General pathology · The pathology laboratory
Special stains in diagnosis: Congo red, Prussian blue, PAS, Ziehl–Neelsen, Gram !
- Technique
When H&E is not enough, a chemical stain gives amyloid, iron, glycogen or microbes a colour of their own.
Updated 2026-11-01
What it is
Special stains are chemical reactions that selectively colour a substance or a microorganism in the section. They are cheap and fast and remain essential to confirm deposits (amyloid, iron, copper, glycogen, mucin), fibrosis and infectious agents (mycobacteria, fungi, bacteria). Unlike immunohistochemistry, they use no antibodies.
Step by step
- H&E shows something that cannot be identified: amorphous pink material, a brown pigment or a granuloma.
- The pathologist asks what it could be: amyloid, iron, a microorganism, mucin.
- The stain that answers that question is chosen and run with a positive control.
- A positive result confirms; a negative one in a granuloma does not exclude infection, because bacilli may be scarce.
- If needed, culture, PCR or immunohistochemistry complete the work-up.
Special stains: what they colour and in which disease
| Stain | What it colours | How it looks | Clinical example |
|---|---|---|---|
| Congo red | Amyloid | Orange-red; apple-green under polarised light | Amyloidosis |
| Prussian blue (Perls) | Iron (haemosiderin) | Blue granules | Hereditary haemochromatosis |
| PAS | Glycogen, neutral mucin, basement membranes, fungi | Magenta | Whipple disease: PAS-positive macrophages |
| PAS with diastase | What remains after glycogen is digested | Resistant magenta | Separates glycogen from mucin or other deposits |
| Ziehl–Neelsen | Acid-fast bacilli | Red bacilli on a blue background | Tuberculosis |
| Fite (modified Ziehl–Neelsen) | Weakly acid-fast bacilli | Red bacilli | Leprosy |
| Gram | Bacteria | Gram-positive violet, Gram-negative pink | Abscesses and other bacterial infections |
| Grocott methenamine silver | Fungal walls | Black fungi | Pneumocystis pneumonia |
| Mucicarmine | Cryptococcus capsule | Red capsule | Cryptococcal meningitis |
| Giemsa | Helicobacter, intracellular parasites | Blue-violet curved rods or amastigotes | Leishmaniasis |
| Masson trichrome | Collagen | Collagen blue or green, muscle red | Cirrhosis |
| Rhodanine | Copper | Reddish granules | Wilson disease |
| Alcian blue | Acid mucin | Blue | Barrett oesophagus: goblet cells |
| Silver (Warthin–Starry) | Spirochaetes | Black spirals | Syphilis |
Clinical notes
- In Colombia tuberculosis and leprosy are still present: Ziehl–Neelsen and Fite stains on skin and lymph node biopsies keep their value.
- Amyloidosis: an abdominal fat pad or rectal biopsy stained with Congo red is a minimally invasive way to confirm it.
- Haemosiderin in the liver with Prussian blue: its pattern (in hepatocytes or in macrophages) points to hereditary or secondary overload.
- Fat dissolves during processing: staining it (oil red O) needs frozen tissue.
Test yourself
Think of the answer, then tap the question to see it.
Which finding under polarised light confirms amyloid on a Congo red stain?
Apple-green birefringence.
Which stain would you use to look for haemosiderin in a liver biopsy?
Prussian blue (Perls), which stains the iron of haemosiderin blue.
Which special stain shows the characteristic macrophages of Whipple disease?
PAS: the lamina propria macrophages are packed with PAS-positive, diastase-resistant material.
Why can fat not be stained in a paraffin section?
Because the solvents used in processing dissolve lipids; frozen tissue is used to stain fat.
Related
- Immunohistochemistry: the principle and the markers to know
- Intracellular accumulations: cholesterol, proteins, hyaline, glycogen
- Pigments in tissue: lipofuscin, hemosiderin, melanin, bilirubin, carbon
- Granulomatous inflammation: caseating vs non-caseating, the causes to know
- Pathologic calcification: dystrophic vs metastatic
- Fatty change (steatosis)
Related conditions
- Amyloidosis
- Hereditary hemochromatosis
- Tuberculosis
- Pulmonary tuberculosis
- Leprosy (Hansen disease)
- Whipple disease
- Pneumocystis pneumonia (PCP)
- Histoplasmosis
- Cryptococcal meningitis
- Wilson disease
- Cirrhosis
- Barrett oesophagus
- Peptic ulcer disease
- Syphilis
- Leishmaniasis
- Multiple myeloma
- Nontuberculous mycobacterial infections (MAC, M. abscessus, Buruli ulcer)
- Abscess, furuncle and carbuncle
Elsewhere in DurgiPedia
- Microscopy and stains: H&E, PAS, trichrome, silver, immunohistochemistry
- From biopsy to slide: fixation, sectioning, frozen section
- Iron metabolism
- Glycogen
- Collagen
Microorganisms
- Mycobacterium tuberculosis
- Mycobacterium leprae
- Pneumocystis jirovecii
- Histoplasma capsulatum
- Cryptococcus neoformans
- Helicobacter pylori
- Treponema pallidum
- Leishmania (Viannia) panamensis, L. braziliensis and L. guyanensis
Educational content on the mechanisms of disease; not a diagnostic or treatment guide and no substitute for a health professional's advice. No doses are given.
Sources: Gram Stain: MedlinePlus Medical Test (MedlinePlus, US National Library of Medicine, 2024); Amyloidosis: MedlinePlus (MedlinePlus, US National Library of Medicine, 2024); Hemochromatosis: MedlinePlus Medical Encyclopedia (MedlinePlus, US National Library of Medicine, 2024); Tuberculosis Overview (StatPearls (NCBI Bookshelf), 2024).
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Representative model, for educational purposes only.