Studying Cells

• Magnification


  how many times larger an image is compared to its actual size
• Resolution

  smallest distance between 2 distinguishable points
• Equation involving magnification

  Image size = Actual size x Magnification
• mm --> um

  x1000
• um --> nm

  x1000
• nm --> um


  divide by 1000
• um --> mm


  divide by 1000
• Optical microscope

  Uses light to form an image
• Optical microscope positives

  cheap

  simple to use

  view live specimens

  colour image
• Optical microscope negatives

  2D image

  limited magnification (x2000)

  limited resolution (200nm) as light has a long wavelength

  cannot observe smaller organelles

  transparent specimens need to be stained
• How an optical microscope works

  Light is sent from the light source below and passes through the specimen which is magnified by the glass lenses
• Electron microscope


  uses electrons to form an image
• Types of electron microscopes

  Transmission Electron Microscope (TEM) and Scanning Electron Microscope (SEM)
• How a TEM works

  electromagnets focus beams of electrons which are fired through the specimen, electrons pass through the specimen, denser parts of the specimen absorb more electrons and appear darker on the image
• TEM positives

  high magnification

  highest resolution

  can see internal structures of cells + organelles
• TEM negatives

  specimens must be very thin

  not in colour

  no live specimens

  expensive

  2D image
• How a SEM works

  beams of electrons (focused by magnets) are fired and scanned across the whole specimen, image formed based on the electrons that are reflected on the surface of the specimen to form a 3D image
• SEM positives

  high magnification

  high resolution

  3D image

  can use thick specimens
• SEM negatives

  cant use live specimens

  black and white image only

  expensive

  lower resolution than TEM
• What is cell fractionation?

  process of separating organelles
• 3 stages of cell fractionation

  Homogenisation

  Filtration

  Ultracentrifugation
• Homogenisation

  breaking open the cell using a blender or homogenizer

  In a buffered, isotonic and ice cold solution

  releases contents of the cell
• Why does the homogenate need to be buffered?

  Maintains the pH of the cell so that enzymes inside the cell don't denature
• Why does the homogenate need to be isotonic?

  Same water potential as the cell so that water does not move into or out of organelles and cause damage
• Why does the homogenate need to be ice-cold?

  to reduce enzyme activity and prevent enzymes breaking down the organelles
• Filtration


  Homogenate is filtered using a gauze to remove any large unbroken cells and debris
• Ultracentrifugation


  Filtrate is placed into a centrifuge and spun at a low speed then faster each time to remove heavier organelles

  Heavier organelles form a pellet at the bottom, remaining organelles stay suspended in the supernatant
• Relative size of organelles

  Nucleus

  Chloroplasts

  Mitochondria

  Lysosomes

  Ribosomes