Anyone else have toxicity issues with Lipofectamine 2000? - (Nov/23/2007 )
I am working with SiHa cells (cervical cancer line) and Lipofectamine 2000. I have tried optimising the transfections but still end up killing around 50% of my cells. Everyone I ask says they never see such cell death with Lipofectamine and their cells, so why are SiHas so sensitive?
I think I may need to switch to something else, anyone have any experience with DEAE-dextran, CaPO4 or PEI?? Or any other suggestions?
Thanks a lot!
I have not worked with this cell line, but I think Lipo2K will top the others on the list. Fugene6 claims to be less toxic, but on my hands, it is slightly weaker agent than the lipo2k in several cell types.
Toxicity is dosage dependent. If you had good transfection efficiency, but also high toxicity, proportionally reduce both components will cut the toxicity. In some cases, that accompanies with reduced transfection efficiency, sometimes it does not. Other thing is to incease the cell density to 85-95% confluncy as suggested by the manufacturer. You may also include 2.5 % FBS to the transfection complexes, once DNA and lipofectamine is mixed and incubated. good luck.
I also have never worked with this cell line. Lipo2000 usually works Ok for me, not much cell death. However, of the last 4-5 years or so, I mostly use Effectene from QIAGEN, high efficiency, low toxicity, need less DNA, no need to change medium. But Lipo2000 IS somewhat better than Effectene with some cell lines (just a few, but still), e.g. NRK. Transfectene from BioRad is another good one. Transfection efficiency depends a lot on cell type, though, so you may want to experiment and optimize. The Enhancer solution (which is included with Effectene) is really good, I sometimes use it with Lipo2000, or with a mixture of Lipo2000-Effectene and get good results.
Higher cell density or simple more cells can help to avoid cell toxicity of lipofectamine.
Thanks everyone! I think my cell density may have been too low - will try to optimise this first with Lipo2000 and if I still have problems I will try the other reagents suggested here
when using lipofectamine your cell density MUST be 90-95%
how did you optimize?
why don't give a try with half the concentration manufacturer has recommended (use half the volume of medium and lipofectamine, dna vol does not matter)