Periodic acid-Shiff (PAS): Staining Protocol

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Periodic acid-Shiff (PAS): Staining Protocol

Sample type required: Deparaffinized and re-hydrated tissue sections on positively charged slides.
Fixative: 10% neutral buffered formalin (NBF)
Step
Reagent
Time
Technical Notes
1
Periodic acid:
0.5% solution
5 minutes

2
Distilled water
3 changes

3
Schiff reagent
15 minutes
Glutaraldehyde should be avoided as a tissue fixative since it is a dialdehyde that will react with this reagent and give false-positive PAS staining.
4
Potassium metabisulfite:
0.55% solution
1 minute in 2 changes
While some technicians omit this reagent step in the protocol with no problems, metabisulfite rinses may be necessary to remove any excess leucofuchsin left over after exposure to the Schiff reagent. Highly chlorinated water can cause these excess molecules to nonspecifically stain other tissue elements in the section.
5
Running tap water
Up to 10 minutes
Warm to hot running water develops the PAS stain and gives a more intense staining reaction than cold water.
Expected results:
The following elements will show positive PAS (magenta/rose red) staining:
  • Glycogen
  • Neutral mucins
  • Some epithelial mucins
  • Basement membranes
  • Fungal walls
Post staining procedure:
  • Tissue sections must be counterstained to fully recognize PAS-positive staining. This can be achieved with the use of either hematoxylin or fast green (see separate pages for specific guidelines).
  • After counterstain, tissue sections should be rinsed well in distilled water, dehydrated with 95% alcohol, followed by 100% alcohols, cleared with xylene (or xylene substitute), and cover-slipped.

PAS demonstrating goblet cells in small intestine