Sustained High-Yield Production of Recombinant Proteins in Transiently Transfected COS-7 Cells Grown on Trimethylamine-Coated (Hillex) Microcarrier Beads

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• 作者：Randall N. Knibbs ; Michael Dame ; Melissa R. Allen ; Yunhong Ding ; William J. Hillegas ; James Varani ; and Lloyd M. Stoolman
• 刊名：Biotechnology Progress
• 出版年：2003
• 出版时间：February 2003
• 年：2003
• 卷：19
• 期：1
• 页码：9 - 13
• 全文大小：156K
• 年卷期：v.19,no.1(February 2003)
• ISSN：1520-6033

文摘

The present study shows that COS-7 cells transiently transfected and maintained onpositively charged (trimethylamine-coated) microcarrier beads synthesize recombinantprotein at higher levels and for longer periods of time than cells transfected andmaintained on polystyrene flasks in monolayer culture. Sustained, high-level synthesiswas observed with secreted chimeric proteins (murine E-selectin- and P-selectin-human IgM chimeras) and a secreted hematopoietic growth factor (granulocyte-macrophage colony-stimulating factor). Studies with green fluorescent protein indicatedthat the transfected cells attached more firmly to the trimethylamine-coated microcarriers than to polystyrene flasks. After 10-14 days in culture, most of the transfectedcells detached from the surface of the polystyrene flasks, whereas most transfectedcells remained attached to the microcarriers. The transiently transfected microcarriercultures produced higher levels of protein per transfected cell due to this prolongedattachment. The prolonged attachment and higher output of transfected cells onmicrocarriers resulted in a 5-fold increase in protein production from a singletransfection over two weeks. Thus, microcarrier-based transient transfection yieldsquantities of recombinant proteins with a significant savings of time and reagentsover monolayer culture.