cell fractionation

Question 1

what is cell fractionation?

Answer 1

Cell fractionation is the process in which different parts and organelles of a cell a separated so that they can be studied in detail

Question 2

describe the process of homogenisation and filtration in cell fractionation

Answer 2

1.sample is placed in an ice cold, isotonic buffer solution
2.cells are blended in homogeniser
3.the homogenate is then filtered through a gauze- this is to separate out any large cell debris or tissue debris that were not broken up
4. this leaves a filtrate full of organelles

Question 3

what is homogenisation?

Answer 3

breaking up cells

Question 4

why must the sample be placed in an ice cold, isotonic buffer solution in cell fractionation?

Answer 4

ice cold to reduce activity of enzymes that break down organelles, isotonic to stop the water from moving into the cells by osmosis, and buffer to stop proteins such as enzymes from being denatured and to maintain the pH

Question 5

why are the cells blended in a homogeniser?

Answer 5

to break the plasma membrane of the cells to release organelles into the solution

Question 6

describe the process of ultracentrifugation

Answer 6

1. Homogenisation- the cells are crushed in a blender to break the cell membranes and release the organelles
2. The homogenised sample is then filtered through a gauze to separate any large cell debris. Organelles are much smaller than the debris so pass through the gauze
   3.The filtrate is placed into a tube and the tube is placed in a centrifuge- a machine that separates materials by spinning
3. The filtrate is spun at a low speed. This causes the largest, heaviest organelles (such as the nuclei) to settle at the bottom of the tube, where they form a thick sediment- a pellet. the remaining solution is the supernatant
4. The supernatant is drained off and placed into another tube, which is spun at a higher speed. This causes heavy organelles (such as the mitochondria) to settle at the bottom and leaves a new supernatant
5. The process is repeated at increasing speeds until all the different organelles have been separated. Each pellet gets lighter and lighter.

Question 7

what is the order of organelles during ultracentrifugation?

Answer 7

Nuclei
Chloroplasts (if carrying out cell fractionation of plant tissue)
Mitochondria
Lysosomes
Endoplasmic reticulum
Ribosomes

Question 8

Why are the cells homogenised?

Answer 8

To break up the plasma membrane and release the organelles into the solution