A Routine and Special Staining in Histopathology LAB practice lecture notes Professor Ali Altimimi.pdf

1.  Connective tissue is one of the major types
of tissue that connects different parts of
tissue and supports the body parts.
 The fibrous part of C.T includes:
• Collagen fibers
• Reticular fibers
• Elastic fibers
• Basement membrane
Connective Tissue Fibers
Histopath Stains
Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London
A Routine and Special Staining in Histopathology LAB practice
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

2.  Collagen fibers : composed of the protein
collagen and provide the greatest strength
of the 3 fiber types. Found in ligaments,
tendons, cartilage and bone.
 Elastic Fibers: composed of the protein
elastin and provide the greatest flexibility
among the fiber types. Found in skin and
walls of blood vessels.
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

3.  Reticular fibers : composed of the protein
collagen. They form a delicate network for
many soft organs and a network around
nerve fibers, fat cells, LNs, skeletal and
smooth ms fibers.
 Basement membrane: C.T elements that
separate the epithelial and endothelial cells
from the underlying connective tissue.
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

4. Collagen fibers demonstrated by:
H & E
Trichrome Stains
PTAH (phosphotungstic acid-
hematoxylin)
Demonstration of Collagen fibers
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

5. 1- Hematoxylin & Eosin
Collagen fibers
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

6. 1- Hematoxylin & Eosin
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

7. Collagen fibers demonstrated by:
H & E
Trichrome Stains
PTAH
Demonstration of Collagen fibers
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

8. 2- Trichrome Stains
 Trichrome stains: 3 dyes used to distinguish
collagen from muscle and aid in the diagnosis of
fibrotic changes, neuromuscular diseases and
tumors of muscle origin.
 Result:
1. Collagen: blue
2. Muscle, Cytoplasm& RBCs: Red
3. Nucleus: blue
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

9. 2- Trichrome Stains
Masson’s
Trichrome
Gomori’s One-Step Trichrome
Stains
 The most common techniques for trichrome staining
 The Masson utilizes all of the mordanting and
staining steps individually , where as the one-step
incorporates all of the staining steps in one staining
solution except the mordant and nuclear stain.
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

10. Masson’s Trichrome Stains
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

11. Masson’s Trichrome Stains
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

12. Masson’s Trichrome Stains
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

13. Gomori’s One-Step Trichrome
Stains
Gomori’s Trichrome (green)
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

14. Gomori’s One-Step Trichrome
Stains
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

15. Collagen fibers demonstrated by:
H & E
Trichrome Stains
PTAH
Demonstration of Collagen fibers
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

16. 3- phosphotungstic acid-hematoxylin
(PTAH)
• PTAH is preferred for demonstrating cross-striations of
skeletal muscle.
• Muscle, cytoplasm, fibrin: Various shades of blue
Nuclei: Various shades of blue
Collagen Red-brown
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

17. 3- phosphotungstic acid-hematoxylin
(PTAH)
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

18. 3- phosphotungstic acid-hematoxylin
(PTAH)
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

20.  Not visualized by H&E.
 commonly demonstrated
by the use of stains
involving silver solutions.
 Impregnation of silver ions
to the fibers and
subsequent reduction of
those silver ions to their
visible metallic form
(argyrophilic).
Demonstration of reticular fibers
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

25. H & E
The Verhoeff-Van Gieson (VVG) stain.
Orcein technique.
Aldehyde fuchsin method.
Demonstration of Elastic fibers
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

26. 1- Hematoxylin & Eosin
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

27. 1- Verhoeff-Van Gieson stain
Elastic fibres (blue/black) , collagen (red/pink) Muscle and
other tissues appear yellow. (Normal human lung).
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

28. 1- Verhoeff-Van Gieson stain
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

29. 1- Verhoeff-Van Gieson stain
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

30. 3- Orcein technique
Aorta
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

31. 2- Orcein technique
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

32. 4- Aldehyde fuchsin method
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

33. H&E The Verhoeff-
Van Gieson
(VVG) stain.
Orcein technique. Aldehyde fuchsin
method.
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

34. H & E
Silver stain
PAS technique
Demonstration of basement
membrane
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

35. 1- Hematoxylin & Eosin
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

36. BM may also be demonstrated using the Periodic Acid-Schiff
(PAS) technique.
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

37. PAS H&E
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London

38. BM are commonly demonstrated with a silver stain
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Prof Ali Altimimi Professor of Pathology & Human Physiology,MbCHB,MSC,PHD,MD.Royal College London