Monday, 13 June 2011
Periodic-Acid Schiff (PAS) Special Stain – Method and Tips
The Periodic-Acid Schiff (PAS) technique (and its numerous variations) is by far the most commonly performed special stain within the histopathology laboratory, therefore knowledge of its method is a vital arrow in any medical scientist’s quiver of knowledge.
The PAS technique is most commonly used to highlight molecules with a high percentage carbohydrate content such as mucins, glycogen, fungi and the basement membrane in skin.
The PAS method works by exposing the tissue to periodic acid. This acts an oxidizing agent which oxidizes vicinal (neighbouring) glycol groups or amino/alkylamino derivatives. This oxidation creates dialdehydes.These dialdehydes when exposed to Schiff’s reagent create an insoluble magenta compound which is similar to the basic fuchsin dye within the Schiff’s reagent.
1% aqueous periodic acid
1. Take sections to water.
2. Expose sections to periodic acid solution for 10-15 mins.
3. Rinse well in tap water.
4. Expose sections to Schiff’s reagent for 10-15 mins.
5. Wash in running tap water for 5-10 mins.
6. Counterstain with a haemtoxylin for approx. 15 secs.
7. Differentiate (if necessary) and blue.
8. Dehydrate, clear and mount.
- Periodic acid and Schiff’s reagent are easily available commercially prepared, the technique for self-made Schiff’s reagent is arduous by comparison but can be found.
- Keep your Schiff’s reagent out of UV light and refrigerated when not in use. Failure to do so will result in the loss of sulphur dioxide in your Schiff’s reagent leading to the solution turning from colourless to a magenta colour resembling the original basic fuchsin colour. When this happens replace your solution. Also keep your periodic acid solution refrigerated when not in use.
- The purpose of washing in running tap water after exposing the sections to Schiff’s is to intensify the magenta colour. This author has found that when the water has runs from a magenta colour to a clear colour the colour isn’t going to intensify any further therefore the washing in running water can be ceased. This may vary from lab to lab.
- There are numerous variations of the PAS technique (eg. PAS + diastase, PAS + Alcian Blue). This will be discussed in a further blog post.
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