Microscopy Flashcards
What are the 4 types of microscope?
- Light microscope
- Scanning Electron microscope (SEM)
- Transmission Electron microscope (TEM)
- Laser Scanning Confocal microscope
How does a light microscope work?
It uses light and a series of glass lenses to form a magnified image, limiting it’s resolution.
What are the advantages of using a light microscope?
- Relatively cheap
- Easy to set up and use
- Can be used to view eukaryotic cells, nuclei and sometimes mitochondria and chloroplasts
- Cells can be viewed in colour
What are the disadvantages of using a light microscope?
- They have a maximum resolution of around 0.2 micrometres / 200 nm
- Cannot be used to observe ribosomes, ER or lysosomes
- Maximum magnification is usually on x1500
How does an SEM microscope work?
A beam of electrons is scanned ACROSS the specimen’s surface. The electrons bounce off the specimen and are detected, forming a 3D image.
What are the advantages of using an SEM microscope?
- Can be used on thick, 3D specimens
- External 3D structures can be observed
What are the disadvantages of using an SEM microscope?
- Lower resolution than TEMs
- Cannot view live specimens
- Colour images are not produced. Images are also digitally processed
- Lower magnification than TEMs (around x500,000)
How does a TEM microscope work?
Electromagnets are used to focus a beam of electrons which is transmitted THROUGH the specimen. Denser parts of the specimen absorb more electrons and appear darker. Specimens require lengthy treatment and artefacts can be introduced.
What are artefacts?
A structural feature which is not part of the actual specimen, and instead produced in preparing the specimen (e.g. an air bubble).
What are the advantages of using a TEM microscope?
- Magnification = x1,000,000 or higher
- Internal cell structures can be seen
- High resolution imagery (more detail).
What are the disadvantages of using a TEM microscope?
- Only thin specimens / sections of an object can be used
- No live specimens as a TEM uses a vacuum
- 2D, no colour
- All water must be removed from the specimen
Why do electron microscopes increase resolution?
A beam of electrons has a shorter wavelength than a beam of light. The max resolution of an electron microscope is around 0.2 nm or 0.0002 micrometres.
How do Laser Scanning Confocal microscopes work?
Cells are stained with fluorescent dyes and a section of tissue is scanned with a thick laser beam (focused light). The light is reflected by the dyes. Multiple depths are scanned to produce an image.
What are the advantages of using a Laser Scanning Confocal microscope?
- Can be used on thick, 3D specimens
- External 3D structures are observed
- Very clear resolution as laser beam can be focused at a very specific depth (cytoskeleton can be viewed)
What are the disadvantages of using a Laser Scanning Confocal microscope?
- Slow process and long time to obtain cell image
- Laser beam can cause photodamage to the cell