Rapid Preparation of Released N-Glycans for HILIC Analysis Using a Novel Fluorescence and MS-Active Labeling Reagent

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Matthew A. Lauber, Darryl W. Brousmiche, Zhengmao Hua, Stephan M. Koza, Ellen Guthrie, Paula Magnelli, Christopher H. Taron, Kenneth J. Fountain
Waters, New England BioLabs
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Application Notes
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Application Notes
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Conventional approaches to the preparation of N-glycans for HILIC-FLR-MS are either laborious, time-consuming, or require compromises in sensitivity. With the development of the GlycoWorks RapiFluor-MS N-Glycan Kit, we address these shortcomings by enabling unprecedented sensitivity for glycan detection while also improving the throughput of N-glycan sample preparation.

With this approach:

  • Glycoproteins are deglycosylated in approximately 10 minutes to produce N-glycosylamines.
  • These glycans are then rapidly reacted with the novel RapiFluor-MS reagent within a 5-minute reaction and are thereby labeled with a tag comprised of an efficient fluorophore and a highly basic tertiary amine that yields enhanced sensitivity for both fluorescence and MS detection.
  • In a final step requiring no more than 15 minutes, the resulting RapiFluor-MS labeled glycans are extracted from reaction byproducts by means of μElution HILIC SPE that has been rigorously developed to provide quantitative recovery of glycans (from neutral to tetrasialylated species) and to facilitate immediate analysis of samples.

Accordingly, an analyst can complete an N-glycan sample preparation, from glycoprotein to ready-to-analyze sample, in just 30 minutes when using the sensitivity enhancing RapiFluor-MS labeling reagent.

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