Oriented immobilization of antibodies on a silicon wafer using Si-tagged protein A

被引:70
作者
Ikeda, Takeshi [1 ,2 ]
Hata, Yumehiro [1 ]
Ninomiya, Ken-ichi [1 ]
Ikura, Yoshiaki [3 ]
Takeguchi, Keigo [3 ]
Aoyagi, Satoka [4 ]
Hirota, Ryuichi [1 ]
Kuroda, Akio [1 ,2 ]
机构
[1] Hiroshima Univ, Grad Sch Adv Sci Matter, Dept Mol Biotechnol, Hiroshima 7398530, Japan
[2] Hiroshima Univ, Res Inst Nanodevice & Bio Syst, Hiroshima 7398527, Japan
[3] Asahi Kasei Corp, Cent R&D Labs, Shizuoka 4168501, Japan
[4] Shimane Univ, Fac Life & Environm Sci, Matsue, Shimane 6908504, Japan
关键词
Immunoglobulin-binding protein; Oriented antibody immobilization; Si-tag; Silica-binding protein; Silicon wafer; Time-of-flight secondary ion mass spectrometry; ION MASS-SPECTROMETRY; ELECTRICAL DETECTION; SURFACE; BIOSENSOR; IMMUNOGLOBULIN; ORIENTATION; FILMS;
D O I
10.1016/j.ab.2008.11.001
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
We previously reported a silica-binding protein, designated Si-tag, which can be used to immobilize proteins on silica surfaces. Here, we constructed a fusion protein of Si-tag and immunoglobulin-binding staphylococcal protein A for oriented immobilization of antibodies on a silicon wafer whose surface is oxidized to silicon dioxide (silica). The fusion protein, Si-tagged protein A, strongly bound to the silica surface with a dissociation constant of 0.31 nM. Time-of-flight secondary ion mass spectrometry analysis of the silicon wafer coated with Si-tagged protein A, combined with principal component analysis and mutual information, demonstrated that protein A is localized on the outermost surface of the bound protein layer. Immunoglobulin G (IgG) was immobilized both on the silicon wafer coated with Si-tagged protein A and, as a control, directly on the intact silicon wafer via physical adsorption. The silicon wafer coated with Si-tagged protein A bound 30-70% more IgG than the uncoated silicon wafer, whereas the antigen-binding activity was 4- to 5-fold higher for the former, indicating that IgG was functionally immobilized on the silicon wafer via Si-tagged protein A in an oriented manner. (C) 2008 Elsevier Inc. All rights reserved.
引用
收藏
页码:132 / 137
页数:6
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