Pre-sessional video - Magnification & Types of Microscopes
- Increasing magnification usually increases resolution as it means smaller things are
moved further apart and our eyes can discern (recognize) them
o Resolution is the ability to discern between two images
- There is a limit to resolution in any type of microscope – if you zoom in enough, the
natural properties of the microscope and the light that we’re looking at limit how
much we can tell apart
o The wavelength of the radiation affects resolution greatly
o Electrons have very short wavelengths which means very high resolution
compared to the wavelength of light
o At other wavelengths of light, there are intermediate levels of resolution –
the higher the frequency, the lower the wavelength gets
There are two groups of microscopes:
1- Optical (Relies on electromagnetic radiation e.g. light)
a. Light microscope
b. Fluorescent microscope (using a wavelength of non-visible light)
2- Electron microscope (Relies on electrons)
a. Scanning electron microscope
b. Transmission electron microscope
Light microscopes:
- The diaphragm condenses the light so that travels through the sample
- Pros:
o Non-destructive to sample
o Live cells can be viewed
o Can see colour & staining
o Quick & easy to use
- Cons:
o Magnification not very high
o Can’t visualise a thick sample as
layers interact with the light and
come out blurred
o Most samples need heavy staining
else light goes through
, Fluorescent microscopy:
- Uses a particular wavelength of fluorescent light
- UV light bounced off a mirror & hits sample & then bounces back
- Not a transmission microscopy anymore, it is an emission microscopy
- Pros:
o Enables fluorescence marking (labelling organelles)
o Improved resolution as higher frequency of
wavelengths so less diffraction
o Distinguishable colours
- Cons:
o UV light damaging to skin
o UV light highly damaging to sample & organism’s
behaviour highly affected
Confocal microscopy:
- The light is shown at various angles of the sample and is
captured to form a 2D or a 3D image
- Pros:
o Has all the advantages of a fluorescent light
microscope
o Clear 2D &3D images
- Cons:
o Very expensive
o Need to be highly trained technician
o Getting a good image is hard
o Laborious sample prep
Transmission electron microscope:
- Electrons are fired from a gun
and are channelled through an
electromagnet that controls the
path and beam of fire of
electrons so that they uniformly
go through the very thin sample
- The image you’re seeing is just
the stain of the sample, not the
organelles themselves
- Due to heavy staining, lots of artefacts are present
o Artefacts are things that are there due to the processing and staining, it is not
actually present in the sample
- Pros:
o Extremely high resolution
o Can view subcellular architecture in detail
- Cons:
- Increasing magnification usually increases resolution as it means smaller things are
moved further apart and our eyes can discern (recognize) them
o Resolution is the ability to discern between two images
- There is a limit to resolution in any type of microscope – if you zoom in enough, the
natural properties of the microscope and the light that we’re looking at limit how
much we can tell apart
o The wavelength of the radiation affects resolution greatly
o Electrons have very short wavelengths which means very high resolution
compared to the wavelength of light
o At other wavelengths of light, there are intermediate levels of resolution –
the higher the frequency, the lower the wavelength gets
There are two groups of microscopes:
1- Optical (Relies on electromagnetic radiation e.g. light)
a. Light microscope
b. Fluorescent microscope (using a wavelength of non-visible light)
2- Electron microscope (Relies on electrons)
a. Scanning electron microscope
b. Transmission electron microscope
Light microscopes:
- The diaphragm condenses the light so that travels through the sample
- Pros:
o Non-destructive to sample
o Live cells can be viewed
o Can see colour & staining
o Quick & easy to use
- Cons:
o Magnification not very high
o Can’t visualise a thick sample as
layers interact with the light and
come out blurred
o Most samples need heavy staining
else light goes through
, Fluorescent microscopy:
- Uses a particular wavelength of fluorescent light
- UV light bounced off a mirror & hits sample & then bounces back
- Not a transmission microscopy anymore, it is an emission microscopy
- Pros:
o Enables fluorescence marking (labelling organelles)
o Improved resolution as higher frequency of
wavelengths so less diffraction
o Distinguishable colours
- Cons:
o UV light damaging to skin
o UV light highly damaging to sample & organism’s
behaviour highly affected
Confocal microscopy:
- The light is shown at various angles of the sample and is
captured to form a 2D or a 3D image
- Pros:
o Has all the advantages of a fluorescent light
microscope
o Clear 2D &3D images
- Cons:
o Very expensive
o Need to be highly trained technician
o Getting a good image is hard
o Laborious sample prep
Transmission electron microscope:
- Electrons are fired from a gun
and are channelled through an
electromagnet that controls the
path and beam of fire of
electrons so that they uniformly
go through the very thin sample
- The image you’re seeing is just
the stain of the sample, not the
organelles themselves
- Due to heavy staining, lots of artefacts are present
o Artefacts are things that are there due to the processing and staining, it is not
actually present in the sample
- Pros:
o Extremely high resolution
o Can view subcellular architecture in detail
- Cons:
