Cegovnik Urska, Novaković Srdjan
Department of Tumor Biology, Institute of Oncology, Zaloska 2, 1000 Ljubljana, Slovenia.
Bioelectrochemistry. 2004 Apr;62(1):73-82. doi: 10.1016/j.bioelechem.2003.10.009.
To achieve the maximal introduction of plasmid DNA into cells and, at the same time, to prevent undesirable cell deaths, electrotransfection conditions should be determined for every single cell type individually. In the present study, we determined the optimal electrotransfection parameters for in vitro transfection of B16F1, SA1, LPB, SCK, L929 and CHO cells. Some of these varying parameters were electric field strength, number of applied pulses and their duration, osmolarity of electroporation buffer, plasmid DNA concentration and temperature at which the electroporation was carried out. The maximal transfection rates at optimal electrotransfection parameters in B16F1, SA1, LPB, SCK, L929 and CHO were 85%, 40%, 60%, 1%, 40% and 65%, respectively. The obtained results confirmed that the electroporation is a useful procedure for an in vitro transfection of the majority of mammalian cells. The method, if optimized, may generate reproducibly high proportion of transfected cells among the cell types that are sensitive to electric field action. Thus, the determined parameters could serve for the subsequent implementations of this method.
为了使质粒DNA最大限度地导入细胞，同时防止不必要的细胞死亡，应针对每种细胞类型单独确定电穿孔条件。在本研究中，我们确定了B16F1、SA1、LPB、SCK、L929和CHO细胞体外转染的最佳电穿孔参数。这些不同的参数包括电场强度、施加脉冲的数量及其持续时间、电穿孔缓冲液的渗透压、质粒DNA浓度以及进行电穿孔时的温度。在最佳电穿孔参数下，B16F1、SA1、LPB、SCK、L929和CHO细胞的最大转染率分别为85%、40%、60%、1%、40%和65%。所得结果证实，电穿孔是大多数哺乳动物细胞体外转染的一种有用方法。如果该方法得到优化，在对电场作用敏感的细胞类型中，可能会可重复地产生高比例的转染细胞。因此，所确定的参数可用于该方法的后续实施。
