Optimized galenics improve in vitro gene transfer with cationic molecules up to 1000-fold

Gene Ther. 1996 Dec;3(12):1074-80.


Reproducible and optimized complex formation between polyanionic DNA and a polycationic vector is a key aspect of nonviral gene transfer systems. To this end, several factors relevant to in vivo delivery have been tested repeatedly on several cell types. Gene transfer with a lipopolyamine (transfectam) in the presence of serum was increased over 10-fold by sequential addition of the lipid to DNA. Paradoxically, high complex concentrations (> 200 micrograms DNA/ml) led to large enhancements too, which points to the fact that formation of productive complexes is a slow process. Each parameter, more than compensates for the decreased efficiency generally observed with nonviral vectors in serum. Transfectam and PEI (polyethylenimine)-mediated transfection also improved after mild centrifugation of the complexes on to the cells. These individual factors were shown to be essentially multiplicative, leading altogether to approximately a 1000-fold transfection increase with a luciferase reporter gene. Finally, 25 cell lines and primary cells (including fibroblasts, hepatocytes and endothelial cells) were successfully transfected over a five orders-of-magnitude efficiency range, From this large set of data, a general relation between the overall transfection level (as measured by luciferase reporter gene expression) and the fraction of transfected cells (histochemically stained for beta-galactosidase) could be inferred. Finally, transfectam and PEI displayed similar trends over this large range of efficiencies, which reinforces the hypothesis of a common transfection mechanism where the key endosome-releasing stop occurs through a "proton sponge' effect.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blood
  • Cations
  • Cells, Cultured
  • Centrifugation
  • DNA
  • Gene Transfer Techniques*
  • Genetic Vectors
  • Glycine / analogs & derivatives*
  • Glycine / pharmacology
  • Humans
  • Mammals
  • Mice
  • Polyethyleneimine / pharmacology*
  • Spermine / analogs & derivatives*
  • Spermine / pharmacology
  • Transfection


  • Cations
  • dioctadecylamidoglycylspermine
  • Spermine
  • Polyethyleneimine
  • DNA
  • Glycine