Affinity chromatography (AFC) is performed on a support that is functionalized with a ligand that shows biological affinity for a particular enzyme or other biological molecule. 

Almost all biomolecules can be purified on the basis of specific interaction between their chemical or biological structure and a suitable affinity ligand. Typical molecular pairs are antigens and antibodies, enzymes and coenzymes, and sugars with lectins. Thus, affinity chromatography distinguishes itself from, e.g., reversed phase and ion exchange chromatography, in that a highly specific interaction with the protein of interest is the cause for separation or purification.

Although affinity chromatography is not specific, in that no enzyme interacts with only one substrate, it is the most selective method for separating proteins. The selectivity is often based on spatial recognition via a 'lock-and-key' mechanism, as in antibody-antigen interactions.

Less selective general ligands have affinity for a whole class of proteins, which often require other chromatographic methods for purification of individual members. The term bioselective adsorption has been proposed as an alternative name for affinity chromatography.