How does affinity chromatography work?

The stationary phase of the chromatographic column is modified to covalently attach a ligand that binds to the target molecule of interest. The ligand can be an antibody, antigen, enzyme, or any molecule capable of binding specifically to the target. The immobilized ligand on the column captures the target molecule, while other molecules in the sample that lack affinity for the ligand pass through or bind weakly. A hydrocarbon chain inhibitor attached to the solid support prevents non-specific interactions and overlap. Non-target molecules are washed away with a wash buffer, removing unwanted contaminants. Since the target molecules have a stronger affinity for the stationary phase, they remain bound. The target molecules are then released from the immobilized ligand using an elution buffer with higher salt concentration. This results in the recovery of highly concentrated and purified target material.