293T transfection problem

[Arastho]

Hi everyone!

I think that I've some problems in my transfections:

I plated:
1 - Cells+Plasmid+Transfection
2 - Cells+Transfection
3 - Cells

From this I realized that the cells from 1 and 2 are very well attached do the plate and they were very difficult to lyse. Anyone has an idea why this is happennig?

I already change the HBS solution!
Thanks

[xangwang]

Hi,
I didn't met that problem.. on the contrary when cells are very sensitive to transfection, they become easier to detach.
how long do you keep them cultivated ?
x.

[Arastho]

After transfection, 48h.
But I'm afraid that I did something wrong: 24h after transfection the protocol says: change medium and wait more 24h before recover your cells. And I didn't change the medium. I wait 48h...

It could be the reason

[xangwang]

Hi,

changing the medium is a good thing when you transfect sensible cells.
For 293T, it doesn't seem necessary to perform a medium change. once again if cells were not healthy they would have been detached and not firmly attached to the dish.
I really don't see what the problem could be,
does anyone has already experienced that kind of matter ?

x.

[eran.hadas]

We transfect 293T cells routinely, using PEI. We never replace medium after transfection and have no problem.
293T cells usually attach poorly and are very easy to get off the plate. Our problem is that we need to be careful not to add the transfection mixture too fast so not to detach the cells.

[Gustaveimj]

ÿþ[

ÿþ[

ÿþ[