Practical 3: H&E and Oil Red O Stain Flashcards
What is embedding
The process by which tissues are solidified in a wax mould to create the wax block for cutting thin sections using a microtome
Why do we use paraffin wax for moulds
It provides a supporting media to enable subsequent sectioning of tissue
What does FFPE stand for
Formalin fixed paraffin embedded tissue
What is cryotomy
The use of a refrigerated microtome to cut frozen sections of tissue
When is cryotomy used
(5)
Lipid staining
Rapid H&E for urgent diagnosis
DNA and/or preservation for molecular analysis by PCR
Special investigations - direct immunofluorescence, enzyme histochemistry, immunohistochemistry
Biobanking - storage for research
Why is cryotomy used in lipid staining
Lipids are dissolved out of tissue if they are exposed to the solvents used in paraffin-processing, therefore lipid staining is always performed on frozen tissues
What is the most common lipid stain?
Oil Red O
What is Oil Red O
A red hydrophobic dye which stains all triglycerides and neutral lipids intensely red
When might you need an Oil Red O stain
Disease cases whereby lipids accumulate e.g. liver disease, muscle wasting or lipid storage diseases
Give another stain for lipids other than Oil Red O
Sudan Black B
What is the different between Oil Red O and Sudan Black B
Sudan Black B stains both hydrophobic and phospholipids black
When might H&E be used with cryotomy?
Patient may be having surgery -> need rapid diagnosis of a legion
Excision margin clearance to ensure diseases tissue fully resected
Lymph node clearance in radical surgery - head and neck surgery, radical hysterectomy, axillary clearance for breast cancer
What is the microtome and how is it used (3)
A mechanical device which advances a tissue block by a small thickness over a blade, which then slices off the section
This is then floated onto water to flatten and expand the section
The section is then collected onto a labeled slide
Slides of 10-20 microns are used for what?
Neurohistology
Lipid staining
Slides of 3-5 microns are used for what
Routine light microscope
Slides of 5-8 microns are used for what
Frozen section
Slides of 0.5 to 3 microns are used for what
Semi-thin resin
Slides of 60-120 microns are used for what
Electron microscopy
List the four major components of the microtome
Block holder
Knife holder or blade holder
Micrometer gauge
Advancement controls
What are the four types of microtome
Rotary
Cambridge Rocker
Sledge
Cryostat
What is a rotary microtome
The most common microtome -> now available as automated microtomes
e.g. Leitz or Jung
What is a Cambridge Rocker
The first type of microtome -> found only in museums
What is a Sledge microtome
A sliding microtome
Used for cutting of large tissue samples, whole organisms or whole brain
What is a cryostat microtome
A freezing microtome for frozen sections
What blades are used for microtomy
Glass or diamond knives
What are microtomy blades usually made of?
Steel
Tungsten carbide
Cobalt
Stellite alloys
