Cell Fractionation

Question 1

Homogenisation (breaking up the cells)

Answer 1

Homogenisation can be done by vibrating the cells or by grinding the cells up in a blender. This breaks up the plasma membrane and releases the organelles into solution.

Question 2

What conditions does the solution have to be in?

Answer 2

Ice- cold- to reduce the activity of enzymes that break down organelles
Isotonic- to prevent the damage to the organelles through osmosis.
Buffer solution- to maintain the pH so the enzymes don’t denature.

Question 3

Filtration (getting rid of the big bits)

Answer 3

Next, The homogenised cell solution is filtered through a gauze to separate any large cell debris. The organelles are much smaller so they can pass through the gauze.

Question 4

Ultracentrifugation (separating the organelles)

Answer 4

To separate a particular organelle:

The cell fragments are poured into a tube. The tube is then put into a centrifuge and is spun at the lowest speed. The heaviest organelle like nuclei get flung to the bottom of the tube by the centrifuge. They form a thick sediment at the bottom which is the pellet.
The rest of the organelles stay suspended in the fluid above the sediment- supernatent

The supernatant is dried off poured into another tube and spun in the centrifuge at a higher speed. Again, the heaviest organelles form a pellet at the bottom of the tube. The supernatant containing the rest do the organelles is drained off and spun in the centrifuge at an even higher speed.

The process is repeated at higher and higher speeds until all the organelle are separated out. Each time the pellet at the bottom of the tube is made up of lighter and lighter organelles .

Question 5

Organelle order pneumonic

Answer 5

The organelle are separated out in order of mass m
Naughty
Clever
Monkeys
Like
Eating
Red
Raspberries

( Nucelus, chloroplast, mitochondria, lysosomes, ER, Ribosomes)