Cell Fractionation

• What is cell fractionation?

  The process where cells are broken up and their different organelles are separated.
• Why is the tissue placed in a cold solution before cell fractionation can begin?

  To reduce enzyme activity that might break down the organelles.
• Why is the tissue placed in a buffered solution before cell fractionation can begin?


  So the pH does not fluctuate and affect enzymes as well as alter the organelle's shape.
• Why is the tissue placed in a solution with the same water potential before cell fractionation can begin?


  To prevent the organelles from bursting or shrinking as a result of osmosis.
• What are the 2 stages of cell fractionation?

  Homogenation and ultracentrifugation.
• What happens in homogenation?

  The cells are broken up by a homogeniser to release the organelles.
• What is a homogeniser?

  A blender.
• What is the resultant liquid produced from homogenisation?

  Homogenate.
• What happens to the produced homogenate?

  It is filtered to remove any complete cells and larger pieces of debris.
• What is ultracentrifugation?


  The process by which the organelles in the filtered homogenate are separated in a centrifuge.
• What does a centrifuge do?

  Spins tubes of homogenate at very high speed to create a centrifugal force.
• What is the first stage of ultracentrifugation?

  The tube of filtrate is spun in a centrifuge at a slow speed.
• What happens when the filtrate is spun first at a slow speed?

  The heaviest organelles, such as nuclei, are forced to the bottom of the tube.
• What is formed when the organelles are forced to the bottom of the tube?

  A thin sediment or pellet.
• What fluid is removed from the top of the tube when the organelles are forced to the bottom to form a pellet?

  The supernatant.
• Why is the supernatant removed?

  To be transferred to another tube and respun in the centrifuge at a higher speed.
• What happens when the supernatant is respun at a higher speed?

  The next heaviest organelle, such as the mitochondria, are forced to the bottom of the tube forming a pellet..