The understanding of glycosylation alterations in health and disease has evolved significantly and glycans are considered to be relevant biomarker candidates. High-throughput analytical technologies capable of generating high-quality, large-scale glycoprofiling data are in high demand. Here, we describe an automated sample preparation workflow and analysis of N-linked glycans from plasma samples using hydrophilic interaction liquid chromatography with fluorescence detection on an ultrahigh-performance liquid chromatography (UHPLC) instrument. Samples are prepared in 96-well plates and the workflow features rapid glycoprotein denaturation, enzymatic glycan release, glycan purification on solid-supported hydrazide, fluorescent labeling, and post-labeling cleanup with solid-phase extraction. The development of a novel approach for plasma N-glycan analysis and its implementation on a robotic platform significantly reduces the time required for sample preparation and minimizes technical variation. It is anticipated that the developed method will contribute to expanding high-throughput capabilities to analyze protein glycosylation.
CITATION STYLE
Adamczyk, B., Stöckmann, H., O’Flaherty, R., Karlsson, N. G., & Rudd, P. M. (2017). High-throughput analysis of the plasma N-glycome by UHPLC. In Methods in Molecular Biology (Vol. 1503, pp. 97–108). Humana Press Inc. https://doi.org/10.1007/978-1-4939-6493-2_8
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