An analytical system for the characterization of highly heterogeneous mixtures of N-linked oligosaccharides

A novel system for characterizing complex N-linked oligosaccharide mixtures that uses a combination of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI–TOF MS), capillary electrophoresis (CE), and high-performance liquid chromatography (HPLC) has been developed. In...

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Bibliographic Details
Published in:Analytical biochemistry 2009-06, Vol.389 (1), p.40-51
Main Authors: Briggs, John B., Keck, Rodney G., Ma, Stacey, Lau, Wendy, Jones, Andrew J.S.
Format: Article
Language:English
Subjects:
Anion Exchange Resins
ANSA
Capillary electrophoresis
Carbohydrate Sequence
Carbohydrates
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Electrophoresis, Capillary
Glycoproteins
Glycoproteins - analysis
Glycoproteins - isolation & purification
High-performance liquid chromatography
Mass spectrometry
Molecular Sequence Data
Oligosaccharides
Oligosaccharides - analysis
Oligosaccharides - isolation & purification
Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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Summary:A novel system for characterizing complex N-linked oligosaccharide mixtures that uses a combination of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI–TOF MS), capillary electrophoresis (CE), and high-performance liquid chromatography (HPLC) has been developed. In this study, oligosaccharides released from recombinant TNK–tPA (tissue plasminogen activator) were derivatized with 5-amino-2-naphthalenesulfonic acid (ANSA). The negative charge imparted by the ANSA label facilitated the analysis of the oligosaccharides by MALDI–TOF MS by allowing the observation of both neutral and sialylated oligosaccharides in a single negative ion mode spectrum. Labeling with ANSA was also determined to be advantageous in the characterization of oligosaccharides by both HPLC and CE. The ANSA label was demonstrated to provide superior resolution over the commonly used label 8-aminopyrene-1,3,6-trisulfonic acid (APTS) in both the CE and HPLC analysis of oligosaccharides. To date, no other labels that enable the analysis of complex oligosaccharide mixtures in a single mass spectral mode, while also enabling high-resolution chromatographic and electrophoretic separation of the oligosaccharides, have been reported. By integrating the structural information obtained by MALDI–TOF MS analysis with the ability of CE and HPLC to discriminate between structural isomers, the complete characterization of complex oligosaccharide mixtures is possible.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2009.03.006