Microscopes

• What does staining do and why is it used?

  it allows chemical reactions in parts of the cell which enhances contrast in the image to identify certain parts of the cell clearly.
• List the features of a good diagram:

  • scale / magnification
  • title
  • drawn in sharp pencil
  • annotations
  • no shading
  • no overlapping lines
• Light/ Optical microscopes have poor resolution due to the long wavelength of light
• In light microscopes, living or dead specimens can be observed and a coloured image is obtained
• In TEM, electrons pass through the specimen to create a black and white 2D image
• In SEM, electrons bounce off the surface of the specimen to create a black and white 3D image
• In LSCM, laser light passes through the specimen and the scattered light is detected to create a 3D or 2D coloured image
• Define Resolution:

  The ability to distinguish two separate points
• In light/ optical microscopes, resolution is determined by the wavelength of light
• In electron microscopes, resolution is determined by the wavelength of the beam of electrons
• Define Magnification:

  The number of times larger the image of a specimen observed is compared to the actual size of the specimen
• State the four main types of sample preparations:

  • dry mount
  • wet mount
  • squash mount
  • smear slide
• Formula for Magnification:

  image size/ actual size
• Staining usually kills the cells
• Transmission electron microscopes(TEMs) use electromagnets to focus the electron beam
• State the three types of microscopes:

  • light / optical microscopes
  • electron microscopes
  • fluorescent microscopes
• What are the two main types of electron microscopes?

  • Transmission Electron Microscope
  • Scanning Electron Microscope
• State the disadvantage of enlarging an image obtained by a light microscope.

  the resolution is still low
• What is the magnification for optical/light microscopes?

  x 1500
• Describe Light/Optical Microscopes:

  • Long wavelength of light from a light beam
  • Magnification: x 1500
  • Resolution: 200 nm (0.2 picometres)
  • Image: coloured, 2D
  • cheap, easy to use, portable
  • The types of specimens it can study: whole, thin, living, dead = stained
• Describe Transmission Electron Microscopes:

  • Electrons pass through specimens from electron beam
  • Electromagnets are used to focus the beam
  • Magnification: x 2 - 50 000 000
  • Resolution: 0.2 - 0.5 nm
  • Image: 2d, black and white = darker areas - electrons were absorbed
  • identifies internal structures
  • expensive, large, difficult to use
  • vacuum
  • The types of specimens it can study: fixed = dehydrated, thin, stained with metal salts
• Describe Scanning Electron Microscopes:

  • Electrons bounce off the surface from electron beam
  • electromagnets are used to focus the beam
  • Magnification: x 15 - 200 000
  • Resolution: 0.2 - 0.5 nm
  • Image: 3D, black and white - contrast due to electrons scattering when bounced off
  • vacuum
  • expensive, large, difficult to use
  • The types of specimens it can study: thick, dead, stained with metal salts
• Why can the specimen observed by scanning electrons can be thick?

  • electrons are bounced off
  • NOT passed through
• Describe Laser Scanning Confocal Microscopes:

  • Magnification:
  • Resolution:
  • Image:
  • The types of specimens it can study:
• State the name of all the stains used in light/optical microscopy, the colour they create, and what they bind to:

  • Acetic orcein = dark red - DNA, chromosomes
  • Eosin = pink - cytoplasm
  • Sudan red = red - lipids
  • Iodine = yellow - cellulose

  = blue-black - starch
• SLIDE PREPARATION:

  EYE PIECE GRATICULE:

  CONTINUE
• Name the stain used to bind to the phosphate group (PO4^3-)of DNA to make chromosomes more visible when using a light microscope ?

  methylene blue
• A student experimented on eight identical samples for her investigation, why did they take reading from all eight samples?

  • To access the repeatability of the data
• millimetres (mm) --- x1000 ---> micrometers (µm)
• What laboratory equipment would you select to make an accurate measurement of length?

  • light microscope
  • graticule
• When calculating any section of M.I.A (calculating magnification), the image size and the actual size should be in micrometers (μm).

  When asked for the image size, convert it back into millimetres(or whatever unit they ask it)
• What can be done to improve an experiment to view a cell under a microscope.

  • add multiple stains to improve contrast
  • use blotting paper to remove excess stain
  • lower cover slip at an angle