[Histonet] Yasue's Silver Nitrate-Rubeanic Acid Method for CalciumOxalate

Tony Henwood AnthonyH <@t> chw.edu.au
Wed Aug 1 18:17:48 CDT 2007

This is the one we have used:

Yasue's Method for Calcium Oxalate

Calcium oxalate can be demonstrated in several organs in patients suffering from hereditary or acquired oxalosis. Calcium oxalate is also common in the thyroid in cases of nodular goitre. Oxalates have been identified in giant cells in various granulomatous lesions such as skin granuloma, tuberculous or chronic granulomatous salpingitis, and sarcoid granuloma (Katoh et al 1993).  Oxalosis is also associated with aspergillosis

Under high power calcium oxalate appears as colourless crystals that exhibit brilliant birefringence under polarised light. They stain black-brown with Yasue's method (Yasue 1969), where as they do not stain with von Kossa's or alizarin red S (Katoh et al 1993).

Yasue (1969) Acta Histochem Cytochem 2:83-95.
Katoh et al (1993) Am J Surg Pathol 17(7):698-705.

Fixation:      10% buffered formalin.

Microtomy:     5μm paraffin sections.
1.	5% Acetic Acid
2.	5% Potassium Hydroxide
3.	5% Silver Nitrate
4.	Rubeanic Acid Solution:

Prepare a saturated solution of rubeanic acid in 100ml of 70% ethanol
Add 2 drops of concentrated ammonium

5.	2% Methyl Green
6.	50% ethanol


1.	Dewax and Hydrate sections
2.	Treat with 5% acetic acid 30min.
3.	Wash in water
4.	Place in 5% Potassium Hydroxide 30min.
5.	Wash in water
6.	Rinse in distilled water
7.	Place in 5% silver nitrate 20min.
8.	Rinse in distilled water
9.	Place in Rubeanic acid solution for 1min
10.	Rinse in 50% ethanol
11.	Rinse in water.
12.	Counterstain in 2% Methyl Green 2min.
13.	Dehydrate, clear and mount


Calcium Oxalate	Brown/black
Nuclei			Green.


Tony Henwood JP, MSc, BAppSc, GradDipSysAnalys, CT(ASC)
Laboratory Manager & Senior Scientist
The Children's Hospital at Westmead,
Locked Bag 4001, Westmead, 2145, AUSTRALIA.
Tel: 612 9845 3306
Fax: 612 9845 3318

-----Original Message-----
From: histonet-bounces <@t> lists.utsouthwestern.edu [mailto:histonet-bounces <@t> lists.utsouthwestern.edu] On Behalf Of djemge <@t> aol.com
Sent: Thursday, 2 August 2007 8:02 AM
To: Histonet <@t> lists.utsouthwestern.edu
Subject: [Histonet] Yasue's Silver Nitrate-Rubeanic Acid Method for CalciumOxalate

Does anyone have the protocol for Yasue's Silver Nitrate Rubeanic Acid method. One of the researchers is doing a calcium oxalate study. I have already given him Pizzolato's, Von Kossa, and Alizarin Red for comparisons. I have heard that the Yasue's method is much more specific. Thanks, Donna

Donna Emge, HT(ASCP)
Northwestern University
Feinburg School of Medicine 
303 E Superior St. Lurie 7-220
Chicago, IL 60611
d-emge <@t> northwestern.edu

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