Ion Exchange Chromatography Resins
Ion Exchange Chromatography is driven by interactions between the charges in the target molecule and the charges of the immobilized ligand on the chromatography resin. Ion exchange chromatography as a whole can be divided into two different sub types. Cation exchange chromatography, in which positively charged species in the mobile phase bind to a negatively charged ligand on the resin; and anion exchange chromatography, in which the binding species in the mobile phase are negatively charged, and the immobilized ligand is positively charged.
After the molecule of interest has been adsorbed, the column is washed to remove any residual unbound species from the solid phase. This is typically done with the same low ionic strength mobile phase in which the column was equilibrated. The bound molecules are then desorbed using a gradient of a second, higher ionic strength mobile phase to steadily increase the ionic strength of the eluent solution. An alternative desorption method involves using a mobile phase of greater or lesser pH than the equilibration mobile phase to give your molecule of interest or the ligand a charge at which they will not interact and your molecule of interest elutes from the resin.
In many cases it may be more advantageous to select conditions at which your target molecule will pass through the column unbound while the impurites will bind. This mode of binding is often referred to as "flow through mode" and it is most typically used with anion exchangers to remove DNA, endotoxins, and other negatively charged species while letting your target molecule to flow through the column.
Ion exchangers are typically classified as being either strong or weak. The terms “strong” and “weak” refer to the acid/base properties of the functional group. If the ligand is derived from a strong acid or a strong base it is referred to as a strong ion exchange resin. If the ligand is derived from a weak acid or a weak base it is referred to as a weak ion exchange resin. The type of ion exchange resin that will work best for a particular process will depend on the variance in the ionization state of the functional groups at different pH values. A strong ion exchange resin will have the same charge density on its surface over a broad pH range, whereas the charge density of a weak ion exchanger changes with pH. The selectivity and the capacity of a weak ion exchanger are different at different pH values. When developing a purification process, it is usually recommended that the chromatographer begin with a strong ion exchanger (e.g., quaternary amine, sulfonic acid, or sulfopropyl ), and then to try a weak ion exchanger (e.g., DEAE or carboxymethyl) if the selectivity is unsatisfactory.