Posted on Leave a comment

PAS Diastase Stain Protocol

If you are going to do a PAS stain then you might as well run a PAS with diastase also.

Safety equipment: Work under a hood with lab coat, gloves, and glasses.

  1. Two sets of slides and 2 control slides are used
  2. De-paraffin slides in xylene (1) —————— 2 minutes (re-use)
  3. De-paraffin slides in xylene (2)—————— 2 minutes (re-use)
  4. Clear slides in 100% alcohol——————— 2 minutes
  5. Clear slides in 100% alcohol——————— 2 minutes
  6. Hydrate slides in 95% alcohol——————- 2 minutes
  7. Hydrate slides in running water—————– 5 minutes
  8. Set 1 (with a control) in 80% alcohol ———— 60 minutes
  9. Set 2 (with a control) in diastase of malt——— 60 minutes
  10. Combine both sets of slides for remaining steps
  11. Wash in running water ————————– 10 minutes
  12. 0.5% periodic acid ——————————- 5 minutes (re-use)
  13. Schiff’s reagent at 37C————————— 15 minutes (re-use)
  14. Wash in running water ————————– 10 minutes
  15. If staining for glycogen, Hematoxylin ———— 20 seconds (re-use)
  16. Dehydrate in 95% alcohol———————— 30 seconds
  17. Dehydrate in 100% alcohol———————- 1 minute
  18. Dehydrate in 100% alcohol———————- 2 minutes
  19. Dehydrate in 100% alcohol———————- 2 minutes
  20. Clear in xylene (3)——————————- 2 minutes (re-use)
  21. Clear in xylene (4)——————————- 5 minutes (re-use)

Results:         Glycogen, mucin, fibrin, or thrombi, colloid droplets, hyaline of arteriosclerosis, hyaline deposits in glomeruli, granular cells in the renal arterioles where preserved, most basement membranes, colloid of pituitary stalks and thyroid, amyloid infiltration may show a positive reaction – Rose to Purplish Red

                        Nuclei – Blue

 

An Expert Histology Service

Posted on Leave a comment

Periodic Acid Schiff (PAS) Stain Protocol

Safety equipment: Work under a hood with lab coat, gloves, and glasses.

  1. De-paraffin slides in xylene (1) —————— 2 minutes (re-use)
  2. De-paraffin slides in xylene (2)—————— 2 minutes (re-use)
  3. Clear slides in 100% alcohol——————— 2 minutes
  4. Clear slides in 100% alcohol——————— 2 minutes
  5. Hydrate slides in 95% alcohol——————- 2 minutes
  6. Hydrate slides in running water—————– 5 minutes
  7. 0.5% periodic acid ——————————– 5 minutes (re-use)
  8. Schiff’s reagent at 37C—————————- 15 minutes (re-use)
  9. Wash in running water ————————— 10 minutes
  10. If staining for glycogen, Hematoxylin ——— 20 seconds (re-use)
  11. If staining for fungus, light green ————— 30 seconds (re-use)
  12. Dehydrate in 95% alcohol———————— 30 seconds
  13. Dehydrate in 100% alcohol———————- 1 minute
  14. Dehydrate in 100% alcohol———————- 2 minutes
  15. Dehydrate in 100% alcohol———————- 2 minutes
  16. Clear in xylene (3)——————————— 2 minutes (re-use)
  17. Clear in xylene (4)——————————— 5 minutes (re-use)

Results:           Hematoxylin – Glycogen, mucin, fibrin, or thrombi, colloid droplets, hyaline of arteriosclerosis, hyaline deposits in glomeruli, granular cells in the renal arterioles where preserved, most basement membranes, colloid of pituitary stalks and thyroid, amyloid infiltration may show a positive reaction – Rose to Purplish Red

                        Nuclei – Blue

Results:          Light Green – Fungi – Red                 Background – Pale Green

 

 

An Expert Histology Service

Posted on 4 Comments

Special stains protocols

I am trying to figure out what to do about the special stains protocols.  There are about 35 of them.  Putting them on the front page does not seem to be efficient.   I would like them all to be searchable and downloadable, maybe on a different page.  What do you think, does it make a difference?

Posted on 3 Comments

Alcian Blue / PAS Stain Protocol

Here is the protocol for alcian blue periodic acid schiff  (AB /PAS).  At the bottom of the post is the link for printing.

Control tissue: Large Intestine

Rat colon PASAB2

  1. De-paraffin slides in xylene (1) ———— 2 minutes (re-use)
  2. De-paraffin slides in xylene (2)———— 2 minutes (re-use)
  3. Clear slides in 100% alcohol————— 2 minutes
  4. Clear slides in 100% alcohol————— 2 minutes
  5. Hydrate slides in 95% alcohol————- 2 minutes
  6. Hydrate slides in running water———-  5 minutes
  7. Place slides in 3% Acetic Acid————- 3 minutes (re-use)
  8. Alcian Blue ———————————– 30 minutes (re-use)
  9. Wash in running water ——————— 10 minutes
  10. Oxidize in 0.5% periodic acid ————- 10 minutes (re-use)
  11. Wash in running water ——————— 5 minutes
  12. Schiff reagent (room temp)—————- 20 minutes (re-use)
  13. Wash in running water ——————— 10 minutes
  14. Dehydrate in 95% alcohol—————— 30 seconds
  15. Dehydrate in 100% alcohol—————- 1 minute
  16. Dehydrate in 100% alcohol—————- 2 minutes
  17. Dehydrate in 100% alcohol—————- 2 minutes
  18. Clear in xylene (3)————————— 2 minutes (re-use)
  19. Clear in xylene (4)————————— 5 minutes (re-use)

Results: Polysaccharides and Mucosubstances containing hexoses or deoxyhexoses with vicinal glycol groups – Magenta to Red

Hyaluronic acid, sialomucins and all but the most strongly acidic sulfated mucosubstances – Blue

PAS Alcian Blue