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Malay - Aim of special stain:
- Identify inorganic calcium salts – most commonly phosphates and carbonates
- Diagnostic applications:
- Abnormal depositions of calcium can be found in necrotic areas of tissue, associated with tuberculosis, infarction, atheroma in blood vessels and malakoplakia of the bladder
- Material supplied for surveys:
- Lung tissue - Mild Interstitial pneumonia with diffuse interstitial calcification, cut at 3µm
- Recommended control:
- Un-decalcified bone, placenta tissue or any other paraffin embedded tissue that contains calcium deposits
Von Kossa staining method (for calcium deposits):
Reagents – commercial source unknown, unless specified:
| 1. 5% Silver nitrate solution | |
| a) Silver nitrate | 5 g |
| b) Distilled water | 100 mL |
| 2. 5 % Sodium thiosulphate solution | |
| a) Sodium thiosulphate | 5 g |
| b) Distilled water | 100 mL |
| 3. Nuclear fast red solution |
| 4. Graded alcohols and Xylene |
Method:
- Deparaffinize and hydrate to distilled water.
- 5% Silver solution, place in bright sunlight, or in front of a 60-watt lamp, place foil (or mirror) behind the jar to reflect the light. Leave for 1 hour or until calcium turns black.
- Rinse in distilled water, 3 changes.
- 5 % Sodium thiosulphate solution for 5 minutes.
- Wash in tap water, rinse in distilled water.
- Nuclear-fast Red for 5 minutes.
- Wash in water.
- Dehydrate, clear, and coverslip.
Results:
Calcium salts – black
Nuclei – red
Disclaimer:
These methods are intended as a guide only. Laboratories that wish to implement these methods should perform internal validation prior to use. The RCPAQAP does not make any claim or warranty for the accuracy or performance of these methods.
References
2015 RCPAQAP Technical General Survey 3.
Bancroft, JD and Gamble, M. Theory and Practice of Histological Techniques. 8th Edition, Churchill Livingstone. London. 2019:217-218.
Susan N. Meloan, HT (ASCP), and Holde Puchtler,MD. Chemical Mechanisms of staining methods. Von Kossa’s Technique: What von Kossa really wrote and a Modified Reaction for Selective demonstration of Inorganic Phospahtes. The Journal of Histotechnology/Vol.8, No.1/March 1985.
George L. Kumar; John A. Kiernan. Dako Education guide. Special stains and H&E. 2nd edition. 2010, Dako North America, Carpinteria, California.
J.A Kiernan J.A. Histological & Histochemical Methods: Theory and Practice 2nd Edition. Butterworth-Heinemann Ltd; 1990:222.
Avery CR, Aterman K. Calcification of the basement-membrane of placental villi. J Pathol. 1971 Mar;103(3):199-200. doi: 10.1002/path.1711030310. PMID: 5567176.