IMMOBILIZATION OF IGG ONTO GOLD SURFACES AND ITS INTERACTION WITH ANTI-IGG STUDIED BY SURFACE-PLASMON RESONANCE

The optical excitation of surface plasmon resonance (SPR) at a metal d ielectric interface has been used to study the binding of immunoglobul in G (IgG) to gold and anti-IgG to immobilised IgG layers. In these st udies both a monoclonal mouse and polyclonal sheep IgG were used as re ceptor layers for anti-IgG. The kinetics of binding were investigated by monitoring the reflectivity of light at an angle close to plasmon r esonance. Both the initial rate of change and final reflectivity were measured during and after protein binding. The amount of protein bound to the surface was found to be less for the monoclonal mouse IgG comp ared to the polyclonal sheep IgG, these two IgG nominally being of the same dimensions and molecular weight. Further, anti-IgG binding produ ced greater changes in reflectivity than the initial IgG layers. By fi tting the full angle-dependent reflectivity data to the Fresnel equati on the effective protein layer thicknesses of IgG and anti-IgG as a fu nction of concentration were determined. Differences in the effective thickness of the bound layer for the two IgG was observed, the mouse I gG having a thinner effective thickness compared with the sheep IgG. T he limitations of direct binding of protein to metal surfaces in SPR b iosensor applications are discussed.