Dermatology, dermatopathology, stains used

1. SPECIAL STAINS
IN
DERMATOLOGY

2. WHAT IS STAIN?
 Staining is an auxiliary technique used in microscopy to enhance contrast in the microscopic
images.
 Stains may be used to define and examine the bulk tissue
1. Cell population
2. Muscle fibers
3. Connective tissues
4. Organelles

3. SPECIAL STAINS
 Used in addition to H & E staining to selectively
 Stain cells & cellular components
Gives information on:
 Presence of certain class of molecules
 Their localization
 Number of molecules

4. STAINS FOR
MUCOPOLYSACCARIDES
1. PAS
2. ALCIAN BLUE
3. MUCICARMINE
4. COLLOIDAL IRON

5. PAS
 Most commonly used special stain
 Stains neutral mucopolysaccharides (Glycogen)
 Basement membranes (e.g SLE)
 Fungi, Parasides
 Fibrin, Inclusion bodies

9. ALCIAN BLUE
 Best stain for mucin
 Alcian blue (pH 2.5)
- Acid mucopolysaccharides (glycosaminoglycain’s)
- Light blue
 Alcian blue (pH 0.5)
- Sulfated mucopolysaccharides (heparin sulfate)
- Blue
 Does NOT stain neutral mucins

10. RESULTS
 Acid mucin & proteoglycans : BLUE
 Nuclei : RED

12. MUCICARMINE
 To demonstrate acid & neutral mucopolysaccharides
- Epithelial mucin
- Capsule of cryptococcus neoformans
Results
 Acid epithelial mucins : Deep rose to red
 Nuclei : Black
 Others tissue elements : Light yellow

14. COLLOIDAL IRON
 A mucin stain for dermal/connective tissue mucin
 Stains acid mucopolysaccharides
RESULTS
 Proteoglycans , Acid mucins : BRIGHT BLUE
 Collagen : RED
 Muscle & cytoplasm : YELLOW

16. STAINS FOR PIGMENTS &
MINERALS
1. Fontana-Masson
2. Von-kossa
3. Alzarin red S
4. Prussian blue stain

17. FONTANA-MASSON
 Stains melanin & argentaffin granules – BLACK (nuclei will be red)
 Useful for quantifying melanocytes (e.g. in vitiligo) & in cases of minocycline pigmentary
alteration
 Also stains cryptococcus
RESULTS
 Melanin , Argentaffin, Chromaffin : BLACK
 Nuclei : RED

18. USES
 To identify melanin & argentaffin granules
 In diagnosis of malignant melanoma
 Argentaffin granules are found in carcinoid tumors

21. VON-KOSSA
 Stains calcium salts – BLACK
 Calcification of vessel walls & elastic tissue
( Calcinonis cutis, pseudoxanthoma elasticum, calciphylaxis, elastosis
& elastofibroma)

24. PRUSSIAN BLUE STAIN (PERLS IRON)
 Hemosiderin and ferric ions
 Useful for identifying iron as the source of pigment
 Stains iron – BLUE
RESULT
 Ferric iron : Blue
 Nuclei : Red

26. ALIZARIN RED
 Stain for calcium salts
RESULTS
 Calcium : ORANGE RED
 Background : GREEN

28. STAINS FOR CONNECTIVE
TISSUE
 Trichrome-Masson
 Verhoeff-Van Gieson

29. TRICHROME-MASSON
 Smooth muscle , cytoplasm, keratin – Red
 Useful for distinguishing leiomyoma’s from dermatofibromas & neural tumors
 Collagen – Blue/green
 Useful in evaluating the characteristics of dermal collagen
 Nucleus - Black

30. USES
 It is used to differentiate b/w collagen & smooth muscle in tumor
 To identified the increased collagen deposition : Keloid
RESULT
 Nuclei : Blue/Black
 Cytoplasm, Muscle, & Erythrocytes : Red
 Collagen : Blue

32. VERHOEFF-VAN GIESON OR WEIGERT
 Used to differentiate collagen and smooth muscle
 Can be used to demonstrate the presence of collagen in pathological conditions( DLE, EN, RA,
SLE, MORPHOEA)
 Stains nuclei : Blue
 Cytoplasm, muscle, fibrin : Yellow
 Collagen : Bright
 Elastic fibers : Black

34. STAINS FOR AMYLOID
 Congo red
 Thioflavin T
 Crystal violet

35. CONGO RED
 Stains amyloid : PINKISH-RED
 Gives apple-green birefringence to amyloid
 The most specific method for amyloid

38. THIOFLAVIN T
 Amyloid shows yellow fluorescence
RESULT
 UV light source : Silver-blue
 Blue light fluorescence : Yellow

40. CRYSTAL
VIOLET
• Stain Amyloid purple-violet

41. STAINS FOR FAT
 Sudan black B
 Sudan orange
 Oil red O

42. SUDAN BLACK B
 Most sensitive of all fat dyes
 Stains neutral fats – Blue-Black
 Stains phospholipid – Grey

44. OIL RED O
 Stains fat red
 Frozen/Fresh tissue ( once tissue is fixed &processed into paraffin blocks, this method does not
work)
 This may be very helpful in seeing the fat globules in sebaceous carcinoma

45. RESULT
Fat : Brilliant Red
Nuclei : Blue FAT
NUCLEI

46. STAINS FOR MAST CELLS
 Giemsa
 Toluidine blue
 Chloroacetate esterase ( Leder stain)

47. GIEMSA
Metachromatically purple
Urticaria
Mastocytosis MAST CELL

48. TOLUIDINE
BLUE
Mast cells stain – Purple
(Metachromatic staining)
Background stain – Blue
(orthochromatic staining)
Mastocytosis
MAST CELL

49. STAINS FOR MICROORGANISMS
 H&E
 Gram
 Giemsa
 Gomori methenamine silver (GMS)
 PAS
 Fontana-Masson
 Warthin-Starry
 Ziehl-Neelson stain

50. H&E
May demonstrate fungi, bacteria

51. GRAM STAINING
 Differentiates bacteria by the chemical & physical properties of their cell well by detecting
peptidoglycan
 Gram +ve bacteria retain the crystal violet dye & thus are stained violet : BLUE
 Gram –ve bacteria : PINK

53. GOMORI METHENAMINE SILVER (GMS)
 Fungal cell walls
 Stains fungi & parasites brown or black with a green background
 Pneumocystis carnii, histoplasma , leshmania

56. GIEMSA STAIN
 Used to stain bacteria & protozoa
 H. pylori, rickettsia & chlamydia
 Bacteria stains : BLUE
 Cytoplasm stains from : Pink to rose
 Nuclei : BLUE
 Eosinophils are also easily detected

58. WARTHIN-STARRY
 A silver nitrate stain
 Spirochetes – BLACK
 Background – Golden yellow

60. ZIEHL-NEELSEN
 Acid fast refers to cell walls containing high lipid content
 Used to stain Mycobacteria, oocysts of Cryptosporidium parvum, Cyclospora, Isospora
 Acid fast cells stain : RED
 Non acid fast cells stain : BLUE

62. ACID FAST STAINING
TECHNIQUES
 Ziehl-Neelsen stain ( classic type)
 Kinyoun
 Auramine-rhodamine (fluorescent dyes)
 Fite : for leprosy bacillus which is less acid fast

64. THANK YOU