Introduction
This staining technique is used in histological diagnosis procedures. First periodic acid-Schiff (PAS) staining was introduced by McManus in 1946 for the demonstration of mucin.
Periodic acid-Schiff (PAS) staining is a special type of stain which is useful in the demonstration of carbohydrates and carbohydrate components or glycoconjugates in tissues.
This stain demonstrates polysaccharides, sialo mucin, neutral mucin, glycogen, basement membrane, α-antitrypsin, reticulin, fungal cell wall components, pancreatic zymogen granules, thyroid colloid, corpora amylacea, and Russell bodies.
It is very helpful in the diagnosis of tumors through the detection of glycogen or mucins.
Moreover, PAS technique is widely used for the demonstration of viable fungi in tissue sections.
This is due to the presence of periodic acid reactive components in the fungal capsule and cell wall.
Common fungal species that can be identified by PAS stain are Candida albicans, Histoplasma capsulatum, Cryptococcus, and Blastomyces.
In addition, this technique is used for the assessing of the thickness of basement membranes.
For this technique, formalin fixed, paraffin embedded tissues as well as frozen sections can be used.
Periodic acid is used as an oxidant. By treating the tissue sections with periodic acid, hydroxyl groups presence in the carbohydrates get oxidized.
By that oxidation, two free aldehyde groups are formed.
Formed aldehyde groups react with the Schiff reagent to form a bright red or magenta colour end product.
Haematoxylin or methyl green is used to stain nuclei of the cells.
Procedure
Solutions necessary
1% Periodic acid solution, Schiff reagent and Carazzi’s Haematoxylin is necessary for the staining procedure.
First weigh 1 g of periodic acid and dissolve them in 100ml of distilled water for the preparation of 1% periodic acid solution.
Then prepare Schiff reagent. For preparation weigh 1 g of basic fuchsin and 1.9 g of sodium metabisulfite (Na2S2O5).
Dissolve them in 100 ml of 0.15 M HCl (Hydrochloric acid solution).
Shake the solution well. For this purpose, a mechanical shaker also can be used.
Keep the solution on mechanical shaker for 2 hours.
Then observe the solution. The solution should be clear and the colour should be yellow to light brown.
Then add 500 mg of activated charcoal as decolorizing agent. Shake the solution well for 1-2 minutes.
By using a No 1 Whatman filter paper, filter the solution into a clean bottle.
This solution should be clear and colourless.
If the solution is still yellow, repeat the charcoal decolorization procedure. Store the solution at 4°C.
Staining method
- First dewax the tissue section and bring it down to the water for the hydration of the tissue section.
- Then treat with 1% Periodic acid solution for 5-10 minutes.
- Then wash with water for several times.
- Next stain with Schiff’s reagent for 2-3 minutes.
- Then the section should be washed in running tap water for 5-6 minutes.
- Stain with Carazzi’s Haematoxylin for 3 minutes.
- Wash in tap water and blue the sections in running tap water for 5 minutes.
- Finally dehydrate, clear and mount with DPX.
Purpose
It is useful in the identification of carbohydrates and carbohydrate components or glycoconjugates in tissues.
It shows polysaccharides, sialo mucin, neutral mucin, glycogen, basement membrane, α-antitrypsin, reticulin, fungal cell wall components, pancreatic zymogen granules, thyroid colloid, corpora amylacea, and Russell bodies in tissues.
Results
Glycogen, neutral mucins, sialo mucins are stained in magenta colour.
Various glycoproteins are also stained in magenta colour. Nuclei in cells stains in blue colour.
The staining procedure can be differed according to the condition of the stain.
Time of staining should be adjusted accordingly.
It is necessary to use a positive control slide.
The staining of the tissue section is dependent to some extent on the time of the treatment with the periodic acid and Schiff reagent.
When staining basement membrane, it is necessary to stain the sections more time.
It should be adjusted accordingly.
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