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CYT 2113 Cytology I

Lesson 9:
Basic Gynecological Cytology

Pap Test
Part of a gynecological exam
An examination under the microscope of cells
scraped from the tip of the cervix

Features to be Evaluated in a Pap test

Presence of abnormal cells
Number and distribution of abnormal cells
Relationship between cells
Cell size and shape

Nuclear size and shape

Nuclear changes and nucleoli
Nuclear-to-cytoplasmic (n:c) ratio
Cytoplasmic features
Background or diathesis

Features of Preneoplastic and

Neoplastic cells

Abnormality in size and shape of cells

Variation in cell size and shape
Increase in nuclear size
Increase in nuclear membrane irregularity

Prominence of nucleoli and irregularity in
shape thereof
Thickening of nuclear membrane
Increase in n:c ratio

Cytoplasm scanty
Mitosis, increased number and abnormal

Common Causes of False-negative

Pap test

Atypical endocervical cells

Crowded cell aggregates
Intermediate cells with nuclear enlargement

Metaplastic-like cells
Necrotic debris
Artifacts such as obscuring blood,
inflammation or air-drying

Common causes of false-positive

Pap test
Atrophic smear
Atypical endocervical or endometrial cells
Multinucleated cells

Perinuclear halo in nonkoilocytes

Squamous metaplasia
Tubal metaplasia

Differential diagnosis of cells with

naked nuclei
Naked nucleus - A nucleus in a cytologic
preparation that is virtually devoid of

Autolysis of cytoplasm in menopause

Degeneration, especially of endocervix
Reserve cells with tamoxifen treatment

Differential diagnosis of giant

multinucleated cells
Folic acid deficiency

Tissue repair
Viral infection

Syncytiotrophoblast (the outer syncytial layer

of the trophoblast that actively invades the
uterine wall forming the outermost fetal
component of the placenta)

Squamous carcinoma
Uterine sarcoma

Cytological features of dark-cell


Crowded with piling up of cells

Hyperchromatic overlapping nuclei
Scant cytoplasm
Increased n:c ratio
Mitosis present
Often difficult to determine whether
squamous or glandular

Differential diagnosis of dark-cell


Reactive endocervical cells

Tubal metaplasia
Atrophy: nuclear membrane smooth
Benign endometrial cells

Atypical squamous cells cannot exclude highgrade squamous intra-epithelial lesion

High-grade squamous intra-epithelial lesion:
nuclear membrane irregular
Adenocarcinoma in situ
Endocervical or endometrial carcinoma

Differential diagnosis of small cells

Lymphocytes in chronic lymphocytic cervicitis

Degenerated cells
Endometrial cells
Reserve cells

Tamoxifen cells
Smaller cell type of squamous-cell carcinoma
Small-cell anaplastic carcinoma

Differential diagnosis of cells with

Repair, regenerative or reactive squamous
Reactive endocervical cells
Viral inclusions
Treatment effect
Metastatic tumour
Nonkeratinizing squamous carcinoma

Differential diagnosis of

Viral infections
Endocervical cells, benign and atypical
Endometrial cells, benign and atypical

Vaginal adenosis
Intrauterine device
Metastatic tumour

Factors Related to Failure of Pap

Test Screening
Instrument and sample
Cytopreparation and interpretation
These factors are interrelated

Patient-related Errors
Including failure of women to get regular Pap
tests or to seek any health care at all
Douching or sexual intercourse can
mechanically remove the superficial cell layers
that the Pap test samples, causing falsenegative results due to sampling error
Some women delay seeking medical attention
even when they have symptoms that they
know are suspicious, such as abnormal vaginal

Clinical Errors
Failure to take a Pap test at all
Failure to take an adequate Pap test
The sample must be obtained under direct
visualization, with considerable pressure
The speculum should not be lubricated
Overzealous rubbing, swabbing or cleaning
the cervix before taking the sample can
remove the abnormal cells, leading to false
negative results

Well-trained health care providers, not

necessarily physicians, take better Pap tests
Failure of the clinician to provide pertinent
clinical data can severely compromise
cytologic interpretation of the Pap test
Failure to follow-up abnormal Pap test results
Failure to perform a biopsy of suspicious
Failure to investigate suspicious clinical

Instrument and Sample Errors

A combination of spatula and brush enhances
sampling compared with either device used
The material from which the sampling device
is important
Cotton swabs and wooden spatulas tend to
trap cells so they unavailable for

Wooden spatulas collect between 600,000

and 1.2 million epithelial cells, but less than
20% are transferred to the glass slide when
making a conventional Pap smear
The transfer of cells to the slides is random
and statistically prone to error because the
abnormal cells are not homogenously
distributed in the sample
Endocervical samples taken by cotton swabs
or with plastic spatulas obtain fewer atypical
cells than do endocervical brushes

The shape of the sampling device is also

important for obtaining an adequate sample,
particularly of the endocervix
Sufficient numbers of well-preserved cells
must be collected and the sample must be
representative, including the transformation
A conventional Pap smear must be thinly
spread and immediately and properly fixed

Cytopreparation and Interpretive

Few abnormal cells (particularly <250)
Small abnormal cells (difficult to find)
Bland abnormal cells (difficult to interpret)
Hyperchromatic crowded groups (difficult:
usually benign, can be malignant)
Obscuration: excess exudate, blood; poorly
Inadequate clinical information

Lesion-related Errors
Some lesions fail to exfoliate sufficient
numbers of cells for detection
Small or inaccessible lesions may be difficult
to sample adequately
The shape of the cells and the quality of the
cervical mucus also affect the sampling
Pap tests are more often inadequate in
women with advanced epithelial

Necrosis, inflammation or bleeding may

obscure, alter or dilute the abnormal cells,
making interpretation difficult
Pap tests repeated within a short time (up to
several weeks) have a particularly high false
negative rate
Apparently it takes some time for the lesion to
regenerate sufficient cells to be detected in
the cytologic sample

A negative following a positive

interpretation can mislead the clinician into
thinking the lesion has regressed or the
previous positive report was wrong
It is also possible that some tumours progress
so rapidly that they develop in between Pap
test screenings