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- 2. a branch ofCytology which deals wit the microscopic study of cells that have been desquamated from the epithelial surfaces. recommended for : Detection of malignant cells or precancerous lesions in the body Detection of asymptomatic or precancerous cervical lesions in women Assessment of female hormonal status in case of sterility and endocrine disorders Determination of genetic (phenotypic) sex Detection of the presence of infectious microorganisms
- 3. CYTOLOGY SPECIMENS 1. peritoneal,pericardial and pleural fluids 2. CSF 3. Nipple discharge 4. Bronchial brushings / washings 5. Sputum 6. Gastric washings 7. Urine sediment 8. Prostatic secretions 9. Cervicovaginal (paps) smear
- 4. • Collection Procedure: -Using standard paracentesis technique. A minimum of 10 mL of specimen is desirable for optimal cytologic evaluation. Heparin may be added to the specimen to reduce clotting. - Place three (3) units of heparin per mL capacity of the collection container. Gently agitate to thoroughly mix the specimen and heparin. - Always Submit the specimen along with the completed cytology request form. - The specimen should be refrigerated until transported to the lab. BODY FLUIDS BODY FLUIDS
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- 8. CSF is usuallyobtained through a lumbar puncture (spinal tap).
- 10. • Gently stripthe sub-areolar area and nipple with the thumb and forefinger. • Place the slide upon the nipple and draw it quickly across the nipple. ***If 1 drop is obtained, get another slide and do the pull- apart technique. • Immediately immersed the slide into a bottle of 95% isopropyl ROH or use spray fixative. NIPPLE DISCHARGE NIPPLE DISCHARGE
- 11. a. bronchial washing b.bronchial lavage c. bronchial brushing
- 12. - Bronchoscopically-directed brushingof identified lesion. •Collection Procedure: - Using standard bronchoscopy technique, identify the lesion in question and obtain a brushing sample of the lesion. - Gently apply the sample on to glass slide and immediately immerse slide into 3% glacial acetic acid alcohol fixative. - The brush tip should also be cut off into the solution. Bronchial Brushings
- 13. Bronchial Washings •Specimen: Bronchscopically-obtained washing(preferable at least 10 mL) of the bronchi in the region of the suspected lesion. •Collection Procedure: -Using standard bronchoscopy technique, lavage the distribution of the bronchus to be sampled. -Collect the wash in a clean container. Label the container with correct patient information and
- 14. NORMAL -obtain at threeconsecutive morning sputum specimens by deep cough method. -Collect the sample in a wide-mouth container containing Saccomano fluid (50% EtOH and 2% carbowax) INDUCED - Inhalation of aerosol solution for 20 mins to produce deep cough sample. - Collect the sample in a wide-mouth container containing Saccomano fluid (50% EtOH and 2% carbowax)
- 16. Washings (Esophageal, Gastric,Other) •Specimen: Endoscopically obtained washing (preferably at least 10 mL) of the region of the suspected lesion. Procedure Patient should fast overnight or for a minimum of six hours prior to the procedure. - Using standard endoscopy technique, lavage the area of interest using a physiologic solution. Aspirate the solution and place in a clean container. - If transport of the specimen will be delayed more than four (4) hours, the specimen should be refrigerated until transported to the lab.
- 17. The endocervical mucuswill prevent air-drying during collection of the subsequent cervical component. Using the extended-tip spatula scrape material from the whole circumference of the cervix. Withdraw the spatula and spread the collected material quickly and evenly onto the slide . Fix Immediately .
- 19. smears should befrom fresh material see requisition form (patient’s ID: name, age; date and type of specimen requested label the slide -Methods of Smear Preparation: 1. streaking 2. spreading 3. pull apart 4. touch or impression smear
- 20. STREAKING used for preparingmucoid secretions vaginal secretions, sputum and gastric content) use a spatula, dissecting needle or applicator stick and streak in a zigzag fashion
- 21. SPREADING - used forthick mucoid secretions (smears of fresh sputum and bronchial aspirates)
- 22. PULL APART - forserous fluids, concentrated sputum, and enzymatic lavage form the GIT, smears of urinary sediment, vaginal pool and breast secretions
- 23. TOUCH IMPRESSION TOUCH IMPRESSION Impressioncytology being Impression cytology being collected From a patient , using collected From a patient , using a sterile glass slide with polished a sterile glass slide with polished edges. edges.
