PAS diastase stain

PAS diastase stain, also known as Periodic acid-Schiff with diastase stain, is a histochemical staining method used in pathology and histology to identify complex carbohydrates in tissues. This technique is particularly useful for highlighting structures such as glycogen, mucosubstances, and fungal cell walls. The PAS diastase stain combines the Periodic acid-Schiff (PAS) reaction with diastase treatment to specifically target glycogen, making it a critical tool in both diagnostic pathology and research.

Overview

The PAS diastase stain procedure involves the application of periodic acid, which oxidizes the carbon-carbon bonds in carbohydrates, creating aldehyde groups. These aldehyde groups then react with Schiff reagent, resulting in a magenta color. The addition of diastase, an enzyme that digests glycogen, before the PAS reaction allows for the differentiation between glycogen and other PAS-positive substances. Structures that lose their magenta color after diastase treatment are identified as glycogen, whereas those that retain the color are typically other types of polysaccharides or mucosubstances.

Applications

The PAS diastase stain is widely used in the diagnosis of various diseases and conditions. In Liver pathology, it helps in identifying glycogen storage diseases by demonstrating abnormal glycogen accumulation within hepatocytes. In Dermatopathology, it is used to detect fungal infections, as the cell walls of fungi are rich in polysaccharides that are PAS-positive. The stain is also employed in the diagnosis of Alimentary tract pathology, particularly in identifying conditions like Whipple's disease, where PAS-positive macrophages are a hallmark feature.

Procedure

The staining procedure involves several steps:

1. Fixation of the tissue sample, typically in formalin.
2. Embedding the tissue in paraffin and sectioning.
3. Deparaffinization and hydration of tissue sections.
4. Treatment with diastase enzyme to digest glycogen.
5. Oxidation with periodic acid.
6. Reaction with Schiff reagent.
7. Counterstaining, often with hematoxylin, to provide a background contrast.
8. Dehydration, clearing, and mounting of the tissue sections for microscopic examination.

Interpretation

Under the microscope, PAS-positive substances appear magenta or purple, while the background and other tissue components are counterstained blue by hematoxylin. The presence or absence of magenta staining after diastase treatment helps in identifying glycogen versus other substances.

Limitations

While the PAS diastase stain is a powerful tool, it has limitations. The specificity of the stain can be affected by the fixation and processing of the tissue. Overdigestion or underdigestion with diastase can lead to false-negative or false-positive results, respectively. Therefore, careful control of staining conditions and interpretation by experienced pathologists is essential.

See Also

• Periodic acid-Schiff reaction
• Histology
• Pathology
• Glycogen storage disease
• Fungal infections