The periodic acid-Schiff (PAS)

technique

The most versatile and

widely used technique for the demonstration of
carbohydrates
or glycoconjugates. The first histochemical
use of this technique was by McManus (1946)
for the demonstration of mucin.
 or
Periodic acid–Schiff (PAS) is a staining method
used to detect polysaccharides such as
glycogen, and mucosubstances such as
glycoproteins, glycolipids and mucins in
tissues.
 PAS reactive - cell &tissue
 Glycogen
 Starch
 Mucin- sialomucin n neutral mucin
 Alpha- antitrypsin
 Reticulin
 Fungi capsules
 Pancreatic zymogen granules
 Thyroid colloid
 Corpora amylacea & Russell bodies
 PRINCIPLE:
• The PAS stain is a histochemical reaction in
that the periodic acid oxidizes the carbon to
carbon bond forming aldehydes which react to
the fuchsin-sulfurous acid which form the
magenta color. ... Use skin, aorta or normal
liver for positive PAS staining.
 Why does PAS stain basement membrane?

• The periodic acid-
Schiff (PAS) staining procedure is most
commonly used in the histology laboratory to
detect glycogen deposits in the liver when
glycogen storage
disease is suspected. ... Basement
membranes, which are present in various
tissues in the body, may also be visualized
through the PAS staining procedure.
 What Colour does PAS stain?

• magenta
• PAS stains carbohydrates and carbohydrate
rich macromolecules a deep red colour
(magenta). Mucus in cells and tissues,
Basement membranes, and Brush borders of
kidney tubules and small and large intestines
Reticular fibres (i.e. collagen) in connective
tissue and Cartilage.
• CONTROL: For staining fungus; use a known
positive such as those used for the GMS. Use
skin, aorta or normal liver for positive PAS
staining.
• FIXATIVE: Any well fixed tissue.
• TECHNIQUE: Cut paraffin sections 4-5 m( 3m for
kidney biopsies).
• EQUIPMENT: Rinse glassware in DI water. Coplin
jar, microwave oven.
 How is Schiff's reagent prepared?
• Schiff reagent may be made by simply adding
1 g. fuchsin,.
• 1.9 g sodium metabisulfite to 100 ml.
• 0.15 N hydrochloric acid,
• shaking at intervals or mechanically for 2
hours,
• decolorizing with charcoal and filtering.
. • Caution: Avoid contact and inhalation
 SAFETY:
• Wear gloves, goggles and lab coat.
• Avoid contact and inhalation.
• Hydrochloric acid: strong irritant to skin, eyes
and respiratory system.
• Target organ effects via inhalation on skin,
respiratory, reproductive and fetal systems.
Corrosive.
 PROCEDURE:
1. Deparaffinize and hydrate to distilled
water.
2. Place slides into 0.5% Periodic acid for 5
minutes.
3. Rinse in distilled water.
4. *Schiff's Reagent, microwave HIGH
power, for 45 - 60 seconds, until deep
magenta.
5. Wash in running tap water for 5 minutes
6. Counterstain in hematoxylin for 3 minutes.
7. Wash in tap water, blue hematoxylin, rinse
in distilled water.
8. Dehydrate in alcohol, clear, and coverslip.
*
Conventional method: Schiff's Reagent, room
temperature for 30 minutes.
 Result- PAS staining
A positive test is indicated by a change from pale
yellow (colourless) to deep red-violet upon
addition of the aldehyde to the rea- gent.

Glycogen,neutral/sialomucin,glycoprotein-
magenta

Nucleus- blue
LM- liver;glycogen(PAS)160x
PM- lung adenocarcinoma(PAS);mucin
 PM- renal corpuscle;PAS

PM- renal corpuscle;PAS

Lung- aspergillus;PAS
PAS stain for glycogen in an alcoholic formalin fixed liver

Treatment with diastase Without diastase

 Diagnostic utility- PAS
 Glycogen storage disease
 adenocarcinoma
 Paget’s ds- breast
 Alveolar soft tissue sarcoma
 Alpha1 anti-trypsin deficiency
 Mycosis fungoides
 Fungal infection
 Carcinoma of bladder,kidney,liver,ovary,
 Ewings Sarcoma,seminoma,juvenile rhabdomyosarcoma
• usefulness of PAS staining in identification of
carbohydrates in normal and malignant lesions of
mammarygland and endocervical gland