ABSTRACT
This chapter focuses on how equilibrium binding constants characterizing non-covalent biomolecular interactions are determined by capillary electrophoresis (CE). It describes binding constant determination in different systems, outlines the theory and different experimental set-ups, and discusses unique problems, pitfalls, and the validity of the results. CE analyses where selectivity for whatever purpose is modulated by reversible molecular interactions may be called affinity CE or capillary affinity electrophoresis. It is clear that there will be interactions characterized by on-and-off rates that are unsuitable for quantitation by CE and that some ligands are uncharged or too small to change the electrophoretic velocity of the analyte–ligand complex as compared with the unbound analyte. CE may be conveniently used to determine the minimum time for equilibration of an analyte–ligand mixture simply by performing repeated injections and determining the time at which the amount of free analyte is not changed anymore.
