EXFOLIATIVE CYTOLOGY SPECIMEN COLLECTION

Body cavity fluids

Exfoliative Cytology ➢ Minimum of 10ml for optimal cytologic evaluation
➢ A branch of cytology which deals with the microscopic ➢ Add heparin to reduce clotting
examination of shed cells from body surface or cells harvested by ➢ Refrigerated or kept on wet ice until transported to the Lab
rubbing or brushing a lesional tissue surface.
➢ First introduce by Papanicolaou in 1941
➢ It’s a simple, pain free, non-invasive and rapid technique
➢ Used for the study of superficial or desquamated cells and
requires other cytological analysis to confirm

Indications
➢ Multiple or large lesions.
➢ Lesion located in the region that present surgical difficulty. 1. Pleural Fluid
➢ Patient with anticoagulant drug and bleeding disorders. ➢ examines a sample of pleural fluid under a microscope to
➢ Older people who cannot tolerate surgical procedure. detect for abnormal cells or microorganism
➢ When herpes or candida are suspected.
➢ Follow up for detection of recurrent cancer

Other Uses of Exfoliative Cytology

1. Detection of malignant cells or precancerous lesions in the body
2. Detection of asymptomatic or precancerous cervical lesions in
women
3. Assessment of female hormonal status in case of infertility and
endocrine disorders 2. Peritoneal Fluid
4. Determination of genetic (phenotype) sex ➢ to determine the cause of ascites, fluid accumulation in the
5. Detection of the presence of infectious microorganisms peritoneal space

Specimens for examination

• peritoneal, pericardial and pleural fluids
• CSF
• nipple discharge
• bronchial brushing/washing
• sputum
• gastric washing
• urine sediments
• prostatic secretions/fluid
• cervicovaginal (Pap) smear
 3. Pericardial Fluid Bronchial Washing
➢ Small needle in inserted into the chest between the rib and ➢ Bronchoscopically-directed washing (10ml) of the bronchi in the
the pericardium. Small amount of fluid is withdrawn region of the suspected lesion
Bronchial Brushing
➢ Bronchoscopically-directed brushing of identified lesion
Bronchial Lavage

4. CSF Fluid
➢ Usually obtained through lumbar puncture (spinal tap),
usually between 3rd and 4th vertebrae.

Sputum
➢ Normal – obtain at three consecutive morning specimens by
cough method
➢ Induced – inhalation of aerosol for 20 mins to produce deep cough
sample

Nipple Discharge/ Lesions

➢ Breast biopsy can help diagnose breast cancer early in the disease
process.
➢ Gently strip the subareolar area and nipple with the thumb and
forefinger
Gastric Secretion
➢ Washing (esophageal, gastric and others)
➢ Endoscopically obtained (10ml) of the region of the suspected
lesion
➢ Fast for a minimum of six hours
Cervicovaginal Specimen Pull Apart
➢ The endocervical mucus will prevent air-drying during collection ➢ For serous fluids, concentrated sputum, and enzymatic lavage
➢ Scrape material from the whole circumference of the cervix form the GIT, smear of urinary sediment, vaginal pool and breast
➢ Spread quickly and evenly onto the slide then fix immediately secretions

Touch or Impression Smear

➢ Impression cytology being collected from patient, using a sterile
glass slides with polished edges

METHODS OF SMEAR PREPARATION

Streaking
➢ Used for preparing mucoid secretions, vaginal secretion, sputum FIXATION
and gastric content ➢ Exfoliated cells decompose rapidly which may destroy cellular
➢ Use a spatula, dissecting needle or applicator stick and streak in a and nuclear details, in turn will give inadequate results for
zigzag fashion diagnosis
Common Fixatives:
1. Equal parts of 95 EtOH and Ether
2. 95% EtOH
3. Carnoy’s fluid
Spreading 4. Equal parts of tertiary butyl alcohol and 1 part 95% EtOH
➢ Used for thick mucoid secretions 5. SCHAUDINN’s fluid
➢ Smears of fresh sputum and bronchial aspirates 6. MeOH – from dried

Papanicolaou or PAP’s Smear

Advantages
➢ Transparent blue staining of cytoplasm is observed
➢ Excellent nuclear staining
➢ Color range is predictable and of great value in identification of
cells
Disadvantages Results
➢ Procedure is lengthy and complicated ➢ Cytoplasm – either bright red or greenish blue
➢ Does not give accurate acidophilic index ➢ Vesicular nucleus – blue
Stains for Pap’s ➢ Pyknotic nucleus – dark blue to black
• Harris Hematoxylin ➢ Bacteria – dark blue
• Orange Green 6 (OG 6) Stain ➢ Mycelia – violet
• Eosin Y (EA 65 or EA50) ➢ Trichomonas vaginalis – pale greenish blue blob of cytoplasm

Trichomonas vaginalis

Bacterial vaginosis
➢ Shift in normal vaginal flora (Lactobacilli) to Gardnerella
Procedure for Pap’s Stain vaginalis
1. Fix in ether-ROH and pass thru 80% ROH, 40% ROH, and distilled
water
2. Stain in Harris Hematoxylin for 4-5mins
3. Wash with H2O
4. Pass thru 0.25% NH4OH in 50% ROH
5. Immerse in 1.5% NH4OH in 70% ROH for 1 min
6. Rinse in 70% ROH and pass thru 80% and 95% ROH
7. Stain with OG 6 for 1.5mins
8. Pass thru 3 changes of 95% ROH
9. Stain with Eosin Y for 3mins
10. Pass thru 3 changes of 95% ROH
11. Dehydrate and clear in
• absolute ROH
• equal parts of ether and absolute ROH
• 2 changes of xylol
12. Mount in Canada Balsam
 Herpes simplex
➢ Swollen nuclei with multinucleation
➢ Ground glass chromatin with prominent nuclear membrane and
clear inclusions (tombstones)
➢ Nuclear moulding

