Kinetic Binding & Receptor Occupancy

4 reasons why Tag-lite is the preferred solution for kinetic work

Growing evidence suggests that ligand–receptor binding kinetics is an overlooked factor in drug discovery. Cisbio has developed Tag-lite® kits & services to address the kinetic binding needs of GPCR researchers around the world.

The non-radioactive Tag-lite® solution is quickly becoming the industry standard for studying receptor-ligand binding interactions. Our Tag-lite offer continues to demonstrate its effectiveness in the lab, taking the lead over traditional SPA and radioligand binding assays for studying kinetic parameters:

  • Absence of separation steps ensures detailed kinetic traces
  • Cell-based solution
  • No radioactive waste
  • Small quantities of labeled ligands
The kinetics of drug-receptor binding, Watch Webinar
Improved binding kinetics analyses with association and dissociation, Download application note.

 

Principle of kinetic binding

Binding kinetics relates to the rate of association and dissociation of a drug-protein complex.  Binding kinetics is thought to be a critical parameter for optimizing the in vivo efficacy of drug candidates. As such kinetics is receiving increasing attention in drug discovery. Tag-lite combines the flexibility of a radioligand binding with the advantages of homogeneous nonradioactive technologies such a SPR. The homogeneous character of Tag-lite allows for the binding events to be continually monitored without ever having to stop the reaction.

Fitted with the proper model (Motulsky & Mahan, Mol Pharmacol 1984), the Tag-lite reveals the kinetic of association and dissociation of 3 compounds, in the form of Kon and Koff (Graph 1: Telenzepine; Graph 2: Atropine; Graph 3: Pirenzepine) (Table 1).

The correlation between Ki at equilibrium and Kd derived from Kon & Koff (Kd = Koff / Kon) was then assessed. The Kd value obtained from the kinetic binding experiment closely matched the Ki obtained at equilibrium (Graph 4).

Product performance

Using Tag-lite and a suitably-adapted model (Motulsky & Mahan, Mol Pharmacol 1984), the Kon and Koff of 3 compounds (Graph 1: Telenzepine; Graph 2: Atropine; Graph 3: Pirenzepine) were determined easily (Table 1).

The correlation between Ki at equilibrium and Kd derived from Kon & Koff (Kd = Koff / Kon) was then assessed. The calculated Kd showed good consistency compared to that obtained in competition experiments at equilibrium (Graph 4).

Compound Atropine Telenzepine Pirenzepine
Assay number n=2 n=3 n=3
Kon (M-1.min-1) 6.82E+06 5.57E+05 6.63E+05
SD 7.70E+05 2.52E+04 1.54E+06
Koff (min-1) 1.92E-02 2.12E-03 6.84E-01
SD 1.05E-03 5.45E-05 1.74E+00
Kd (=Koff/Kon) (M) 2.81E-09 3.80E-09 1.03E-06
Ki (equlibrium competitions) (3h) 3.325E-09 3.069E-08 5.87E-07

 

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