Determination of Affinity and Kinetic Rate Constants Using Surface Plasmon Resonance

Abstract

Surface plasmon resonance (SPR) is a relatively new technique extremely useful for studying macromolecular interactions between proteins, proteins and DNA, or proteins and lipids. Biospecific interaction analyses using SPR provides valuable information about the strength, speed, and stoichiometry of the interaction in real time and without the use of labels. An excellent review on the commercial SPR instrument called BIAcore™ has been published recently (1). In general the device is a biosensor that measures mass accretion/loss within a finite surface volume as a function of time. The initial step is to stably immobilize a known quantity of one of the interactants (ligand) on the surface. Following this, the second or free-flowing interactant (analyte) is made available for binding to its putative surface-linked homologue through diffusion from a pool or source that steadily flows over the immobilized ligand surface. By replacing analyte solution with buffer the source becomes a sink taking away complex-dissociated analyte from the ligand surface, resulting in a detectable loss of mass.