At 11:18 21/03/02 -0500, vous avez écrit:
>Just wondering, I've seen vibratomes for years but never really understood
>what people are using them for. Anyone?

They are mainly used for immunocytochemistry and histoenzymology.
Some people use them for pre-embedding immunostaining for EM. The advantage 
of the vibratome is that you don't have to embedd or freeze, and thus 
ultrastructure (including membranes) is well conserved. You immunostain the 
vibratome sections as floating sections with minimal or no detergents (and 
a lot of patience !!!) and peroxidase-DAB as signal. After the immunostain, 
you osmicate the section and embedd them in resin. The osmicated and 
embedded sections can still be viewed under the microscope to locate 
interesting areas, which you then cut out and process for ultrathin EM 
sections.
The problem with this procedure is that antibodies penetrate only for 1 or 
2 micrometers using this procedure, so you better catch those first 
ultrathins ...
One major use of this procedure is for antigens that are present in a few 
isolated cells, e.g. somatostatin in cortex. By standard EM you will have 
to do a lot of ultrathins to have one that contains a somatostatin neuron. 
With the pre-embedding you process a whole coronal section and then locate 
the few somatostatin cells which you then recut for EM.

You can also cut unfixed, living tissue on the vibratome. But if you want 
to use those sections for cell culture, the vibratome is usually to slow 
and a McIlwain tissue chopper is less regular in section thickness, but 
also much much faster.

In our lab, some (IMHO very courageous) people use the vibratome for 
standard optical immunostaining. They cut 25-50 micrometer brain sections, 
permeabilize them with lots of detergent and process the floating sections 
for ICC. The procedure seems to work quite well.

I for my part don't like the vibratome. Actually, I get criminal urges if 
someone (my boss ...) suggest I should use it. If I have to do 
pre-embedding EM I use well cryoprotected tissue frozen sections. They work 
just as well, but are less bothersome to produce (1 frozen section = one 
turn of the handle, 1 vibratome section = about 1 minute and lots of nerves)

Paul
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Paul Klosen, PhD
CNRS UMR 7518 Neurobiologie des Fonctions Rythmiques et Saisonnieres
Universite Louis Pasteur  12, rue de l'Universite
F-67000 Strasbourg, FRANCE
Tel. 03.90.24.05.01  Fax. 03.90.24.05.28
========================klosen@neurochem.u-strasbg.fr=========================