Rapid Antibody Glycoengineering in CHO Cells Via RNA Interference and CGE-LIF N-Glycomics.
Rapid Antibody Glycoengineering in CHO Cells Via RNA Interference and CGE-LIF N-Glycomics.
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DOI：
10.1007/978-1-0716-1685-7_7
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发表时间：
2021-10
影响因子：
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通讯作者：
Pavlos Kotidis;Masue M Marbiah;Roberto Donini;Itzcóatl A. Gómez;Ioscani Jimenez Del Val;S. Haslam;K. Polizzi;C. Kontoravdi
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文献类型：
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作者：
Pavlos Kotidis;Masue M Marbiah;Roberto Donini;Itzcóatl A. Gómez;Ioscani Jimenez Del Val;S. Haslam;K. Polizzi;C. Kontoravdi
聚糖分布对单克隆抗体的体内功能和半衰期的影响长期以来一直促使生产细胞的基因工程以实现增强功效、安全性和稳定性的结构。为了促进糖工程的IgG生产的中国仓鼠卵巢细胞，我们提出了一个快速的协议，涉及使用RNA干扰敲低感兴趣的基因，再加上毛细管凝胶电泳和激光诱导荧光检测（CGE-LIF）的快速，高通量聚糖分析。我们将这种方法应用于Fut 8基因，该基因负责添加核心岩藻糖，这是增加抗体依赖性细胞毒性的典型靶标。
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.