期刊
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
卷 405, 期 8, 页码 2481-2493出版社
SPRINGER HEIDELBERG
DOI: 10.1007/s00216-012-6690-3
关键词
Anthranilic acid; 2-aminobenzamide; Mass spectrometry; N-glycosylation; Reversed-phase chromatography; Monoclonal antibody
N-Glycosylation is a common post-translational modification of monoclonal antibodies with a potential effect on the efficacy and safety of the drugs; detailed knowledge about this glycosylation is therefore crucial. We have developed a reversed-phase liquid chromatographic-mass spectrometric method, with different fluorescent labels, for analysis of N-glycosylation, and compared the sensitivity and selectivity of the methods. Our work demonstrates that anthranilic acid as fluorescent label in combination with reversed-phase liquid chromatography-mass spectrometry is an advantageous method for identification and quantification of neutral and acidic N-glycans. Our results show that mass spectrometry-based quantification correlates with quantification by fluorescence. Chromatographic discrimination between several structural glycan isomers was achieved. The sharp peaks of the eluting anthranilic acid-labeled N-glycans enabled on-line mass spectrometric analysis of even low-abundance glycan species. The method is broadly applicable to N-glycan analysis and is an orthogonal analytical method to the widely established hydrophilic-interaction liquid chromatography of 2-aminobenzamide-labeled N-glycans for characterization of N-glycans derived from biopharmaceuticals.
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