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Molecular Biotechnology

, Volume 40, Issue 2, pp 136–143 | Cite as

The Kinetics of Polyethylenimine-Mediated Transfection in Suspension Cultures of Chinese Hamster Ovary Cells

  • Martin Bertschinger
  • Arnaud Schertenleib
  • Jean Cevey
  • David L. Hacker
  • Florian M. Wurm
Research

Abstract

The kinetics of polyethylenimine (PEI)-mediated gene transfer at early times after transfection of Chinese hamster ovary (CHO) cell in suspension were investigated using a novel in vitro assay. Addition of an excess of competitor DNA to the culture medium at various times after the initiation of transfection inhibited further cellular uptake of PEI–DNA particles. Using this approach, a constant rate of particle uptake was observed during the first 60 min of transfection at a PEI:DNA ratio of 2:1 (w/w) and a cell density of 2 × 106 cells/ml under serum-free conditions. The uptake rate declined considerably during the next 2 h of transfection. Both the rate and the level of PEI–DNA uptake in serum-free minimal medium were found to be dependent on the PEI–DNA ratio, the cell density at the time of transfection, and the extent of particle aggregation. These studies of the early phase of PEI-mediated transfection are expected to lead to further opportunities for optimization of gene transfer to suspension cultures of mammalian cells for the purpose of large-scale transient recombinant protein production.

Keywords

Transfection Polyethylenimine Plasmid DNA Mammalian cells 

Notes

Acknowledgments

We thank Dr. Lucia Baldi for critically reading the manuscript. Financial support was provided by the Swiss Innovation Promotion Agency (KTI/CTI).

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Copyright information

© Humana Press 2008

Authors and Affiliations

  • Martin Bertschinger
    • 1
    • 2
  • Arnaud Schertenleib
    • 1
  • Jean Cevey
    • 1
  • David L. Hacker
    • 1
  • Florian M. Wurm
    • 1
  1. 1.Laboratory of Cellular BiotechnologyInstitute of Bioengineering, École Polytechnique Fédérale de Lausanne (EPFL)LausanneSwitzerland
  2. 2.Glenmark Pharmaceuticals SALa Chaux-de-FondsSwitzerland

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