Abstract
To develop a novel high-throughput tool for monitoring carbohydrate–protein interactions, carbohydrate or glycoprotein microarrays have been prepared for binding with lectins. The interaction events are marked by attachment of fluorescent dyes and gold nanoparticles. The attachment of the fluorescent dyes and gold nanoparticles is achieved by standard avidin–biotin chemistry. The detection principle is fluorescence or resonance light scattering (RLS). The electroless deposition of silver onto the gold particles has been employed for RLS signal enhancement. Well-defined recognition systems, three monosaccharides (Man-α, Glc-β, and Gal-β) or three glycoproteins (Asf, RNase A, and RNase B) with two lectins (ConA and RCA120), are chosen here to establish the microarray-based assay, respectively. Highly selective recognition of carbohydrate–protein down to 25.6 pg/mL for RCA120 in solution and 8 μM for Gal-β and 32 ng/mL for Asf on the microarray spots is demonstrated.
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References
Schena, M. (2003) Microarray Analysis. Hoboken, NJ: Wiley-Liss.
Wang, D., Liu, S., Trummer, B. J., Deng, C., and Wang, A. (2002) Carbohydrate microarrays for the recognition of cross-reactive molecular markers of microbes and host cells. Nat. Biotechnol. 20, 275–281.
Varki, A. (1993) Biological roles of oligosaccharides: all of the theories are correct. Glycobiology 3, 97–130.
Feizi, T. (2000) Progress in deciphering the information content of the ‘glycome’-a crescendo in the closing years of the millennium. Glycoconj. J. 17, 553–565.
Blixt, O., Han, S., Liao, L., Zeng, Y., Hoffmann, J., Futakawa, S., and Paulson, J. C. (2008) Sialoside analogue arrays for rapid identification of high affinity siglec ligands. J. Am. Chem. Soc. 130, 6680–6681.
Huang, C. Y., Thayer, D. A., Chang, A. Y., Best, M. D., Hoffmann, J., Head, S., and Wong, C. H. (2006) Carbohydrate microarray for profiling the antibodies interacting with Globo H tumor antigen. Proc. Natl. Acad. Sci. U. S. A. 103, 15–20.
Liang, P. H., Wang, S. K., and Wong, C. H. (2007) Quantitative analysis of carbohydrate-protein interactions using glycan microarrays: determination of surface and solution dissociation constants. J. Am. Chem. Soc. 129, 11177–11184.
Gao, J., Liu, D., and Wang, Z. (2008) Microarray-based study of carbohydrate-protein binding by gold nanoparticle probes. Anal. Chem. 80, 8822–8827.
Taton, T. A., Mirkin, C. A., and Letsinger, R. L. (2000) Scanometric DNA array detection with nanoparticle probes. Science 289, 1757–1760.
Nam, J. M., Thaxton, C. S., and Mirkin, C. A. (2003) Nanoparticle-based bio-bar codes for the ultrasensitive detection of proteins. Science 301, 1884–1886.
Katz, E., and Willner, I. (2004) Integrated nanoparticle-biomolecule hybrid systems: synthesis, properties, and applications. Angew. Chem. Int. Ed., 43, 6042–6108.
Sun, L., Liu, D., and Wang, Z. (2007) Microarray-based kinase inhibition assay by gold nanoparticle probes. Anal. Chem. 79, 773–777.
Turkevich, J., Stevenson, P. C., and Hillier, J. (1951) A study of the nucleation and growth processes in the synthesis of colloidal gold. Discuss. Faraday Soc. 11, 55–75.
Frens, G. (1973) Controlled nucleation for the regulation of the particle size in monodisperse gold suspensions. Nat. Phys. Sci. 241, 20–22.
Wang, Z. X., Levy, R., Fernig, D. G., and Brust, M. (2005) The peptide route to multifunctional gold nanoparticles. Bioconjugate Chem. 16, 497–500.
Acknowledgments
The author would like to thank the NSFC (Grant No. 20675080 and 20875087) for financial support.
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Wang, Z., Gao, J. (2010). Microarray-Based Study of Carbohydrate–Protein Binding. In: Li, J. (eds) Functional Glycomics. Methods in Molecular Biology, vol 600. Humana Press. https://doi.org/10.1007/978-1-60761-454-8_10
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DOI: https://doi.org/10.1007/978-1-60761-454-8_10
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