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Sensors 2018, 18(3), 784; https://doi.org/10.3390/s18030784

A New Method for Immobilization of His-Tagged Proteins with the Application of Low-Frequency AC Electric Field

Department of Environmental Engineering Science, Graduate School of Science and Technology, Gunma University, Kiryu, Gunma 376-8515, Japan
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Received: 29 January 2018 / Revised: 28 February 2018 / Accepted: 1 March 2018 / Published: 5 March 2018
(This article belongs to the Special Issue Protein-Based Biosensors)
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Abstract

Continued advancement of protein array, bioelectrode, and biosensor technologies is necessary to develop methods for higher amount and highly oriented immobilization activity of proteins. In pursuit of these goals, we developed a new immobilization method by combining electrostatic transport and subsequent molecular diffusion of protein molecules. Our developed immobilization method is based on a model that transports proteins toward the substrate surface due to steep concentration gradient generated by low-frequency AC electric field. The immobilization of the maximum amounts can be obtained by the application of the AC voltage of 80 Vpp, 20 Hz both for His-tagged Green Fluorescent Protein (GFP) and Discosoma sp. Red Fluorescent Protein (DsRed), used as model proteins. The amounts of the immobilized His-tagged GFP and DsRed were approximately seven-fold higher than that in the absence of the application of low-frequency AC electric field. Furthermore, the positively and negatively charged His-tagged GFP at acidic and alkaline pH were immobilized by applying of low-frequency AC electric field, whereas the non-charged His-tagged GFP at the pH corresponding to its isoelectric point (pI) was not immobilized. Therefore, unless the pH is equal to pI, the immobilization of electrically charged proteins was strongly enhanced through electrostatic transport and subsequent molecular diffusion. View Full-Text
Keywords: protein immobilization; mass transport phenomenon; self-assembled monolayer; surface treatment; low-frequency alternating current (AC) electric field protein immobilization; mass transport phenomenon; self-assembled monolayer; surface treatment; low-frequency alternating current (AC) electric field
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).
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Takahashi, S.; Kishi, K.; Hiraga, R.; Hayashi, K.; Mamada, Y.; Oshige, M.; Katsura, S. A New Method for Immobilization of His-Tagged Proteins with the Application of Low-Frequency AC Electric Field. Sensors 2018, 18, 784.

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