Cytopath Flashcards
SCC on Pap
Approximately __% of women with an LSIL Pap result prove to have HSIL (CIN2/CIN3) on biopsy.
__% of untreated LSILs progress to invasive squamous cancer.
Most LSILs regress. Approximately 18% of women with an LSIL Pap result prove to have HSIL (CIN2/CIN3) on biopsy. Less than 1% of untreated LSILs progress to invasive squamous cancer.
CELL SIZE
The size is compared to a normal cell or its nucleus (RBC, PMN, lymphocyte, intermediate squamous cell)
Small:
Medium:
Large:
Giant cell:
CELL SIZE
The size is compared to a normal cell or its nucleus (RBC, PMN, lymphocyte, intermediate squamous cell)
Small: 2 – 2.5x
Medium: 3 – 6x
Large: 6 – 10x
Giant cell: > 10x size of a lymphocyte
NUCLEOLI
- Nucleoli are mostly protein and __
- Chromocenters stain __
- Conspicuous:
- Prominent nucleoli:
- Macronucleoli:
NUCLEOLI
- Nucleoli are mostly protein and stain red
- Chromocenters stain blue
- Conspicuous: seen at 40x / high power
- Prominent nucleoli: seen at 10x / scanning power
- Macronucleoli: about the size of RBCs
STAINS / FIXATIVES
Stains / Fixatives used for FNAs
STAINS / FIXATIVES - I
Stains / Fixatives used for FNAs
- Papanicolaou stain
- Alcohol fixation
- Nuclear stain – hematoxylin
- Cytoplasmic counterstains – OG, EA
- Alcohol and xylene rinses
- Rapid Romanowsky stain
- Air dried smears
- Solutions I, II and III
- H & E stain (alcohol fixed smears)
- Histochemical and Immunohistochemical stains: fixation depends upon stain required
DQ / ROMANOWSKY GIEMSA STAIN
DQ / ROMANOWSKY GIEMSA STAIN
- Azure B
- Eosin Y
- Romanowsky-Giemsa effect
STAINS / FIXATIVES
Effects of Alcohol Fixation
STAINS / FIXATIVES
Effects of Alcohol Fixation
- Alcohol can act as a solvent
- Example: RCC
- DQ retains fat and subsequently can be stained with Sudan III
- Alcohol fixation dissolves fat
- Alcohol immersion may cause cell loss
- Solution: spray fixatives, coated slides
PAPANICOLAOU STAIN
Advantages?
PAPANICOLAOU STAIN
Staining method which depends on degree of cell maturity and cellular metabolic activity
- Advantages:
- Nuclear detail
- Cytoplasmic transparency
- Cell differentiation (differential cytoplasmic staining)
PAPANICOLAOU STAIN
The main steps?
PAPANICOLAOU STAIN
The main steps are:
- Fixation
- Hydration
- Nuclear staining with hematoxylin
- Dehydration
- Cytoplasmic staining with Orange G and EA
- Rinsing, clearing and mounting
PAPANICOLAOU STAIN
FIXATION
PAPANICOLAOU STAIN
FIXATION
- 95% ETOH or equivalent
- Wet fixation (slide immersion)
- Coating or spray fixatives
- Alcohol and carbowax mixture
- Carbowax must be dissolved before staining
PAPANICOLAOU STAIN
STAINS
PAPANICOLAOU STAIN
STAINS
- Nuclear stain
- Hematoxylin
- Chromatin patterns of normal and abnormal cells
- Cytoplasmic counterstains
- Orange G and EA
- Provide cytoplasmic transparency
- Clear visualization through areas of overlapping cells, mucus and debris
PROGRESSIVE VS REGRESSIVE METHODS
PROGRESSIVE VS REGRESSIVE METHODS
- Applies primarily to hematoxylin component
- Progressive: stained until required nuclear optical density is achieved
- Regressive: Overstains entire cell Acid bath to extract excess Greater variability of staining Less “forgiving” % of dilute HCl difficult to control Timing is critical to remove only excess hematoxylin Some cell loss (less suitable for non-GYN samples which do not adhere as well to the slides)
ADEQUACY
Minimum number of squamous cells?
