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Fungal Descriptions

Laboratory Methods

	Antifungal Susceptibility Testing

	Culture Techniques and Media

	Microscopy Techniques and Stains

	Specimen Collection and Processing

School of Molecular & Biomedical Science
THE UNIVERSITY OF ADELAIDE
AUSTRALIA 5005

Contact:
Dr David Ellis

Email

Telephone:
+61 8 8161 6459
Facsimile:
+61 8 8161 7589

Periodic acid-Schiff (PAS) and PAS Digest stain.

For the demonstration of glycogen and neutral mucins. Useful for the detection of fungal elements in tissue sections.

Mechanism: Certain tissue elements are oxidized by periodic acid to produce aldehydes. These aldehydes will react with Schiff's reagent to produce a magenta colored compound. In the PAS Digest, the glycogen is digested by salivary amylase present in saliva.

Solutions.

1. 1% Periodic acid (50% vol.)

Periodic acid 2 ml

Distilled water 98 ml

2. Schiff's reagent:

Basic Fuchsin (C.I. 42500) 1 g

Potassium metabisulphite 2 g

Distilled water 200 ml

HCl concentrate 2 ml

Deactivated Charcoal 1-2 g

Boil distilled water, add basic fuchsin slowly, mix and cool to 50C. Add potassium metabisulphite. Mix and cool to room temperature before adding HCl. Keep in the dark overnight for bleaching to occur. Add charcoal and filter through coarse filter paper, then fine filter paper. Store in fridge.

Method.

1. Take sections to water.

2. For PAS digest only

Digest glycogen with saliva for 15 minutes

Wash well in water.

3. Treat with 1% periodic acid for 5 minutes

4. Rinse in water.

5. Treat with Schiff's reagent for 10 minutes.

6. Wash in running tap water for 5 minutes (this develops the color).

7. Counterstain nuclei lightly with Mayer's Hematoxylin for 1 minute.

8. Wash and "blue up" in Li2Co3.

9. Dehydrate, clear and mount in D.P.X.

Results: PAS positive material - magenta; Nuclei - blue; PAS digest material - colorless.