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论文类型：期刊论文

发表时间：2014-11-21

发表刊物：JOURNAL OF CHROMATOGRAPHY A

收录刊物：SCIE、EI、PubMed

卷号：1369

页面范围：98-104

ISSN号：0021-9673

关键字：HIC; Displacement chromatography; Cyclodextrin; Antibody purification; Salt-independent; Host-guest interaction

摘要：Hydrophobic interaction chromatography (HIC) offers an orthogonal selectivity to ion exchange chromatography and the combination of the two processes can provide a potential cost-effective alternative to protein A chromatography in industrial antibody purification. However, the application of HIC is limited by its close dependence on high concentrations of kosmotropic salts to achieve desired separation. These salts can cause antibody precipitation and induce the corrosion of manufacturing facilities. Here, we report a new strategy of salt-independent HIC, which can capture antibody at the physiological salt concentration and allow the recovery of bound proteins through cyclodextrin (CD)-based displacement elution. Hydrophobicity-intensified HIC media with different coupling amount of phenyl ligands were prepared and assessed for their antibody binding capacity and selectivity. beta-CD was investigated for its supermolecular interaction with phenyl ligands and elution capacity as a displacer. The results clarified a nearly linear correlation between binding capacity of human immunoglobulin G (IgG) and phenyl coupling density in the range of 44-159 mu mol/mL. The host-guest interaction between beta-CD and the phenyl ligands revealed a modest binding strength (K-a = 4.1 x 10(3) M-1), and 15 mM beta-CD solution showed a general effectiveness as displacement eluent for these HIC media, with IgG recovery varying with the ligand density. This strategy allowed the direct purification of human IgG from serum with satisfactory purity. The whole procedure of this method, including loading and elution, can be performed under physiological conditions. We expect such a salt-independent mode of HIC could be used as a capture or intermediate step in industrial antibody purification. (C) 2014 Elsevier B.V. All rights reserved.