Affinity chromatography makes use of specific binding interactions between molecules. It is this specificity that makes this mode of chromatography especially attractive as a capture step to isolate the target molecule from crude extracts. A particular ligand is chemically immobilized or “coupled” to a solid support so that when the crude extract is passed over the column, those molecules having a specific binding affinity to the ligand become adsorbed. After the non-specifically binding components are washed away, the target molecule is desorbed from the support, resulting in its purification from the original sample.
There are many custom designed affinity ligands available to the chromatographer. TOYOPEARL affinity chromatography resins are functionalized with chemically active groups or group-specific ligands. Resins with activated functional groups are ready to directly couple a protein or other ligand. Resins with reactive groups require carbodiimide coupling or reductive amination to achieve a stable covalent linkage.
TOYOPEARL polymeric AFC resins are available in a variety of group specific ligands. In addition, activated and reactive forms are available for the immobilization of target specific ligands. In AFC the mechanism of binding and elution varies depending on the immobilized ligand used. In some cases such as with TOYOPEARL AF-Chelate-650M (for Immobilized Metal Affinity Chromatography or IMAC) the binding and elution mechanism may change through the use of different metals or chemical modifiers. Dynamic binding capacity (DBC) values for AFC materials vary with the ligand and are typically on the low side. However, because AFC has high selectivity and most of the capacity is used for the target, with feedstock impurities remaining unbound, AFC resins are typically used for the Capture and Intermediate purification steps of a manufacturing process.