Nature Methods 2, 851 - 856 (2005)
Published online: 21 October 2005; | doi:10.1038/nmeth803
Evanescent-field fluorescence-assisted lectin microarray: a new strategy for glycan profilingAtsushi Kuno1, 4, Noboru Uchiyama1, 4, Shiori Koseki-Kuno1, Youji Ebe2, Seigo Takashima3, Masao Yamada2, 3
& Jun Hirabayashi11
Research Center for Glycoscience (RCG), National Institute of Advanced Industrial Science and Technology (AIST), AIST Tsukuba Central 2; Tsukuba, Ibaraki 305-8568, Japan. 2
Moritex Co., Yokohama, Kanagawa 225-0012, Japan. 3
Nippon Laser & Electronics Lab., Nagoya, Aichi 456-0032, Japan. 4
These authors contributed equally to this work.
Correspondence should be addressed to Jun Hirabayashi jun-hirabayashi@aist.go.jp Glycans have important roles in living organisms with their structural diversity. Thus, glycomics, especially aspects involving the assignment of functional glycans in a high-throughput manner, has been an emerging field in the postproteomics era. To date, however, there has been no versatile method for glycan profiling. Here we describe a new microarray procedure based on an evanescent-field fluorescence-detection principle, which allows sensitive, real-time observation of multiple lectin-carbohydrate interactions under equilibrium conditions. The method allows quantitative detection of even weak lectin-carbohydrate interactions (dissociation constant, K
d > 10-6 M) as fluorescent signals for 39 immobilized lectins. We derived fully specific signal patterns for various Cy3-labeled glycoproteins, glycopeptides and tetramethylrhodamine (TMR)-labeled oligosaccharides. The obtained results were consistent with the previous reports of glycoprotein and lectin specificities. We investigated the latter aspects in detail by frontal affinity chromatography, another profiling method. Thus, the developed lectin microarray should contribute to creation of a new paradigm for glycomics.
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