Understanding binding affinity is key to appreciation of the intermolecular interactions driving biological processes, structural biology, and structure-function relationships. It is also measured as part of the drug discovery process to help design drugs that bind their targets selectively and specifically.

Binding affinity is the strength of the binding interaction between a single biomolecule (e.g. protein or DNA) to its ligand/binding partner (e.g. drug or inhibitor). Binding affinity is typically measured and reported by the equilibrium dissociation constant (KD), which is used to evaluate and rank order strengths of bimolecular interactions. The smaller the KD value, the greater the binding affinity of the ligand for its target. The larger the KD value, the more weakly the target molecule and ligand are attracted to and bind to one another. Binding affinity is influenced by non-covalent intermolecular interactions such as hydrogen bonding, electrostatic interactions, hydrophobic and Van der Waals forces between the two molecules. In addition, binding affinity between a ligand and its target molecule may be affected by the presence of other molecules.

There are many ways to measure binding affinity and dissociation constants, such as ELISAs, gel-shift assays, pull-down assays, equilibrium dialysis, analytical ultracentrifugation, SPR, and spectroscopic assays. Isothermal titration calorimetry (ITC) is a direct, label-free analytical technique which measures the binding affinity between any two molecules that interact with each other, such as a protein and a ligand. ITC can measure KD values in the millimolar and nanomolar range, and can also determine the binding stoichiometry and binding thermodynamics important in the characterization of intermolecular interactions. ITC is considered the “Gold Standard’ of interaction analysis as it enables the study of a broad range of interactions and delivers highly quantitative KD values.