Western Blotting Blues

Patrick bier13 at mail.com
Tue Dec 11 13:05:40 EST 2001


> "kalyan" <kalsriv at mailcity.com> wrote in message
> news:e7b6a13.0112100859.499dda77 at posting.google.com...
> > Hi All
> >  I have been doing the western Blots with the proteins from mammalin
> > cells. Interestingly, what I saw that after first western Blot, that I
> > did immidiately I got good bands. But after the storage of protein
> > extract for 7 to 8 days, I saw no bands at all, working with same
> > antibody and conditions. Predictably it is because of the degradation
> > of protein at the storage time. I kept it at -70 degree after the
> > solublisation with 5X sample buffer after the lysis and extraction of
> > the protein.
> >
> > Can someone suggest the best storage condition for the protein before
> > and after the solublisation with sample buffer.

I always store my proteins in the lysisbuffer (after i pulled the
extract through a needle to break down the DNA)in wich i also have
protease inhibitors. This mix i put in the - 80 (so I don't fast
freeze with Nitrogen)We never have the problems your revering to. I
think you can store proteins for months or even years with this
technic. But i never tryedto store my proteins in samplebuffer.

Hope you are pleased with this answer

Patrick




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