Microscopy Flashcards
Who saw the first cells and when
Robert Hooke
In 1665
He saw the cell wall from a dead cell of the bark of an oak tree
How do light microscopes work
Visible light is passed through a specimen then through glass lenses
The lenses refract the light
The image of the specimen is magnified as it’s projected to the eye or camera.
Methods used in microscopy to increase the contrast of a cell
Brightfield (stained)
Phase-contrast
Fluorescent
How does brightfield (stained) increase contrast in a cell
Stained:
Adding dye to cell creates contrast.
Staining kills the cell.
How does phase-contrast increase contrast in a cell
Uses varied densities in the cell to enhance contrast.
Advantage: can use on living and unpigmented cells.
How does fluorescents increase contrast in a cell
Label specific molecules in the cell with fluorescent dyes or antibodies.
Fluorescent substances absorb UV and emit visible light.
Why is resolution better in electron microscopes that light microscopes
Electrons have shorter wavelengths than visible light
How do electron microscopes work
They focus a beam of electrons through the specimen
How does a scanning electron microscope work
An electron beam scans the surface, that’s usually coated with a thin layer of gold.
The beam excites the electrons on the surface.
A device detects and translates the pattern into electronic signals.
How do transmission electron microscopes work
Specimen is stained with heavy metal atoms.
These attach to certain cellular structures, enhancing electron density.
Electrons pass through and scatter more and transmit less in denser regions.
Does a low speed centrifuge produce large or small component pellets?
Larger component pellets.
Resolution of a light microscope
200nm - 0.2 micrometers
Resolution of an electron microscope
1nm -0.1nm (high resolution)
Why do you stain a specimen in a TEM microscope with heavy metals
They attach to certain cellular structures and enhance the electron density.
Electrons going passing through will scatter more in denser regions, transmitting fewer electrons.
Disadvantages of a TEM
Specimens must be dead - vacuum so they’ll die anyways.
Preparation can cause artifacts.
