1.0 Purpose
To establish a consistent procedure for the PAS staining of tissue sections.
2.0 Scope
Applicable to all tissue sections received in the Surgical Pathology section for PAS staining.
3.0 Introduction
Periodic Acid-Schiff (PAS) staining is used to detect glycogen in tissues.
4.0 Principle
Periodic acid selectively oxidizes glucose residues, creating aldehydes that react with Schiff reagent to produce a purple-magenta color. A basic stain is often used as a counterstain. PAS staining highlights structures with a high proportion of carbohydrate macromolecules (glycogen, glycoprotein, proteoglycans) typically found in connective tissues, mucus, glycocalyx, and basal laminae.
PAS staining assists in diagnosing various medical conditions:
- Certain fungi
- Liver and renal biopsies
- Paget's disease
- Alveolar soft part sarcoma
- Staining macrophages in Whipple's disease
- Alpha-1-antitrypsin deficiency (positive staining in periportal liver hepatocytes)
- Aggregates of PAS-positive lymphocytes in epidermis in Mycosis fungoides and Sezary syndrome (Pautrier microabscesses).
5.0 Analytical Measuring Range and Limit of Detection
Not Applicable
6.0 Responsibility
- Technical staff
- Pathologist
- Technical staff
- Pathologist
7.0 Abbreviations
- PAS: Periodic Acid-Schiff
- PAS: Periodic Acid-Schiff
8.0 Requirements
Kit Reagents
- Schiff's reagent
- Xylene
- Absolute alcohol
- Periodic acid
Instrumentation and Software
Not Applicable
Disposables
Not Applicable
Kit Reagents
- Schiff's reagent
- Xylene
- Absolute alcohol
- Periodic acid
Instrumentation and Software
Not Applicable
Disposables
Not Applicable
9.0 Precautions
- Handle all specimens with gloves and masks.
- Handle all instruments carefully to prevent injury.
10.0 Limitations & Interferences
Not Applicable
11.0 Documentation
All documents are maintained in:
- Daily routine register
- Special stains log/register
- Handle all specimens with gloves and masks.
- Handle all instruments carefully to prevent injury.
10.0 Limitations & Interferences
Not Applicable
11.0 Documentation
All documents are maintained in:
- Daily routine register
- Special stains log/register
12.0 Instructions
12.1 Pre-Analytical Steps
12.1.1 Preparation of 1% Periodic Acid
1. Rinse the reagent bottle with distilled water.
2. Take 100 ml of distilled water in the reagent bottle.
3. Add 1 g of periodic acid and mix well.
4. Store at room temperature.
12.1.2 Preparation of Schiff's Reagent
1. Rinse a conical flask with distilled water and add 200 ml of distilled water.
2. Add 1 g of basic fuchsin and dissolve by heating to boiling point.
3. Cool the solution to 50°C and add 2 g of potassium metabisulphite.
4. Allow to cool to room temperature and add 2 ml of concentrated HCl. Mix well.
5. Add 2 g of activated charcoal, mix well, and keep in a dark place for 24-48 hours.
6. Filter after 48 hours and store at 4°C.
12.2 Analytical Steps
1. Bring sections down to water by dipping in two changes of xylene to remove the wax, and two changes of alcohol to remove xylene.
2. Wash in running tap water.
3. Treat with 1% periodic acid for 10 minutes.
4. Wash in running tap water.
5. Treat with Schiff's reagent for 30 minutes.
6. Wash in running tap water until the section becomes pink (approximately 10 minutes).
7. Counterstain with Harris's hematoxylin.
8. Dehydrate, clear, mount, and label.
9. Submit the slides to the pathologist for reporting.
12.1 Pre-Analytical Steps
12.1.1 Preparation of 1% Periodic Acid
1. Rinse the reagent bottle with distilled water.
2. Take 100 ml of distilled water in the reagent bottle.
3. Add 1 g of periodic acid and mix well.
4. Store at room temperature.
12.1.2 Preparation of Schiff's Reagent
1. Rinse a conical flask with distilled water and add 200 ml of distilled water.
2. Add 1 g of basic fuchsin and dissolve by heating to boiling point.
3. Cool the solution to 50°C and add 2 g of potassium metabisulphite.
4. Allow to cool to room temperature and add 2 ml of concentrated HCl. Mix well.
5. Add 2 g of activated charcoal, mix well, and keep in a dark place for 24-48 hours.
6. Filter after 48 hours and store at 4°C.
12.2 Analytical Steps
1. Bring sections down to water by dipping in two changes of xylene to remove the wax, and two changes of alcohol to remove xylene.
2. Wash in running tap water.
3. Treat with 1% periodic acid for 10 minutes.
4. Wash in running tap water.
5. Treat with Schiff's reagent for 30 minutes.
6. Wash in running tap water until the section becomes pink (approximately 10 minutes).
7. Counterstain with Harris's hematoxylin.
8. Dehydrate, clear, mount, and label.
9. Submit the slides to the pathologist for reporting.
12.3 Quality Control and Assurance
Known positive controls are run with each batch. Results are reported when controls are acceptable.
Calibration and Calibration Verification
Not Applicable
12.4 Storage of Samples
Slides are stored for 10 years.
13.0 Reference Range
Not Applicable
Known positive controls are run with each batch. Results are reported when controls are acceptable.
Calibration and Calibration Verification
Not Applicable
12.4 Storage of Samples
Slides are stored for 10 years.
13.0 Reference Range
Not Applicable
14.0 Critical Value
Not Applicable
Not Applicable
15.0 Reporting of Results and Interpretation
Magenta pink color is interpreted as positive staining in the appropriate context. Glycogen is diastase-sensitive, and mucin is resistant.
16.0 Contingency Plan
In the event of reagent shortages, manpower issues, or instrument breakdowns, suitably trained backup staff are available.
Magenta pink color is interpreted as positive staining in the appropriate context. Glycogen is diastase-sensitive, and mucin is resistant.
16.0 Contingency Plan
In the event of reagent shortages, manpower issues, or instrument breakdowns, suitably trained backup staff are available.
17.0 References
- Bancroft: Theory and Practice of Histological Techniques by John D Bancroft and Marilyn Gamble, 2007 (6th ed).
- Bancroft: Theory and Practice of Histological Techniques by John D Bancroft and Marilyn Gamble, 2007 (6th ed).
END OF THE DOCUMENT
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