Serum-Free Transfection of CHO Cells with Chemically Defined Transfection Systems to Generate Transient and Stable Cell Lines

摘要

In modern mammalian cell culture technology the demand for recombinant proteins in technological and clinical application gains increasing importance. Novel transfection and clonal selection methods allow the generation of stable or transient cell lines expressing recombinant proteins at high concentrations (Meissner et al., 2001). To generate such cell lines a broad variety of methods have been developed that facilitate the uptake and integration of DNA. Unfortunately, most of the systems generating stable expression systems are cost and time-consuming. While the request for recombinant proteins in fundamental and clinical application escalates (Muller et al., 2005), faster and more efficient transfection systems are required.Thus, we developed and improved a transfection system based on defined lipo-somes, so-called LUVs (=large unilamellar vesicles) consisting of DOTAP/DOPE or CHEMS/DOPE (Even-Chen and Barenholz, 2000). The cationic lipids are attractive due to their relatively easy preparation and extensive characterization (Martin et al., 2005). Secondly polyethylenimine (=PEI) a polycation was used for DNA transfer. PEI complexes the DNA and delivers it to the ER and/or nucleus via caveo-somes enabling to by-pass the degradation trough endocytosis. Both systems were used as chemical defined transfection systems in combination with a protein-free cultivated host cell line. Our studies focused on the improvement of our systems and the comparison to other available transfection systems.