Gene transfer by electroporation, lipofection, and DEAE-dextran transfection: compatibility with cell-sorting by flow cytometry

Cytometry. 1992;13(1):23-30. doi: 10.1002/cyto.990130106.

Abstract

The aim of this work was to define a transfection procedure that is compatible with the sorting and propagation of cells that transiently express a heterologous gene. Three requirements were established for the procedure and were met with COS monkey kidney cells that express a recombinant glutathione S-transferase (GST) gene. The transfection procedure used had to generate (i) populations in which at least 10% of the cells expressed recombinant GST, (ii) cellular morphological homogeneity throughout the population, and (iii) viable cells with at least a 5% colony-forming ability. Of the transfection techniques tested, only electroporation satisfied all three requirements. Usually 20-22% of the cells that survived electroporation expressed recombinant GST 3 days after electroporation as measured by flow cytometry, and 25% of the cells that survived electroporation formed colonies in cloning assays. Transfection with DEAE-dextran and chloroquine did enable 40% of the surviving cells to express GST, but only 0.01% of the cells that survived transfection formed colonies in cloning assays. Finally, with lipofection, only 1% of the surviving cells expressed recombinant GST, although 25-40% of the cells that survived transfection formed colonies. These studies define the merits and limitations of transfection techniques relative to the analysis and sorting of transfected cells by flow cytometry.

Publication types

  • Comparative Study
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Cell Line
  • Cell Separation
  • DEAE-Dextran / pharmacology
  • Electricity
  • Flow Cytometry*
  • Genetic Techniques*
  • Phosphatidylethanolamines
  • Transfection*

Substances

  • Phosphatidylethanolamines
  • 1,2-dielaidoylphosphatidylethanolamine
  • DEAE-Dextran