Evaluation of desialylation during 2-amino benzamide labeling of asparagine-linked oligosaccharides |
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Authors: | Udayananth Aich Deanna C. Hurum Lipika Basumallick Srinivasa Rao Chris Pohl Jeffrey S. Rohrer Sebastian Kandzia |
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Affiliation: | 1. Thermo Fisher Scientific, Lakeside Drive, Sunnyvale, CA 94085, USA;2. Thermo Fisher Scientific, Oakmead Parkway, Sunnyvale, CA 94085, USA;3. GlycoThera, 30625 Hannover, Germany |
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Abstract: | Labeling of released asparagine-linked (N-linked) oligosaccharides from glycoproteins is commonly performed to aid in the separation and detection of the oligosaccharide. Of the many available oligosaccharide labels, 2-amino benzamide (2-AB) is a popular choice for providing a fluorescent product. The derivatization conditions can potentially lead to oligosaccharide desialylation. This work evaluated the extent of sialic acid loss during 2-AB labeling of N-linked oligosaccharides released from bovine fetuin, polyclonal human serum immunoglobulin G (IgG), and human α1-acid glycoprotein (AGP) as well as of sialylated oligosaccharide reference standards and found that for more highly sialylated oligosaccharides the loss is greater than the <2% value commonly cited. Manufacturers of glycoprotein biotherapeutics need to produce products with a consistent state of sialylation and, therefore, require an accurate assessment of glycoprotein sialylation. |
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Keywords: | N-linked glycans Oligosaccharides Glycoprotein Sialic acids 2-Amino benzamide HPAE&ndash PAD |
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