Western - tiny tissue amounts
H.Veldman at lab.azu.nl
Mon Feb 28 10:29:12 EST 2000
Jörg Kempfert schreef:
> First thanks for your interest.
> The main problem for us is: how to crush those small pieces of tissue (here talking
> about 20mg each) and then to get the cytosol.
> Would be very kind if you could help us.
> Joerg Kempfert
> pathos schrieb:
> > In article <38B664E6.97A9B956 at uke.uni-hamburg.de>, =?iso-8859-1?Q?J=F6rg?=
> > Kempfert <kempfert at uke.uni-hamburg.de> wrote:
> > > We want to do some western blot analyses with really small tissue
> > > amounts. Does anybody have experience with this kind of problem? It
> > > would be very nice if someone could help us in this case.
> > >
> > > Looking forward to get your hints.
> > What is the problem? I routinely use very small pieces of tissue for westerns.
> > --
> > Peter Pediaditakis
> > University of Pittsburgh
> > Dept. of Pathology
I used to do SDS-PAGE - Western blotting on small (20-30mg) pieces of frozen muscle
biopsies. I took up the material in a 20 X volume of SDS-PAGE sample treatment buffer
(about 0.5ml) in a 4ml RIA tube. For homogenizing I used an Ultra-Turrax with a
dispersing tool for 0.3-3ml volumes at full speed twice for 15 sec.
Hope this helps
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