Dec 01, 2023

Public workspaceHematoxylin and Eosin Stain - FOR PARAFFIN-EMBEDDED TISSUE SECTIONS

DOI
dx.doi.org/10.17504/protocols.io.kxygx3q9zg8j/v1
  • Michael X. Henderson1
  • 1Van Andel Institute
Michael Henderson
Open access
DOI: dx.doi.org/10.17504/protocols.io.kxygx3q9zg8j/v1
Protocol Citation: Michael X. Henderson 2023. Hematoxylin and Eosin Stain - FOR PARAFFIN-EMBEDDED TISSUE SECTIONS. protocols.io https://dx.doi.org/10.17504/protocols.io.kxygx3q9zg8j/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: November 30, 2023
Last Modified: May 31, 2024
Protocol Integer ID: 91708
Keywords: ASAPCRN
Funders Acknowledgement:
Aligning Science Across Parkinson's
Grant ID: ASAP-020616
Abstract
This protocol details Hematoxylin and Eosin Stain for paraffin-embedded tissue sections.
Attachments
911-2361.pdf
911-2361.pdf
81KB
Guidelines
Results:

1. Nuclei are stained blue. 2. Cytoplasm stained various shades of pink, identifying different morphologic tissue components.
Materials
Reagents


Stock Solutions:

1% stock Eosin Eosin Working Solution

AB
CompoundQuantity
Eosin Y 1 gm
80% EtOH100 ml


Eosin Working Solution

AB
CompoundQuantity
Stock Eosin45 ml
80% EtOH 135 ml
Glacial Acetic Acid 900 ul


0.1% Acid Alcohol Solution

AB
CompoundQuantity
ddH2O 500 ml
100% EtOH500 ml
HCl1 ml


ReagentEpredia™ Shandon™ Harris HematoxylinFisher ScientificCatalog #6765001

Hematoxylin and Eosin Stain
Hematoxylin and Eosin Stain
21m
Deparaffinize slides and hydrate to ddH2O:

AB
Compound Time
Xylenes2 x 5 min
100% EtOH2 x 1 min
95% EtOH1 min each
80% EtOH1 min each
70% EtOH1 min each
ddH2O1 min each

Filter undiluted Harris Hematoxylin and Eosin using Whatman #4 filter paper.
Immerse sections in filtered Harris Hematoxylin for Duration00:05:00 .

5m
Rinse in 2 changes of ddH2O.
Wash
Differentiate in 0.1% acid alcohol solution:

AB
Compound Time
EtOH fixed tissue2 sec
NBF fixed tissue4 sec

Rinse in tap H2O for 10- Duration00:15:00 or until sections turn bright blue.

15m
Wash
Immerse in Eosin for Duration00:01:00 .

1m
Briefly rinse in tap H2O.
Wash
Dehydrate in:

AB
Compound Time
95% EtOH2 x 30 sec
100% EtOH 2 x 1 min
Xylene 2 x 5 min

Coverslip with Cytoseal.