How can I recover affinity values from the kinetic curves?

  • Kinetics parameters

    • ka is the association rate constant (M-1-s-1), in some cases, it is also called kon
    • kd is the dissociation rate constant (s-1), in some cases, it is also called koff

SPR kinetics data (interactions) can be modeled simply in order to evaluate the affinity between 2 molecules. The model generally used supposes that the reactions involve one ligand (L) and one analyte (A) which interact with a 1:1 stoichiometry:

This model gives the following formula for our interaction kinetic curves, one for the association response (Ra) and one for the dissociation response (Rd), the response being in our case the reflectivity variation:

Ra(t)=Req•(1-exp[-kobs•(t-t0)])   (association)

Rd(t)=Rd(t1)•exp[-koff•(t-t1)]      (dissociation)

with kobs = ka•Ci+kd, the observed on-rate constant of the response during association, Ci the injected analyte concentration and kofff = kd, the off-rate constant of the response during dissociation, t0 and t1 are indicating respectively the beginning of the association and of the dissociation.

Briefly, the kinetics parameters ka and kd are calculated by fitting each part of the interaction (association and dissociation) with a mono-exponential curve, preferentially by using the ScrubberGen software (Scrubber-based).

  • Affinity parameters

    • KA is the affinity constant (M-1)
    • KD is the equilibrium constant (M)

Knowing the kinetics parameters, the affinity constant between 2 molecules can be deduced:

KA = 1/KD = ka/kd

The affinity parameters can also be calculated from equilibrium values with a Langmuir fit.