Optical Spectroscopy in Studies of Antibody–Hapten Interactions

This article describes the use of optical spectroscopy in studying antibody–hapten interactions and in determining the equilibrium binding constants. Along with equilibrium binding data, spectroscopic tools often deliver structural information on bindinginduced conformational changes of antibodies (or haptens). Structural implications of results from example antibody–hapten systems are included. Fluorescence spectroscopy has been particularly useful in the area of ligand binding, and thus steady-state fluorescence quenching and fluorescence polarization are the primary techniques under discussion. A brief description of fluorescence correlation spectroscopy is also provided. Absorption techniques, including circular dichroism, are mentioned to a lesser extent. A basic description of the mathematical models involved in the analysis of binding equilibria is provided along with references to more complete works. Simulated and experimental data are used to illustrate the various experimental protocols and the appropriate analytical methods. Typical sources of errors and experimental precautions are indicated throughout the general discussion.