[Histonet] Caveats Re: Bouins in microwave for Massons trichrome

[gayle callis]

Dear Histonetters, 

 

Microwaving Bouins can be an absolute disaster as experienced  by me.   It
went all over the interior of microwave (borrowed one at that!) and created
an unbelievable mess of picric acid that then dried out,  and formaldehyde
going into every nook and cranny of the microwave interior along with
terrible fumes.  I spent a good deal of time    All I can say is DON"T
MICROWAVE BOUINS!!  

 

Other caveats:  

 

#1 is already mentioned

 

#2 Microwaving sections in Bouins can lead to spurious staining results.
Too short a time can create poor staining with this protocol.   As stated by
Liz Chlipala,  use a heated oven or waterbath at 56°C - 60°C for one hour to
ensure the section is properly exposed Bouins acid components (picric and
acetic acids) is a must for staining the connective tissue fibers.
This is the classic way of doing Massons Trichrome and still works the best.
A microwave must be properly vented in a hood, or it is a toxic mess of
fumes, let alone having to clean up nasty picric acid that will dry out in
crevices/corners of MW.   You can use a one zip lock baggie,  left open at
one end of top to vent fumes, and collect any boil over, but staining
problems have to considered due to short exposure to Bouins.   This is
something I learned from Jerry Fredenburgh many years ago, and after bulk
staining several hundred decalcified bone sections, the classic method,
correct time in Bouins gave the best and most consistent results.  

 

#3  If you are doing Mass Tri the next day, deparaffinize the sections,
rehydrate and immerse in Room Temperature Bouins and let sections sit
overnight.   No heating means:   no toxic formalin fumes from either MW or
heated oven,  problem and decalcified bone sections stay on slide as these
can dislodge due to  mechanical/physical forces from MW heating and
sometimes with heating in an oven, and is safer for user.  This became our
favorite method, and no more spillage/toxic fume problems.   

 

If anyone wants an excellent Massons Trichrome method, the classic one I
have from AFIP lab is superb.  It also gives hints on achieving best results
with proper differentiation of connective tissue fibers - something we often
do not think about when doing this stain.   

 

Take care

 

Gayle M. Callis

HTL/HT/MT(ASCP)