Analytical affinity chromatography: I. Local equilibrium theory and the measurement of association and inhibition constants

Abstract

Affinity chromatography can be used as an analytical tool to measure the binding constants for biological interactions. Most chromatography theories are based on the assumption that the equilibrium isotherm is linear, a poor assumption for most biological interactions. This paper presents a local-equilibrium theory for nonlinear zonal and frontal elution chromatography. Results are compared to other commonly-used affinity chromatography elution equations. The effect of isotherm nonlinearity is shown to be relatively small, provided the product of the association constant and feed concentration is less than 1. The major effect of nonlinearity is zone spreading; this can be very important when measuring rate constants. The measurement of binding kinetics by analytical affinity chromatography is discussed in Part II.

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