Antibody-ligand interactions for hydrophobic charge-induction chromatography: a surface plasmon resonance study.
This article describes the use of surface plasmon resonance (SPR) spectroscopy to study antibody-ligand interactions for hydrophobic charge-induction chromatography (HCIC) and its versatility in investigating the surface and solution factors affecting the interactions. Two density model surfaces presenting the HCIC ligand (mercapto-ethyl-pyridine, MEP) were prepared on Au using a self-assembly technique. The surface chemistry and structure, ionization, and protein binding of such model surfaces were characterized by X-ray photoelectron spectroscopy (XPS), near-edge X-ray absorption fine structure (NEXAFS), contact-angle titration, and SPR, respectively. The influences of the surface and solution factors, e.g., ligand density, salt concentration, and solution pH, on protein adsorption were determined by SPR. Our results showed that ligand density affects both equilibrium and dynamic aspects of the interactions. Specifically, a dense ligand leads to an increase in binding strength, rapid adsorption, slow desorption, and low specificity. In addition, both hydrophobic interactions and hydrogen bonding contribute significantly to the protein adsorption at neutral pH, while the electrostatic repulsion is overwhelmed under acidic conditions. The hydrophobic interaction at a high concentration of lyotropic salt would cause drastic conformational changes in the adsorbed protein. Combined with the self-assembly technique, SPR proves to be a powerful tool for studying the interactions between an antibody and a chromatographic ligand.
Fang
F
Cheng
§School of Life Science and Biotechnology, Dalian University of Technology, Dalian 116023, China.
Ming-Yang
MY
Li
Han-Qi
HQ
Wang
∥Department of Chemical and Biological Engineering, Zhejiang University, Hangzhou 310027, China.
Dong-Qiang
DQ
Lin
Jing-Ping
JP
Qu
D002845:Chromatography / Q000379:methods*; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D020349:Surface Plasmon Resonance / Q000379:methods*
D008024:Ligands