Setting optimal parameters for in vitro electrotransfection of B16F1, SA1, LPB, SCK, L929 and CHO cells using predefined exponentially decaying electric pulses.
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.
Department of Tumor Biology, Institute of Oncology, Zaloska 2, 1000 Ljubljana, Slovenia.
Urska
U
Cegovnik
Srdjan
S
Novaković
D000818:Animals; D002021:Buffers; D016466:CHO Cells; D045744:Cell Line, Tumor; D002470:Cell Survival; D006224:Cricetinae; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004560:Electricity; D004563:Electrochemistry; D018274:Electroporation; D066298:In Vitro Techniques; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D013696:Temperature; D013997:Time Factors; D014162:Transfection / Q000379:methods*; D014410:Tumor Stem Cell Assay; D001616:beta-Galactosidase / Q000378:metabolism
D002021:Buffers; D004247:DNA; D001616:beta-Galactosidase