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Hydrophilic Interaction Chromatography-Based High-Throughput Sample Preparation Method for N-Glycan Analysis from Total Human Plasma Glycoproteins
Abstract
Many diseases are associated with changes in the glycosylation of plasma proteins. To search for glycan biomarkers, large sample sets have to be investigated for which high-throughput sample preparation and analysis methods are required. We here describe a 96 well plate-based high-throughput procedure for the rapid preparation of 2-aminobenzoic acid (2-AA) labeled N-glycans from 10 microL of human plasma. During this procedure, N-glycans are released from glycoproteins and subsequently labeled with 2-AA without prior purification. A hydrophilic interaction chromatography (HILIC)-based solid phase extraction method is then applied to isolate the 2-AA labeled N-glycans, which can be analyzed by MALDI-TOF-MS, HPLC with fluorescence detection, and CE-MS. The relative standard deviation for the intrabatch repeatability and the interbatch repeatability of the sample preparation method remained below 7% and below 9%, respectively, for all peaks observed by HPLC. Similar results were obtained with MALDI-TOF-MS, where 47 N-glycans could be measured consistently. The 2-AA labeled N-glycans were additionally analyzed by a CE-ESI-Q-TOF-MS method, which featured high resolution and mass accuracy, allowing the unambiguous determination of the N-glycan compositions. Up to four times, 96 human plasma samples can be handled in parallel, which, together with the versatility of the 2-AA label, makes this procedure very attractive for glycomics analysis of larger sample cohorts.
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Cited by
Oligosaccharide Analysis by Mass Spectrometry: A Review of Recent Developments
Kailemia, M. J., Ruhaak, L. R., Lebrilla, C. B., & Amster, I. J. (2014) · Analytical Chemistry · doi:10.1021/ac403969n · Cited by 377
Self-citationHuhn, C., Selman, M. H. J., Ruhaak, L. R., Deelder, A. M., & Wuhrer, M. (2009) · PROTEOMICS · doi:10.1002/pmic.200800715 · Cited by 301
Self-citationHILIC analysis of fluorescence-labeled N-glycans from recombinant biopharmaceuticals
Melmer, M., Stangler, T., Schiefermeier, M., Brunner, W., Toll, H., Rupprechter, A., Lindner, W., & Premstaller, A. (2010) · Analytical and Bioanalytical Chemistry · doi:10.1007/s00216-010-3988-x · Cited by 104
Giménez, E., Ramos‐Hernan, R., Benavente, F., Barbosa, J., & Sanz‐Nebot, V. (2011) · Rapid Communications in Mass Spectrometry · doi:10.1002/rcm.5114 · Cited by 36
Metadata
- Authors
- Ruhaak, L. R., Huhn, C., Waterreus, W.-J., de Boer, A. R., Neusüss, C., Hokke, C. H., Deelder, A. M., & Wuhrer, M.
- Venue
- Analytical Chemistry
- Publisher
- American Chemical Society (ACS)
- Published
- 2008-07-02
- Volume
- 80
- Issue
- 15
- Pages
- 6119-6126
- ISSN
- 0003-2700 (ISSN-L)
- 1520-6882
- OpenAlex
- W2043272559
- Cited by
- 208
- References
- 0
- FWCI
- 4.71
- Type
- journal article
- Language
- en
- Open access
- closed
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- Ruhaak, L. R., Huhn, C., Waterreus, W.-J., de Boer, A. R., Neusüss, C., Hokke, C. H., Deelder, A. M., & Wuhrer, M. (2008). Hydrophilic Interaction Chromatography-Based High-Throughput Sample Preparation Method for N-Glycan Analysis from Total Human Plasma Glycoproteins. Analytical Chemistry. https://doi.org/10.1021/ac800630x
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