- 24. FIXATION exfoliated cells decomposerapidly which may destroy cellular and nuclear details, in turn will give inadequate results for diagnosis. •COMMON FIXATIVES 1. equal parts of 95% EtOh and ether 2. 95% EtOH 3. Carnoy’s fluids 4.equal parts of tertiary butyl alcohol and 1 part 95% EtOH 5. SCHAUDINN’S FLUID – sat. aq. Hg2Cl, absolute HoAc 6. MeOH – for dried films
- 25. 1. PAPANICOULAOU orPap’s Smear Advantage: - transparent blue staining of cytoplasm is observed - excellent nuclear staining - color range is predictable and of great value in identification of cells Disadvantage: - procedure is lengthy and complicated - does not give accurate acidophilic index
- 26. Stains for Pap’s: 1.Harris Hematoxylin 2. OG 6 Stain Orange Green 6, 0.5 solution in 95% ROH 100 ml Phosphotungstic acid 0.015gm 3. EA 50 light green SF, yellowish 0.1% solution in 95% ROH 45ml Bismarck brown 0.5 in 95% ROH 10 ml Eosin Y, 0.5% in 95% ROH 45 ml Phosphotungstic acid 0.2 gm Lithium Carbonate. Sat. aq. Solution 1 drop *** EA 50 is comparable to EA 36 *** EA 65 differs from EA 50 or EA 36 only with respect to the concentration of the light green stock solution
- 27. Procedure for Pap’sStain: 1. Fix in ether-ROH and pass thru 80% ROH, 40% ROH and distilled H2O. 2. Stain in Harris Hematoxylin for 4-5 minutes. 3. Wash with H2O. 4. Pass thru 0.25% HCl in 50% ROH. 5. Immerse in 1.5% NH4OH in 70% ROH for 1 minute. 6. Rinse in 70% ROH and pass thru 80% and 95% ROH.
- 28. Procedure for Pap’sStain: 7. Stain with OG 6 for 1.5-a minutes. 8. Pass thru 3 changes of 95% ROH. 9. Stain with EA 65 or EA 50 for 3 minutes. 10. Pass thru 3 changes of 95% ROH. 11. Dehydrate and clear in: a. absolute ROH, b. equal parts of ether and absolute ROH, c. 2 changes of xylol 12. Mount in Canada Balsam.
- 29. Results: Cytoplasm – eitherbright red or greenish blue vesicular nucleus – blue pyknotic nucleus – dark blue to black bacteria – dark blue mycelia – violet Trichimonas vaginalis – pale greenish blue blob of cytoplasm
- 30. Cytoplasm – eitherbright red or greenish blue vesicular nucleus – blue pyknotic nucleus – dark blue to black
- 31. Trichimonas vaginalis –pale greenish blue blob of cytoplasm
- 32. Altered nuclear-cytoplasmic ratios Hyperchromasia Increasedmitotic activity Atyical mitoses CHANGES IN MALIGNANCY CHANGES IN MALIGNANCY
- 33. Multinucleate cells – withirregular hyperchromatic or bizarre nuclei should be suspicious, Anisokaryosis – considerable variation in nuclear size and shape is common in malignant cells. Giant single nucleus (polyploidy) – often seen in malignant cells, polypoidic cells may also occur in benign conditions, especially in thyroids of older women.
- 34. CYTOPLASMIC CHANGES - cellsof squamous carcinomas frequently show a tendency to cytoplasmic eosinophila. - adenocarcinoma cells may enclose, endometrial and colonic cancers) - cytoplasmic vacuolation is common in adenocarcinoma cells, but may be also be seen in endometrial cells following cutterage.
- 35. In general: - malignantcells show reduced cohesiveness, possibly related to a defect of the intercellular “zippers” i.e., desmosomes. -Cancer cells are larger than their normal counterparts and frequently show bizarre and grotesque shape. Occasionally: - exfoliated cells from epithelial tumors assume a greatly elongated, fibrocyte-like appearance.
- 36. CELL PATTERN •Examine forthe small groups or clusters of cells -Oblivious patterns e.g. acini in adenocacinoma arising in glandular tissues -Rosettes in neuroblastomas and ependymoma -Stratification in squamous epithelial growths -Whorls in mesotheliomas If there are no patterns focus up and down the on cell clumps -In strips epithelium, irregular stratification of anisokaryotic, hyperchromatic cells are helpful in diagnosing carcinoma of cervix and bronchus.
- 37. OTHER CRITERIA 1. Inbronchial secretion smear, abundant lymphocytes are common in presence of malignancy, but may also be found in certain inflammatory conditions and in leukaemia. 2. The presence of old blood and blood pigments is a minor indirect clue, but has many other etiologies.
- 38. VAGINAL CYTOLOGY Vaginal cytologyis a type of endocrine assay. Tracking changes in the morphology of desquamated vaginal epithelial cells provides a convenient means of assaying changes in estrogen levels.
- 39. •Vaginal smears maybe taken regularly and often. •Hormonal changes are best mirrored in the upper third of the vagina. •They can also be taken from the lateral walls because their more accessible and less likely to be contaminated by cellular debris or discharge.
