Pas staining

it is useful for pathology lab

1. PAS staining Dr.Sundaresan P athology

2. What Does PAS Stain? (1) Polysaccharides : The main polysaccharide identified via histology staining in human and animal tissue sections is glycogen. This is present in numerous tissues, including skeletal muscle, cardiac muscle, liver, and kidney . (2) Neutral mucus substances:   These can include glycoproteins, glycolipids, and neutral mucins , which are produced by epithelial cells in different organs . (3) Tissue basement membranes: These PAS-positive thin layers of reticular connective tissue anchor and support epithelium and endothelium to underlying connective tissue. (4) Fungal organisms: The cell walls of some fungal organisms contain high levels of carbohydrate , and also stain PAS-positive. However, this only works on living fungi.

3. General Principles of the Stain The first reaction -periodic acid acting as an oxidising agent to oxidise the carbon-to-carbon bonds between two hydroxyl groups . This produces Schiff reactive aldehyde groups. second reaction- This comprises a mixture of basic fuchsin , hydrochloric acid, and sodium metabisulphite . The basic fuchsin in the mixture reacts with newly formed aldehyde groups in the tissue to produce a bright magenta colour . Finally, when the section is rinsed in water, bound fuchsin molecules in the tissue then produce a bright magenta colour . Haematoxylin is then typically used as a counter stain to visualise other tissue elements . When PAS is used to demonstrate fungal organisms, however, a light green counter stain is preferred .

4. PROCEDURE : 1 . Deparaffinize and hydrate to distilled water. 2. Place slides into 1% Periodic acid for 10 minutes. 3. Rinse in tap water. 4. Schiff's Reagent for 15-30 minutes. 5. Wash in running tap water for 5 minutes. 6. Counterstain in hematoxylin for 5 minutes. 7. Wash in tap water, differentiate,blue and wash in distilled water. 8. Dehydrate in alcohol, clear, and coverslip.

5. Who Uses PAS Stain? Glycogen storage diseases: (liver -(hepatocytes,  alpha-1-antitrypsin disease ,) muscles- ( demonstrate glycogen), or kidney- ( renal cell carcinoma  ). Tumors : Glycogen granules can also be present (pancreas-( acinar cell carcinoma), lung-( alveolar proteinosis ), and bladder). Adenocarcinomas , which often secrete neutral mucins . Alveolar soft part sarcoma . [2] Staining  macrophages  in  Whipple's disease . [3]   α1- antitrypsin deficiency  if periportal liver hepatocytes stain positive . Erythroleukemia , a leukemia of immature red blood cells. These cells stain a bright fuchsia. Pulmonary alveolar proteinosis . Fungal infection.  the cell walls of fungi stain magenta ;(

6. Breast cytology:  PASD positive cells with structure and producing nuclear indentation, particularly in dissociated or atypical cells, correlate with malignant histology. Fungi:  stains fungal cell walls; PAS+ granule, spores is diagnostic of microsporidia . Hematopathology :   ALL Kidney:   renal cell carcinoma (stains glycogen, removed by diastase) Liver:  routine stain for hepatocytes , alpha-1-antitrypsin disease Lung:   alveolar proteinosis Muscle biopsies:   demonstrate glycogen Pancreas:   acinar cell carcinoma (PASD+) Parotid glands:  zymogen granules are PAS+ Prostate:  Cowper’s glands are PASD + Skin:   eosinophilic globoid bodies ( Kamino bodies) in Spitz nevus are PASD+ Small intestine:  stains Whipple’s disease bacteria Testis :  stains intratubular germ cell neoplasia and seminoma (PAS+, PASD negative) Tumors :   adenocarcinoma of various sites ( mucin is PASD+), alveolar soft parts sarcoma (PASD+ crystalline structures), clear cell tumors (stains glycogen), glycogen rich carcinomas , balloon cell melanoma , granular cell tumor (cytoplasmic granules ).

7. Histologic feature Glycogen , basement membranes, mucus substances, fungal organisms . Nuclei Other tissue elements Colour with PAS Stain Magenta Blue (with haematoxylin counter stain ) Green (with light green counter stain)

8. Periodic Acid – Schiff [PAS] Reaction Giant multinucleate late normoblasts (left). Granular PAS positivity in proerythroblasts and homogeneous positivity in the later normoblasts

9. alpha-1-antitrypsin deficiency

10. Diagnosis moderately differentiated adenocarcinoma Description Dysplastic epithelium with little  mucus formation.

11. granular cell tumor

12. Positive PAS stain acute megakaryocytic leukemia AML, M7. Positive PAS stain in ALL PAS positivity in M6. Not the intense staining of the large abnormal erythroblast.