alizarin red and alcian blue

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From:MontagueDonnaC@exchange.uams.edu
To:histonet@pathology.swmed.edu
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Date:Mon, 30 Aug 1999 11:28:23 -0500
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Hal and others:

The reference we used here was:

"Differential Staining of cartilage and Bone in Fetal Mouse Skeleton by
Alcian Blue and Alizarin Red S"
Minoru Inouye (1976) Congenital Anomolies 16:171-173

For full term mouse fetuses weighing 1.0 -1.6 g

1) Fix 95 % ethanol, 4 days or longer. Remove skin, viscera and adipose
tissue at the time of sacrifice or within a few days after fixation before
hardening of the specimens.
2) Place in acetone one day or longer to remove fat.
3) Stain 2 - 3 days in the following staining solution at 37 oC
	0.3 % filtered alcian blue 8GS (CI 74240) in 70 % ethanol	5 ml
	0.1 % filtered alizerin red S (CI 58005) in 95 % ethanol	5 ml
	Acetic acid, glacial 					5 ml
	70 % ethanol						85 ml
(use approximately 10 ml staining solution per fetus)
4) Wash in water (gently running tap water 3 - 5 minutes)
5) Clear in 1 % KOH (aq) 12 - 48 hours unti skeletons are clearly visible
through the surrounding skin.
6) Place in 20 % glycerin (aq) containing 1 % KOH 1 - 5 days.
7) Clear specimens through 50 % glycerin followed by 80 % glycerin and store
in 100 % glycerin.

Results:
	Cartilage = blue
	Ossified skeleton = purple to red (reddens as it ages in the 100 %
glycerin)

We used this stain for pups of dams fed lead in their drinking water.
Companion pups were subjected to specific lead staining using the
Rhodizonate Technique of Bancroft and Stevens, 1990.

Hope this helps, Donna Montague, UAMS Orthopaedic Research, Little Rock, AR,
USA



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