H&E staining
Show general tissue architecture, with nuclei blue and cytoplasm pink.
Our ruleSimulations prepare you for the bench but don't replace hands-on practice.
1Two dyes, one picture of the tissue
Haematoxylin and eosin is the stain every other histological stain is compared against. Haematoxylin, oxidised to haematein and combined with aluminium as a mordant, forms a positively charged dye–metal complex that binds the phosphate groups of nucleic acids: chromatin, nucleoli and RNA-rich cytoplasm turn blue to purple.
2Regressive staining: overstain, then take it out
In a regressive method such as Harris haematoxylin, the section is deliberately overstained so every nucleus is saturated, and then briefly differentiated in acid alcohol. The acid breaks the dye–tissue bond, but much faster where the dye is held weakly (cytoplasm, matrix) than in chromatin, so a few seconds leave crisp nuclei on a clean background.
Red-purple
The haematoxylin–aluminium lake is reddish in acid, which is how it comes out of the differentiator.
Bluing
Scott's tap water substitute or dilute ammonia converts it to the insoluble blue form; distilled water does not.
Three shades
Red cells brightest, cytoplasm intermediate, collagen palest: a sign the eosin is balanced.
Eosin washes out
Eosin is soluble in the lower alcohols, so dehydration must be quick.
3What goes wrong around the stain
Many bad H&E slides have perfectly good timings. Paraffin left on the section blocks the water-based dyes and leaves pale blotches.
Key terms
Practise these as flashcards →A metal ion (aluminium for Harris haematoxylin, iron for Weigert's) that links the dye to the tissue.
Deliberate overstaining followed by removal of the excess with a differentiator.
Selective removal of excess dye, here with 0.5–1% hydrochloric acid in 70% ethanol.
Converting the red, soluble haematoxylin lake to the blue, insoluble form in a weak alkali.
Stained by the basic dye (haematoxylin-like, blue) or by eosin (pink).
A second stain in a contrasting colour that shows the tissue around the main target.
When to use it
- First look at any tissue section: architecture, cell types, inflammation, necrosis and tumour.
- Checking fixation, processing and sectioning quality before special stains or immunohistochemistry.
- Scoring histological changes in an animal study against a reference section.
Compared with alternatives
When H&E cannot tell collagen from muscle, or glycogen, mucin, mineral or lipid from cytoplasm.
When the question is about a specific protein, not general morphology.
Section faults such as compression, chatter and folds are usually first seen on the H&E.