Special Stains used in Histopathology and Biopsy

Stains used for special staining

1. SPECIAL STAINS
Lecture No. 4 Path 301

2. The “special stains” refer to a
large number of alternative
staining techniques that are
used when theH&E does
not provide all the
information the pathologist
or researcher needs

4. Fat stain
•Lipids cannot be demonstrated in routine tissue sections, as during processing the
tissue and clearing, lipids will be dissolved
•Lipids are routinely demonstrated in frozen sections or cryostat sections
•Special fat stains used are
•Oil Red O
•Sudan III
•Sudan IV
•Sudan Black

5. Thing to remember
For these lipid stains, frozen
formalin-fixed sections are
used instead of paraffin-
embedded sections

6. Photomicrograph of liver of goat with diffuse hepatic lipidosis showing fat droplets stained
orange yellow (Sudan III).

8. Periodic Acid Schiff (PAS)

11. Periodic acid-Schiff stain showed many
fungal hyphae under the microscope

12. Mucins are a family of high molecular weight, heavily glycosylated proteins
(glycoconjugates) produced by epithelial tissues in most animals

15. Amyloidosis (Protein aggregation
disease)
•Abnormal deposit of insoluble polymeric protein fibrils (amyloid
starch like) in tissues and organa
•Fibrils formed by the aggregation of misfolded proteins,
normally soluble proteins and it is usually extra cellular
•Congo Red and Thioflavin S are the two major histological
stains used to detect any form of amyloid. These dyes bind to
the characteristic β-pleated sheet conformation of amyloid

17. Connective tissue
•Collagenis a strong protein and is a main component of ligaments and
tendon. It is also responsible for skin elasticity and is the most visible type
which can be visualized with an H&E.
•Elasticfibers are located in the skin and walls of blood vessels. They are
composed of elastin -a protein that is flexible and allows many tissues in
the body to resume their shape after stretching or contracting and are not
visualized by an H&E
•Reticular fibers are composed of collagen and form a delicate framework
around nerve fibers, fat cells, lymph nodes, and smooth and skeletal muscle
fibers. They are also not visualized by an H&E.

18. Routine stain for liver
and kidney biopsies

19. Standard applications Of MT stain: Masson's
trichrome staining is widely used to study muscular
pathologies (muscular dystrophy), cardiac pathologies
(infarct), hepatic pathologies (cirrhosis) or kidney
pathologies (glomerular fibrosis). It can also be used to
detect and analyze tumors on hepatic and kidney
biopsies

22. Liver section stained with
a Gomori’sTrichrome that
colors collagen, green rather
than blue

23. Gomoritrichrome
stain Severe
histological
degeneration, with
loss of collagen

24. Verhoef stain/ Elastic Stain
Elastic Stain is used to identify the
atrophy of elastic tissue as in emphysema,
evidence of vascular diseases
(arteriosclerosis), and the invasion of
tumors into vessels.
Elastic fibers and nuclei being stained
black, collagen stained red and
cytoplasmic elements stained yellow

25. Skin stained with Elastic Stain
Elastin fibers and elastic lamina in
histological specimens are stained
black, while remaining tissue
elements are stained as follows:
nuclei -blue/black, collagen -red,
other tissue elements -yellow.

26. Reticulin fibers support the body
and are common in the liver,
spleen and kidneys.
Characteristic reticulin patterns
can help diagnose cirrhosis of the
liver, early fibrosis in bone marrow
and tumors
Black Stained Reticulin Fibers
Reticulin Fiber Stain

27. A mast cell is a type Of white blood cellsresidentcell of connective tissue that contains many
granules rich in histamine and heparin. Important in allergic and parasitic infection

28. Photomicrograph of
toluidine blue stain
showing mast cells
(Toluidine blue stain, x100)

32. Nucleic Acid (Feulgen Stain)
This Feulgen stain demonstrates DNA (magenta) in breast carcinoma tissue.
DNA Magenta
Background Light
green

36. Acid fast stains /Ziehl–Neelsen
staining
Acid fast stains areused to
differentiate acid fast organisms
such as mycobacteria. Acid fast
bacteria have a high content of
mycolic acids in their cell walls

37. Mycobacterium
tuberculosis with an
acid fast stain (pink
rods)

38. Romanowsky Stains
•Romanowsky Stains are the stains (a mixture of
oxidized methylene blue (azure) dyes and Eosin )
that are used in hematology and cytological studies,
to differentiate cells in microscopic examinations of
blood and bone marrow samples. These stains are
also applied to detect the presence of parasites in
the blood such as malaria parasites. For example
•Giemsa stain
•Wright and Wright-Giemsa stain
•May-Grunwald stain
•Leishman stain

39. Giemsa-stained blood smear
showing intra-erythrocytic
Pyriform (Pear-shape) of
Babesia bovisin pairs

40. Giemsa-stained blood
smear show Babesia
bigemina

41. Giemsa-stained blood smear showing intra-erythrocytic
(Pear-shape) of Babesia bovisin pairs

42. Giemsa-stained blood
smear showing intra-
erythrocytic Theileria

43. Giemsa-stained section of small
intestinal mucosa showing
clusters of Giardia that stain
purple (arrows) in the crypts. The
background is stained faint pink
by eosin.

44. Grocott methenamine silver(GMS)
stain
GMS is commonly used for the
identification of fungi on cytosmears
and tissue sections. It imparts a black
color to the fungal profiles and a pale
green color to the background. It
stains all pathogenic and
nonpathogenic fungi

47. Von Kossastainis widely used in histology
to detect thepresence of abnormal
calcium deposits in the body. The
principle of this coloration is based on the
transformation of calcium salts into silver
salts: calcium ions, bound to phosphates,
are replaced by silver ions brought by a
solution of silver nitrate. Placed under a
light source, the silver phosphates undergo
a photochemical degradation, leading to the
observation of metallic silver deposits.

49. Fontana-Masson staining for melanin
Melanin granules in the skin stained Black