Magnification

 Living things are composed of Cells. Cells are very small

(ususally between 1 and 100 μm) and can only be seen by
magnification with a microscope. A distinction is made
between Magnification and Resolution: Magnification is how
large the image is compared to real life, whereas Resolution is
the amount of information that can be seen in the image -
defined as the smallest distance below which two
discrete objects will be seen as one.
 To work out the size of an object viewed with a microscope,
a Graticule is used. It is a small transparent ruler that
becomes superimposed over the image. As the same sample
may look to be different sizes under different
magnifications, the Graticule must be calibrated.
Metal Foam in Scanning Electron Microscope, magnification 10x. Credit SecretDisc

 Actual Size, Image Size and Magnification are related by the

formula:

Image Size = Actual Size × Magnification

The Light Microscope

 Light Microscopes, or Optical Microscopes, as they are
more correctly termed, use light and several lenses in order to
magnify a sample. Light from the Condenser Lens, and then
through the Specimen where certain wavelengths are filtered
to produce an image. The light then passes through
the Objective Lens, which focuses it and can be changed in
order to alter the magnification. Finally, the light passes
through the Eyepiece Lens, which can also be changed to
alter the magnification, and into the eye.

 The maximum magnification of light microscopes is

usually ×1500, and their maximum resolution is 200nm,
due to the wavelength of light. An advantage of the light
microscope is that it can be used to view a variety of samples,
including whole living organisms or sections of larger plants
and animals. It is also relatively inexpensive.

 There are two types of light microscope. Compound

Microscopes contain several lenses and magnify a sample
several hundred times. Dissecting Microscopes on the other
hand have a low final magnification but are useful when a
large working distance between the objectives and the stage is
required (e.g. during dissection). They have two eyepieces to
produce a 3D stereoscopic view.
 Many specimens require preperation before being viewed by
a light microscope, as some may not be coloured or might
distort when cut. Samples are Stained with coloured stains
that bind to certain chemicals or cell structures. For example,
 Acetic Orcein stains DNA dark red. Samples may also
be Sectioned - embedded in wax; this helps with preserving
structure while cutting.

The Electron Microscope

 Light microscopes are great and all, but sometimes their
(relatively) low magnification and resolution are insatisfactory
for viewing very small things, like Organelles within cells. In
these circumstances, and Electron Microscope may be used.
Electorns have a much lower wavelength than light (100000
times shorter in fact, at 0.004nm) which means that they can
be used to produce an image with resolution as great
as 0.1nm. Electron Microscopes can have magnifications
of ×500000.
 There are different types of Electron Microscope.
A Transmission Electron Microscope (TEM)produces a 2D
image of a thin sample, and has a maximum resolution
of ×500000.

 A Scanning Electron Microscope (SEM) produces a 3D

image of a sample by ‘bouncing’ electons off and dectecting
them at multiple detectors. It has a maximum magnification of
about ×100000.
 …

 The preparation of a sample for electron microscopy is a

complex process. It may involve
 Chemical Fixation: Stabilising an organism/sample’s
mobile macrostructure
 Cryofixation: Freezing the sample very rapidly to
preserve its state
 Dehydration: Removing the water form a specimen, for
example, by replacing it with ethanol
 Embedding: Embedding in resin, ready to be sectioned
 Sectioning: Cutting the sample into thin strips that are
semitransparent to electrons, for example with a diamond
knife
 Staining: Using heavy metals to scatter electrons and
produce contrast
 Freeze Fracturing: Freezing the sample rapidly, and
then fracturing it, for example, when viewing cell
membranes
 Mounting: Placing the sample on a copper grid
 It is advantageous to use an Electron Microscope in many
situations because they offer a much higher resolution that
Light Microscopes, so they can be used to image very small
objects in detail, and also because of the 3D images that
SEMs offer. However, samples must be placed in a
vacuum as electrons are deflected by particles in the air, they
are very expensive to buy and maintain, and preparing the
samples requires a lot of skill to do.