Histopathology - special stains

The Alcian Blue technique is used to demonstrate of mucin and also mast cells. The alcian blue stain is most commonly used on tissue samples obtained from the gastrointestinal (GI) tract This technique can be used in conjunction with other staining methods such as Haematoxylin and Eosin, Van Giesons and Periodic Acid Schiffs. Acidic mucoploysaccharides are stained blue by the Alcian Blue technique, ie mucus, cartilage etc.

Congo red is used for staining in amyloidosis, and for the cell walls of plants and fungi, and for the outer membrane of Gram-negative bacteria. Apple-green birefringence of Congo red stained preparations under polarized light is indicative of the presence of amyloid fibrils.

Giemsa method can be used to stain impression smears, blood films and tissue sections. It is commonly used to demonstrate micro-organisms.

The Gomoris Grocott Hexamine Silver technique is used for the demonstration of fungi and basement membrane. The majority of fungi are Periodic Acid Schiffs +ve, but the Gomori”s Hexamine Silver technique is widely used. Some fungi can also be stained by the Gram methods, with hyphae staining Gram positive and spores staining Gram negative.

PAS positive structures, Basement membranes, fungi, Pneumocytis carinii, chitin, glycogen and starch. Black/brown. Mucin Grey to rose. Background Light green

Gram positive organisms are deep purple/blue, Gram negative organisms are magenta/pink, and the background is green.

Luxol fast blue stain (LFB) is used to visualize myelin in nerve tissue. Staining with LFB makes it possible to visualize the myelin and the fatty substance of the brain called white matter which includes the axons of the nerve cells. This stain is often combined with PAS and Haematoxylin. The above two pictures show LFB without counterstain (Blue and white) and LFB with PAS counterstain (Blue and pink).

Myelin and erythrocytes are blue, and the background is pink.

As the name implies three stains (tri) are used selectively staining muscle collagen fibres, fibrin and erythrocytes types of tissue. Standard applications: Masson's trichrome staining is widely used to study muscular pathologies (muscular dystrophy), cardiac pathologies (infarct), hepatic pathologies (cirrhosis) or kidney pathologies (glomerular fibrosis).

Nuclei are purple/black; Muscle fibres, erythrocytes cytoplasm and fibrin are red; and Connective tissue and collagen and green.

The Modified ZN (MZN) technique is used by this laboratory to positively identify Chlamydia.

Chlamydia comes up as Magenta, Nuclei green and other tissue constituents are green.

The Periodic Acid Schiff’s Technique is performed by this laboratory for the demonstration of fungi, and carbohydrates particularly glycogen. PAS staining is mainly used for staining structures containing a high proportion of carbohydrate macromolecules (glycogen, glycoprotein, proteoglycans), typically found in e.g. connective tissues, mucus, the glycocalyx, and basal laminae.

Hyphae come up as dark pink.

The Perles Prussian Blue technique is used by this laboratory for the demonstration of Haemosiderin (ferric iron). Deposits of ferric iron in haematoxylin and eosin-stained sections appear as golden green, coarsely to finely granular, intracellular material known as haemosiderin.

Haemosiderin (ferric iron) is blue, and the background/nuclei is red.

The Rhodanine technique is used by this laboratory for the demonstration of copper deposits. The reaction utilizes the property of copper to bind with high affinity to protein copper deposits in tissue sections.

Copper comes up as bright red.

Toluidine blue is used for staining mast cells and Nissli substance. Use of toluidine blue in tissue sections is done with the aim to highlight components, such as mast cells granules, mucins, and cartilage.

Orthochromatic colour, Nissli substance, nuclei and cytoplasm are blue, Metachromic colour is red, and Granules of mast cells are purple.

It is the simplest method of differential staining of collagen and other connective tissue. Used often to demonstrate muscle.

Nuclei come up as black/brown; Collagen and connective tissue come up as red; and Cytoplasm, smooth and striated muscle, keratin, erythrocytes come up as yellow.

The Von Kossa staining technique is used to stain Calcium salts that are normally present in bone and teeth, and in some pathological conditions, deposits of calcium that are formed in tissues normally devoid of them.

Nuclear chromatin, cytoplasm, collagen comes up as blue/purple; Keratin and erythrocytes come up as pink and Calcium deposits come up as black.

This is a silver technique for the demonstration of spirochaetes in paraffin sections. Warthin-Starry (WS) staining is an ancillary stain used in the detection of Helicobacter sp., spirochaete and other microorganisms in tissue sections.

Spirochaetes and other bacteria come up as black and background tissues come up as yellow/brown.

The Ziehl Neelson (ZN) technique is used to positively identify Acid fast mycobacteria. This stain is used as a routine confirmation stain for Johnes disease in sheep and cattle but is also used to identify the presence of Tuberculosis.

Acid-fast bacilli come up as magenta coloured, and Nuclei and other tissue components come up as blue.