@article { ISI:000338488800044,
	title = {Simple Capillary Electrophoresis-Mass Spectrometry Method for Complex Glycan Analysis Using a Flow-Through Microvial Interface},
	journal = {ANALYTICAL CHEMISTRY},
	volume = {86},
	number = {13},
	year = {2014},
	month = {JUL 1},
	pages = {6479-6486},
	abstract = {A flow-through microvial is used to interface capillary electrophoresis and mass spectrometry (CE-MS) to develop a method for simultaneous profiling both neutral and sialylated glycans without derivatization or labeling. The CE separation was performed at near-zero electroosmotic flow in a capillary with neutral, hydrophilic coating, using 50 mM ammonium acetate in 20\% methanol (pH 3.1) as the background electrolyte. The method was optimized with reversed CE polarity and negative ion ESI-MS. Enzymatically released N-glycans from human immunoglobulin G (IgG) were used as the test sample. The approach was also used to study the more complex N-glycans from recombinant human erythropoietin (rHuEPO) expressed in Chinese hamster ovary (CHO) cells. Glycoscreening of rHuEPO was performed using a triple quadrupole MS and an ultrahigh resolution TOF-MS. The high sensitivity and high mass accuracy of the TOP-MS revealed the presence of more than 70 glycans. Three mono- and di-sialylated tetra-antennary N-glycans and one mono-sialylated tri-antennary N-glycan of rHuEPO are reported for the first time. Further glycan heterogeneity was identified of the highly sialylated N-glycans of rHuEPO by extensive acetylation, Neu5Ac/Neu5Gc variation and the presence of N-acetyl-lactosamine repeats. For comparative purposes, porous graphitic carbon-based LC-MS/MS was also used to glycoprofile rHuEPO. This work demonstrates the potential of CE-MS to provide a comprehensive glycosylation profile with detailed features of the secondary glycan modifications. The CE-MS based method eliminates the need to label the N-glycans, as well as the requirement to desialylate before analysis, and could complement other established techniques for glycan characterization of therapeutic glycoproteins.},
	issn = {0003-2700},
	doi = {10.1021/ac5010212},
	author = {Jayo, Roxana G. and Thaysen-Andersen, Morten and Lindenburg, Petrus W. and Haselberg, Rob and Hankemeier, Thomas and Ramautar, Rawi and Chen, David D. Y.}
}
