Microscopy

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A microscope is an instrument that produces a magnified image of an object.

Optical (light) microscope:

  • Uses light
  • Long wave length
  • So can only distingush between two objects that are 0.2 micrometers or further apart

(http://tse2.mm.bing.net/th?id=OIP.px5F7nw46nWVpE0xCf6rogEsEZ&w=205&h=192&c=7&qlt=90&o=4&pid=1.7)

Objective lens -

  • Placed near to the specimen

Eyepiece lens -

  • Which the specimen is viewed

Electron microscope:

  • Uses electrons
  • Has a very short wave length
  • Has a high resolving power - resolve objects that are 0.1 nanometers apart
  • Electrons are negatively charged so the beam can be focussed using electromagnets
  • Electrons are absorbed or deflected by the molecules in the air, so anear-vacuum has to be created within the chamber of an electron microscope in order for it to work effeciently

TEM - tranmission

  • Consists of an electron gun that produces a beam of electrons that is focused onto the specimen by a condenser electromagnet
  • The beam passes through a thin section of the specimen
  • Parts of the specimen absorb electrons and therefore appear dark
  • Other parts of the specimen allow the electrons to pass through so they appear light
  • An image cane be produced on a screen and this can be photographed to give a photomicrograph

Although the resolving power is 0.1 micrometers, this cannot always be achieved in practise because:

  • Difficulties preparing the specimen limit the resolution that can be achieved.
  • A higher energy electron beam is required and this may destroy the specimen

SEM - scanning

  • Directs a beam of electrons on to the surface of the specimen from above, rather than penetrating it from below.
  • The beam is then passed back and forth across a portion of the specimen in a regular pattern
  • The electrons are scattered by the specimen and the pattern of the scattering depend on the countours of the specimen surface
  • We can build a 3-D image by computer analysis of the pattern of scattered electrons and secondary electrons produced
  • The basic SEM has a lower resolving power than the TEM, being around 20 nanometers

The main limitations of both electron microscopes are:

  • The whole system must be in a vucuum and therefore living specimens cannot be observed
  • A complex 'staining' process is required and even the image is not in colour
  • The specimen must be extreamely thin
  • The image may contain artifacts. Artefacts are things that result from the way the specimen is prepared. Artefacts may appear on the finished photomicrograph but are not part of the natural specimen. It is therefore not always easy to be sure that what we see

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