| A high-throughput method for quantification of glycoprotein sialylation. | |
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MedLine Citation:
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PMID: 20692221 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Sialic acid can improve qualities of therapeutic glycoproteins such as circulatory half-life, biological activity, and solubility. In production of therapeutic glycoproteins, a high-throughput method is required for process monitoring and optimization to ensure consistent and optimal sialic acid content. Current methods for quantifying sialic acid, however, require chromatographic separation that is time-consuming and cannot rapidly analyze many samples in parallel. Here we present a novel high-throughput method for quantifying glycoprotein sialylation. Using chemical reduction, enzymatic release of sialic acid, and chemical derivatization of the sialic acid, the method can accurately, rapidly (15 min), and specifically analyze many samples in parallel. It requires only 45 μl of sample and has a quantitation limit of 2 μM sialic acid. It has also been validated for monitoring sialylation of recombinant interferon gamma (IFN-γ) produced in Chinese hamster ovary (CHO) cell culture. This method is useful for various applications in upstream and downstream bioprocesses. |
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Authors:
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Lam Raga A Markely; Boon Tee Ong; Kong Meng Hoi; Gavin Teo; Michelle Y Lu; Daniel I C Wang |
Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2010-08-06 |
Journal Detail:
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Title: Analytical biochemistry Volume: 407 ISSN: 1096-0309 ISO Abbreviation: Anal. Biochem. Publication Date: 2010 Dec |
Date Detail:
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Created Date: 2010-09-22 Completed Date: 2011-01-18 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 0370535 Medline TA: Anal Biochem Country: United States |
Other Details:
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Languages: eng Pagination: 128-33 Citation Subset: IM |
Copyright Information:
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Copyright © 2010 Elsevier Inc. All rights reserved. |
Affiliation:
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Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Animals Cricetinae Cricetulus Glycoproteins / chemistry* Glycosylation Interferon-gamma, Recombinant / chemistry*, isolation & purification, metabolism N-Acetylneuraminic Acid / analysis* Neuraminidase / metabolism Nitriles / chemistry Oxidation-Reduction Spectrometry, Fluorescence / methods* |
| Chemical | |
Reg. No./Substance:
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0/Glycoproteins; 0/Interferon-gamma, Recombinant; 0/Nitriles; 109-77-3/dicyanmethane; 131-48-6/N-Acetylneuraminic Acid; EC 3.2.1.18/Neuraminidase |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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