Re: [Histonet] Re: overprocessing

From:John Kiernan

Dear Carlos (whoever and wherever you are),

You are quite right, and your observations are
supported by published literature that has been
through the academic (= accept nothing without
proof) peer-review process. 

Hardness is difficult to evaluate, but anyone can
measure changes in volume or linear dimensions.
This was all done decades ago, and I'll be happy
to send literature references to anyone who's
seriously interested. 

Cross-linked polymers 
(such as the gels used in PAGE) shrink rather
suddenly when water removal (by evaporation or
alcohol) hits a critical level. I tried this in
the lab after reading a Scientific American
article about 20 years ago. There was a rapid
change from soft & slimy to hard & gritty, exactly
as Sci. Am. said it would be. I can't find the ref
for this pre-computers Sci. Am. article. 

Shrinkage of animal tissues is less than that of
highly hydrated polyacrylamide gels or gelatin. A
linear measurement in a stained and mounted
paraffin section may be about 70% of the in vivo
measurement, but the % can vary with the organ. 

I can send references to anyone who's intersted in 
following this up. 

John Kiernan
London, Canada
carlos wrote:
> Hi Kemlo,
> I must disagree re hardening......if only in my experience of the following:
> I have processed gelatin ( 1%)blocks after formalin fixation (  1cm blocks
> fixed for 2, 5, 10 and 60 days) and find that they all suffer from shrinkage
> and hardening. My processing is via gradual alcohols to alc/xylene 1:1, then
> xylene x2 and wax x2. Two hrs in each.
> Be grateful for enlightenment.
> Best wishes
> carl
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