| Chapter title |
A Rapid Procedure for the Purification of 8-Aminopyrene Trisulfonate (APTS)-Labeled Glycans for Capillary Electrophoresis (CE)-Based Enzyme Assays
|
|---|---|
| Chapter number | 18 |
| Book title |
Protein-Carbohydrate Interactions
|
| Published in |
Methods in molecular biology, April 2017
|
| DOI | 10.1007/978-1-4939-6899-2_18 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-6898-5, 978-1-4939-6899-2
|
| Authors |
Danyluk, Hayden J., Shum, Leona K., Zandberg, Wesley F., Hayden J. Danyluk, Leona K. Shum, Wesley F. Zandberg |
| Editors |
D. Wade Abbott, Alicia Lammerts van Bueren |
| Abstract |
Purified glycan standards are required for glycan arrays, characterizing substrate specificities of glycan-active enzymes, and to serve as retention-time or mobility standards for various separation techniques. This chapter describes a method for the rapid separation, and subsequent desalting, of glycans labeled with the highly fluorescent fluorophore 8-aminopyrene 1,3,6-trisulfonate (APTS). By using fluorophore-assisted carbohydrate electrophoresis (FACE) on polyacrylamide gels, which utilizes equipment readily available in most molecular biology laboratories, many APTS-labeled glycans can be simultaneously resolved. Excising specific gel bands containing the desired APTS-labeled glycans, followed by glycan elution from the gel and subsequent solid-phase extraction (SPE), yields single glycan species free of excess labeling reagents and buffer components. This chapter describes a FACE/SPE procedure ideal for preparing glycans for capillary electrophoresis (CE)-based enzyme assays, as well as for the purification of rare, commercially unavailable glycans from tissue culture samples. |
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