Effectene™ Transfection Reagent From Qiagen

Transfection reagents and protocols have become very widespread and utilized in many aspects of DNA research. There are many applications that require the introduction of vectors and expression systems into various cell lines. There are also many primary cell lines that have been notoriously tricky to transfect, with reagents often causing cytotoxicity. The Effectene™ Transfection Reagent is specialized and has been optimized to provide highly efficient stable and transient transfection of primary and cultured cell lines. It works well for adherent and suspension cell cultures. The Qiagen website also provides excellent technical assistance with a cell line database and protocols from customers that can be found at www.qiagen.com/transfectiontools/.

The Effectene™ Transfection Reagent is a non-liposomal, lipid-based reagent. The reagent comes with a DNA-condensing enhancer and buffer. The ratio of the Effectene™ to the DNA-enhancer complex can be varied and optimized to obtain the highest transfection efficiencies. The enhancer is first added to the DNA in a brief 5-minute incubation step to allow the DNA molecules to condense. The Effectene™ Reagent is then added, providing a cationic lipid coating of the DNA. This Effectene™-DNA complex forms in just 10 minutes. All of this can be done very conveniently at room temperature. While the complex is being allowed to form, the cells are washed and fresh growth medium is added. The Effectene™-DNA complex is then added to a small amount of growth medium in which the cells are currently being cultured (very convenient as it requires no Opti-Mem or special serum-free media and antibiotics and other growth factors can continue to be used). The media is then added to the cells and incubated for 24-48 hrs. Stable transfections can then be passaged at a 1:5 or 1:10 ratio into the appropriate selection medium.

I have used Effectene™ many times for stable and transient transfection of primary RGC and hippocampal cells as well as 293 cell lines. I have had great success with all of my transfections, much greater than with any other transfection reagent. Only a small amount of DNA is required (1.0 ug for a 60 mm dish compared to the 5-15 ug needed for other liposome or calcium-phosphate based transfections). This is very useful when using expensive vectors and reagents. The cells also don’t need to be at full confluency and I have never had them die as a result of the transfection. The Qiagen handbook has a variety of different optimization tips and protocols for various sizes of dishes and for various ratios of Effectene™-DNA complexes. I have optimized the protocol somewhat for our primary cell lines to account for the low cell number, the size of dishes and the sensitivity of the cells. The protocol suggests that the media does not need to be removed or replaced but I usually wash the cells with PBS and replace with growth and maintenance media the next day (often a 1:1 mixture of the previously collected conditioned media and fresh growth media). This protocol is so fast and easy that it can easily be done in less than 30 minutes.

I always recommend Effectene™ to others for cell lines and transfections that have previously given them problems and always get positive feedback. This is a great product and almost guaranteed to work.