In an aqueous environment most biomolecules are charged due to the presence of ionic groups within their structure. Depending on the pH of the environment these biomolecules carry a net positive charge, a net negative charge or no net charge. The pH value at which a biomolecule carries no net charge is called its isoelectric point (pI). The pI is highly specific for each type of biomolecule. When exposed to a pH below its pI a biomolecule will carry a positive charge.
It is therefore possible to separate out biomolecules according to their charge characteristics using a column of separation media that consists of beads covered in positively or negatively charged groups. A positively charged bead, known as an anionic exchanger, will tend to bind to biomolecules with a net negative charge, and a negatively charged bead, known as a cationic exchanger, will tend to bind to biomolecules with a net positive charge. The binding of the biomolecules to the beads is fully reversible and their removal (elution) is usually achieved through the flow of increasing amounts of sodium chloride salt down the column. The sodium or chloride ions compete with the binding of the biomolecules to the charged beads causing the biomolecules to be released and allowing them to be eluted out of the bottom of the column. The order in which the biomolecules are eluted is dependant upon their net charge; with the weakest charged coming off first.
Ion Exchange chromatography is a binding technique with very high binding capacities, high flow characteristics and potentially excellent resolution. It is therefore perfect for the separation of large volumes of sample (fluid feed) and fits well into the early or capture step of a purification methodology.