期刊
ANALYTICAL BIOCHEMISTRY
卷 407, 期 1, 页码 128-133出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ab.2010.07.029
关键词
Carbohydrate; Sialic acid; Glycoprotein; Mammalian cell culture; High-throughput screening
资金
- MIT Portugal
- Singapore MIT Alliance
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 mu l of sample and has a quantitation limit of 2 mu M sialic acid. It has also been validated for monitoring sialylation of recombinant interferon gamma (IFN-gamma) produced in Chinese hamster ovary (CHO) cell culture. This method is useful for various applications in upstream and downstream bioprocesses. (C) 2010 Elsevier Inc. All rights reserved.
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