K1

To determine the association rate constant k+1 a constant target concentration is incubated with a constant marker concentration. The experiment is stopped after different periods of time and the amount of marker bound to the target quanti-

Fig. 7.3 Kinetic binding experiments: (a) Typical association experiment where binding between target (T) and marker (M) reaches steady state after approx. 25 min. (b) Typical dissociation experiment with a half-life of target-marker complex (TM) of approx. 4.5 min.

fied. In dissociation assays, constant concentrations of target and marker are first incubated until equilibrium is reached. Next, dissociation is started under conditions suppressing the association reaction almost completely, e.g. by ''infinite'' dilution or addition of a large excess of a competitor. The course of the dissociation is registered by measuring the amount of the bound marker after different periods of time. The association rate constant k+1 and dissociation rate constant k_1 can be determined from the association and dissociation diagrams (Fig. 7.3) compiled from the experimental data [7, 16, 17, 21].

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