DEAE-Dextran Transfection of Lymphocyte Cell Lines
Adapted from Transcriptional Regulation in Eukaryotes: Concepts, Strategies, and Techniques, 2nd edition, by Michael F. Carey, Craig L. Peterson, and Stephen T. Smale. CSHL Press, Cold Spring Harbor, NY, USA, 2008.INTRODUCTION
In this procedure, negatively charged DNA binds to the positively charged diethylaminoethanol (DEAE) moieties of a DEAE-dextran polymer. The soluble complex is added to the cells, which take it up by nonspecific endocytosis. The method described here uses an initial incubation with a DNA/DEAE-dextran mixture, followed by a second incubation in growth medium supplemented with chloroquine. Chloroquine is thought to enhance transfection efficiencies by neutralizing lysosomal hydrolases, which can degrade the DNA. This protocol describes transfection of lymphocyte cell lines, but it can be used for transient transfection of both adherent and nonadherent cells. However, it rarely is used for stable transfection experiments because toxicity prevents long-term maintenance of the transfected cells. One notable advantage of the DEAE-dextran method is that a relatively small amount of DNA is needed for each experiment.
