[Histonet] HYALURONIC ACID detection

[John Kiernan]

Some properties of hyaluronic acid.

(1) Stains with alcian blue 8G at pH 2.5 
    but not at pH 1.0.
(2) Staining prevented by prior treatment with
    streptococcal hyaluronidase. (Testicular
    hyaluronidase is cheaper & more easily
    available but can also remove chondroitin
    sulphates. Chondroitin sulphates are
    stained by alcian blue at both pH 1 and 2.5.
(3) Staining by alcian blue is reversibly 
    blocked by prior methylation of the section.
    Stainability is restored if methylation is
    followed by saponification. The staining of
    chondroitin sulphates with alcian blue is 
    irreversibly blocked by methylation.
(4) Negative with classical PAS method but
    becomes PAS-positive if a special 
    oxidation sequence is used (Scott &
    Harbinson 1969 Histochemie 19:155-161).

If you have pure hyaluronic acid there will
be no difficulty staining it. If, as is usually
the case, other macromolecular carbohydrates
or glycoproteins (collagen, for example) are
present in exactly the same place, you'll need
to do all the above methods and more. The best
account of carbohydrate histochemistry that I
know is Chapter 15 in Vol 2 of Pearse's
"Histochemistry". It's quite heavy going, 
but this is a field where you really do need
to be in command of every step of every method.

Hope this helps.
-- 
-------------------------------
John A. Kiernan
Department of Anatomy and Cell Biology
The University of Western Ontario
London,   Canada   N6A 5C1
   kiernan[AT]uwo.ca
   http://publish.uwo.ca/~jkiernan/
   http://instruct.uwo.ca/anatomy/530/index.htm
_______________________________
Hernan Aldana wrote:
> 
> Dears histonetters
> I like to demonstrate the presence of hyaluronic acid in fixed eyes. Can
> anybody help me with some advices?
> Histochemical or immunohistochemical. Please help me.
> Thanks in advance to all
> 
> Dr. Hernán J. Aldana Marcos
> Facultad de Medicina. Universidad de Morón
> Machado 914. B1708JPD. Buenos Aires. Argentina
> e-mail alternativo hernanjavier <@t> yahoo.com
> web: http://hjaldanamarcos.bravepages.com
> http://www.ht.org.ar
> 
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