA008Dopamine D1Dopamine(+)-Butaclamol[35S]-GTPγS
FBA009Dopamine D2DopamineSpiperone[35S]-GTPγS
FBA010Dopamine D3DopamineSpiperone[35S]-GTPγS
FBA011Histamine H1HistaminePyrilamine[35S]-GTPγS
FBA022Muscarinic M1Oxotremorine MPirenzepine[35S]-GTPγS
FBA023Muscarinic M44-DAMPMcN-A-343[35S]-GTPγS
FBA012Opioid DOP (δ)DPDPENaltrindole[35S]-GTPγS
FBA013Opioid KOP (κ)U-69593nor-Binaltorphimine[35S]-GTPγS
FBA014Opioid MOP (µ)DAMGONaloxone[35S]-GTPγS
FBA015Nociceptin ORL1NociceptinBAN ORL24[35S]-GTPγS
FBA016Platelet-Activating Factor (PAF)PAFWEB 2086[35S]-GTPγS
FBA001Serotonin 5-HT1ASerotoninWAY 100635[35S]-GTPγS
FBA002Serotonin 5-HT1A8-OH-DPATWAY 100635[35S]-GTPγS
FBA003Serotonin 5-HT2CSerotoninSB242084[35S]-GTPγS
FBA017Sphingosine-1-Phosphate (SP1P)SP1PW146[35S]-GTPγS
Primary & Secondary Functional Binding Assays

Compounds are typically subjected to primary functional screening assay at targets (receptors) selected by investigators. In primary screening assays, compounds are tested in quadruplicate at a final concentration of 10 μM or at specific concentrations upon request for agonist and antagonist activities. Results are normalized and transformed to percentage values. For agonists, the reference agonist activity at 10 μM is set as 100% and basal activity with buffer as 0%. For antagonists, the basal activity with buffer only is set as 100% inhibition and the activity of the EC80 of the reference agonist as 0% inhibition. For allosteric potentiators, the activity of the EC20 of a reference agonist is set as 0% potentiation. Compounds with a minimum of 30% agonist activity, or 50% antagonist activity, or 30% potentiation above control are selected for secondary screening using concetnration-response assays.

In secondary functional assays, potential agonist hits are subjected to full concentration-response (8 concentrations in quadruplicate) assay to determine their efficacy and potency. Potential antagonist hits are subjected to full concentration-response (8 concentrations in quadruplicate) assay to determine IC50 values against EC50to EC80 concentrations of a reference agonist. Potential antagonist hits and positive allosteric modulators are further characterized if necessary.

Both primary and secondary functional binding assays are carried out in a final of volume of 125 μl per well in appropriate binding buffer. [35S]-GTPγS is at a concentration specific for the target receptor (unless stated otherwise). Basal and nonspecific binding are determined in the absence and presence of 10 μM of GTPγS. Agonist & Antagonist activity is determined by receptor specific compounds.In brief, plates are incubated at a specific temperature in the dark for a specific amount of time (receptor specific). Reactions are stopped by vacuum filtration onto buffer soaked 96-well Unifilters using a Filtermate harvester, followed by a wash with cold wash buffer. Scintillation cocktail is then added onto air-dried filters. Radioactivity is counted in a Microbeta counter.

Key details of our functional binding assay services:
  • Gold standard filtration assays.
  • Industry Standard.
  • Multiple assays formats to choose from upon request.
  • Single concentration screening or dose-response for IC50 or EC50 determinations.
  • Determine full, partial, or inverse agonist and antagonist activity
  • Quick turnaround times of 1-3 weeks depending on project types.
  • Plotted EC50 curves.
  • Proven skills and expertise to develop difficult targets.
  • Reliable, quick turnaround times.
  • High quality data with strict quality controls.
  • Flexible custom services to supplement our off-the-shelf services to meet your needs, providing a cost-effective way to expand your capabilities and capacity.