Ultramicrotomy

Microscopy Methods in Microbiology
Microroscopy Methods in Microbiology Part 2
Microroscopy Methods in Microbiology - Introduction
Observation of Microbes in the Living State
Observation of Microbes in Smears
Dyes and Staining
Useful pH Range and Colour Change of Some Indicator Dyes
Factors Affecting Staining
Microscopy and Micrometry
Microscopy and Micrometry - Introduction
Behaviour of Light
Light Theories
Resolving Power
Refraction and Reflection
Bright Field Microscopy
Use of Bright Field Microscope

Ultramicrotomy

Ultramicrotomy
Ultramicrotome is employed using I glass or diamond knife which is capable of taking sections in the range of 50-300 nm, required for electron microscopy. Detailed instructions on section cutting are normally provided by the manufacturer of ultramicrotome and the techniques are very well documented (10, 12, 19, 20). Therefore, only general points will be described here. Glass knife can be prepared from a strip of glass using Knife-Maker.If high quality sections are required, diamond knife should be used which are available with a water trough attached. Trough can be prepared by wrapping a short strip of Scotch insulating tape (no.33) around the back of the knife and sealing it with paraffin wax.

Block can be made ready for the sectioning by trimming it at the tip with a razor blade or black trimmer so that the smallest cutting face is available. Fix the block to a microtome and cut the sections. Sections float onto a surface of liquid held in trough and remain together in a form of ribbon. Freshly distilled water is generally used to fill the trough. The thickness of the sections cut can be judged by the interference colours generated in the ribbon by reflection of light. Grey and pale gold to light silver colour is generated by the sections having thickness of about less than 10nm and 80nm, respectively. Green, blue and purple colours indicate that the sections are too thick they are useless for electron microscopy. The floating sections are then fished from the tough with a small circular copper gird.

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