Microscopy Flashcards
What are the two lenses of a compound light microscope?
The objective and eyepiece lens
Where is the objective lens positioned?
Near the specimen
Where is the eyepiece lens positioned?
Near your eye
How do the objective and eyepiece lens work together and what does this allow for?
The objective lens produces a magnified image, which is magnified again by the eyepiece lens. This allows for much higher magnification and reduced chromatic aberration
What is the process of slicing very thin slices of a specimen known as?
Sectioning
Why are stains used on specimens in light microscopy?
Stains increase contrast as different components within a cell take up stains to different degrees. The increase in contrast allows components to be more visible and more easily identified.
What is differential staining?
Can distinguish between two types of organism
Give 2 differential staining techniques
Gram stain and acid-fast technique
What is the formula for magnification?
Magnification = Size of image / Actual size of object
What are transmission electron microscopes (TEMs)?
Where a beam of electrons is transmitted through a specimen and focussed to produce an image
What are scanning electron microscopes (SEMs)?
Where a beam of electrons are sent across the surface of a specimen and the reflected electrons are collected to produce the image
What is the resolving power of TEMs?
0.5nm
What is the resolving power of SEMs?
3-10nm
What type of image will SEMs give?
3D
In electron microscopes, what do the specimens have to be?
Dead
3 difference between light and electron microscopes
In electron a vacuum is required, black and white images are produced and specimens must be dead, whereas in light a vacuum is not required, the natural colour or staining of specimen is seen and specimens can be living or dead
3 pros of light microscopes
Inexpensive
Small and portable
Simple sample preparation
What is the resolving power of light microscopes?
200nm
What is the magnification of light microscopes?
Up to x2000
What is the magnification of electron microscopes?
Over x500,000
How do laser scanning confocal microscopes work?
It moves a single spot of focused light across a specimen (point illumination). This causes fluorescence, which is sent back up through a pinhole opening and detected by the detector
What happens to the out of focus light and what does this mean for the resolution in confocal microscopy?
The out of focus light is blocked and does not pass through the pinhole, and so is not detected. This light from other parts of the specimen would reduce resolution and cause blurring, however this is prevented
What is the definition of resolution?
The shortest distance between two points that can be distinguished
Why are we able to see with a greater resolution with electron microscopes compared to light microscopes?
Electron microscopes use electrons instead of light and electrons have a shorter wavelength than light, which produces images with a higher resolution
What is the definition of magnification?
Magnification is how much bigger the image is compared to the original object viewed with the naked eye
