Immobilisation 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 dielectric interface has been used to study the binding of immunoglobulin G (IgG) to gold and anti-IgG to immobilised IgG layers. In these studies both a monoclonal mouse and polyclonal sheep IgG were used as receptor layers for anti-IgG. The kinetics of binding were investigated by monitoring the reflectivity of light at an angle close to plasmon resonance. 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 compared to the polyclonal sheep IgG, these two IgG nominally being of the same dimensions and molecular weight. Further, anti-IgG binding produced greater changes in reflectivity than the initial IgG layers. By fitting the full angle-dependent reflectivity data to the Fresnel equation the effective protein layer thicknesses of IgG and anti-IgG as a function of concentration were determined. Differences in the effective thickness of the bound layer for the two IgG was observed, the mouse IgG having a thinner effective thickness compared with the sheep IgG. The limitations of direct binding of protein to metal surfaces in SPR biosensor applications are discussed.

Surface plasmon resonance studies of immunoreactions utilizing disposable diffraction gratings.

The specificity of protein binding between immuno-gamma globulin (IgG) and anti-IgG has been investigated by means of surface plasmon resonance measurements, with a view to determining whether a perspex replica of a holographic diffraction grating can be used efficiently as the momentum-coupling device. The replicas are easy to fabricate, of low cost, and may be useful as disposable sensing heads in a biosensor. The majority of all published work concerning surface-plasmon-based biosensors has detailed the use of prisms as the momentum-coupling devices. Therefore, the relative advantages and disadvantages of both systems are discussed in this paper, including a sensitive optical technique which is beyond the scope of prism geometries.