Buck Institute Morphology H & E staining protocol
Stella Breslin
Abstract
Histological Stain for FFPE slides
RESULTS: Nuclei-blue; Cytoplasm, red blood cells and connective tissue-shades of pink
Before start
Slides can be placed in 60 C oven prior to deparaffinization.
Steps
Hematoxylin and Eosin Staining
Deparaffinization and rehydration: 2 x xylene 7 min, 100% ethanol 4 min, 95% 4 min, 80% 4 min, 70% 4 min, bring to water.
Hematoxylin staining for 30 seconds - 4 min. using filtered dye (usually 2 mins) - Modified Mayes’s Hematoxylin HXMMHPT StatLab Medical Products
Rinse in ddH20 x2
10% acetic acid rinse (acid into water) – 1 min
rinse in ddH20 x2
TBS (1x) rinse or Scott’s Tap Water (1L + 3.5g Sodium Bicarb + 20g Magnesium Sulfate pH 8.0) - 2 mins
Rinse in ddH20
Rinse in 70% ETOH - 1 min
Stain Eosin for 0.5 to 2 minutes (keep an eye on intensity of pink) – Eosin Y Stain STE0150 StatLab Medical Products
Rinse in 90% EtOH
Rinse in 2 x 95% EtOH, 2 x 100% EtOH, xylene
Coverslip with Permount Mounting Medium EMS #17986-01
