We are using Hela cells for checking localization of proteins by Immunofluorescence and transfecting these cells with PEI, but efficiency is just 30 %. How to improve it? Should we use another lipofectamine?
Try a HeLa-specific transfection reagent. Lipofectamine is great, but it's a generic that can have trouble sometimes with picky cells. This reagent (https://altogen.com/product/hela-transfection-reagent-cervix-adenocarcinoma-cells/) is tailored to HeLa, and will probably boost your results as well. You'll need complex condensers if you have larger proteins to be transfected, but otherwise the cell markers should be enough to make sure most cells get transfected.
We have used both calcium phosphate and JetPrime for HeLa cells and get very good results. We tend to go with JetPrime more often. Here is the link http://www.polyplus-transfection.com/transfection-reagents/1_gene-expression/1_broad_spectrum/jetprime%C2%AE/
we use human low-density lipoprotein. J Lipid Res. 2010 Jul;51(7):1704-18. Epub 2010 Feb 19
We have used FuGENE HD from Roche to transfect HeLa cells with 75-80% efficiency. See attached procedure.
Hi... we have used used both FuGENE HD and Turbofectin 8.0 from ORIGene to transfect plasmids into HeLa cell lines, with a transfection efficiency of around 70%
Hi You can try using Turbofect or lipofectamine 2000. the efficiency in this is great. Also these transfections can be done with serum containing medium.
Use Opti MEM for transfection.
Effecten also works great with HeLa, similar protocol to lipofectamine
turbo fect works fine, 6well plate, 2,5ug DNA in 0,5ml optimem add 4ul TF 20-30min RT add to the cells (usually plate teh day before at 3-4. 105 cell/well in DMEM WO antibio +FBS
I tried three different reagents: Fugene (Promega), Fugene (Roche) and Jetprime (Polyplus). The last one was the best to transfect DNA into Hela cells.
Dear Disha,
Recently we have launched a transfection reagent dedicated to HeLa cells. HeLaFect presents a high compaction level of nucleic acid associated with low toxicity (biodegradable). More information are avaliable in the link below.
I would be very glad to send you a sample if you would like to try it; you can contact me via ResearchGate or directly at [email protected]
good luck with your experiment,
Cedric
http://www.ozbiosciences.com/transfection-cell-specific/77-hela-cells-transfection-helafect-reagent.html
3.10e5cells /well/6w plates/ in DMEM W/O Atbio the day before transfection
0,5ml optimem + 2,5ug DNA+4ul turbofect ..30min RT
add this to 1,5ml complete DMEM...2h at 37° , remove the media....around 60-70% efficiency
Lipofectamine 2000 is a must and works well in general, but it has some degree of toxicity. Lipofectamine 3000 is much better from this point of view, even if less efficient in our experience (mainly related to connexins). Recently we found Nanovex Transfectosome to be a very good transfectant with no toxicity side effects. If you decide to use it, ask for the N version + enhancer.
I have used both Fugene and JetPrime as transfection reagents in HeLa cells. While JetPrime is highly efficient, it tends to be very cytotoxic so you need to change the media around 4 hours or less after transfection. I also saw a high variation of transfection efficiency within samples transfected with the same plasmid. It would usually depend on what experiments you are doing but since I've been doing some comparison among different conditions in my assay, it's better to get consistent transfection efficiency. Which is why I'm sticking with Fugene for now.
Try a HeLa-specific transfection reagent. Lipofectamine is great, but it's a generic that can have trouble sometimes with picky cells. This reagent (https://altogen.com/product/hela-transfection-reagent-cervix-adenocarcinoma-cells/) is tailored to HeLa, and will probably boost your results as well. You'll need complex condensers if you have larger proteins to be transfected, but otherwise the cell markers should be enough to make sure most cells get transfected.
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