- 41. -SUP{ERFICIAL CELLS -Large (30-60u) -Polyhedralflat cells -Cytoplasm: may be acidophilic or basophilic -Presence of small dark pyknotic nuclei (less than 6u)
- 42. Anucleate cells areabnormal which may be derived from: 1. smear contamination by the cells from the vulva 2. epidermization of the vagina or cervix resulting from prolapse 3. leukoplakia of the cervix 4. ruptured membranes in pregnant women 5. marked hyper-estrinism
- 43. INTERMEDIATE CELLS - medium large(20-30u) - Polyhedral or elongated - Cytoplasm: basophilic with vacuoles - Vesicular nuclei (6-9u)
- 44. - Boat-shaped intermediatecells with a strong tendency to fold and curl their edges. - Expression of the combined estrogen-progesterone effect -found in the latter half of menstrual cycle, during pregnancy, menopause - -may also be found as a result of abnormal androgen stimulation, either endogenous or exogenous
- 45. -PARABASAL CELLS -Round tooval cells -Smaller than intermediate (15-25 u) -Thick -“sunny-side up” like cells -Have strong basophilc cytoplasm and vesicular nucei (6-9 u ) -Found from 2 weeks of age to puberty, after childbirth, abortion or miscarriages and after menopause.
- 46. -ENDOCERVICAL CELLS -Slightly cylindricalappearance -occurs in groups and strips of three or more cells -cytoplasm: deeply basophilic the that of the parabasal cells
- 47. -ENDOMETRIAL CELLS -Found duringmenstruation period ( in groups) and 1-4 days after the cessation of the period (single) -Endometrial stromal cells: seen in tight clusters of small, oval dark cells; Glandular cells: slightly larger. -Nucleus: small and moderately dark -Cytoplasm: basophilic and maybe vacuolated
- 48. - “Lactobacillus acidophilus” -Gram+ slender rod bacteria - -Predominant organism of the vaginal normal flora: establishes the low pH that inhibits the growth of pathogens -Stains pale blue to lavender -Energy is obtained by the fermentation of glycogen derived from disintegrating epithelial cells -Numerous in the luteal phase and during pregnancy
- 49. CYTOLYSIS IN VAGINALCYTOLOGY - Infection - estrogen -low vaginal pH (below 4.2) Occurs in the last trimester of pregnancy, more common in diabetic patients shows large numbers of naked nuclei and very abundant Doderlein bacilli.
- 50. The BETHESDA SYSTEM SpecimenAdequacy Satisfactory Limited Unsatisfactory General Categorization: Negative for Intraepithelial lesion or malignant cell Epithelial cell abnormality Descriptive Diagnosis: Atypical squamous cells of unknown significance Low grade squamous intraepithelial lesion High grade squamous intraepithelial lesion Squamous Cell Carcinoma Glandular cell abnormality Atypical glandular cells Adenocarcinoma Others
- 51. Pap smear specimensare considered satisfactory for interpretation if there are: •Adequate numbers of well-visualized squamous cells present •Adequate numbers of well-visualized endocervical cells or squamous metaplastic cells (from the transformation zone). •Less than 50% of the cells obscured by blood or inflammation •Properly labeled specimens Specimen Satisfactory
- 52. Pap smear specimensare considered unsatisfactory for interpretation if there are: •Inadequate numbers of well-visualized squamous cells present •Inadequate numbers of well-visualized endocervical cells or squamous metaplastic cells (from the transformation zone). •More than 75% of the cells obscured by blood or inflammation •Improperly labeled specimens Usually, these smears are recommended for repeat sampling.
- 53. Specimens subjected torejection 1. Specimen is submitted without a requisition. 2. Specimen is not labeled with the patient name. 3. The patient name (or other identifying information) on the specimen and requisition do not correspond. 4. The specimen is labeled appropriately but the requisition is not labeled. 5. The specimen slide(s) is (are) irreparably broken. 6. Specimen is submitted from an unauthorized source.
- 54. NEGATIVE FOR NEGATIVEFOR INTRAEPITHELIAL INTRAEPITHELIAL LESION LESION
- 55. * Atypical squamouscells -of undetermined significance (ASC-US) -cannot exclude HSIL (ASC-H) Atypical squamous cell (ASC-US)
- 56. * Low gradesquamous intraepithelial lesion (LSIL) (encompassing: HPV / mild dysplasia / CIN 1) Low Grade SquamousIntraepithelial Lesion
- 57. •High grade squamousintraepithelial lesion (HSIL) (encompassing: moderate and severe dysplasia, CIS, CIN 2 and CIN 3) •- with features suspicious for invasion (if invasion is suspected) Severe Dysplasia or CIS
- 58. * Squamous cellcarcinoma
- 59. Adenocarcinoma in situof the cervix (upper left), next to normal glandular epithelium (lower right).
- 60.
- 61. REFFERENCES REFFERENCES Pathology.jhu.edu Pathology.jhu.edu www.pathologyoutlines.com www.pathologyoutlines.com icytologywordpress.com icytologywordpress.com er.wikipedia.org er.wikipedia.org