2 replies to this topic

[calimero]

hi!!!!
I have a very big problem with my 293T cells in culture!!!!!
I use them for simple transfection by using CaCl reagent, so i need to change the medium 2h before the transfection, but after the "very gently" change with fresh medium, 293t strart to detach and to become rounded.
this is a problem appeared in the last 2mounths!!
I changed the fbs, the flask, I tested them for mycoplasm, I used a low passage batch of cells...what else??
please help me!!
thanks in advance!!

[SciCell]

calimero, on 09 December 2010 - 09:52 AM, said:

hi!!!!
I have a very big problem with my 293T cells in culture!!!!!
I use them for simple transfection by using CaCl reagent, so i need to change the medium 2h before the transfection, but after the "very gently" change with fresh medium, 293t strart to detach and to become rounded.
this is a problem appeared in the last 2mounths!!
I changed the fbs, the flask, I tested them for mycoplasm, I used a low passage batch of cells...what else??
please help me!!
thanks in advance!!

I have this problem with my 293 cells too. So I modify the protocol and seed less number of cells than recommended in the protocol. Some times I seed 2 or 3 6 well plates with different seeding density and pick the best one for my transfection. This is optimizing the protocol right?

[calimero]

THANK YOU FOR your answer...but I think that, in my case, number of cells is not my problem.I tried different concentration of cells with the same result!!
sigh!!!