Liquid-based:
Conventional:
ADEQUACY
Minimum number of squamous cells
- Liquid-based: 5000
- Conventional: 8000-12000
UNSATISFACTORY?
UNSATISFACTORY
- Lack of patient identification
- Unacceptable specimen
- Slide broken beyond repair
- Insufficient squamous component > 75% epithelial cells obscured
- Blood, inflammation, drying artifact, other
Gyn CATEGORIES
CATEGORIES
- NILM (negative for intraepithelial lesion or malignancy)
- Epithelial cell abnormality
- Squamous
- Glandular
- AIS
- Adenocarcinoma
- Other
- Other
- Endometrial cells in woman > 40yr*
PARABASALS
STICKY HISTIOCYTES!
LUS, ENDOMETRIUM
- Large and small tissue fragments
- Glands
- Stroma (oval, spindle-shaped)
IUD Changes
ASC (ASCUS)
- Management?
- ASC rates less than _% of all PAP cases
- Labs with high-risk populations:
- ASC/SIL ratio should not exceed __ (median ratio in US labs is __)
ASC (ASCUS)
- Suspicion of SIL – LSIL
- Reflex HPV testing (if +, colposcopy + directed biopsy)
- ASC rates less than 5% of all PAP cases
- Labs with high-risk populations:
- ASC/SIL ratio Should not exceed 3:1 (median ratio in US labs is 1:5)
ASC-H
- __% of all Pap tests
- Management?
ASC-H
- 0.3% of all Pap tests
- Higher rate of histologic CIN 2/3 than ASC-US
- Refer for coloposcopy regardless of HPV status
ENDOMETRIAL CARCINOMA
- 3-D groups
- Nuclear enlargement
- Nucleoli
- Hyperchromasia
- Scant cytoplasm
- Cytoplasmic vacuoles
- Finely granular (watery) diathesis
ENDOCERVICAL vs. ENDOMETRIAL CA
ENDOCERVICAL vs. ENDOMETRIAL CA
Endocervical CA
- More cells
- Larger
- Columnar
- Preserved
- Rosettes
- Crowded sheets
- Granular
- PMNs rare
Endometrial CA
- Less cells
- Smaller
- Rounded
- Degenerated
- Balls
- Molded groups
- PMNs common
PSAMMOMA BODIES
- Infrequently found in cervical smears
- Associated with both benign and malignant conditions
- IUCD Endosalpingiosis
- TB endometritis
- Benign endometrial and ovarian lesions
- Serous papillary carcinoma of the ovary or peritoneum and endometrial malignancies
Chlamydia
COCKLEBURRS CRYSTALS
- Radiate arrays of crystalline material often surrounded by macrophages
- Thick club-like spokes
- More likely to be seen in smears from pregnant women
- No significance
- D/D:
- Haematoidin crystals (finer crystalline rays)
- ‘Sulphur granules’ of Actinomyces
Barr Body
LOW ESTROGEN STATES
LOW ESTROGEN STATES
- Premenarche
- Postpartum
- Post menopause
- Turner syndrome
- S/P bilateral oophorectomy
- Basal
- Parabasal cells
- Transitional cell metaplasia (grooves)
Vaginal
PINWORM (Enterobius vermicularis) EGGS
- rectovaginal fistula
Coccidioidomycosis in a liquid-based (ThinPrep) Pap test
- Large round and tear-shaped fungal spherules
- some endospores characteristic for coccioidomycosis
Schistosoma haematobium in a conventional Pap test.
- Several ova can be seen with a terminal spine (circles) that are scattered among numerous inflammatory cells.
