Cell Fractionation Flashcards
Suggest why cell fractionation would be carried out:
If you wanted to look at an individual organelle under a microscope rather than a whole cell.
What is the purpose of cell fractionation?
It separates organelles.
What are the three steps in cell fractionation?
1) Homogenisation,
2) Filtration,
3) Ultracentrifugation.
What is homogenisation?
Breaking up the plasma membrane to release organelles into solution.
What is left after filtration?
A solution containing a mixture of (smallish) organelles.
Why must the homogenised solution be kept ice-cold?
To reduce the activity of enzymes that break down organelles.
Why must the homogenised solution be kept isotonic?
It maintains the same concentration to maintain turgor pressure.
Suggest one way of homogenisation?
Vibrating the cells or Grinding them in a blender.
Explain the process of filtration?
The homogenised solution is filtered through a gauze to seperate any large cell debris or tissue debris.
What passes through the gauze in filtration?
Organelles as they are small enough to pass through.
Give an example of something that would not pass the filter during filtration.
Connective tissue.
Why should a buffer solution be added to the homogenised solution?
To minimise the effect of pH changes.
What is the most dense organnelle?
Nuclei.
What should be done to the homogenised cell to make sure it stays in good condition?
Ice-cold, Isotonic and a buffer solution added.
Explain the process of ultracentrifugation.
1) Cell fragments are poured into a tube, this is put into a centrifuge and spun at a low speed.
2) The supernatant is drawn off, poured into another tube and centrifuged at a higher speed.
3) This process repeats and the pellet at the bottom is made up of lighter and lighter organelles.
