[Histonet] immunohistochemistry and frozen sections

[Vincenzo La Manna]

Hi Histonetters,

I've been struggling for 4 months on my 5 microns
frozen sections trying to find estrogen receptors.
I've tried both immunoperoxidase kit from Vector
laboratories either indirect immunofluorecsence (FITC
Goat anti mouse Secondary Ab from Sigma) but I've not
yet obtained good results.
My primary Ab is a mouse monoclonal to bovine
Estrogen receptor from Cymbus biotechnology.
Sections come from tissue that is supposed to be a
non-target tissue (mid laminar region from cow's
hooves)blocking with 2% goat normal serum (sigma)
whash in tbs buffer
All different methods for antigen retrieval have been
tested but sections look poor regarding histologic
quality and I still have very big problems with non
specific binding an staining (indirect
immunofluorescence).
It looks like frozen sections are too tender to
efford the whole processing.
Every suggestion or reference is welcome,
thank U

La Manna Vincenzo
Department of Agriculture and Forestry, University of
Aberdeen, 581 King Street, AB24 5UA, Aberdeen , UK.
Telephone; 01224 274259
Fax; 01224 273731
e-mail v.lamanna <@t> abdn.ac.uk