Continued advancement of protein array, bioelectrode, and biosensor technologies will necessitate development of methods that allow for increased protein immobilization capacity and more control over protein orientation. Toward these ends, we developed a method involving modification of chitosan with nitrilotriacetic acid (NTA) to achieve immobilization of a larger amount of His-tagged protein than is possible with current methods. The immobilization capacity of our method was evaluated using His-tagged GFP (Green Fluorescent Protein) as a model protein. The average immobilization density on modified glass was about 32 ng/mm2. Our method is suitable for use on a variety of solid surfaces, including glassy carbon, silicon wafers, polycarbonate, and beaten gold.
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M. Oshige, K. Yumoto, H. Miyata, S. Takahashi, M. Nakada, K. Ito, M. Tamegai, H. Kawaura and S. Katsura, "Immobilization of His-Tagged Proteins on Various Solid Surfaces Using NTA-Modified Chitosan," Open Journal of Polymer Chemistry, Vol. 3 No. 1, 2013, pp. 6-10. doi: 10.4236/ojpchem.2013.31002.
 F. Rusmini, Z. Zhong and J. Feijen, “Protein Immobilization Strategies for Protein Biochips,” Biomacromolecules, Vol. 8, No. 6, 2007, pp. 1775-1789.
 H. Ge, “UPA, a Universal Protein Array System for Quantitative Detection of Protein-Protein, Protein-DNA, Protein-RNA and Protein-Ligand Interactions,” Nucleic Acid Research, Vol. 28, No. 2, 2000, p. e3.
 L. J. Holt, K. Bussow, G. Walter and L. M. Tomlinson, “By-Passing Selection: Direct Screening for AntibodyAntigen Interactions Using Protein Arrays,” Nucleic Acid Research, Vol. 28, No. 2, 2000, p. e72.
 A. Pollak, H. Blumenfeld, M. Wax, R. L. Baughn and G. M. Whitesides, “Enzyme Immobilization by Condensation Copolymerization into Cross-Linked Polyacrylamide Gels,” Journal of the American Chemical Society, Vol. 102, No. 20, 1980, pp. 6324-6336.
 G. MacBeath and S. L. Schreiber, “Printing Proteins as Microarrays for High-Throughput Function Determination,” Science, Vol. 289, No. 5485, 2000, pp. 1760-1763.
 J. A. Camarero, “Recent Developments in the Site-Specific Immobilization of Proteins onto Solid Supports,” Biopolymers, Vol. 90, No. 3, 2008, pp. 450-458.
 M. A. Gauthier and H. A. Klok, “Peptide/Protein-Polymer Conjugates: Synthetic Strategies and Design Concepts,” Chemical Communications, No. 23, 2008, pp. 2591-2611. doi:10.1039/b719689j
 M. K?hn, “Immobilization Strategies for Small Molecule, Peptide and Protein Microarrays,” Journal of Peptide Science, Vol. 15, No. 6, 2009, pp. 393-397.
 M. Frasconi, F. Mazzei and T. Ferri, “Protein Immobilization at Gold-Thiol Surfaces and Potential for Biosensing,” Analytical and Bioanalytical Chemistry, Vol. 398, No. 4, 2010, pp. 1545-1564.
 S. Cosnier and M. Holzinger, “Electrosynthesized Polymers for Biosensing,” Chemical Society Reviews, Vol. 40, No. 5, 2011, pp. 2146-2156. doi:10.1039/c0cs00090f
 D. Samanta and A. Sarkar, “Immobilization of Bio-Macromolecules on Self-Assembled Monolayers: Methods and Sensor Applications,” Chemical Society Reviews, Vol. 40, No. 5, 2011, pp. 2567-2592.
 A. Holmberg, A. Blomstergren, O. Nord, M. Lukacs, J. Lundeberg and M. Uhlén, “The Biotin-Streptavidin Interaction Can Be Reversibly Broken Using Water at Elevated Temperatures,” Electrophoresis, Vol. 26, No. 3, 2005, pp. 501-510. doi:10.1002/elps.200410070
 M. L. Lesaicherre, R. Y. Lue, G. Y. Chen, Q. Zhu and S. Q. Yao, “Intein-Mediated Biotinylation of Proteins and Its Application in a Protein Microarray,” Journal of the American Chemical Society, Vol. 124, No. 30, 2002, pp. 8768-8769. doi:10.1021/ja0265963
 A. Girish, H. Sun, D. S. Yeo, G. Y. Chen, T. K. Chua and S. Q. Yao, “Site-Specific Immobilization of Proteins in a Microarray Using Intein-Mediated Protein Splicing,” Bioorganic & Medicinal Chemistry Letters, Vol. 15 No. 10, 2005, pp. 2447-2451. doi:10.1016/j.bmcl.2005.03.079
 T. Kinpara, R. Mizuno, Y. Murakami, M. Kobayashi, S. Yamaura, Q. Hasan, Y. Morita, H. Nakano, T. Yamane, and E. Tamiya, “A Picoliter Chamber Array for Cell-Free Protein Synthesis,” Journal of Biochemistry, Vol. 136 No. 2, 2004, pp. 149-154. doi:10.1093/jb/mvh102