Module Flashcards
(156 cards)
What is histology?
The study of tissues and organs, examining the architecture of these structures at the microscopic level and forming links between structure and function of the different tissue and cell types.
What are the disciplines of histology?
- Anatomy –> Structures
- Physiology –> Tissues and cell types (systems)
- Biochemistry –> Cellular reactions/pathways
What are the 4 types of tissues?
Connective, epithelial, muscle and nerve.
What are the general rules of tissue collection?
- Dissection from autopsy/abattoir/ euthanised organisms/plant tissue.
- Biopsy from a clinician.
- Use sharp instruments
- Limited handling
- Within 10 min of slaughter
- Size (1cmx2cmx5mm) maximum
- Into the correct volume of fixative.
Why do we need to fix tissues?
To preserve the chemical and physical characteristics of the tissue. If not done properly it can lead to irreversible damage to the tissue.
Time is important to fix the tissue asap.
Limit loss and diffusion of soluble substances.
How are tissues fixed?
Tissue elements will be: crossed-linked (stabilised and preserved); coagulated/precipitated; chemically unaffected but trapped.
What are the types of tissue fixation?
-Types of fixation = physical or chemical
Physical = heat/microwave/cryopreservation (liquid nitrogen cooled isopentane)
- Chemical = Dehydrant coagulant fixatives (ethanol, methanol, acetone), Non-coagulant fixatives (4% formaldehyde -10% NBF)
Describe tissue embedding in paraffin.
The processed tissue is now removed from the cassette and placed into a metal/plastic mould and covered in liquid paraffin.
The back of the cassette is placed on top and more wax is added.
These are allowed to set to give you paraffin embedded, processed and fixed tissue ready for sectioning.
Describe tissue sectioning in paraffin, the microtome.
The paraffin block is secured into the microtome - set to cut at a certain thickness
~30μm for trimming in the block – removing excess wax.
~3-7 μm for sectioning (takes a lot of practice)
Different tissues types, structures, shapes how well it was fixed and processed plus cutting techniques = bad/good quality sections.
Describe tissue embedding frozen.
Following dissection, the tissue is cut into sections. Place in plastic moulds and fill with embedding matrix (OCT) – Optimum Cutting Temperature.
Float sample on liquid nitrogen cooled isopentane until it solidifies.
Describe the tissue sectioning, frozen OCT in the cryostat.
Set-up is very important
Cutting Temp will need to be varied for different tissue types.
Depends on water and fat content.
Describe the 2 stains: hematoxylin and eosin.
Hematoxylin – Basic, +ve compound and binds to acidic, -ve charges therefore DNA/RNA.
It stains nuclei blue-black showing good intranuclear detail.
Eosin – Acidic, -ve compound and binds basic +ve charges therefore many proteins.
Stains cytoplasm, most connective and muscle tissue fibres different shades and intensities of pink, orange and red.
What is immunohistochemistry?
Target cellular and subcellular localisation of antigens (e.g. proteins) using antibodies.
Antibodies bind small and large antigens with high affinity and specificity.
What is the protein localisation, 2 step indirect method?
- Fixation and Permeabilization - Cells or tissue are fixed to preserve cellular structures and immobilize proteins. Permeabilization is performed to allow antibodies to access intracellular proteins.
- Blocking - The sample is treated with a blocking solution (e.g., 1-5% BSA or serum) to reduce nonspecific antibody binding.
- Primary Antibody Binding - The sample is incubated with a primary antibody that specifically binds to the target protein.
This antibody is usually not labelled and is specific to the protein of interest. - Secondary Antibody Binding - A labelled secondary antibody is added, which binds specifically to the primary antibody.
The secondary antibody is conjugated with a detectable marker, such as: Fluorophores for fluorescent microscopy, Enzymes for for colorimetric detection, Gold particles for electron microscopy. - Visualization - The sample is analysed using an appropriate detection system:
Fluorescence microscopy for fluorophore-labelled antibodies.
Chromogenic substrates for enzyme-labelled antibodies in light microscopy.
Electron microscopy for gold-labelled antibodies.
What is histopathology?
The study of tissues affected by disease, can be very useful in making a diagnosis and in determining the severity and progression of a disease.
Can we view viruses and bacteria under a light microscope?
Bacteria yes, viruses no.
Describe acute inflammation.
Dilation of arterioles - increase in the blood.
Increase in the permeability of capillaries allowing antibodies to enter the tissue.
Migration of leukocytes from the blood into the tissues cells cross the endothelial cells, move out into the tissue.
What is scarring and fibrosis.
Seen when the cells of a tissue are damaged or killed and regeneration of the normal tissue architecture cannot take place
Laying down collagen – not well detected by H&E, better with Masson’s trichrome (stains collagen blue) or with Gieson (stains collagen red/pink).
What is hyperplasia?
Can be a normal physiological response to demand placed on a tissue.
What is dysplasia?
Cells may lose some of their morphological characteristics and/or functions. The tissue becomes disordered, often with an increase in the numbers of immature cells, and greater variability between cells.
What is neoplasia?
Cervical Intraepithelial Neoplasia Increased (CIN). Hyperchromasia
Variation in nuclear size.
Loss of polarity.
Increased mitosis; abnormal mitosis
What is histopathology?
Histopathology is the study of tissues affected by disease, used for diagnosis and assessing disease severity and progression.
Why is understanding normal tissue structure and function important in histopathology?
It helps in interpreting the changes that occur in tissues during disease.
What are the main types of diseases diagnosed using histopathology?
Cancer, inflammatory diseases, benign abnormal growths, and infections.