L1. Microscopes Flashcards
What is the res of a light microscope
0.2um
How to examine dead cells under a light microscope
- Make tissue rigid by fixing in formalin and embedding in paraffin wax or freezing into a block
- Cut into 1-10um sections using a microtome
- Mount on glass slide
- Stain eg. Using H&E
What does H&E stand for?
Haematoxylin and Eosin
What does haematoxylin stain
Stains nuclei blue
What does Eosin do
Stains cytoplasm pink
What is immunochemistry
Fluorescent or enzyme labelled antihodies that bind to specific structures or proteins in a cell
How does a fluorescent microscope work ?
Uses probes or antibodies
Different parts of the cell emit different frequencies of light
Show as different colours
How does a light microscope show different parts of a living cells
The microscope optically exploits the distortion that the different parts of the cell show creating contrast.
Eg. Some dense parts of the cell distort the light a lot so appear darker
How do light microscopes work?
- White light goes through a material
- Wavelength of light changes
- Some wavelengths are refracted or reflected
- Density of material changes the angle of light
What can differential interference contrast show
Cells as they grow
What is the benefit of confocal microscopy
Can examine 3D specimens
Improves resolution
By removing out of focus light
What does TURF stand for
Total interference reflection flourescence
How does TURF work
Indirectly excites samples to capture the surface of cells and removing light from other focal points
What does atomic force microscopy measure
Rigidity of cells
Is a tumour cell more or less rigid than a normal cell
More
What is the res of an electron microscope?
1nm
What can electron microscopes be used for
To see sub cellular structures eg. Orgenelles
What stain is used in an electron microscope
Heavy metals
How does a scanning EM work
Detects where electrons are scattered and where they’re reflected off of the sample
What is the res of SEM
10nm
What is shadowing
Add metals and chemicals at a bleak angle, the specimen will create a shadow on the bare side
What is freeze fracture
Physically break apart the specimen and treating the exposed area with Pt and C forming a replica which is vuewied under the microscope
What is negative staining
Stains background not the sample to provide contrast
What is cyroelectron microscopy
Freeze sample quickly to cryogenic temp
Water vitrifies and doesn’t damage the sample
Many images collected and combined
