293 transfection

[Immuno AG]

ggggggggIn article <319A1FD3.BF5 at nrc.uab.edu>, rdudley at nrc.uab.edu says...
>
>I am transfecting 293 cells (human epithelial kidney) with Lipofectaime 
>fron Gibco-BRL.  The first time worked splendidly, but subsequent 
>transfections (we're using transients) have yielded mediocre results.  
>There are several superstitions I have heard, one of which is something 
>to the effect of, if the primary culture exceeds approx. 70% 
>confluency, subsequent generations of cells will be affected negatively 
>in terms of transfection.  The solution is to thaw an aliquot of early 
>generations and start again.  I have tried several other solutions--I 
>was wondering if anyone could provide some insight into this one.  My 
>primary did reach approx. 90% confluency on one ocassion.  I have many 
>early generations frozen away, so if this is true, there's not much of 
>a problem.  Thanks to anyone with info, and I'll post what I find out 
>later.
>
>rich
>
>--- --- ---
>Richard J. Dudley
>Neurobiology Research Center
>University of Alabama School of Medicine