DETERMINATION OF ASSOCIATION RATE CONSTANTS BY AN OPTICAL BIOSENSOR USING INITIAL RATE ANALYSIS

We show that initial rate analysis can be successfully applied to anal yze experimental binding data generated by an optical biosensor. The i nitial rates of binding obtained from linear regression are concentrat ion dependent, and plots of initial rate against ligate concentration yield a straight line that passes through the origin. The slope of thi s graph is the product of the association constant times the maximal b inding capacity of the immobilized ligand. This latter parameter is ea sily obtained from a single binding curve at high ligate concentration , allowing rapid determination of the association rate constant. The a ssociation rate constant obtained in this manner is found to be in goo d agreement with that obtained by the more customary method of nonline ar regression analysis of the entire binding profile. Initial rate ana lysis is more simple than fitting the full association profile and nee ds less data collection time. It also requires fewer assumptions about the functional form of the association profile. This can be advantage ous when fitting biosensor-derived data, which often show complex asso ciation kinetics. Furthermore, it avoids the potential complication of second-order kinetics which may be found at low ligate concentrations with high-affinity interactions. (C) 1997 Academic Press.