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High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography
N-Glycosylation is the most complex post-translational modification of proteins and involved in many physiological processes and is therefore of major interest in academic research and in the biopharmaceutical industry. Reliable, robust, reproducible, and selective analysis of N-glycans is essential...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Springer Berlin Heidelberg
2018
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6338698/ https://www.ncbi.nlm.nih.gov/pubmed/30478517 http://dx.doi.org/10.1007/s00216-018-1495-7 |
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author | Wilhelm, Johannes G. Dehling, Marco Higel, Fabian |
author_facet | Wilhelm, Johannes G. Dehling, Marco Higel, Fabian |
author_sort | Wilhelm, Johannes G. |
collection | PubMed |
description | N-Glycosylation is the most complex post-translational modification of proteins and involved in many physiological processes and is therefore of major interest in academic research and in the biopharmaceutical industry. Reliable, robust, reproducible, and selective analysis of N-glycans is essential to understand the multitude of biological roles of N-glycosylation. So far, hydrophilic interaction liquid chromatography analysis of 2-AB or 2-AA derivatized N-glycans has been the standard method. In this work, the superiority of reversed-phase chromatography for complex N-glycosylation analysis is demonstrated. Separation of N-glycans derivatized with anthranilic acid on polar-embedded stationary alkyl phases with sub-2-μm particles results in outstanding selectivity and resolution. In combination with the highly mass spectrometry–compatible mobile phase, even very complex glycan mixtures can be separated, identified, and quantified precisely and accurately. The presented methodology can be applied broadly from basic research to analytical control and release testing of biological drug products and can be implemented in analytical laboratories with minimal effort. [Figure: see text] ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s00216-018-1495-7) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-6338698 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Springer Berlin Heidelberg |
record_format | MEDLINE/PubMed |
spelling | pubmed-63386982019-02-08 High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography Wilhelm, Johannes G. Dehling, Marco Higel, Fabian Anal Bioanal Chem Research Paper N-Glycosylation is the most complex post-translational modification of proteins and involved in many physiological processes and is therefore of major interest in academic research and in the biopharmaceutical industry. Reliable, robust, reproducible, and selective analysis of N-glycans is essential to understand the multitude of biological roles of N-glycosylation. So far, hydrophilic interaction liquid chromatography analysis of 2-AB or 2-AA derivatized N-glycans has been the standard method. In this work, the superiority of reversed-phase chromatography for complex N-glycosylation analysis is demonstrated. Separation of N-glycans derivatized with anthranilic acid on polar-embedded stationary alkyl phases with sub-2-μm particles results in outstanding selectivity and resolution. In combination with the highly mass spectrometry–compatible mobile phase, even very complex glycan mixtures can be separated, identified, and quantified precisely and accurately. The presented methodology can be applied broadly from basic research to analytical control and release testing of biological drug products and can be implemented in analytical laboratories with minimal effort. [Figure: see text] ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s00216-018-1495-7) contains supplementary material, which is available to authorized users. Springer Berlin Heidelberg 2018-11-26 2019 /pmc/articles/PMC6338698/ /pubmed/30478517 http://dx.doi.org/10.1007/s00216-018-1495-7 Text en © The Author(s) 2018 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. |
spellingShingle | Research Paper Wilhelm, Johannes G. Dehling, Marco Higel, Fabian High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography |
title | High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography |
title_full | High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography |
title_fullStr | High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography |
title_full_unstemmed | High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography |
title_short | High-selectivity profiling of released and labeled N-glycans via polar-embedded reversed-phase chromatography |
title_sort | high-selectivity profiling of released and labeled n-glycans via polar-embedded reversed-phase chromatography |
topic | Research Paper |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6338698/ https://www.ncbi.nlm.nih.gov/pubmed/30478517 http://dx.doi.org/10.1007/s00216-018-1495-7 |
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