15] Determination of rate and equilibrium binding constants for macromolecular interactions by surface plasmon resonance

Abstract

This chapter focuses on the determination of rate and equilibrium binding constants for macromolecular interactions by surface plasmon resonance. The characterization of the kinetics and thermodynamics of macromolecular interactions is increasingly important for developing an understanding of the molecular basis of events such as cell adhesion and viral infection, and it may ultimately aid in the rational design of antagonists of such interactions. Surface plasmon resonance (SPR) detectors, such as the BIAcore instrument, Pharmacia, allow for the direct visualization of these macromolecular interactions in real time, and thus the data obtained contains information on the rate and equilibrium binding constants that describe the interaction being investigated. SPR is an optical phenomenon that occurs as a result of the total internal reflection (TIR) of light at a metal film-liquid interface. TIR is observed in situations where light travels through an optically dense medium such as glass and is reflected back through that medium at the interface with a less optically dense medium such as buffer, provided the angle of incidence is greater than the critical angle required for the pair of optical media.