Hematoxylin & Eosin staining
Staining is one of most important process in which tissue sections are coloured by various techniques. Haematoxylin and Eosin is the most popular & widely used histological stain. So in this article we are going to discuss
- What is the H&E Staining
- Principle of H&E Staining
- when we use this stain?
- Advantages / Disadvantages etc …
As the name implies stain combination with 2 types of dies ,
Hematoxylin & Eosin. Hematoxylin is extracted from the hard wood of the tree Haematoxylin campechianum found in Mexico & Central America. this tree also call as , logwood or bloodwood .
Eosin is a ynthetic acidic dye. It use along with the Haematoxylin in staining to demonstrate the cytoplasm of different types of cells,connective tissues, muscles, RBCs etc. It stain basic structures in red or pink color.
This Eosin is available in market as
- Eosin Y: yellowish colour and it is alcohol and water soluble
- Eosin S: alcohol soluble
- Eosin B: blush in colour and water soluble
Principle of Hematoxylin & Eosin Stain
Haematoxylin is a basic dye which stain acidic components of the cell , such as nuclei in purple- black colour.
Eosin is a acidic dye which stain basic components such as cytoplasm, connective tissues in pink- orange colour.
However extracted hematoxylin is not a stain and it is necessary to undergo oxidation process to become a natural dye.
This oxidation results the product called haematin and it can be achieved by 2 ways.
(Neutral oxidation & Chemical oxidation )
When we consider neutral oxidation ,
- Natural oxidation- Can be occurred by exposing to the sunlight and air & this haematin retain its staining ability for a long period
Examples: Ehrlich’s, Delafield’s Haematoxylin
When we consider chemical oxidation ,
- Chemical oxidation can be done by using chemicals when preparing the stain.But remember this product retain the staining ability for short period.
Examples: Sodium Iodate in Mayer’s Haematoxylin,
Mercuric oxide in Harris Haematoxylin
Iodine in cole’s hematoxylin
However , The oxidized product haematin it self have poor nuclear staining properties. To enhance the dye-cell component binding, mordants are being used.
What are Mordants ???
Mordents are metallic ions such as , Tungsten , Molybdenus , Lead , Alumonium , Ion
Hematin is a week anionic basic dye , therefore it has weak affinity for acidic cellular structures …
Combine with a “mordant” , dye component get positive charge & increase affinity to selected tissue eliments. Cationic dye , metal complex – bind to the anionic nuclear chromatin.
Haematoxylin stains can be categorized based on the types of mordants used.
Such as :-
- Alum Haematoxylin which contain potash alum or ammonium
Examples: Mayer’s, Harris, Coles Haematoxylin
- Iron Haematoxylin
Examples: Weigert’s, Verhoeff’s Haematoxylin
- Tungston Haematoxylin
- Molybdenum Haematoxylin
- Lead Haematoxylin
What are the uses of Hematoxylin & Eosin Stain ?
Hematoxylin & Eosin is the most widely used stain in medical diagnosis and is often the gold standard.
As a example, when a pathologist looks at a biopsy of a suspected cancer, with Hematoxylin & Eosin.
Result of Hematoxylin & Eosin Stain
Nuclei , Calcium salts ,urates & Bacteria – stain in blue color
Also Cytoplasm, Connective tissues , erythrocytes – stain in shaded pink color.
Advantages of Hematoxylin & Eosin Staining
1 Provide excellent nuclear details – Outline , chromatin pattern & nucleoli
2 Provide Important information helpful for diagnosis – keratinization , oncocytes & psammoma bodies
3 not very expensive stain
4 can be used to diagnose a wide-range of histopathologic conditions.
Disadvantages of Hematoxylin & Eosin Staining
1 Long staining time
2 H&E staining does not always provide enough contrast to differentiate all tissues & cellular structures