Histopathology Flashcards
What is a histopathologist?
They deal with tissues and cross-examine sections noting the architecture of tissue and what it tells us about a condition
What is a cytopathologist?
They deal with cells
They’re often taking cells from a patient, preparing them for examination and then delivering diagnosis
What tissue samples does a histopathologist work with?
Biopsies
Resection specimens
Frozen sections
Post-mortems
What are biopsies?
Small sections of tissue removed from the patient and placed in formalin solution which preserves the tissue by cross-linking proteins
They’re embedded in paraffin wax to allow thin sections to be cut by a microtome
These are mounted onto a glass microscope slide for further preperation
What questions can microscopic examination of the biopsy answer?
Is the tissue normal?
Is the tissue inflamed and what’s the likely cause?
Is the tissue cancerous and if it is what type of cancer?
What is the importance of chemical stains?
Often employed with the tissue section to aid the identification of cells within the biopsy
What are examples of stains and what are their use?
Haemotoxylin and Eosin (H&E) staining can be used to identify the nuclei and cytoplasmic granules of leukocytes within tissues
Ziehl- Neelsen stain will stain acid-fast bacteria red aiding diagnosis of Tb
What are resection specimens?
Taken from tissue thats been removed as part of surgical procedure and can be processed for a biopsy
Resections are mostly used to look at stage of disease
What questions can be answered with a resection?
For cancer:
How far is the disease progressing?
Has the cancer penetrated the bowel wall, spread to lymph nodes or liver?
How else can tissue from resection specimens be used?
Can be donated to biobanks and used to perform genomic studies of disease process
What are frozen sections?
Taken during surgical procedures and are examined in real time while patient is being operated upon
Freshly taken tissue is frozen by machine called cryostat, cut then mounted onto glass slide and stained for biopsy
procedure can give rapid diagnosis in minutes
Tissue must be fresh and free of preservatives such as formalin
What questions can we answer from frozen sections?
Is the tissue cancerous?
Has all the cancerous tissue been removed?
Is there another pathological process going on?
What are fine needles used for and how?
Used for fine needle aspirates
Fine needle can be used to get into lesson and suck out (aspirate) the cells which can then be analysed for a smear
Very powerful technique as needle can penetrate relativeness inaccessible tissues e.g. thyroid nodule and assess suspect mass without need of surgery
What is a downside to fine needle aspirates?
Cytopathologists only look at cells and are unable to look at architecture of tissue
What is the timescale for a result for different histopathological samples?
Frozen section: 30 mins
Biopsies: 2-3 days
Resection specimens: 5-7 days
What can immunohistochemistry be used for?
Can be used to identify key molecules within tissue
How can we identify an endothelial tumour?
An antibody recognising endothelial cell marker CD31 on surface of endothelial cells allows us to identify the biopsy as an endothelial tumour
What can high levels of antibodies recognising dsDNA be associated with?
System lupus erythematosus
SJrogen’s syndrome
Rhematoid arthritis
What do manufactured antibodies do?
Can be used to specifically detect molecules in process of immunohistochemistry
Attachments (conjugates) to Fc region of antibody can be made permitting a wide variety of uses
What are examples of antibody conjugates?
Enzymes: peroxidase, alkaline phosphatase
Fluorescent probes: allow rapid measurement of levels of molecules in sample. Multiplexing (using several antibodies with different fluorescents) can allow measurement of several molecules in single sample
Magnetic beads: e.g. purification of cell types. With use of magnets, cells can be depleted of contaminants
Drugs: the biological kadcyla, an anti-HER2 antibody linked to cytotoxic chemical emtansine
How are antibodies used as a diagnostic tool?
Unique specificity of antibodies for their target is beneficial for testing
Antibodies can be used to detect proteins and other biological molecules such a s carbohydrates and lipids
This can be direct detection of antigen or via detection of unlabelled antibody by secondary antibody conjugated with a fluorescent probe or enzyme
What are examples of antibody uses for diagnostic tools?
Blood group serology
Immunoassay e.g. detection of hormones, circulating antibody/antigen
Immunodiagnosis e.g. infectious diseases and antibody levels
What is an ELISA test?
Enzyme linked immunosorbant assay
- Clinical samples are allowed to adhere to a plastic plate
- Then they’re probed with a specific antibody raised against the molecule of interest
- Antibody is conjugated with an enzyme which can turn a colourless substance a particular colour
What is flow cytometry?
Allows detection of specific cells, most notably lymphocyte subpopulations using fluorescent conjugated specific antibodies
What lymphocyte populations may be detected by flow cytometry?
Anti- CD3+ = pan T cell markers Anti - CD4+ = T helper cells Anti- CD8+ = Cytotoxic T cells Anti- CD19+ = B cells anti- CD56+ = NK cells
How does flow cytometry work?
- Cells are labelled with a different conjugated antibody
- Then run as a stream of single cells through a laser beam
- Colour of light emitted and the forward or side scatter of the laser beam can tell us the identity of the cell surface molecule expressed and size and granularity of the cells
