Fwd: [Cell-biology] Re: Lung fibroblast methotrexate toxicity

Sonal Patel via cellbiol%40net.bio.net (by stylosweetu from gmail.com)
Sat Jul 14 15:06:23 EST 2012


Hi

I am using MTX as selective agent. Alongwith colonies I also get lot of
background in KO only is there any way I can reduce it. I have already
tried different concentrations of MTX it doesnt help.

Thanks,
Sonal.


On Thu, Jul 12, 2012 at 4:53 PM, PiLS <pils from invalid.ca> wrote:

> Sonal Patel <stylosweetu from gmail.com> à ploppé:
> > I am trying to find out effect of methotrexate on lung fibroblast WT
> cells
> > and KO where gene of my interest is knocked out . After 3 weeks on
> staining
> > with crystal violet I get lot of background cells in KO alongwith
> colonies.
> > Can anyone suggest me how to remove the background cells.
>
> Hello,
>
>

> can you provide more info about what you are trying to achieve? Are you
> using the MTX as a selection agent or are you trying to study its
> effects on KO cells?
>
> In general if you have incomplete selection (assuming that's your
> problem) reducing the concentration of serum and/or increasing the
> concentration of the selection agent gives good results.
>
> --
> PiLS
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