Delivery of episomal vectors into primary cells by means of commercial transfection reagents

Biochem Biophys Res Commun. 2015 May 29;461(2):348-53. doi: 10.1016/j.bbrc.2015.04.037. Epub 2015 Apr 15.

Abstract

Although episomal vectors are commonly transported into cells by electroporation, a number of electroporation-derived problems have led to the search for alternative transfection protocols, such as the use of transfection reagents, which are inexpensive and easy to handle. Polyplex-mediated transport of episomal vectors into the cytoplasm has been conducted successfully in immortalized cell lines, but no report exists of successful transfection of primary cells using this method. Accordingly, we sought to optimize the conditions for polyplex-mediated transfection for effective delivery of episomal vectors into the cytoplasm of primary mouse embryonic fibroblasts. Episomal vectors were complexed with the commercially available transfection reagents Lipofectamine 2000, FuGEND HD and jetPEI. The ratio of transfection reagent to episomal vectors was varied, and the subsequent transfection efficiency and cytotoxicity of the complexes were analyzed using flow cytometry and trypan blue exclusion assay, respectively. No cytotoxicity and the highest transfection yield were observed when the ratio of transfection reagent to episomal vector was 4 (v/wt) in the cases of Lipofectamine 2000 and FuGENE HD, and 2 in the case of jetPEI. Of the three transfection reagents tested, jetPEI showed the highest transfection efficiency without any cytotoxicity. Thus, we confirmed that the transfection reagent jetPEI could be used to effectively deliver episomal vectors into primary cells without electroporation.

Keywords: Delivery; Electroporation; Episomal vector; Primary cells; Transfection reagent.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Cell Survival / drug effects
  • Cells, Cultured
  • Electroporation / methods
  • Genetic Vectors / administration & dosage*
  • Genetic Vectors / chemistry
  • Genetic Vectors / genetics
  • Green Fluorescent Proteins / genetics
  • Lipids / chemistry
  • Lipids / toxicity
  • Mice
  • Plasmids / administration & dosage*
  • Plasmids / chemistry
  • Plasmids / genetics
  • Transfection / methods*

Substances

  • FuGene
  • Lipids
  • Lipofectamine
  • enhanced green fluorescent protein
  • Green Fluorescent Proteins