Hydrophobic interaction chromatography (HIC) is a powerful tool for the purification of biomolecules. The technique USES the interaction between the hydrophobic region on the protein surface and the weakly hydrophobic groups in the stationary phase for chromatography separation. HIC is a good complement to ion exchange and dimensional exclusion chromatography, especially when protein isomers are present in the sample, or when the target sample is similar to the isoelectric point or molecular weight of the impurity and is difficult to separate. HIC can be used after affinity chromatography by selectivity difference when affinity filler is difficult to distinguish proteins with similar recognition sites.