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Language: English

Conflict of Interest: In relation to this article, we declare that there is no conflict of interest.

Publication history: Received May 20, 2013
Accepted July 2, 2013

This is an Open-Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/bync/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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A comparative study on antibody immobilization strategies onto solid surface

Ji Eun Lee Jeong Hyun Seo Chang Sup Kim Yunkyeoung Kwon Jeong Hyub Ha¹ Suk Soon Choi² Hyung Joon Cha^†

Department of Chemical Engineering, Pohang University of Science and Technology, Pohang 790-784, Korea ¹School of Environmental Science and Technology, Pohang University of Science and Technology, Pohang 790-784, Korea ²Department of Biological and Environmental Engineering, Semyung University, Jecheon 390-711, Korea

hjcha@postech.ac.kr

Korean Journal of Chemical Engineering, October 2013, 30(10), 1934-1938(5), 10.1007/s11814-013-0117-5

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Abstract

Antibody immobilization onto solid surface has been studied extensively for a number of applications including immunoassays, biosensors, and affinity chromatography. For most applications, a critical consideration regarding immobilization of antibody is orientation of its antigen-binding site with respect to the surface. We compared two oriented antibody immobilization strategies which utilize thiolated-protein A/G and thiolated-secondary antibody as_x000D_ linker molecules with the case of direct surface immobilization of thiol-conjugated target antibody. Antibody immobilization degree and surface topography were evaluated by surface plasmon resonance and atomic force microscope, respectively. Protein A/G-mediated immobilization strategy showed the best result and secondary antibody-mediated immobilization was the worst for the total immobilization levels of target antibodies. However, when considering realto-ideal ratio for antigen binding, total target antigen binding levels (oriented target antibody immobilization levels) had the following order: secondary antibody-mediated immobilization>protein A/G-mediated immobilization>direct thiol-conjugated immobilization. Thus, we confirmed that protein A/G- and secondary antibody-mediated strategies, which consider orientation of target antibody immobilization, showed significantly high antigen binding efficiencies compared to direct random immobilization method. Collectively, the oriented antibody immobilization methods using linker materials could be useful in diverse antibody-antigen interaction-involved application fields.

Keywords

Antibody Immobilization Orientation Thiolation Protein A/G Secondary Antibody Surface Plasmon Resonance Atomic Force Microscopy

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