KineticTitration

Can I run the biomolecular interaction measurement any faster? I do not have time to find suitable regeneration solution, can I still run multiple concentrations?

 

What is KineticTitration?

KineticTitration reduces time required to run a full biomolecular interaction experiment by half (depending on assay type and number of concentrations). During KineticTitration, analyte samples are flown over the surface in a series from low to high concentration without dissociation and regeneration steps between the sample injections. The dissociation rate is measured after the last analyte sample.

When is KineticTitration useful? 

KineticTitration is suitable for all biomolecular interaction measurements. It is especially useful when:

  • surface is difficult to regenerate
  • regeneration is detremental to the ligand
  • less time is available for assay development

How to measure affinity and kinetics of molecular interactions using traditional multi-cycle method and using KineticTitration?

KineticTitration: Analyte samples are injected in a series over the immobilized ligand, from the lowest to the highest concentration. There are no regeneration or dissociation injections during this time. After the last analyte sample molecule is injected and plateau reached, the dissociation rate is measured. Affinity and kinetic constants are calculated by fitting the measured data using TraceDrawer™ for MP-SPR Navi™. See top figure.

Traditional multi-cycle method: Analyte sample is injected over the surface with immobilized ligand. After sample injection (association), the analyte is let to dissociate. Rest of the bound analyte is removed by injecting a suitable regeneration solution. After that, next sample concentration is injected in a similar manner as the first sample. Affinity and kinetic constants are calculated using TraceDrawer™ for MP-SPR Navi™. See lower figure. Suitable regeneration solutions depends on the ligand molecule and it is essential that ligand activity is not reduced after regeneration. Assay optimization has to be performed in order to find a suitable regeneration solution.

 

KineticTitration_v2_small.png

Example: Small molecular weight drug Tolcapone (273 Da) binding to human serum albumin (HSA) as measured using KineticTitration.

 

Tolcapone_binding_kinetictitration.png

Tolcapone to HSA is a two-state binding.

Seven samples of tolcapone were injected from the lowest to the highest concentration in the range of 0.25 - 25 µM. 

 See our Application Note #155 on KineticTitration!


 

KineticTitration is available only in 4-channel MP-SPR Navi 420A ILVES!

  • See more about affinity and kinetics here.
  • SPR measurements of small molecules, interactions with lipids and biomaterials are typically hindered by bulk effect, an optical artefact. See how MP-SPR solves bulk effect with our PureKinetics™ feature.
  • What is the difference between SPR and MP-SPR? Find out here.