[Histonet] Re. Decalcification with formic acid sodium
gayle.callis at bresnan.net
Mon Jul 27 18:21:38 CDT 2015
Dorothy and Carl,
Comments about your Histonet replies on formic acid decalcification.
The Morse solution referred to by Dorothy can be picked up online by typing
in the DOI number: 10.1.1.4689.3439.pdf or title, Morse A. Formic
acid-sodium citrate decalcification and butyl alcohol dehydration of teeth
and bones for sectioning in paraffin. 1945 J Dental Res 1945:24:143. You
will find the reference to Evans and Krajian paper on formic acid/sodium
citrate along with the original recipe for their solution (equal parts of
85% (stock) formic acid and 20% sodium citrate). Morse modified the Evans
Krajian method (1 part diluted stock formic acid i.e. 90% diluted 1:1 with
water for 45% formic acid) plus 1 part 20% sodium citrate. The Morse paper
was excellent and well worth reading. Interestingly, in 1962, our lab used
the Morse solution for decalcifying teeth although it was never referred to
by that name but simply formic acid/sodium citrate. The formic acid
content in Morse's solution is half the concentration of formic acid in
Evans/Krajian solution. It seems both work equally well and the higher
concentration of formic acid should increase the decalcification rate
somewhat. Morse also did chemical decalcification endpoint testing.
Carl is correct about not mixing citric acid with formic acid as citric acid
is not going to act as a buffer salt. However, you will find in the
literature that citric acid is very mild and has been used as a decalcifying
agent for EM studies. Carl is also correct in that sodium formate can be
used as a buffering salt instead of sodium citrate. We have worked with
both of the buffering salts/formic acid formulations and found they works
equally well for decalcification. I have some publications on file
comparing acid versus EDTA for cartilage and IHC studies and learned some
researchers referred to buffered formic acid methods as "acidic buffers" .
The latter terminology could be confusing to people in the business of
decalcifying bones and teeth. but no more so than the acronyms manufacturers
give their solutions. It pays to read the MSDS for any decalcifying
solution, and even compare this information to what is in histology
textbooks as part of our education.
I have found the discussions on this topic enlightening. I will be happy
to send the pdf of the Morse method to those interested in reading it. I
have not been able to access the 1930 Evans Krajian method yet. What is
important is knowing these older, classic formic acid methods are still
tried and true with the added advantage of being available commercially for
There was a paper
Talking about formic acid (Morse solution) can get as good result as EDTA in
Mouse knee joints:
done lots of decalcified FFPWS for assessment of articular cartilage
See Histonet images for a Tol blue image.
Decal in 10 % EDTA for 3 days on a rocker at RT.
Sure....5days if you are worried.
No difference in Immuno-reactivity, imho.
If you want to use buffered Formic acid, use Formic acid; sodium formate.
Use of citric acid with Formic acid does not make a buffer.
It's just mixing two relatively mild acids.....
However, I am sure that Prof Kiernan can further enlighten us.
Carl Hobbs FIBMS
Histology and Imaging Manager
Guys Campus, London Bridge
Kings College London
020 7848 6813
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