Cell Fractionation Flashcards
What happens before cell fraction?
Before cell fractionation can being, the tissue is placed in a cold, buffered solution, same water potential as the tissue.
How do you preparing the cell solution before cell fraction?
This solution is:
- cold – to prevent enzyme activity that might break down the organelles
- Isotonic: the same water potential as the tissue – to prevent movement of water, this prevents organelles bursting or shrinking as a result of osmotic gain or loss of water
- buffered – maintain pH so that the pH does not fluctuate. Any change in pH could alter the structure of the organelles or affect the functioning of enzymes
What are the two stages of cell fractionation?
Homogenation:
Ultracentrifugation:
What happens first homogenation or ultracentrifugation?
Firstly homogenation then ultracentrifugation
What is the process of homogenation?
What is the process of ultracentrifugation?
Homogenation is
Tissue is cut up and kept in a cold, isotonic, buffered solution first. Then the cell is further broken up in a homogeniser (blender). This releases the organelles from the cell. The resultant fluid, known as homogenate.
ultracentrifugation is
Ultracentrifugation is the process by which the fragments in the filtered homogenate are separated in a machine called a centrifuge. This spins tubes of homogenate at very high speed in order to create a centrifugal force. For animal cells, the process is as follows:
-The tube of filtrate is placed in the centrifuge and spun at a slow speed
-The heaviest organelles, the nuclei, are forced to the bottom of the tube, where they form a thin sediment or pellet
-The fluid at the top of the tube (supernatant) is removed, leaving just the sediment of nuclei.
-The supernatant is transferred to another tube and spun in the centrifuge at a faster speed than before
- The next heaviest organelles, the mitochondria, are forced to the bottom of the tube
- The process is continued in this way so that, at each increase in speed, the next heaviest organelle is sedimented and separated.
What organelle drops to the bottom first,last and MIDDLE and why?
Sediment 1: the nuclei, as it is the heaviest organelle
Supernatant 1: mitochondria, lysosomes, ribosomes
Sediment 2: the mitochondria
Supernatant 2: lysosomes,
Sediment 3: lysosomes
Supernatant 3: ribosome as it the lightest organelle
What is the point of cell fractionation?
They allowed a detailed study of the structure and function of organelles, by showing what isolated components do.
What is cell fractionation?
Cell fractionation is the process where cells are broken up and the different organelles are separated out
Before the cell is fractionated the cell is placed in a cold, buffered solution of the same water potential as the tissue. Why is the solution:
- Cold?
- Same water potential as the tissue? ( a isotonic solution)
- Buffered?
- to reduce enzyme activity that might break down the organelles
- to prevent organelles from bursting or shrinking as a result of osmotic gain or loss of water
- so that pH does not fluctuate. Any change in pH could alter the structure of organelles or affect the functioning of enzymes
- What is homogenation?
2. What is ultracentrifugation?
- When celjr are broken up by a homogeniser (blender). This realises the organelles from the cell.
- Ultracentrifugation is the process by which the fragments in the filtered homogenate are separate in a machine called a centrifuge. This spins at very high speeds in order to create a centrifugal force.
- In homogenation what is the resultant fluid known as?
2. What happens to the resultant fluid in homogenisation
- the homogenate
2. the homogenate is filtered is then filtered to remove any complete cells and large pieces of debris.
What is ultracentrigation?
Is the process by which the fragments in the filtered homogenarate are separated in a machine called centrifuge. The centrifuge spins at very high speeds in order in order to create a centrifugal force
