Fibroblasts won't restart after freeze

[k.harvey at erasmusmc.nl]

Hi 

We grew primary stromal cell lines (which basically have a fibroblast
apperance), we found using 20% CM (filtered) worked well and also
gelatin coating the plate worked well for us. We also observed a crisis
period and found just plating them in progressively smaller plates until
they restarted to expand works.

Hope this helps and good luck
Kirsty

"D.K." wrote:
> 
> In article <c09b237b.0305071848.6bea6062 at posting.google.com>, ladasky at my-deja.com (John Ladasky) wrote:
> >Hi, folks,
> >
> >I am growing six primary human fibroblast lines.  I have vials of all
> >of them in liquid nitrogen.  When I thaw these vials, four of the six
> >will eventually start to grow.  The other two have viable cells but
> >remain stubbornly quiescent.  I can pass them and they will re-adhere,
> >but they have no interest in dividing.  They were frozen down at the
> >earliest possible passage number, below passage ten in each case.  Two
> >of my six lines that regrow nicely were actually frozen down above
> >passage fifteen.
> >
> >Does anyone have any advice for coaxing a few cell divisions out of
> >these lazy cells?  Conditioned medium, perhaps?  More FBS?  Human
> >serum?  FGF?
> 
> Sounds tough. I think your best chance would be conditioned medium
> from those primary lines that do grow.
> 
> Just out of curiosity - when they adhere, do they spread nicely
> or just sit there?
> 
> DK
>