CORNFLAKING
CORNFLAKING
- Trapped air bubbles on superficial squamous cells
- Reverse
- Return slide through xylene and alcohol to water rinses
- Restain
- Recoverslip
LIQUID-BASED CYTOLOGY
ThinPrep
SurePath
Advantages:
LIQUID-BASED CYTOLOGY
ThinPrep
SurePath
Advantages:
- Duplicate slides
- Cell blocks
- Testing (HPV, chlamydia, gonorrhea)
- Automated screening
Sputum collection and fixation
SPUTUM
- Formerly most common respiratory tract specimen
- Used in symptomatic patients
- Collect multiple samples over several days
- Early morning deep cough specimens
- Sputum induction
- Fresh, 70% ethanol fixation
- Pick and smear, Saccomanno method (50% ethanol 2% carbowax)
Lung PERCUTANEOUS FNA CONTRAINDICATIONS
PERCUTANEOUS FNA CONTRAINDICATIONS
- COPD
- Emphysema
- Bleeding diathesis
- Patient: uncooperative, coughing
- Severe pulmonary hypertension
- AV malformation
- Suspected echinococcal cyst
Lung
Lung SQUAMOUS METAPLASIA
SIALADENOSIS
- Diffuse, often bilateral swelling
- Peripheral autonomic neuropathy
- Acinar hypertrophy, fat
- Malnutrition, DM, alcoholism, antihypertensive medications
Urine Cytology specimens/fixation/stain
URINE
- Fresh, or refrigerated, 50% ethanol fixative
- Pap stain preferred, 100 – 300 ml, 3
samples over several days optimum
• Voided urine: take 2nd voided urine of
day, after hydrating for 2 – 3 hours +/-
jumping up and down, mid stream, clean
- Bag urine unsuitable
- Do not diagnose malignancy in cells
without intact nuclear membranes
Urine FISH
Leydig cell tumor
Pancreas SIMPLIFIED WHO CLASSIFICATION
Pancreas SIMPLIFIED WHO CLASSIFICATION
- Ductal adenocarcinoma
- Acinar cell carcinoma
- Pancreatic endocrine neoplasm
- Solid-pseudopapillary neoplasm
- Pancreatoblastoma
- Mucus-producing cystic neoplasm
– Intraductal papillary mucinous neoplasm
– Mucinous cystic neoplasm
- Serous cystadenoma
- Nonepithelial tumors
- Metastases
Pancreatic DUCTAL ADENOCARCINOMA
- • Necrotic background
- • Cellular aspirates, predominantly ductal cells
- • Disordered sheets of cells – “drunken honeycombs”
- • Loss of polarity
- • Pleomorphism
- • Squamoid cytoplasm
- • “Tombstone cells” – large, tall columnar cells
- • Nucleomegaly (greater than RBCs)
- • Anisonucleosis (4:1 or greater ratios)
- • Irregular nuclear membranes: grooves, folds, clefts (“popcorn”,
- “tulip nuclei”)
- • Intranuclear cytoplasmic invaginations
- • Abnormal chromatin, thick nuclear membranes
- • Nucleoli
- • Mitoses +++
Pancreatic DUCTAL ADENOCARCINOMA
Special studies:
Pancreatic DUCTAL ADENOCARCINOMA
Special studies:
- Mucicarmine+
- PASD+ mucin in tumor cells
- EMA, Keratin (AE1/AE3), CK 7, polyclonal CEA, CAM 5.2 +
- Some CD10, CK 20 +
- Focal chromogranin, pancreatic enzyme markers +
- CA 19-9 +
- K-ras mutation detection
Pancreas
INTRADUCTAL PAPILLARY MUCINOUS NEOPLASM (IPMN)
- Rare, M > F
- 60 – 70 years old
- More commonly in head of pancreas
- Radiological and clinical input is essential
- Single or multiloculated cysts
- Dilated pancreatic ducts
- Abundant mucin flowing from patulous ampulla at endoscopy
- Cytology
- Rounded, papillary cell islands and fragments
- Mucinous cells
- Potential for malignant transformation – obvious malignant cytologic features
- Positive Stains:
- EMA, CK, Mucin, MUC 2, MUC 1, PCNA
- Ki 67 increased inmalignant tumors
- P53 + in borderline tumors and carcinomas
- Better prognosis than usual pancreatic cancer
Pancreas
MUCINOUS CYSTIC TUMOR
- Rare, indolent, potentially malignant
- Middle-aged women
- Cytology:
- Intracellular and extracellular mucin
