Using Microscopes

  • Created by: Freja
  • Created on: 12-04-21 15:07
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  • Using Microscopes
    • Preparing A Slide
      • Dry mount
        • Sectioning- solid specimens are cut into very thin slices with a sharp blade
        • The specimen is placed on the centre of the slide and a cover slip is placed over the sample
      • Wet Mount
        • Specimens are suspended in a liquid such as water or an immersion oil
        • Add the cover slip at an angle, to avoid getting air bubbles, as they will obstruct the view
    • Using Stains
      • Stains increase contrast
        • The increase in contrast allows components to become visible so that structures can be identified
      • Different components take up stains to different degrees
      • How Stains are Used
        • Stains can be added to the specimen and left to dry
        • The sample can be heat fixed, which involves passing the sample and microscope slide through a flames, and this adheres the sample to the microscope slide and will allow the stain to be taken up when it is added
      • Differential Staining
        • Used to identify specific components of a cell and can identify different organisms that otherwise would be too difficult to identify
        • Crystal Violet/ Meth Blue are positively charged dyes
          • They attract to negatively charged materials in the cytoplasm, so part of the cell becomes stained and so becomes visible
        • Nigrosin/ Cargo red are negatively charged dyes
          • Are repelled by negatively charged cytosol, so the dye does not enter the cells so they stay outside leaving the cells unstained and this causes contrast between the unstained cells and the stained background
      • Gram Staininng
        • Differential Staining can also be used to identify between two different types of bacteria
        • Can be Gram Positive or Gram Negative
    • How to use a Light MIcroscope
      • 1) Start by clipping the side containing the specimen onto the stage 2) Select the lowest powered objective lens 3) Use the coarse adjustment knob to bring the stage up to just below the objective lens
      • 4) Look down the eyepiece (containing the ocular lens). Use the coarse adjustment knob to move the stage downwards away from the objective lens until the image is roughly in focus
      • 5) Adjust the focus with the fine adjustment knob, until you get a clear image of what's on the slide              6) If you need to see the slide with greater magnification, swap to a higher-powered objective lens and refocus
    • How to Use an Eyepiece Graticule and Stage Micrometer
      • They can be used to find the size of a specimen
      • An eyepiece graticule is fitted onto the eyepiece and its like a transparent ruler with numbers but not units
      • The stage micrometer is placed on the stageand it is a microscope slide with an accurate scale and units
        • It is used to work out the value of the divisions on the eyepiece graticule at a particular magnification
      • 1) Line up the eyepiece graticule and stage micrometer   2) Each division on the stage micrometer is 0.1mm long 3)At this magnification, 1 division on the stage micrometer is the same is 4.5 divisions on the eyepiece graticule
        • 4) To work out the size of 1 division on the graticule, you need to divide 0.1 by 4.5: 1 division =0.1/4.5 =0.022mm       5) So if you look at an object under the microscope at this magnification and it is 20 eyepiece divisions, it measure 20x 0.022= 0.44mm
      • 1 Graticule Division= number of micrometers/ number of graticule division
      • Measurement (um)= graticule divisions x magnification factor


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