[Histonet] Help-Fried tissue

[Kemlo Rogerson]

Don't panic Mr Mainwaring!!!

You will need to do a little experimentation and only change one
criteria at a time; I'd first look at fixation as it is the foundation
for processing. You get brittle biopsies if you fix for long times in an
alcoholic fixative for example Carnoys; you may wish to try something a
tad gentler such as 10% aqueous buffered formalin. Either reduce the
time in the alcoholic fixative or use the aqueous one then see what
happens. 

If you are happy that it's not the fixative then look at the processing;
too much time in the alcohols and clearing agents could cause
brittleness. Make sure you have good, fresh solvents and minimise the
dehydration times or reduce heat if you are using it. Finally look at
the time you put the biopsies in paraffin wax and at what temperature.
>From my experience brittleness is usually caused by (in order of
commonness);

Using an inappropriate fixative maybe for too long; or perversely too
short a period and processing has an adverse effect on the unfixed
proteins. Fixation provides a matrix by binding proteins together and
allowing them to resist the strong coagulation effects of alcohol; if
this doesn't occur then it's like trying to fix them in alcohol; it
cause proteins to be strongly coagulated and a tendency towards
brittleness.

Too long, at too high a heat, in alcohols, clearing agent or wax.
Usually if you get the fixation correct on small biopsies the rest
follows, usually!!! The use of heat can cause proteins to become brittle
by coagulating them and disrupting the matrix; this is exacerbated by
heat.

Try it in a systematic way, one thing at a time, see what happens; OK? 

 
Kemlo Rogerson        
Pathology Manager
DD   01934 647057 or extension 3311
Mob 07749 754194; Pager 07659 597107;
Don't be afraid to take a big step when one is indicated. You can't
cross a chasm in two small jumps. --Buckminster Fuller 

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