Efficient large volume lentiviral vector production using flow electroporation.
Efficient large volume lentiviral vector production using flow electroporation.
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DOI：
10.1089/hum.2011.088
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发表时间：
2012-02
影响因子：
4.2
通讯作者：
S. Witting;Linhong Li;A. Jasti;C. Allen;K. Cornetta;J. Brady;R. Shivakumar;M. Peshwa
中科院分区：
文献类型：
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作者：
S. Witting;Linhong Li;A. Jasti;C. Allen;K. Cornetta;J. Brady;R. Shivakumar;M. Peshwa
慢病毒载体开始成为人类基因治疗的可行选择。在这里，我们描述了一种方法，它结合了悬浮细胞系的便利性与可扩展的，非化学基础的，符合GMP的转染技术称为流动电穿孔（EP）。优化无血清适应HEK 293 FT细胞的Flow EP参数，以限制毒性并最大化滴度。使用第三代基于艾滋病毒的慢病毒载体系统（用水泡性口炎糖蛋白包膜假型化），小体积和大体积转染均产生超过1×10（8）感染单位/mL的滴度。因此，实施大规模临床慢病毒生产的一个很好的选择是悬浮细胞系的流EP。
Lentiviral vectors are beginning to emerge as a viable choice for human gene therapy. Here, we describe a method that combines the convenience of a suspension cell line with a scalable, nonchemically based, and GMP-compliant transfection technique known as flow electroporation (EP). Flow EP parameters for serum-free adapted HEK293FT cells were optimized to limit toxicity and maximize titers. Using a third generation, HIV-based, lentiviral vector system pseudotyped with the vesicular stomatitis glycoprotein envelope, both small- and large-volume transfections produced titers over 1×10(8) infectious units/mL. Therefore, an excellent option for implementing large-scale, clinical lentiviral productions is flow EP of suspension cell lines.