title: A new method for multilayered, site-directed immobilization of antibody on polystyrene surface.
abstract: Polystyrene is a common substrate material for protein adsorption in biosensors and bioassays. Here, we present a new method for multilayered, site-directed immobilization of antibody on polystyrene surface through the linkage of a genetically engineered ligand and the assembly of staphylococcal protein A (SPA) with immunoglobulin G (IgG). In this method, antibodies were stacked on polystyrene surface layer by layer in a potential three-dimensional way and exposed the analyte-binding sites well. Enzyme-linked immunosorbent assay (ELISA) revealed that the new method showed a 32-fold higher detection sensitivity compared with the conventional one. Pull-down assay and Western blot analysis further confirmed that it is different from the ones of monolayer adsorption according to the comparison of adsorption capacity. The differentiated introduction of functional ligands, which is the key of this method, might offer a unique idea as a way to interfere with the dynamic behavior of a protein complex during the process of adsorption.
authors.affiliation: College of Chemical Engineering, Xiangtan University, Xiangtan 411105, Hunan Province, China. Electronic address: fengbo5460@hotmail.com.
authors.forename: Bo
authors.initials: B
authors.lastname: Feng
authors.affiliation: College of Chemical Engineering, Xiangtan University, Xiangtan 411105, Hunan Province, China.
authors.forename: Caiyun
authors.initials: C
authors.lastname: Wang
authors.affiliation: College of Chemical Engineering, Xiangtan University, Xiangtan 411105, Hunan Province, China.
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Xie
authors.affiliation: College of Chemical Engineering, Xiangtan University, Xiangtan 411105, Hunan Province, China.
authors.forename: Xi
authors.initials: X
authors.lastname: Feng
authors.affiliation: College of Chemical Engineering, Xiangtan University, Xiangtan 411105, Hunan Province, China.
authors.forename: Yuqin
authors.initials: Y
authors.lastname: Li
authors.affiliation: College of Life Science, Wuhan University, Wuhan 430072, Hubei Province, China.
authors.forename: Zhijian
authors.initials: Z
authors.lastname: Cao
mesh_terms: D000327:Adsorption; D000914:Antibodies, Viral / Q000737:chemistry* / Q000276:immunology*; D001665:Binding Sites; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006514:Hepatitis B Surface Antigens / Q000737:chemistry* / Q000276:immunology*; D011137:Polystyrenes / Q000737:chemistry*; D011485:Protein Binding; D013499:Surface Properties
chemical_list: D000914:Antibodies, Viral; D020099:Coated Materials, Biocompatible; D006514:Hepatitis B Surface Antigens; D011137:Polystyrenes
keywords: Affinity ligand; Antibody immobilization; Immunoglobulin G; Polystyrene surface; Staphylococcal protein A