Optical and Electron Microscopy

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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. Light rays are transmitted, the light source usually being a bulb, through the object, and then through the two convex lenses where refraction occurs. At the objective lens an inverted image is produced, see figure 1, the size of which depends on the magnifying power of the objective lens and the distance of the object from the lens. The eyepiece allows the eye to see an enlarged but inverted image (image2 in figure 1) of the object, magnified to between x25 to x1500 depending on the total magnifying power of both lenses.

Figure 1:

   Note that there are actually more lenses in the compound microscope through which light passes. These correct imperfections such as aberrations - the inability of a lens to be light of different colours to focus at the same point (chromatic aberration), or the failure of a lens to focus light coming from the edge of the lens and light coming from the centre of the lens at the same point (spherical aberration).

Biology Essay continued:

Preparation of biological material for optical microscopy:

   

   Before the specimen is prepared the microscope being used may have to be calibrated so that dimensions of small objects, such as the diameter of a cell, in the specimen can be measured using an eyepiece graticule. Calibration involves measuring the length of one eyepiece unit under low, medium and high magnifying power, so that dimensions can be recorded in millimetres. This is done using stage micrometers, tiny scales of changeable lengths, mounted on a slide.

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   To find the length of one eyepiece unit under low power, a 10 mm stage micrometer is used. This is a scale 10 mm long, divided up into tenths of one millimetre. When viewed under a microscope using the eyepiece graticule, the zeros of both scales are lined up. When a mark on both the micrometer scale and the eyepiece graticule scale lined up, a measurement in millimetres is obtained for a certain number of eyepiece units. The stage micrometer value is then divided by the number of eyepiece units to give the length of one eyepiece unit in ...

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