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Optical and Electron Microscopy

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Introduction

Biology essay:

Optical and Electron Microscopy:

Introduction:

   Microscopes allow us to see things so small they normally can’t be seen by the naked eye. The intention of all microscopic studies is to produce an image that is a copy of the object, or the specimen.

   There are two main aspects to a microscope, which determine the image - magnification and resolution. The magnification is the number of times the image is linearly larger than the object. So, small objects will appear larger in the image under higher magnification than under lower magnification. The resolution, or resolving power, is the microscope’s ability to differentiate between small objects that are close together. Theoretically the electron microscope has 100,000 x better resolving power than optical microscopes, but in practise the resolution of an electron microscope is at best 1 nm (nanometre), i.e. two objects less than 1 nm apart will be seen as one.  

Optical Microscopy:

   The main instrument, which utilizes light to produce a magnified image, is the compound microscope. A compound microscope uses the magnifying power of two convex lenses, the eyepiece and the objective lens, which itself has three variable magnifying powers – low, medium and high, to produce an image.

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Middle

embedding, would not mix if any water were present. Dehydration is usually accomplished by gradually increasing the concentration of added alcohol, usually ethanol, until ‘absolute alcohol’ is reached – nearly pure alcohol.

3. Clearing:

   In most cases alcohol is immiscible with the embedding, see next stage, or mounting media, so it is replaced by a clearing agent, such as xylol or xylene, which does mix. The agent also makes the material transparent, which is necessary, as light rays must travel through the specimen into the eye.

4. Embedding:

   Before the material can be cut into thin sections to allow light through, it must be embedded using a supporting medium so that the slices do not collapse. In optical microscopy, the material is impregnated with molten wax, which consequently sets.

Biology Essay Continued:

5. Sectioning:

   Once the wax has settled, the material can be cut up into thin sections. This is either done using a razor or a microtome. The latter being a machine that cuts extremely thin slices, usually between 3 and 20 micrometres. A freezing microtome may be used which avoids the need of embedding as the specimen is frozen and therefore firm. A razor can

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Conclusion

Scanning Electron Microscopy: (SEM)

   In SEM the whole specimen is exposed to a beam of electrons. An image is then created from the electrons reflected from the surface of the object. Although the resolution isn’t as good as with TEM, scanning electronmicrographs show depth of focus and the object is three-dimensional. Another advantage is that SEM allows larger specimens to be examined.

   When comparing optical microscopy and electron microscopy, it is clear that each has its own advantages and disadvantages over the other. Obviously, an electron microscope produces an image of greater magnification and resolution than optical microscopes due to the very nature of light, but electron microscopy doesn’t produce coloured images. EM is also very expensive and preparation takes a lot of time. Another disadvantage in EM is that the specimen has to be nonliving, which can be a cause for irritation as studying living material under high magnification and resolution would help us to learn more about cell structure and function, for example.

   To conclude, the choice of one type of microscopy over the other is dependant on the requirements of the given study. For example, if one wanted to observe the structure of mitochondria one would choose EM over optical microscopy because it has greater magnification and resolving powers.  

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