Practical histo- and cytopathology by Hussein A. Abid

1. PRACTICAL PATHOLOGY
HISTO- & CYTOPATHOLOGY
Hussein A. Abid
Medical Lab Specialist
Middle Technical University (teacher)
American Society for Microbiology (member)
Iraqi Medical Laboratory Association (president)

2. Terminology
• Biopsy: examination of tissue taken from living
body (gross & microscopic examination).
• Autopsy: examination of dead body
• Disease: any abnormality in the structure or
function of an organ or tissue.
2

3. TYPES OF BIOPSY
1. Incisional biopsy: a portion of tissue from a large
lesion is taken-only diagnostic.
2. Excisional biopsy: the entire lesion is removed with
a margin of adjacent normal tissue-diagnostic &
therapeutic.
3. Punch biopsy: by biopsy forceps in the uterus
,cervix, oral cavity, esophagus.
4. Core needle biopsy: by wide bore needle used
percutaneously for sampling of internal organs.
5. Curettage biopsy: for diagnosis of uterine
diseases. 3

4. Multiple excised masses ( excisional biopsy)
4

5. Large bulky well defined mass removed totally
(excisional biopsy) 5

6. Segment of colon removed with central mass
(excisional biopsy) 6

7. Kidney (excisional biopsy)
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8. Uterus with enlarge ovaries (excisional biopsy)
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9. GENERAL PRINCIPLES FOR GROSS
EXAMINATION
• Proper identification and orientation of the specimen.
• Place the specimen on a cutting board & record all the
following data:
1. Type of specimen
2. Dimension (in centimeters)
3. Weight (in grams)
4. Shape
5. Consistency
6. Surgical margins whether included or not involved by
the tumor.
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10. HISTOPATHOLOGICAL TECHNIQUES
• Deals with tissue deals with preparation of tissue
for histopathological examination.
• The aim of these technique is to preserve
microscopic anatomy of tissues and to cut
tissue in very thin sections (4-5 microns) this is
achieved by passing tissue in a series of
process.
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11. • Tissue processing can be done manually
or mechanically & includes the following
processes:
1.Fixation
2.Dehydration
3.Cleaning
4.Embedding
5.Cutting
6.Staining
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13. 13

14. Tissue processor used for biopsy processing through
passing the biopsy into multiple steps
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15. FIXATION
• Most fixatives act by denaturating or
precipitating cellular proteins which form
meshwork that hold other structures &
prevent autolysis.
• The most widely used fixative is 10%
formalin.
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16. DEHYDRATION AND CLEARING
• Dehydration: is removal of water molecules
from tissues and is achieved by graded alcohol.
• Cleaning: alcohol replace water in the tissues,
removal of alcohol from tissues is by Xylene
which creates empty tissue spaces to be
infiltrated by wax.
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17. EMBEDDING WITH WAX
• Paraffin wax is used for
embedding of tissue which
form tissue blocks after
cooling the it can be
trimmed into thin sections
(4-5 microns) using microt-
ome, the sections are placed on glass slides and
become ready for staining
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18. Embedding with wax 18

19. 19

20. 20

21. Tissue blocks are trimmed into thin sections (4-5 microns)
using the microtome
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22. 22

23. Sections are placed in water bath for opening of folded
tissue
23

24. Sections are placed on glass slides and become ready for
staining
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25. STAINING
• Hematoxylin and eosin (H & E): is the most
widely used stain in histopathology
 Nuclei appear dark blue
 Collage and cytoplasm appear pink
 Keratin appears pink to red
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26. 26

27. 27

28. 28

29. 29

30. 30

31. 31

32. STAINING
Special stains
• PAS (periodic acid schiff) stain glycogen and
mucin
• Congo-red for amyloid
• Sudan-black for fat
• Giemsa stain for Helicobacter pylori
32

33. H. pylori stained dark blue with
Giemsa stain
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34. Amyloid deposits stain orange-red with Congo
Red stain
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35. FROZEN SECTION
• In this technique the tissue is frozen rapidly (using
cryostat) to -20 ºC, ten sections are cut and stained
(without passing in the steps of tissue processing) so
that tissue can be examined microscopically within 5-
10 minutes of removal from the body.
• It allows rapid diagnosis of the nature of the lesion
whether benign or malignant to decide the next step in
surgery.
• All laboratory staff should be informed and all
preparations should be completed before arrival of
tissue. 35

36. CYTOLOGY
• Is the study of cell (normal or diseased altered
cell) obtained from various sites of the body. It
allows rapid diagnosis often within minutes.
• Cells examined by this process are collected by
one of the following methods:
 Exfoliated cell
 Cells removed by brushing or scraping
 Removal of cells from deep tissue (by
aspiration) 36

37. EXFOLIATIVE CYTOLOGY
• Spontaneous shedding of cells derived from
lining of cavities where they can be removed by
non invasive methods e.g. vaginal smear,
sputum examination, voided urine, body
fluids, nipple discharge
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38. ABRASIVE CYTOLOGY
• Cells of specimen are obtained from superficial
scraping of the lesion e.g. cervical scraping
(pap-smear), buccal mucosal smear, skin
scraping of various lesions.
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39. FINE NEEDLE ASPIRATION
CYTOLOGY (FNAC)
• Cells are aspirated from deep non surfaces
organs or masses e.g. beast mass, thyroid
nodules, palpable lymph nodes, internal organs
like liner and kidney.
39

40. TECHNIQUE OF CYTOLOGY
1. Use a needle and syringe to obtain cells from a mass.
 Exfoliated material is sprayed on a slide directly.
 The needle is gently introduced through the skin into
the mass (while the mass is fixed in between fingers
and thumb) and is moved in different directions while
applying negative pressure onto the syringe with
continuous suction of material through out the
process, then the needle should be withdrawn gently
from the mass.
2. Smearing collected material on a glass slide 40

41. TECHNIQUE OF CYTOLOGY
3. Immediate immersion of the slide in a fixative (95%
ethanol) to avoid dryness of material.
4. Applying a stain.
5. Examine under the microscope.
41

42. Palpable lymph node in the neck can be examined using
fine needle aspiration cytology
42

43. Aspiration of material from cervical mass while applying
negative pressure
43

44. Spraying aspirated material on a glass slide 44

45. Immediate fixation of material in 95% ethanol 45

46. Stain the slides 46

47. Cytology is the study of cell
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48. 48

49. INDICATIONS OF CYTOPATHOLOGY
1. Diagnosis of malignancy.
2. Diagnosis of precancerous changes e.g. cervical atypia.
3. Detection of inflammation and pathogenic agents e.g.
fungal or parasitic infection of vagina
4. Study of hormonal patterns and gonadal function e.g.
examination of squamous cells in vaginal smear which
are under influence of ovarian hormones to assess
ovarian function in infertility.
5. Identification of sex chromosome in newborn with
ambiguous genitalia, buccal smear is used as a source of
cells. 49

50. LIMITATIONS OF CYTOLOGY
1. The nature of lesion is not so obvious as in a
histological section.
2. Difficulty in identification of the exact site of
lesion e.g. malignant squamous cells shaded in
sputum may arise from buccal mucosa, pharynx,
larynx and bronchi.
3. The size of lesion cannot be approximated by
cytology.
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