Microscopy

• Pros vs. Cons - Light Microscope?

  Pros ConsAffordable - Low resolution

  • You can view living specimens - Low magnification
  • It's easy to use

  Can be used to view whole cells and tissue, cant be used to view orgenelles.
• How do Electron Microscopes work?

  They fire a beam of electrons with a wavelength of less than 1 nm to illuminate the specimen. Because of this smaller wavelength, we can see much more detail of the cell's ultrastructure.
• Pros vs. Cons - Electron Microscopes?

  Pros Cons

  • High resolution - Expensive
  • High magnification - Can only view dead specimens

  - Require specialist training

  - They also produce artifacts
• How does a Transmission Electron Microscope (TEM) work?

  The electrons pass through a very thin prepared sample. Electrons pass through the denser parts of the same sample less easily giving the denser parts higher contrast, which in turn creates the final 2D image of the sample.
• How does a Scanning Electron Microscope (SEM) work?

  The electron beam is directed onto a sample. The electrons don't pass through the specimen, but rather "bounce off" the specimen, which produces a 3D image view of the surface of the specimen.
• How does a Laser Scanning Confocal Microscope work?

  A specimen is treated with a fluorescent dye, a laser is then used to illuminate the specimen, which causes the components that have been treated with fluorescent dye to fluoresce. The emitted light from the specimen is then filtered through a pinhole aperture. This means any out-of-focus light is blocked producing a clearer image. The main purpose of the LSCM is to look at objects at different depths in thick specimens.
• What are the ways to prepare a specimen for examination under a microscope?

  Stages of preparation

  1. Fixing (Stabilising a sample or stain, stops it from moving)
  2. Sectioning (Cutting a sample into thin slices)
  3. Staining (Highlighting certain fetures)
  4. Mounting (Placing the sample on a slide)
• What are the different Stains and what can they be used for?

  Methylene Blue > All-purpose stain

  Acetic Orcein > Stains chromosomes

  Eosin stains > Stains cytoplasm

  Iodine > Stains starch in plant cells & highlights plant cell walls
• What is Differential staining?

  Differential staining can distinguish between 2 types of organisms that would otherwise be hard to identify. It can also be used to distinguish between different organelles.
• What is Magnification and Resolution?

  Magnification: How many times larger the image is in comparison to the actual size of the image being viewed.

  Resolution: The smallest distance at which 2 objects can be seen as separate entities.
• What are the magnification and resolution of the different microscopes?

  Light Microscope - Magnification 2,000 Resolution 200 nm

  TEM - Magnification 500,000 Resolution 0.5 nm

  SEM Microscope - Magnification 100,000 Resolution 3-10 nm
• What is the conversion from cm to nm?

  1cm > 10mm > 10,000mm > 100,000nm

  x10 x1000 x1000
• What equation do you use for microscopes?

  Image size = Actual size x Magnification

  
  

  Image size > Use a ruler

  Actual size > Given in the question/Scale bar/Graticule

  Magnification > In the question/corner