Activation and diffusion in the kinetics of adsorption and molecular recognition on surfaces. Enzyme-amplified electrochemical approach to biorecognition dynamics illustrated by the binding of antibodies to immobilized antigens

An electrochemical method is proposed for investigating the dynamics of rec ognition between a biomolecule and an immobilized receptor. It involves red ox labeling of the solute molecule and monitoring the binding by the electr ochemical response of the electrode onto which the receptor is immobilized. With large biomolecules, as, for example, antigens and antibodies, leading to small surface concentrations, simple redox labeling may prove insuffici ent to obtain detectable responses. Redox enzymes are then advantageously u sed as labels thanks to the signal amplification offered by their catalytic properties. The applicability of the method is illustrated by the reaction of an immobilized monolayer of goat IgG antigen (or of one Fab fragment) w ith an antigoat antibody labeled with glucose oxidase. Particular care is t aken to free the kinetic data from the effect of diffusion. The latter fact or may interfere whatever the detection technique. A full account of the co mbination between recognition kinetics and diffusion is therefore given in terms of a kinetic zone diagram leading to diagnostic criteria and data pro cessing procedures that allow a proper extraction of the recognition thermo dynamic and kinetic constants. The theory applies as well to the dynamics o f adsorption of nonbiological molecules on surfaces.