Dear Kathy,

As always, each antibody will have it's own particular requirements. 
I do a lot of immuno on human brain which has been fixed for at least 
two weeks in strong formalin, and often don't need any type of 
antigen retreival.

On the other hand there are many antibodies I can never use because 
the antigens are so sensitive to formalin fixation. Are you parafin 
embedding? or cutting frozen sections after fixation. We do a minimum 
of fixation (2 hours post fix after perfusion of rats) for frozen 
sections because they usually have more sensitive antigens involved.

Everything is dependent on the antibody you are using or the antigen 
you are trying to visualise. You need to do very careful research 
when selecting the antibody. make sure to get the manufacturers 
specification sheets that say very clearly it works on formalin 
fixation parafin embedded tissue, and they should also tell you if 
you need antigen retreival. Also try and find out if other people are 
using this antibody on similar tissue and what they do.

Hope this helps, Cathy

---------------------------------------------------
Cathy Gorrie
Scientific Officer
Neural Injury Research Unit,
School of Medical Sciences,
University of New South Wales
Sydney, N.S.W. 2052

Phone: 61-2-9385 2462
Fax   : 61-2-9313 6252
e-mail: c.gorrie@unsw.edu.au


At 1:48 PM -0500 13/12/01, Kathleen Cormier wrote:
>Hello All,
>
>I was wondering, I am curently getting back into immuno's in a research
>setting, ack! It's been a long time, and my brain is fuzzy...how long is
>too long for for immuno's. The general rule of thumb here
>is 24 hrs best, no more than 36 hours. Has anyone else noticed this
>excessive sensitivity  to formalin? We do do antigen retrival, and some
>digestion techniques on all the slides. Is this just the nature of the
>beast? Has anyone found a nice easy to use all around fixative that works
>well with immuno and that does not render the tissue hard as a rock to
>section? Thanks all!
>
>Kathy
>Histo Supervisor
>Division of Comparative Med
>MIT