- Subepithelial stroma, resembles ovarian stroma
- Mucinous background, moderate cellularity
- Regularly honeycombed epithelial sheets
- Papillary structures, psammoma bodies may be present
- Mucinous epithelial cells – goblet, signetring cells
- Well differentiated – resemble benign endocervical cells
- Mucinous macrophages
- Stroma may be present
- Special studies:
- • EMA +
- • CK 7, 18, 18, 19 +
- • CEA +
- • CA 19-9+
- • DUPAN-2 +
- • Stromal component: vimentin, SMA, desmin, ER, PR, inhibin +
- • MUC 2 +
- • MUC 1 +
Pancreas
PANCREATIC PSEUDOCYST
CALCIUM OXALATE CRYSTALS in lung
CALCIUM OXALATE CRYSTALS
- Aspergillus niger fungal infection
- Produces large amounts of oxalic acid
- Toxic to the blood vessels
- Fatal pulmonary hemorrhages
- Consequently calcium oxalate crystals in sputum or lung specimens is also an indication of an Aspergillus infection of the lung
CLIA Dates
1967
1988
1990-92
1994
CLIA Dates
1967 CLIA Act
1988 Ammended CLIA
1990-92 Rules published in Federal Register
1994 Enforced
CLIA 88 Personnel Standards
- Technical Supervisor
- General Supervisor
- Cytotech
CLIA 88 Personnel Standards
- Technical Supervisor
- MD/DO
- ABP/ASC w state license
- Must review all non-gyn
- Must confirm react/repair and ECA
- General Supervisor (Debbie)
- TS or CT w 3 yr cyto experience
- Day to day supervision of lab
- Document daily workload
- Cytotech
- Graduate from school accredited by CAAHEP
CLIA 88 Workload Limits
- Conventional smear:
- liquid based =
- Location guided
- field of view (FOV) =
- FOV + full manual =
- No less than __ hrs
- __ slides/hr max
- Reassessed q __
- CA & NY __ slide limit
CLIA 88 Workload Limits
- Conventional smear: 100 slides
- liquid based = 0.5
- Location guided
- field of view (FOV) = 0.5 (if negative)
- FOV + full manual = 1.5
- No less than 8 hrs
- 12.5 slides/hr max
- Reassessed q 6 mo
- CA & NY 80 slide limit
CLIA 88 10% Rescreen
CLIA 88 10% Rescreen
- Prospective rescreen of negatives
- TS, GS or designee
- Include % high risk cases
- Document results and remedial measures
CLIA 88 5 yr Retrospective
CLIA 88 5 yr Retrospective
- Review previous negatives in current cases of HSIL or cancer
- Types of error
- None
- Screening - tech
- Interpretation - pathologist
- Document stats
- Only report if it affects current management
CLIA 88 Cyto/Histo Correlation
- What cases need it?
- What’s the #1 cause for non-correlation?
CLIA 88 Cyto/Histo Correlation
- Mandated for HSIL & cancers
- Good QA
- # 1 reason for non-correlation is sampling
CLIA 88 Statistics (4)
CLIA 88 Statistics
- Anual gyn & non-gyn
- Breakdown of gyns, including unsats
- # cases with + 10% rescreen or 5 yr lookback
- Cyto/Histo correlation
CLIA 88 Performance eval and workload limits
- for who?
- how often?
- by who?
- using what?
Performance eval and workload limits
- q 6 mo for techs
- by TS (MD)
- use stats
CLIA 88 Proficiency Testing
- where?
- what’s passing?
- what if you fail (1,2,3)?
Proficiency Testing
- on site
- 10 slide test, 90% to pass
- 1st fail: retest 10 slides in 45 days
- 2nd fail: 20 slide test, all paps reexamined
- 3rd fail: 35 hr CME, 20 slide test
CLIA 88 Regulatory
- Who enforces CLIA?
- Must notify CAP if:
Regulatory
- CMS enforces CLIA
- Contract ASCT for complaints
- CAP > CMS > COLA > JC
- CMS does 10% reinspection after CAP
- Must notify CAP if:
- negative media
- investigation
CLIA 88 Retention
- Glass exfoliative
- FNA
- Report
- Accesion log/worksheet
CLIA 88 Retention
- Glass exfoliative - 5 yr
- FNA - 10 yr
- Report - 10 yr
- Accesion log/worksheet - 2 yr
