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Data from an interactionFollowing an interactionFlow systemsTypes of interaction
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The extent to which different molecules interact with a single partner immobilized on a sensor surface reveals the specificity of an interaction.

Simple yes/no answers are required for a wide variety of applications:
search for binding partners
screen for inhibitor specificity
test for cross-reactivity
look for activity after purification
test cell culture lines for the expression of a given protein
Specificity, no response / response | 25sec
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Kinetics: rates of reaction

The kinetics of an interaction, i.e. the rates of complex formation (ka) and dissociation (kd), can be determined from the information in a sensorgram.

If binding occurs as sample passes over a prepared sensor surface, the response in the sensorgram increases. If equilibrium is reached a constant signal will be seen.

Replacing sample with buffer causes the bound molecules to dissociate and the response decreases.

Biacore evaluation software generates the values of ka and kd by fitting the data to interaction models.

The traditional approach to a kinetics assay is to inject a series of analyte concentrations over the sensor surface, one at a time, with regeneration of the surface between each analysis cycle. More recently, additional approaches have been developed to provide simpler assay development and overcome potential problems with some interaction partners, to support specific applications and to support increased throughput for kinetic assays. 

 • Single-cycle kinetics - more>>

 • 2-over-2 kinetics - more>>


Kinetics, rapid association and dissociation / slow association and dissociation | 25sec
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Affinity: the strength of binding

The affinity of an interaction is determined from the level of binding at equilibrium (seen as a constant signal) as a function of sample concentration.
Affinity can also be determined from kinetic measurements. For a simple 1:1 interaction, the equilibrium constant KD is the ratio of the kinetic rate constants, kd/ka.

Affinity, strong interaction / weak interaction | 25sec
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Concentration analysis

Concentration is determined by monitoring the interaction of a molecule with a prepared sensor surface in the presence of a target molecule in solution (solution inhibition) or excess analyte (surface competition). Depending on the Biacore system used, concentrations are calculated either by interpolation of the binding responses on a calibration curve, or using the more recently developed Calibration-free concentration analysis (CFCA) method. Read more about CFCA>>

Concentration can be determined for purified molecules or for molecules in complex mixtures such as serum or food samples.
Concentrations in the nanomolar range can be measured.
The response reflects the concentration of the molecule in relation to the amount that can interact with the immobilized interaction partner. Values can therefore differ from results achieved by UV absorbance or general protein assays that measure only total amounts.
A variety of concentration assay formats are used in the Qflex Kits of Biacore Q, a system designed for rapid determination of food quality and safety.

Multiple interactions during complex formation

Complex formation can be monitored as each component is incorporated into a multimolecular complex. For example:

The ability of different antibodies to bind simultaneously to an antigen can be used to map epitopes on the antigen.
Multiple Binding, changes in response | 7sec
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