General Microarray Technique for Immobilization and Screening of Natural Glycans
Citations Over TimeTop 10% of 2007 papers
Abstract
We here present a printed covalent glycan microarray for protein-binding studies, using low-femtomole quantities of glycans. Glycans, either natural glycans, which were released from glycoproteins and glycolipids from natural sources, or synthetic glycans, were labeled with common fluorescent labels (e.g., 2-aminobenzamide or 2-aminobenzoic acid) by reductive amination and purified by HPLC. The purified glycoconjugates were covalently immobilized on commercial epoxide-activated glass slides via the secondary amine group that links the glycan moiety with the fluorescent tag. This immobilization procedure is generally applicable to reductively aminated glycans with different established fluorescent labels and allows the spatial arrangement of oligosaccharides. The microarray comprised a variety of natural glycans from various biological sources and synthetic glycans and provided informative binding fingerprints for the lectin concanavalin A as well as 14 monoclonal antibodies. Recognized glycans were characterized by tandem mass spectrometry revealing binding motifs. This natural glycan array allowed the characterization of the specificity of carbohydrate-binding proteins for oligosaccharide ligands from sparse biological sources. Moreover, it was applied for the characterization of the microarray glycans by using known carbohydrate-binding proteins.
Related Papers
- → Approach to the comprehensive analysis of glycoproteins isolated from human serum using a multi-lectin affinity column(2004)276 cited
- → A lectin array-based methodology for the analysis of protein glycosylation(2006)110 cited
- → The Lectin Riddle: Glycoproteins Fractionated from Complex Mixtures Have Similar Glycomic Profiles(2010)50 cited
- → Determination of the glycoprotein specificity of lectins on cell membranes through oxidative proteomics(2020)40 cited
- → Microassay Analyses of Protein Glycosylation(2009)