Solid Supports in Enzyme-Linked Immunosorbent Assay and Other SolidPhase Immunoassays

A very large proportion of modern immunoassays involve the use of synthetic solid phases to immobilize one of the reactants. These solid-phase immunoassays (SPIs) therefore involve ligand–receptor interactions that occur within a reaction volume close to the solution/solid phase interface. As a consequence, the immunochemistry/biochemistry of these ligand–receptor interactions differs from that of their counterparts in solution. Furthermore, the immobilization process can significantly alter the biological activity of the reactant; most adsorbed proteins on polystyrene or silicone are partially or largely denatured. Therefore the use of alternative methods of immobilization is attractive but may result in little increase in the amount of total functional reactant. However, all commonly used solid phases do not have the same properties or the same capacity for reactant immobilization or experience the same level of nonspecific binding. Empiricism plays a major role in SPIs. Derivations of mass law equations for measuring the antigen capture of solid-phase antibodies, for determining the affinity of solid phase for protein adsorption, and for estimating antibody affinity are reviewed.