PAS Staining Procedure (Fungus)

Purpose:
This standard operating procedure (SOP) establishes the standard for staining fungus using the PAS staining procedure.

Reagents and Materials:

Reagents

0.5% Periodic Acid
Periodic Acid 2.5gm
Distilled water 500 ml

Schiff’s
Commercially prepared by Fisher Co.
Expiration date provided by manufacturer.
Store in refrigerator.

Hematoxylin
Commercially prepared by Anatech Co.
Expiration date and lot number on bottle.
Store at room temperature.

Materials
Tissue slides and control slides for special stain requested.
Microwave to bake slides.
Staining rack, 20 slide capacity.
Coverslips and synthetic mounting media.

Hazards and Precautions:
See Material Safety Data Sheets.

Sample Preparations:
Paraffin tissue section slides cut at 4 microns.

Calibration and Standardization:
A tissue section containing fungus is stained with each procedure.

Procedure:

1. Deparaffinize and hydrate to distilled water.

2. 0.5% Periodic Acid – 5 minutes.

3. Distilled water rinse.

4. Schiff’s reagent – 15 minutes.

5. Tap water – 5 minutes.

6. Hematoxylin (Harris) – 4 minutes.

7. Proceed as routine staining omitting Eosin.

8. Dehydrate, clear and coverslip.


Make fresh:
Light green 10ml
Distilled water 40ml

11. Distilled water rinse.

12. Dehydrate, clear and coverslip.


Positive control: Dog lung with Blastomyces dermatitidis (Figure 1)

Results:
Fungus – red


Figure 1. Blatomyces dermatitidis; budding yeast (arrows) in dog lung, left 400X, right 1000X.

Effective: July 1, 2004
Cherie Chapman
James R. Turk

References:

Sheehan DC, Hrapchak BR: Theory and Practice of Histotechnology, 2nd Ed, CV Mosby, St. Louis, 1980, pp. 168

Armed Forces Institute of Pathology Laboratory Methods in Histotechnology, eds. Prophet EB, Mills B, Arrington JB, Sobin LH, American Registry of pathology, Washington D.C. , 1992, pp. 151.