Centrifugation

• Centrifugation

  1. Homogenise tissue/use a blender

  2. Place in a cold, isotonic, buffered solution

  3. Filter homogenate

  4. Ultracentrifugation - separates organelles in order of density/mass
• Homogenisation

  • Disrupts cell membrane, breaking open cells and releasing contents/organelles
• Filtering homogenate


  • Remove large, unwanted debris e.g. whole cells, connective tissues
• Ultracentrifugation

  1. Centrifuge homogenate in a tube at a high speed

  2. Remove pellet of heaviest organelle as respin supernatant at a higher speed

  3. Repeat at increasing speeds until separated out, each time pellet made of lighter organelles (nuclei -> Chloroplasts/mitochondria -> Lysosomes -> ER -> Ribosomes)
• Why do we use a cold, buffered, isotonic solution during centrifugation?

  • Cold = to reduce enzyme activity -> so organelles not broken down /damaged
  • Buffered = to keep pH constant -> so enzymes don't denature
  • Isotonic = So water doesn't move in or out of organelles by osmosis -> so they don't burst