Hello,
When doing retroviral transfection of 293T cells, according to the protocol there are two media; one has pH 7.1 and this one will be mixed with the plasmid DNA and the other medium has pH 7.9 (and this medium contains Fetal calve serum to added to 293T cells
I couldn't find actually why these numbers have to be exact ? and Why they are toxic and should be added for duration of less than 16 hours.!
Any explanation,
Thanks
Hi Alpha,
I'm not sure about the two media. I usually just use OPTI-MEM (reduced serum media) to mix with the plasmid DNA and transfection reagent (in my case, Fugene). Incubate the mix for 15 mins and pipet slowly to 293T cells which were seeded a day before in DMEM medium. This is what we usually do in our lab and it worked for my experiments.
Do you use any transfection reagent or you do calcium phosphate transfection?
Regards,
Carla
why not simply using any chemical transfectant? Most of them are very easy to use, their protocols do not require different pH or media, and 293T cells are not problematic at all.
Good luck! Sandra
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