Silver Nitrate Staining Protocol

Silver Nitrate has had a long usage in historical staining techniques and is still used in modern pathology. Initially, early researchers used silver nitrate to enhance the visibility of the tissue structure while studying it; this was done by applying solid silver nitrate on a tissue and then studying it. The stain substance has been developed for many compounds, and confirmatory tests are needed when silver nitrate is used.
Grocott-Gomori's Methenamine Silver (GMS) stain is a histological silver staining. Its initial application to assess missing tissues and diseases in the liver and the rectum and then used for the identification of carbohydrates in fungal microorganisms.

Preparation of solution. Chromic acid (1.4%) is composed of 4 g chromium trioxide and 100 ml distilled water; Silver solution is composed of 23 ml 3% methenamine / hexamine, 1.25 ml 5% silver nitrate, 3 ml 5% borax (sodium tetraborate) and 25 ml distilled water; 3.2% sodium chloruretic (yellow gold chloride) is composed of 1.0 g gold chloride and 500 ml distilled water; 4.2% sodium thiosulphate (hypo) is composed of 2.0 g sodium thiosulphate and 100 ml distilled water; Working light green stock solution is composed of 10 ml of 1% light green in 1% acetic acid and 40 ml distilled water.
Hydrate sections with distilled water.
Then oxidize the section with 4% aqueous chromic acid at room temperature and leave for 1 hour, wash in water for a few seconds.
Treat the sections with 1% sodium metabisulphite for 1 min, wash in smoothly running tap water for 3 mins, and rinse thoroughly in distilled water.
Put the slides in pre-heated working silver solution in a water bath at 60°C for 15 to 20 mins until the section turns yellowish-brown.
Rinse well in distilled water.
Add to the sections 0.2% gold chloride and leave for 2 mins, and rinse well in distilled water.
Treat the sections with 2% sodium thiosulphate for 2 mins, wash with smoothly running tap water for 5 mins.
add the counterstain into the light green for 15 secs.
Rinse the excess light green solution off the slide with alcohol.
Dehydrate, clear and mount.

Wear gloves, goggles and laboratory coats. Keep hot uncapped solutions under the fume hood. Avoid contact and inhalation of dyes and chemicals.
Chromic acid is corrosive to skin and mucous membranes and highly toxic affecting the kidneys.
Sodium metabisulfite ais toxic if ingested and can cause gastrointestinal distress and irritability. It also irritates the skin, eye and the mucous membrane.
Silver nitrate is an irritant to the skin and eyes.
The oxidizer if ingested caused production of a violent gastrointestinal discomfort and it is possibly carcinogenic since it's a tumorigenic agent.
Sodium thiosulfate is toxic in ingestion and it causes stomach, skin, eyes and respiratory tract irritation.
Light Green SF Yellowish is considered to be a possible carcinogen.

For research use only. Not for any other purpose.

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