Hematoxylin & Eosin (H&E) 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. Stain slides in Harris Hematoxylin————–  10 minutes (re-use)
  8. Rinse slides in running water———————  2 minutes
  9. Differentiate in acid alcohol———————-   1 quick dip (re-use)
  10. Rinse in water————————————–    3 dips
  11. Dip in ammonia water—————————-    15-30 seconds (re-use)
  12. Wash in running tap water———————–    10 minutes
  13. Place in 95% alcohol——————————    2 minutes
  14. Stain in eosin—————————————    1 minute (re-use)
  15. Dehydrate in 95% alcohol————————   1 minute
  16. Dehydrate in 100% alcohol———————-   1 minute
  17. Dehydrate in 100% alcohol———————-   2 minutes
  18. Dehydrate in 100% alcohol———————-   2 minutes
  19. Clear in xylene (3)———————————   2 minutes (re-use)
  20. Clear in xylene (4)———————————   5 minutes (re-use)

Results: Nuclei –Blue   Cytoplasm – Pink     Blood – Red


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