Candidiasis
➢ “Shish kebab” appearance

Changes in Malignancy
➢ Altered Nuclear-cytoplasmic ratios
➢ Hyperchromasia
➢ Increased mitotic activity
➢ Atypical mitoses

Actinomycosis
➢ Common in women with Intrauterine device (IUD)

Nuclear Changes
➢ Multinucleate cells
• with irregular hyperchromatic or bizzare nuclei should
be suspicious
➢ Anisokaryosis
• variation in nuclear size and shape
➢ Giant single nucleus (polyploidy)
• may occur in benign conditions
 Vaginal Cytology
➢ Vaginal cytology is a type of endocrine assay
➢ Tracking changes in the morphology of desquamated vaginal
cells provides a means of assaying changes in estrogen levels
➢ Hormonal changes are best mirrored in the upper third of the
vagina
➢ Can also be taken from the lateral walls because of easy access
and less contamination

Superficial Cells
➢ Large, polyhedral flat cells (30-60u)
➢ Cytoplasm maybe acidophilic or
basophilic
➢ Presence of small dark blue pyknotic
nuclei (< 6u)

Intermediate Cells
➢ Medium large, polyhedral or elongated cells (20-30u)
➢ Basophilic with vacuolated cytoplasm
➢ Vesicular nuclei (6-9u)
Cytoplasmic Changes
➢ Cells of squamous cell carcinomas frequently show a tendency to
cytoplasmic eosinophilia
➢ Adenocarcinoma cells may enclose endometrial and colonic
cancers
➢ Cytoplasmic vacuolation is common in adenocarcinoma

Parabasal Cells
➢ Round to oval cells (15-25u)
➢ Thick sunny side-up like cells
➢ Have strong basophilic cytoplasm and vesicular nuclei (6-9u)
➢ Found from 2 weeks of age to puberty
Endocervical Cells Bethesda System
➢ Slightly cylindrical appearance ➢ Specimen Adequacy:
➢ Occurs in groups and strips of three or more cells • Satisfactory
➢ Cytoplasm deeply basophilic than the parabasal cells • Limited
• Unsatisfactory
➢ General Categorization:
• Negative for intraepithelial lesion or malignant cells
• Epithelial cell abnormality
➢ Descriptive Diagnosis:
• (ASCUS) Atypical Squamous Cell Of Unknown
Significance
• (LSIL) Low grade squamous intraepithelial lesion
• (HSIL) High grade Squamous Intraepithelial Lesion
• Squamous cell carcinoma
Endometrial Cells • Glandular cell abnormality
➢ Found during menstruation peroid (in groups) • Atypical glandular cells
➢ Endometrial stromal cells seen in tight clusters of small, oval • Adenocarcinoma
dark cells • Others
➢ Nucleus small and moderately dark
➢ Cytoplasm basophilic and maybe vacuolated Specimen Adequacy
Satisfactory/ Adequate:
➢ Adequate numbers of well-visualized squamous cells present
➢ Adequate number of well-visualized endocervical or squamous
metaplastic cells (transformation zone)
➢ Less than 50% of cells obscured by blood and inflammation
➢ Properly labeled specimens

Lactobacillus acidophilus
➢ Gram + slender rod bacteria
➢ Vaginal normal flora: provides low pH that inhibits growth of
pathogens
➢ Stains pale blue to lavender
➢ Numerous in luteal phase and during pregnancy
Unsatisfactory/ Inadequate: Atypical Squamous Cell of Unknown Significance (ASC-US)
➢ Low number of squamous cells present ➢ Cannot exclude HSIL (ASC-H)
➢ Inadequate numbers of metaplastic or endocervical cells
➢ More than 70% of cells obscured by blood and inflammation
➢ Improperly labeled smears
➢ Recommend repeat sampling

Low grade squamous intraepithelial lesion (LSIL)

➢ Nuclear enlargement x3
➢ Hyperchromasia
Specimen for Rejection ➢ Optically clear cytoplasmic halo
➢ Submitted without a requisition ➢ Nuclear irregularities
➢ Not labeled with the patient’s name
➢ Patient name on the specimen and requisition do not correspond
➢ Specimen is labeled inappropriately
➢ Specimen slides are irreparably broken
➢ Specimen is submitted from an unauthorized sourced

Negative for Intraepithelial Lesion or Malignant cells (NILM)

High grade squamous intraepithelial lesion (HSIL)

➢ Severe dysplasia or (CIS) Carcinoma In Situ
➢ Suspicious for invasion
Squamous Cell Carcinoma Cervical Adenocarcinoma
➢ Cervical Squamous Cell Carcinoma
➢ Gross and Microscopic

Endometrial Carcinoma

Cytology of CIN seen in Pap Smear

➢ There is progressive reduction of size of Cytoplasm, and increase
in Nuclear Cytoplasm Ratio on the grade of lesion progresses,
which reflects there is loss of cellular differentiation on the
surface of cervical lesion