13 replies to this topic

[caro]

Hi there

Has anyone tested different transfection reagents for transfection siRNA, like X-tremeGENE or RNAiFect?? Is there a big difference compared to normal transfection reagents eg FuGene??
I am goin to transfect my epiphelial cells in january and would like to test only the best reagents to save money and time.

Regards Caro

[pcrman]

Hi Caro,

Yes, before you start your RNAi experiments, find a good transfection reagent and stick to it. I have used such reagent from ambion and invitrogen and have not compared side by side the effect. But the protocol for Invitrogen's lipofectamin 2000 seems much simpler and that is why I later have only used it for all my transfections.  

Tom tuschl has some good comments on different transfection reagents:
What about other transfection reagents? There are amazing differences in the efficiency of the cationic liposome reagents available on the market. Old, classical reagents are normally bad and only kept in the program of manufacturers to satisfy "conservative" customers, e.g. Lipofectamine 2000 is about 10 times more efficient than the still distributed Lipofectamine. We are just testing different transfection reagents.

To read more, click here http://www.rockefell...schl/sirna.html

Edited by pcrman, 16 December 2004 - 11:39 AM.

[lydia]

Open Biosystems has a transfection reagent called Arrestin that is comparable to Lipofectamine but cheaper.  I have not tested it but have heard good efficiencies from others in the lab.

[postdoc2130]

We and others have good experiences on Epoch Biolabs' GenCarrier-1/2 reagents. They are basically similar to Lipofectamine2000 (efficiency and protocol) but much cheaper. In some primary cells, it even outperforms Lipofectamine2000.

[badcell]

I've just tested two reagents from Ambion (siPORT Lipid and SiPORT Amine) side by side with Lipofectamine 2000. In my cells (a mouse beta-cell line) I got high levels of transfection using siPORT Amine (>80% cells transfected), but the results with the two lipidic reagents were dismal (approx 25% cells transfected). Other people in my lab gets 90% transfection levels with lipofectamine 2000 in fibroblasts cell lines. It all depends on the cell line. As pcrman said, best thing to do is test several reagents at the beginning of your experiment and stick with the best.

[expresson_help]

since you're using epithelial cells I'd recommend NeoPhectin which we sell, that is very good for difficult to transfect cell types such as endothelial and primary cells. Find more information at http://www.expresson...neophectin.html

David
ExpressOn

Edited by expresson_help, 18 January 2005 - 05:10 AM.

[fred_33]

hi
you may look at

http://cpmcnet.colum...orts2001/c3.htm
http://pen2.igc.gulb..._monolayers.pdf

option:
http://research.dfci.....ent cells.pdf

hope that helps

[miRNA_diabetes]

I have tried transfecting hepatocytes with labelled miRNA using Metafectene-pro as the transfection agent. It worked great for me with >90% transfection efficiency!!
I hope to expect the same with unlabelled miRNA.  

[Julie123]

Hello Pcrman,

I am using lipofectamine2000 to cotransfect siRNA and plasmid in HeLa?have you tried this cell line before?It doesn't look that i have good knockdown.What advice you can give me?
thx

[Julie123]

pcrman, on Dec 16 2004, 11:38 AM, said:

Hi Caro,

Yes, before you start your RNAi experiments, find a good transfection reagent and stick to it. I have used such reagent from ambion and invitrogen and have not compared side by side the effect. But the protocol for Invitrogen's lipofectamin 2000 seems much simpler and that is why I later have only used it for all my transfections.  

Tom tuschl has some good comments on different transfection reagents:
What about other transfection reagents? There are amazing differences in the efficiency of the cationic liposome reagents available on the market. Old, classical reagents are normally bad and only kept in the program of manufacturers to satisfy "conservative" customers, e.g. Lipofectamine 2000 is about 10 times more efficient than the still distributed Lipofectamine. We are just testing different transfection reagents.

To read more, click here http://www.rockefell...schl/sirna.html

Hello pcrman,

Which cells you transfected with lipofectamine 2000?I tried to cotransfect siRNA and plasmid but it is not working:(I am using HeLa cells.What do you advise me?

[ribonerd]

Ambions transfection reagents are not good for most "challenging" cell lines. I love their siRNAs, but for transfection I use RNAiMax (Invitrogen) or Lipofectamine 2000- for pDNA delivery

[krenny7]

for the transfection of siRNA and mRNA biontex transfection reagent works great. I tried their test samples for free then I am using the products. check this link https://www.biontex-...p?id=2&subID=18

[jcroz]

Does anyone have suggestions for a good transfection reagent for primary cells? I'm trying to transfect primary monocytes.

[EEO]

RNAiMax works well for most cell lines!