Cytopathology-4 DR. MAHA AL-SEDIK
Introduction: • Cancer of the cervix is one of the most preventable cancers, • Because most of the cellular changes which may lead to carcinoma can be detected and accordingly treated at an early stage before progression.
In most cases, cervical carcinoma develops slowly ( over a period of up to 10 years ) • It passes into different pre-neoplastic conditions before it reaches the cancer stage; termed dysplasia or cervical intraepithelial neoplasia (CIN).
Microscopic examination of the exfoliated cells from the uterine cervix through what we call the Pap Smear test should therefore lead to the detection of these pre-neoplastic lesions in their earliest stages.
Pap Smear test : Is a simple procedure in which a small number of cells are collected from the cervix and sent to the laboratory where they are tested for anything abnormal. No anesthesia is required, and the diagnosis could be obtained within few minutes.
Advantages of Pap Smear: ● It is painless and simple. ● Does not cause bleeding. ● Does not need anesthesia. ● Can detect cancerous and precancerous lesions. ● Can identify non-specific and specific inflammations. ● Can be carried out as an outpatient procedure.
Who to screen? It is recommended that all women between the ages of 18 and 70 years, who have ever had sexual activity, are advised to have the test every 2 years.
Screening frequency: Yearly until three consecutive normal pap smears, then may decrease frequency to every two or three years.
When to stop routine screening • Age 70 • Three consecutive normal pap smears. • No abnormal pap smears in last 10 years. • Hysterectomy for benign disease.
Original Squamous Epithelium: • Vagina and outer ectocervix. • Columnar Epithelium: • Upper and middle endocervicalcanal. • Single layer of columnar cells arranged in folds.
Squamocolumnar Junction: • Where squamous and columnar epithelium meets. • Most often found on ectocervix. • Less apparent over time with maturation of epithelium • Can migrate proximately as columnar epithelium is replaced by metaplastic squamous epithelium.
New SquamocolumnarJunction: • Border between squamous epithelium and columnar epithelium • Found on ectocervix or in endocervical canal • Transformation zone: • Also called ectropion, area between original SC junction and new SC junction due to regenerative metaplastic response. • Site of > 90% of squamous cell carcinomas and dysplasia.
brush Slide Ayre spatula
Steps: 1- Insert Speculum to visualize entire cervix if possible and carefully remove any obscuring discharge.
3- Sample endocervix with gentle cytobrush rotation: • Insert ~ 2 cm (until brush is fully inside canal) • Rotate only 180 degrees (otherwise will cause bleeding)
5- Fix rapidly with spray or liquid fixative: The cell samples are smears and must be well fixed in 95% alcohol.
Papanicolaoustaining: The originalstainingprocedurewasdevelopedbyGeorge N. Papanicolaou. The three main advantages of this staining procedure are: (1) Good definition of nuclear detail. (2) Cytoplasmic transparency. (3) Indication of cellular differentiation of squamous epithelium.
There arefourmainstepsinthestainingprocedure: • Fixation. • Nuclearstaining. • Cytoplasmicstaining. • Clearing.
Fixation :95% alcohol. • Nuclearstaining: • The haematoxylin nuclear stain is a natural stain which has been used for over 100 years in histology. It has affinity for chromatin, attaching to sulphate groups on the D.N.A. molecule.
Cytoplasmicstaining.: • 1-First OG-6 counterstain ( orange 6) . It stains keratin. Its original role was to stain the small cells of keratinizing squamous cell carcinoma.
2- Cytoplasmic staining E.A. 50 (Eosin Azure) and rinsing off in 95% alcohol. Eosin gives a pink colour to: (a) cytoplasm of mature squamous cells (b) nucleoli (c) cilia
Clearing. • Clearing, in xylol results in cellular transparency . • Xylolis the commonest clearing agent . • Xylolis colourless, chemically non-reactive and has almost the same refractive index as glass which is important to give the best possible transparency of the image.
1 70° ethanol2 mn or more 2 water2 mn 3 haematoxylinstain 4 mn
4 water5 mn 5 95° ethanol2 mn 6 Orange G 3 mn
95° ethanol2 mn 7 EA-50 4 mn 8 9 95° ethanol2 mn
Vaginal smear showing epithelial cells. Epithelial cells are large. Notice the neutrophils in the slide for size comparison.