D.J. Lowrie Jr.
Department of Cell Biology, Neurobiology, and Anatomy
University of Cincinnati College of Medicine, Cincinnati, OH 45267
Transfection of mammalian cell lines is enhanced by the use of liposome-based reagents. Here we compare the transfection efficiency of green fluorescent protein into human adrenalcarcinoma cells using both LipoTAXI transfection reagent and LipofectAMINE reagent.
Our lab has recently been interested in the transient transfection of green fluorescent proteins into mammalian cells. We have been using the pEGFP-C3 and pEGFP-N3 vectors, which contain sequences encoding the enhanced version of green fluorescent protein (EGFP) whose expression is driven by the cytomegalovirus (CMV) promoter. Convenient restriction sites in these vectors allow fusion of a protein of interest to either the amino- or carboxy-terminus of EGFP.
The cell line used for many of our experiments is the human adrenalcarcinoma
cell line, SW13, which contains vimentin intermediate filaments. A subclone
derived from this cell line lacks vimentin expression and is thought to contain
no intermediate filament network.1 Although we have previously had
success using this cell line for transfection studies using calcium phosphate,2,3
two recent observations have led us to look for new methods that would improve
our efficiency. First, we have found that some of our constructs have produced
lower transfection efficiencies than others, making it difficult to obtain a
suitable number of transfected cells to draw conclusions. Second, we have
noticed heterogeneous subcellular localization of some of our fusion proteins in
the same set of transfected cells. Presumably, this is due to varying l