[Histonet] RE: goldners trichrome

From:"Prior, Andrew"

We've used this protocol in the past with some success on 15Ám ground MMA sections. It was adapted from a Nottingham University method. As usual, you may need to tweak the times to suit your sections.
Good Luck.

Andrew Prior
Smith &Nephew Research Centre
York Science Park
YO10 5DF

Staining Protocol:


*	Ponceau/Acid Fuchsin Stock Solution:
1.5 g - Ponceau 2R.
0.5 g Acid Fuchsin.
2 ml Acetic Acid (glacial).
98 ml Distilled Water.

*	Azophloxine Stock Solution:
0.5 g Acid Red 1.
0.6 ml Acetic Acid (glacial).
99.4ml Distilled Water.

*	Ponceau Working Solution:
12ml Ponceau/Acid Fuchsin Stock Solution.
8ml Azophloxine Stock Solution.
80 ml 0.2% Acetic Acid.

*	Orange G Solution:
1 g Orange G.
1.5g Phosphomolybdic Acid.
250 ml Distilled Water.

*	Light Green Solution:
1 g Light Green (Yellowish).
1 ml Acetic Acid (glacial).
500 ml Distilled Water.

1)	Slides in distilled water for 15 minutes.
2)	Stain in Weigert's Haematoxylin for 30 minutes.
3)	Wash in running tap water for 2 minutes.
4)	Differentiate in 0.5% acid alcohol (30 seconds).
5)	Wash in water for 20 minutes.
6)	Stain in Ponceau Working Solution for 5 minutes.
7)	Rinse in 1% Acetic Acid for 15 seconds.
8)	Stain in Orange G Solution for 20 minutes.
9)	Rinse in 1% Acetic Acid for 10 seconds.
10)	Stain in Light Green Solution for 20 minutes.
11)	Rinse in water.
12)	Blot dry.

Expected Results:
Nuclei = blue/black.
Mineralised bone = green.
Muscle = green.
Osteoid = red.
Collagen = red.


-----Original Message-----
From: histonet-bounces@lists.utsouthwestern.edu [mailto:histonet-bounces@lists.utsouthwestern.edu] On Behalf Of histonet-request@lists.utsouthwestern.edu
Sent: 27 February 2008 18:05
To: histonet@lists.utsouthwestern.edu
Subject: Histonet Digest, Vol 51, Issue 40

Message: 5
Date: Wed, 27 Feb 2008 10:19:52 -0500
From: Simoskevitz@Osteotech.com
Subject: [Histonet] goldners trichrome
To: histonet@lists.utsouthwestern.edu
Content-Type: text/plain; charset=US-ASCII


I was asked to do a Goldners Trichrome in Methymethacylate embedded bone .
Do you have a good protocol for this or do you know where I could find one.

Thank you for any help you can give me.

Ricki Simoskevitz
Osteotech Inc.
Eatontown, NJ 07724
(732) 542-2800 X6328
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