[Histonet] Phosphotungstic Acid Hematoxylin stain without usi
ng Zenker's
Tony Henwood
AnthonyH <@t> chw.edu.au
Mon Mar 22 18:22:18 CST 2004
Dear Boswell,
The following Cherukian's modification works very well.
Tony Henwood JP, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: 612 9845 3306
Fax: 612 9845 3318
http://www.histosearch.com/homepages/TonyHenwood/default.html
http://us.geocities.com/tonyhenwoodau/index.html
Document Procedure:
Phosphotungstic Acid Haematoxylin
Principle:
Cherukian's modification employs an eosin solution that stains the
erythrocytes red and differentiates them from the blue fibrin.
Fixation: 10% buffered formalin.
Microtomy: paraffin sections at 5um.
Controls:
Use brain sections and section of muscle. A good stain will demonstrate the
dendrites as blue where as in a bad stain they appear light grey to salmon
in colour. Nuclei, fibrin, platelets and muscle will be blue, red cells and
collagen appear red. Muscle striations should be well defined.
Reagents:
1. Eosin: - Warning: Flammable - see MSDS
Eosin Y, water soluble (CI 145380) 0.5g
Distilled Water 10ml
80% Ethanol 190ml
Working: 10ml stock and
Before use add 50ul glacial acetic acid.
2. 1% Periodic Acid
3. PTAH solution
Haematoxylin (CI 75290) 0.5g
Phosphotungstic Acid 10g
Distilled water 500ml
Dissolve solid ingredients in separate portions of the water. Use gentle
heat for Haematoxylin. Combine solutions when cool. Add 0.088g potassium
permanganate to ripen. The stain is ready to use.
Procedure:
1. Dewax and hydrate sections to 80% alcohol.
2. Place slides in eosin for 30 seconds.
3. Wash slides in distilled water for a few seconds.
4. Place slides in 1% periodic acid for 20 minutes.
5. Wash slides in water for 3 minutes.
6. Place slides in PTAH for 30-90 minutes in 60oC oven. Check from 30
minutes on.
7. Dehydrate, clear and mount.
Results:
Dendrites, nuclei, fibrin, platelets and muscle - blue
Red blood cells and collagen - red.
Notes:
Reference:
1. Cherukian, C.J., Histologic. 8(4); 105, (1977).
2. Luna, L., Histologic. 5(2); 66, (1975).
-----Original Message-----
From: Boswell [mailto:kcboswell <@t> grandecom.net]
Sent: Saturday, 20 March 2004 7:30 AM
To: Histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Phosphotungstic Acid Hematoxylin stain without using
Zenker's
Does anyone have a procedure for Phosphotungstic Acid Hematoxylin stain
without using Zenker's. We have looked everywhere and are unable to find
it. Thank you.
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