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Rapid Antibody Glycoengineering in CHO Cells Via RNA Interference and CGE-LIF N-Glycomics

Part of the Methods in Molecular Biology book series (MIMB,volume 2370)

Abstract

The impact of the glycan distribution on the in vivo function and half-life of monoclonal antibodies has long motivated the genetic engineering of producer cells to achieve structures that enhance efficacy, safety and stability. To facilitate glycoengineering of IgG-producing Chinese hamster ovary cells, we present a rapid protocol that involves the use of RNA interference for the knockdown of genes of interest coupled with capillary gel electrophoresis and laser-induced fluorescence detection (CGE-LIF) for fast, high-throughput glycan analysis. We apply this methodology to the Fut8 gene, responsible for the addition of core fucose, which is a typical target for increasing antibody-dependent cellular cytotoxicity.

Key words

  • Fucosylation
  • Gene knockdown
  • RNAi
  • siRNA
  • Antibody-dependent cell cytotoxicity
  • Capillary gel electrophoresis
  • Chinese hamster ovary cells

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Acknowledgments

PK thanks the Department of Chemical Engineering, Imperial College London, for his scholarship. RD thanks the U.K. Biotechnology and Biological Sciences Research Council for his studentship. MM is funded by the U.K. Biotechnology and Biological Sciences Research Council (Grant reference: BB/S006206/1). IAG thanks the Irish Research Council (Scholarship No. GOIPG/2017/1049) and CONACyT (Scholarship No. 438330).

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Kotidis, P. et al. (2022). Rapid Antibody Glycoengineering in CHO Cells Via RNA Interference and CGE-LIF N-Glycomics. In: Davey, G.P. (eds) Glycosylation. Methods in Molecular Biology, vol 2370. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1685-7_7

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  • DOI: https://doi.org/10.1007/978-1-0716-1685-7_7

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