KINETIC CHARACTERIZATION

Agile R100 is a kinetic binding data platform that can be run at your benchtop with just a 10 µL drop of sample, pipetted directly onto the biosensor surface. Now you can get to data in a matter of minutes.

Agile R100 Features:

  • Direct measurement of Kfor structure-activity relationship (SAR) research
  • EC/IC50 ranking verification with an orthogonal method for accurate results
  • Measurements in as little as 2 minutes to capture binding of fast-acting drugs
  • 8+ hour run-time to gain accurate off-rates of tight binders
  • Lower LOD in fM range to measure tight binders that bottom out on other platforms
  • Kinetic characterization of small molecules ≥10 Da

What Kinetics Does Agile R100 Provide?

Agile R100 takes data continuously throughout your experiment, enabling you to gain accurate affinity measurements (KD) and rate constants (kon and koff).

Why Is Affinity Data Important?

During pharmacological or biological research, biochemical processes are often examined for how they inhibit or stimulate the system. To understand the potency of a particular compound, half maximal inhibitory concentration (IC50) or half maximal effective concentration (EC50) values are often back-calculated from dose‑response curves using an end-point assay such as an ELISA. IC50 and EC50 values have limitations. They are dependent on the experimental conditions under which they’re determined, which means they can only be compared against each other when the assays conditions are identical. In contrast, the equilibrium dissociation constant (KD), a measure of affinity, is intrinsic to the interaction of the binding pair and is independent of experimental conditions. In this manner, kinetic characterization as a few advantages: 1) Affinity measurements provide flexibility and time savings in comparing different systems that IC50 and EC50 values can’t provide. 2) KD tends to be more reproducible because it is less dependent on experimental conditions.

What Can Off-rates Tell Me?

Drugs with slow dissociation rates or off-rates (koff) often have extended duration of action, and affinity may not be an accurate proxy to discover which compounds will provide beneficial efficacy durations. While slow dissociation is often connected with high binding affinity, this is not always the case, and without a dependable kinetic binding platform, it’s impossible to know for sure. By characterizing on- and off-rates at the same time as affinity, compounds with desirable kinetic profiles for target binding can be chosen. Many compounds can bind to numerous off-target proteins as well as the intended target. Monitoring the dissociation rates of the compound from all possible targets may be advantageous. Kinetic profiles can differentiate compounds with improved toxicity and selectivity, by enabling selection of those that have both slow off-rates for chosen targets and fast off-rates from unintended ones.

CytokineTNFα Binding to Three Small Molecule Compounds

label-free Agile R100 kinetic characterization

Overlaid dose-response curves generated with COOH Agile biosensors show full kinetic characterization of small molecules SPD304, Evans Blue, and Trypan Blue diluted in 1X PBS binding to immobilized cytokine TNFα. Agile Plus software provides kon, koff, and KD.