title: Ultrasensitive detection and quantification of acidic disaccharides using capillary electrophoresis and quantum dot-based fluorescence resonance energy transfer.
abstract: Rapid and highly sensitive detection of the carbohydrate components of glycoconjugates is critical for advancing glycobiology. Fluorescence (or Förster) resonance energy transfer (FRET) is commonly used in detection of DNA, in protein structural biology, and in protease assays but is less frequently applied to glycan analysis due to difficulties in inserting two fluorescent tags into small glycan structures. We report an ultrasensitive method for the detection and quantification of a chondroitin sulfate disaccharide based on FRET, involving a CdSe-ZnS core-shell nanocrystal quantum dot (QD) streptavidin conjugate donor and a Cy5 acceptor. The disaccharide was doubly labeled with biotin and Cy5. QDs then served to concentrate the target disaccharide, enhancing the overall energy transfer efficiency, with unlinked QDs and Cy5 hydrazide producing nearly zero background signal in capillary electrophoresis using laser-induced fluorescence detection with two different band-pass filters. This method is generally applicable to the ultrasensitive analysis of acidic glycans and offers promise for the high-throughput disaccharide analysis of glycosaminoglycans.
authors.affiliation: Department of Chemistry and Chemical Biology, ‡Chemical and Biological Engineering, §Biology, and ∥Biomedical Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute , Troy, New York 12180, United States.
authors.forename: Yuqing
authors.initials: Y
authors.lastname: Chang
authors.forename: Chao
authors.initials: C
authors.lastname: Cai
authors.forename: Lingyun
authors.initials: L
authors.lastname: Li
authors.forename: Jianjun
authors.initials: J
authors.lastname: Miao
authors.forename: Ebru
authors.initials: E
authors.lastname: Ucakturk
authors.forename: Guoyun
authors.initials: G
authors.lastname: Li
authors.forename: Mellisa
authors.initials: M
authors.lastname: Ly
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Linhardt
mesh_terms: D002809:Chondroitin Sulfates / Q000737:chemistry; D004187:Disaccharides / Q000032:analysis*; D019075:Electrophoresis, Capillary; D031541:Fluorescence Resonance Energy Transfer; D015394:Molecular Structure; D045663:Quantum Dots
chemical_list: D004187:Disaccharides; D002809:Chondroitin Sulfates