Techniques in diagnostic pathology Flashcards
What are the different types of fixatives
formalin,bouin(mercury based), carnoy(alcohol based), 3%glutaraldehyde
What step do you do after fixation for non frozen tissue?
Embed tissue in wax, cut from wax and dewax with xylene, deparaffinze and rehydrate with alcohol and stain
When do you administer adjuvant therapy?
When incomplete resection, vascular invasion, advanced stage
What’s the difference between ptx and pt0
PTX can’t be assessed histologically
pt0- no histological evidence of primary tumour
Difference between V1 and V2
venous invasion microscopic and venous invasion macroscopic
How to deal with frozen tissue
Put OCT
What factors do we evaluate when looking at an invasive carcinoma?
Histological type, grade, tumour size, lymph node stage, vascular grade
What does histological grade asses? And how is it done?
degree of differentiation
By looking at morphological characteristics such as size, nuclear pleomorphism, mititic count, tubule formation
Does histological grade correlate to patient survival?
yes
What methods are often used to study tissues and is each one associated iwth fresh, fixed tissue or both?
PCR-FFPE(300-500bp) or fresh tissue Pyrosequencing RT-PCR-FFPE(100BP) or fresh tissue FISH-fresh or fixed spectral karyotype-fresh flow cytometry- fresh expression profiling-both CGH-both, but needs amplification step for FFPE tissue Western blotting-fresh MS-fresh metabolomics-fresh
Why is immunocytochemistry important?
Evaluating subtypes of tumour differentiation of tumours secretory products identification of metastasis prognostic markers
How is antigen retrieval done?
Unmasking epitopes by formaldehyde fixation
digestion with proteolytic enzymes
microwave energy
cook in heat and pressure
What are the problems with ICH?
antigens might crosslink, not be specific, they can be lost, cross reactivity, appropriate concentration of antibody has to be used, entrapment of normal tissue in tumour cells
release of soluble proteins
Give examples of IHC diagnostic markers
CK20-squamous cell&adenocarcinoma
PAS-mucin
CEA-present in carcinoma
CK7-Lung.breast, ovary cancer, ductal, glandular,transitional epithelium
Give the therapy most appropraite to the following breats cancer patients and prognosis ER,PR +/+ ER-, ER+, ER low
ER,PR +/+ better response
ER-, poor prognosis, give chemotherapy
ER+, endocrine therapy, better prognosis
ER low both endocrine and chemotherapy
What does CISH do? And why is is it better than FISH?
Allows direct comparison between morphology and gene amplification. Doesn’t need fluorescent microscopy and is quick
What’s electron microscopy for?
soft tissue tumours, muscle disease, peripheral nerve tumours, renal pathology, tumour differentiation, paedriatic tumours,diagnostic ultrastructural features
Whta’s immunofluorescence and confocal microscopy used for?
To see if proteins are being co-expressed, interacting, metabolism, cell membrane kinetics,
What does PAS, mason fontana, oil red O, PAS, alcian blue
PAS-detection of mucin in poorly differentiated tumours
Mason fontana-detection of melanin in metastatic deposits
Oil red O-lipid and specifc markers differentiating between tumours
PAS, alcian blue- metaplasia in GIST
What is intra-operative smears
swap taken from CBS fluid or spine to assess lesions by seeing nuclei abnormality, blood vessel sizes, necrosis
What are the different methods of doing immunohistochemistry analysis?
Peroxidase antiperoxidase immune complex peroxidase enzyme AB conjugate labelled antigen labelled bridge biotinylated primary AB Biotinylated peroxidase avidin biotinylated peroxidase complex
How is spindle cell sarcoma diagnosed using IHC? which markers do they look for?
Vementin-IF of mesenchymal cells LCA-leukemia common antigen AE1-cytokeratin CD31-endothelial cells CD34-soft tissue
What are the charactersitics of spindle cell lesions? What is it diagnosed as?
Proliferation rate is low, express BCL-2, cd34, CD99 but not EMA,SMA(smooth muscle actin),s100, hmb45(found in melanoma)
solitary fibrous tumour
Markers for lymphoma
CD3-T-cells
CD20-B-cells
BCL-2-follicular lymphoma
MIB-1- cell proliferative marker
What does cytogenetics study normally?
pedriatic tumours, hematological malignancies, clonality studies
What kind of probes are used for FISH?
centromere, telomere probes, chromosome paints, microdeletion probes, gene amplification, translocation junction
What kind of information do we get from FISH?
Microdeletions, copy number of chromosomes and genes, DM&HSR, translocation
How is neuroblastoma diagnosed?
Loss of material from 1P which has tumour suppressors
amplification of n-myc(>10) located at 2p24
gain of 17q
overexpression of BCL2
What are the characteristics of ewing sarcoma?
t(11;22)q(24:12)
Fli1-EWS fusion gene
fli-DNA binding domain
EWS-promoter region
11q24- DNA binding domain+transcription activator
22Q12-RNA bidning domain and promoter region
How is molecular studies done usually?
PCR, RNA extraction, proetin extraction
