Biacore T100 Software Handbook BR-1006-48 Edition AE
153
Kinetics and affinity analysis 9
9
Kinetics and affinity analysis
Biacore T100 offers three main functions for analysis of interaction kinetics and
affinity:
•
Kinetics and affinity measurements on the sensor surface, which
determine the interaction characteristics between ligand and analyte.
Kinetic parameters are evaluated from the association and dissociation
phases of the sensorgram, and affinity either from the kinetic parameters
or from plots of steady-state analyte binding levels (R
eq
) against
concentration.
•
Thermodynamic analysis, which relies on measurement of either kinetics
or affinity at different temperatures.
•
Affinity in solution, where the interactants are mixed in known
concentrations in solution and allowed to reach equilibrium. Biacore T100
is then used to determine the concentration of free interactant in
equilibrium with the complex.
Kinetic constants derived from surface interaction measurements in multiple
separate experiments can be collated and summarized in the separate Biacore
T100 Kinetic Summary software (Section 9.6).
This chapter describes how to evaluate surface-bound kinetics and affinity. If
there are multiple sample series in the same data set, evaluation can be
performed either in
single mode
(where each sample series is evaluated
separately with interactive control over several aspects of the evaluation), or in
batch mode
(where multiple series are evaluated automatically using default
settings).
Thermodynamic analysis and measurement of affinity in solution are described
in Chapters 10 and 11 respectively.
Kinetics and affinity are normally determined from the binding characteristics
of a series of analyte concentrations. These concentrations may be injected in
separate cycles with surface regeneration between the cycles (
multi-cycle
analysis
) or sequentially in a single cycle with no regeneration between
injections (
single-cycle analysis
), as illustrated in Figure 9-1. Results from these
two approaches are evaluated in the same way, using the same tools and fitting
models, and may even be evaluated together in a single evaluation. Significant
differences in evaluation appearance between the approaches are illustrated
where relevant in this chapter.
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