Gresch Oliver, Altrogge Ludger
Lonza Cologne GmbH, Cologne, Germany.
Methods Mol Biol. 2012;801:65-74. doi: 10.1007/978-1-61779-352-3_5.
Primary cells are a valuable tool for researchers and are often preferred over transformed or immortalized cell lines since they are biologically more relevant and resemble the in vivo situation much closer. Unfortunately, efficient gene transfer in primary cells is still limited. Whereas viral strategies are time consuming and involve safety risks, nonviral methods are often inefficient for most primary cells. Nucleofection has been proven to overcome these limitations. Here, we describe the Nucleofection protocol for efficient transfection of human umbilical vein endothelial cells. Using a combination of a cell type-specific solution and electrical conditions, transfection efficiencies up to 90% can be achieved while survival rate is more than 70%.
原代细胞对研究人员来说是一种有价值的工具，并且由于其在生物学上更具相关性且更接近体内情况，因此通常比转化细胞系或永生化细胞系更受青睐。不幸的是，原代细胞中的高效基因转移仍然有限。虽然病毒策略耗时且存在安全风险，但非病毒方法对大多数原代细胞来说往往效率低下。核转染已被证明可以克服这些限制。在此，我们描述了用于高效转染人脐静脉内皮细胞的核转染方案。通过结合细胞类型特异性溶液和电转条件，可实现高达90%的转染效率，同时存活率超过70%。
