Isolation and recovery of acidic oligosaccharides from polyacrylamide gels by semi-dry electrotransfer.
Acidic oligosaccharides derived from glycosaminoglycan heparin were separated by polyacrylamide gradient gel electrophoresis (PAGE). The gel could be visualized using Alcian Blue dye to give a pattern of highly resolved, well defined bands. The particular banding pattern obtained was the result of a heparinase catalyzed depolymerization which afforded oligosaccharide products that differed in size by one disaccharide unit. The separated oligosaccharides could be recovered prior to staining by electroelution onto a positively charged nylon membrane by a semi-dry transfer procedure. Subsequent elution and quantitative recovery of individual oligosaccharides from the membrane was achieved. By using multiple membrane layers a second separation dimension was obtained, resulting in increased oligosaccharide purity proportional to transfer depth. Preparative gradient polyacrylamide gel electrophoresis followed by semi-dry electro-transfer and recovery represents a novel method for the preparation of homogeneous acidic oligosaccharides.
Division of Medicinal and Natural Products Chemistry, College of Pharmacy, University of Iowa, Iowa City 52242.
A
A
al-Hakim
R J
RJ
Linhardt
D000423:Alcian Blue; D004591:Electrophoresis, Polyacrylamide Gel; D006025:Glycosaminoglycans / Q000032:analysis; D006493:Heparin / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D015394:Molecular Structure; D009844:Oligosaccharides / Q000302:isolation & purification*; D013194:Staining and Labeling
D006025:Glycosaminoglycans; D009844:Oligosaccharides; D006493:Heparin; D000423:Alcian Blue