[Histonet] tissue quality controls for ish and ihc

[Nancy.Walker <@t> sanofi-synthelabo.com]

Hello,

I'm struggling with ideas about immunohistochemistry and in situ
hybridization controls to make sure tissue has been not over fixed and also
that the mRNA has been well preserved. I work on detecting mRNA or protein
for targets for orphan receptors (most of which we have very little info
where the target is. In the past working on rodent brain tissue, we got
nice results "assuming" that everything was perfect. Now I'm working on
human tissue arrays where I don't control the fixation and processing so
I'd like to have an internal control. Even in rodent tissue I think it
would be a good idea to control the tissue.

I'm making a riboprobe for vWF because it's found in all tissues (low in
liver gut and muscle) and I like the idea of having something specific to a
particular cell type (vessel endothelial cells), compared to something like
GAPDH.

What do you think about an oligo dT probe? This detects poly (A) mRNA, the
advantage is that it would be universal for all species. I found it in the
BioGenex catelogue on page 157, it is supposedly used to assess mRNA
preservation. Any comments?? For our riboprobe ISH we are thinking of
making an oligo with a T7 promotor followed by a tail of T (25 or more).
Then we'll transcribe with a mix of UTP's radioactive and normal  (1:20)
and this will hybridize to all mRNA (they have poly A tails).

Biogenex  also promote a monoclonal antibody (V9) to Vimentin, on page 113
(reference cited: Battifora, H., Am J Clin Pathol 96:669, 1991). I ordered
the article, does anyone have it? Does any vimentin work or is this clone
special??? This sounds nice does any one have any comments?

yours truely,
Nancy Walker
Molecular Biology Scientist

Sanofi-Synthelbo Research
B.P. 37 Labége Innopole
31676 LABEGE CEDEX FRANCE

nancy.walker <@t> sanofi-synthelabo.com
tel : (33)561004179  fax :(33)561004001