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What is Tissue Histology?

Tissue histology is tissue morphology. This means that a histologist will examine all tissue samples grossly. All tissue has a definite “normal” appearance as seen inside and outside its normal environment. This tissue must be described as color, shape, and size before it can be processed for microscopic evaluation. This description (morphology) is part of the tissue histology. The second part is the microscopic description (morphology) of the tissue after it has been stained. The typical stain done on all tissues is the hematoxylin and eosin (H&E). This shows the basic structure of the tissue. All tissue components that are basophilic are stained with eosin and will have a pink hue. All the tissue components that are acidophilic are stained purplish blue with the hematoxylin. These are typically the nuclei that turn blue. This stain allows the identification of a particular tissue type, (muscle, lung, and so on). The examination and description of the microscopic structures constitutes the tissue histology.

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Oil Red O Stain Protocol

This is one of the stains done on frozen tissues for lipids or fat globules.

Safety equipment: Work under a hood with lab coat, gloves, and glasses.
Use on frozen sections only

1. Hydrate slides in running water———————– 10 minutes
2. Stain in working Oil Red O ————————— 10 minutes
3. Rinse in running water ——————————- 2 minutes
4. Rinse in 60% isopropanol —————————– 5 minutes
5. Rinse in running water ——————————- 2 minutes
6. Counterstain with Harris Hematoxylin —————– 20 seconds
7. Blue in running water ——————————– 5 minutes
8. Coverslip with glycerin jelly or aqueous mounting media

Results:
Fat Globules – Orange Red
Nuclei – Blue

Notes:
• For tissues that do not stay on slides, do not rinse slides in water prior to staining.
• Dextrin must be made for new working solution.
• Dextrin is only good for 1 month.

 

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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

 

 

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Congo Red Stain Protocol For Amyloid

Run a control slide.                                     Control tissue – Tissue with amyloid

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. 1% Congo red ————————————— 60 minutes (re-use)
  8. Rinse in water ————————————— 1 minute
  9. Alkaline Alcohol ————————————- until background clear (re-use)
  10. Wash in running water —————————- 5 minutes
  11. Harris Hematoxylin ——————————— 30 seconds (re-use)
  12. Wash in running water —————————- 10 minutes
  13. Dehydrate in 95% alcohol———————— 30 seconds
  14. Dehydrate in 100% alcohol——————— 1 minute
  15. Dehydrate in 100% alcohol——————— 2 minutes
  16. Dehydrate in 100% alcohol——————— 2 minutes
  17. Clear in xylene (3)——————————– 2 minutes (re-use)
  18. Clear in xylene (4)——————————– 5 minutes (re-use)
  19. Coverslip

Results:         Amyloid – Pink to red                     Nuclei – Blue

With polarizing microscope

Amyloid – Green Birefringence

 

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