Abstract: The invention discloses a Non-Equilibrium Capillary Electrophoresis of Equilibrium Mixtures (NECEEM) method and NECEEM-based practical applications. The NECEEM method is a homogeneous technique, which, in contrast to heterogeneous methods, does not require affixing molecules to a solid substrate. The method of the invention facilitates 3 practical applications. In the first application, the method allows the finding of kinetic and thermodynamic parameters of complex formation. It advantageously allows for revealing two parameters, the equilibrium dissociation constant, Kd, and the monomolecular rate constant of complex decay, koff, in a single experiment. In the second practical application, the method of this invention provides an approach for quantitative affinity analysis of target molecules. It advantageously allows for the use of affinity probes with relatively high values of koff.

Abstract: The invention discloses a Non-Equilibrium Capillary Electrophoresis of Equilibrium Mixtures (NECEEM) method and NECEEM-based practical applications. The NECEEM method is a homogeneous technique, which, in contrast to heterogeneous methods, does not require affixing molecules to a solid substrate. The method of the invention facilitates 3 practical applications. In the first application, the method allows the finding of kinetic and thermodynamic parameters of complex formation. It advantageously allows for revealing two parameters, the equilibrium dissociation constant, Kd, and the monomolecular rate constant of complex decay, koff, in a single experiment. In the second practical application, the method of this invention provides an approach for quantitative affinity analysis of target molecules. It advantageously allows for the use of affinity probes with relatively high values of koff.