western blot help!!!

James Bassuk bassuk at u.washington.edu
Mon Jun 2 01:19:06 EST 2003


On Fri, 9 May 2003, Péter Fedorcsák wrote:

> Hi,
>
> re-wet your membrane in methanol (it won't wet in anything else), and
> wash in changes of your buffer.
> Regards,
> Peter
>
>
> In article <20030419165956.10325.qmail at ww02.hostica.com>,
>  karthij96 at yahoo.com wrote:
>
> > i have just started doing westerns recently.  i am facing problems with the
> > PVDF membrane that
> >
> > we use.  i soak it in methanol  & before it dries i soak it in ddwater & then
> > in the western transfer
> >
> > buffer.
> >
> > The transfer looks good--as i am able to see all the lanes & even the marker
> > .  The problem is
> >
> > that after i block the membrane in 5% milk ( carnation non-fat dry milk
> > powder &  made in 1X
> >
> > TBS) for an hour or even overnite in a cold room---the membrane refuses to
> > wet---it looks as if
> >
> > its dry, only the egdes seem to be wet.
> >
> >  Anyone with a solution---will be greatly appreciated.
> >
> > thank you
> >
> > aditi
> >
> > http://biowww.net/mynews/tree.php?group_name=bionet_molbio_proteins&begin=0
>

try a different brand of PDVF --- this is bizarre.

Jim




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