Introduction
Masson-Fontana method is a special staining method used in Histological diagnostic procedures.
This staining method is useful for the identification and histological visualization of Argentaffin cells and Melanin in paraffin or frozen sections.
Moreover, Masson-Fontana method is useful for the identification of Cryptococcus neoformans.
For this staining procedure it is necessary to use formalin fixed tissue sections.
Fixatives such as chromate and mercuric chloride should be avoided as the tissue sections fixed with these fixatives are not suitable for Masson-Fontana staining.
Melanin is a pigment that normally can be found in skin, eye, substantia nigra of the brain.
Moreover, in pathological conditions such as in benign nevus cell tumors and malignant melanoma conditions, melanin can be found.
Argentaffin cells are the cells that take up silver stain. Melanin also has argentaffin properties.
Principle
Melanin is considered as a powerful reducing agent.
This property is used for the demonstration of melanin in tissue sections in histological diagnostic procedures.
Melanin and Argentaffin cells bind the silver from ammoniacal silver solution and reduce them into metallic silver.
For this purpose, it does not need any extraneous reducing agent.
- For this staining procedure, three solutions are needed.
- They are 5% Hypo (Sodium Thiosulphate) solution, Ammoniacal silver solution and 0.2% Safranin.
- For the preparation of Ammoniacal silver solution add concentrated ammonia drop by drop to 20ml of 10% aqueous silver nitrate.
- After adding every single drop mix the solution well until the formed precipitate almost re dissolved.
- There can be observed a faint opalescent after reaching the end point.
- Finally add 20ml of distilled water.
- Then mix it well and filter.
- This solution can be stored in dark container up to one month until use.
Method
First bring the tissue sections down to the water.
Then treat the tissue section with ammoniacal silver solution in a dark container or in a coplin jar covered with an aluminum foil for 30-40 minutes at 56°C or overnight at room temperature.
Wash in distilled water well for several changes.
Then treat the section with 5% Sodium thiosulphate solution for 1-2 minutes.
Wash in running water for 2-3 minutes.
After that counter stain with 0.2% Safranin for few seconds.
Then wash with distilled water.
Finally dehydrate, clear and mount with DPX.
Observe under the microscope.
Purpose
Identification and histological visualization of melanin in histological sections
Results
Argentaffin Cells and melanin stain in black colour, Nuclei stain in red colour, cytoplasm can be seen in light pink colour and if cryptococcus present, cryptococci cell wall will be stained in black colour.
Other than Safranin, 0.5% aqueous neutral red or 0.1% aqueous nuclear fast red can be used as counter stains.
For this staining procedure it is necessary to use thoroughly cleaned glassware.
Because, silver solution can be reacted with any debris left in glassware.
When staining in 56°C, it should be removed at the given time. If not, it may give rise to fine deposits over the section.
Faintly stained tissue sections or sparsely positive cells can be difficult to interpret.
A positive control slide should be stained with every batch of staining.
Any cell containing melanin can be used as normal positive control. Always results should be interpreted with a positive control slide.
The staining procedure can be differed according to the condition of the stain. Time of staining should be adjusted accordingly.
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