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Imaging glycans in zebrafish embryos by metabolic labeling and bioorthogonal click chemistry. 2011

Hao Jiang, and Lei Feng, and David Soriano del Amo, and Ronald D Seidel Iii, and Florence Marlow, and Peng Wu
Department of Biochemistry, Albert Einstein College of Medicine, Yeshiva University, USA.

Imaging glycans in vivo has recently been enabled using a bioorthogonal chemical reporter strategy by treating cells or organisms with azide- or alkyne-tagged monosaccharides. The modified monosaccharides, processed by the glycan biosynthetic machinery, are incorporated into cell surface glycoconjugates. The bioorthogonal azide or alkyne tags then allow covalent conjugation with fluorescent probes for visualization, or with affinity probes for enrichment and glycoproteomic analysis. This protocol describes the procedures typically used for noninvasive imaging of fucosylated glycans in zebrafish embryos, including: 1) microinjection of one-cell stage embryos with GDP-5-alkynylfucose (GDP-FucAl), 2) labeling fucosylated glycans in the enveloping layer of zebrafish embryos with azide-conjugated fluorophores via biocompatible Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC), and 3) imaging by confocal microscopy. The method described here can be readily extended to visualize other classes of glycans, e.g. glycans containing sialic acid and N-acetylgalactosamine, in developing zebrafish and in other living organisms.

UI MeSH Term Description Entries
D008845 Microinjections The injection of very small amounts of fluid, often with the aid of a microscope and microsyringes. Microinjection
D011134 Polysaccharides Long chain polymeric CARBOHYDRATES composed of MONOSACCHARIDES linked by glycosidic bonds. Glycan,Glycans,Polysaccharide
D004625 Embryo, Nonmammalian The developmental entity of a fertilized egg (ZYGOTE) in animal species other than MAMMALS. For chickens, use CHICK EMBRYO. Embryonic Structures, Nonmammalian,Embryo, Non-Mammalian,Embryonic Structures, Non-Mammalian,Nonmammalian Embryo,Nonmammalian Embryo Structures,Nonmammalian Embryonic Structures,Embryo Structure, Nonmammalian,Embryo Structures, Nonmammalian,Embryo, Non Mammalian,Embryonic Structure, Non-Mammalian,Embryonic Structure, Nonmammalian,Embryonic Structures, Non Mammalian,Embryos, Non-Mammalian,Embryos, Nonmammalian,Non-Mammalian Embryo,Non-Mammalian Embryonic Structure,Non-Mammalian Embryonic Structures,Non-Mammalian Embryos,Nonmammalian Embryo Structure,Nonmammalian Embryonic Structure,Nonmammalian Embryos,Structure, Non-Mammalian Embryonic,Structure, Nonmammalian Embryo,Structure, Nonmammalian Embryonic,Structures, Non-Mammalian Embryonic,Structures, Nonmammalian Embryo,Structures, Nonmammalian Embryonic
D005456 Fluorescent Dyes Chemicals that emit light after excitation by light. The wave length of the emitted light is usually longer than that of the incident light. Fluorochromes are substances that cause fluorescence in other substances, i.e., dyes used to mark or label other compounds with fluorescent tags. Flourescent Agent,Fluorescent Dye,Fluorescent Probe,Fluorescent Probes,Fluorochrome,Fluorochromes,Fluorogenic Substrates,Fluorescence Agents,Fluorescent Agents,Fluorogenic Substrate,Agents, Fluorescence,Agents, Fluorescent,Dyes, Fluorescent,Probes, Fluorescent,Substrates, Fluorogenic
D006154 Guanosine Diphosphate Fucose A nucleoside diphosphate sugar formed from GDPmannose, which provides fucose for lipopolysaccharides of bacterial cell walls, and for blood group substances and other glycoproteins. GDP Fucose,Guanosine Diphosphofucose,Diphosphate Fucose, Guanosine,Diphosphofucose, Guanosine,Fucose, GDP,Fucose, Guanosine Diphosphate
D000480 Alkynes Hydrocarbons with at least one triple bond in the linear portion, of the general formula Cn-H2n-2. Acetylenic Compounds,Alkyne,Acetylenes
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D001386 Azides Organic or inorganic compounds that contain the -N3 group. Azide
D015027 Zebrafish An exotic species of the family CYPRINIDAE, originally from Asia, that has been introduced in North America. Zebrafish is a model organism for drug assay and cancer research. Brachydanio rerio,Danio rerio,B. rerio,D. rerio,Zebra Fish,Zebra Fishes,Zebra danio,Zebrafishes,D. rerios,Fishes, Zebra,Zebra danios,danio, Zebra
D057930 Click Chemistry Organic chemistry methodology that mimics the modular nature of various biosynthetic processes. It uses highly reliable and selective reactions designed to "click" i.e., rapidly join small modular units together in high yield, without offensive byproducts. In combination with COMBINATORIAL CHEMISTRY TECHNIQUES, it is used for the synthesis of new compounds and combinatorial libraries. Click Chemical Reactions,Click Chemical Techniques,Chemical Reaction, Click,Chemical Reactions, Click,Chemical Technique, Click,Chemical Techniques, Click,Chemistries, Click,Chemistry, Click,Click Chemical Reaction,Click Chemical Technique,Click Chemistries,Reaction, Click Chemical,Reactions, Click Chemical,Technique, Click Chemical,Techniques, Click Chemical

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