Histopathology Flashcards
What does a histopathologist do?
Deals with tissues, examines the sections, and notes the architecture of the tissue and identify what it tells us about a particular condition
What does a cytopathologist do?
Deals with cells, takes them from the patient and prepares them for examination and then delivers their expert diagnosis on the cell sample.
What are the four tissue samples a histopathologist will work with?
- Biopsies
- Resection specimens
- Frozen Sections
- Post-Mortems
What are real-time examinations used for?
for surgical processes
What are biopsies?
Small sections of tissues removed from a patient
What solution are biopsy tissues placed in for preservation?
Formalin
How are biopsy samples preserved?
The formalin they are preserved in cross links the proteins and fixes the tissue
What are biopsy tissues embedded in to allow very thin sections to be cut?
Paraffin wax (2-3 micrometers thick)
What cuts the biopsy tissue into very small sections?
Microtome
What stain is used to identify the nuclei and cytoplasmic granules of leukocytes within tissues?
Haemotoxylin and Eosin (H&E)
What is used to stain acid fast bacteria?
Ziehl-Neelsen stain
What stain is used to aid the diagnosis of tuberculosis infection?
Ziehl-Neelsen
How long does the results of a biopsy take to reach the clinician?
2-3 Days
How long does it take for the results of a frozen section to reach the clinician?
30 Minutes
How long does it take for results of a resection specimen to reach the clinician?
5-7 days
What are resection specimens?
specimens are taken from tissue that has been removed as part of a surgical procedure and can be processed as for a biopsy.
What are resection specimens primarily used to do?
Look at the stage of disease
Why may tissue samples from resection specimens be donated to biobanks?
They may be used to inform genomic studies of the disease process
What is a frozen section?
A sample taken during a surgical procedure and examined by pathologists in real time
What is used to freeze the tissue?
Cryostat (a machine)
What happens to the frozen tissue from the frozen section?
it is frozen, cut, mounted on slides and stained as for biopsies
What are the requirements for tissue samples to be frozen?
Fresh and free of preservatives like formalin
What is a fine needle aspirate?
When a fine needle is inserted into a lesion and used to aspirate the cells out the lesion for anaylsis
Why is fine needle aspiration useful?
The needle can easily penetrate relatively inaccessible tissues without the need for surgery
e.g., thyroid nodule
What is the downside of fine needle aspiration?
the cytopathologist is only looking at cells and is unable to comment upon the likely architecture of the tissue.
Give an example of when both histopathology and cytopathology are used?
In the diagnosis of reactive lymphadenopathy
fine needle aspirate of enlarged lymph nodes and skin biopsy
What does Oil Red O stain?
Lipids
What does congo red stain?
ECM components like elastin and collagen
Which molecules are conjugated to the Fc region of an antibody for their use in flow cytometry?
Fluorophores
When requesting a test from the pathology lab to aid with a diagnosis, it is important to appreciate just how long the various procedures take. Put the following pathology lab tests in order of the time taken to get the result back to the clinician (quickest to slowest).
- Resection tissue
- Biopsy
- Frozen section
3 - 2 - 1
The immunological lab offers how many different tests, and of those how many are used to detect the level of specific antibodies circulating in patients?
86
53
What are examples of the antibody tests offered by the immunological lab?
dsDNA
systemic lupus erythematosus (SLE)
Sjogren syndrome
Rheumatoid arthritis
What is Kaposi’s sarcoma?
Rare cancer and is often indicative of immunodeficiency (AIDS)
Spindly resembling cells evidently penetrates the collagen fibres
What four forms can antibody conjugations come in?
- Enzymes
- Fluorescent probes
- Magnetic beads
- Drugs
What 2 types of detection can antibodies be used for?
Indirect - antibody with marker binds to another antibody which binds to antigen
Direct - antibody with marker binds to antigen
What are conjugations?
Attachments to the Fc region of the antibody
What are two examples of enzyme attachments?
Peroxidase and alkaline phosphatase
When may enzyme attachments be used clinical?
The detection of oestrogen receptors on breast cancer tissue receptors
What is multiplexing and what does it allow?
When several antibodies with different fluorescence are used, allows the measurements of several molecules in a single clinical sample
What part of the antibody are conjugates attached to?
The Fc region
How does enzyme conjugation work?
Antibody binds to antigen, and colourless substrate is added, the colourful product can be detected wherever the immune complex is (formed)
Why do fluorescent probes allow for the rapid measurement of the levels of molecules within a sample?
because the colour is already there and unlike enzyme conjugation does not need to be formed
Which is a good choice for an antibody conjugate when you have a small sample?
Fluorscent probes as you can performe multiplexing
What can magnetic beads be used for?
The purification of cell types
Give an example of how magnetic beads are used as a clinical conjugate?
deplete the bone marrow of T cells before bone marrow graft
What can drugs as an antibody conjugate be used to do?
Directly deliver the drugs to the cells that are malfunctioning
Describe an example of the clinical use of drugs as an antibody conjugate?
Kadcyla, anti-HER antibody - used in the treatment of breast cancer as HER2 is over-expressed in over 30% of breast cancers
It is linked to the cytotoxic chemical emtansine
What makes antibodies useful as a diagnostic tool?
The unique specificity of the antibodies
What other molecules can antibodies detect?
Proteins, carbohydrates and lipids
Describe how indirect detection of antibodies works?
The antibody with the antigen bound to it is detected by a secondary antibody which is conjugated with an enzyme of fluorescent probe
What can manufactured antibodies be used for?
Blood group serology
Immunoassays
Immunidiagnosis
What can immunoassays be used for?
Detection of hormones or circulating antibodies/antigens
What can be detected via immunodiagnosis using manufactured antibodies?
infectious diseases
antibody levels
IgE antibodies suggestive of allergic phenotype
Infectious diseases due to presence of anti-disease antibodies like HIV
What is immune complex glomerulonephritis?
Disease caused by the presence of IgG-antigen immune complexes and Complement C3 from kidney biopsy
What does flow cytometry notably allow the detection of?
Lymphocyte subpopulations
How does flow cytometry work?
- Cells labelled with differently conjugated antibodies
- These cells then run as a stream of single cells through laser beams
- The colour of the light emitted and the scatterpatterns of the laser beam can denote the identify of the cell surface molecules expressed and the size and granularity of the cells and therefore the type and of cell
Why is flow cytometry highly informative to?
The nature of a particular inflammatory reaction, as the tagged antibodies allow the detection of specific white blood cells like CD4 helper cells etc
Anti CD3+ antibodies are used to detect what cell?
T cell - pan T cell marker
Anti CD4+ antibodies are used to detect what cell?
T helper cells
Anti CD8+ antibodies are used to detect what cell?
Cytotoxic T cells
Anti CD19+ antibodies are used to detect what cell?
B cells
Anti CD56+ antibodies are used to detect what cell?
NK cells
What does ELISA mean?
Enzyme Linked ImmunoSorbent Assay
What is the purpose of ELISA?
Used to determine the precise concentrations of molecules in a sample
How does ELISA work?
- Clinical samples (e.g. adheres to a plastic plate)
- Probe with specific antibody raised against the molecule of interest.
- Enzyme conjugation generates a coloured product.
- Wash away excess, unbound antibody
- Reference to standard curve (absorbance of light by coloured product) , determine precise concentrations of the molecule in the sample.
