The kinetics of polyethylenimine-mediated transfection in suspension cultures of Chinese hamster ovary cells.
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 x 10(6) 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.
Laboratory of Cellular Biotechnology, Institute of Bioengineering, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland.
Martin
M
Bertschinger
Arnaud
A
Schertenleib
Jean
J
Cevey
David L
DL
Hacker
Florian M
FM
Wurm
D000818:Animals; D016466:CHO Cells; D018929:Cell Culture Techniques; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA; D007700:Kinetics; D010957:Plasmids; D011094:Polyethyleneimine; D014162:Transfection / Q000379:methods*
D011094:Polyethyleneimine; D004247:DNA