Cell Fractionation & Ultracentrifugation Flashcards
What is cell fractionation? [1 mark]
When cells break up to separate the different organelles.
What is homogenisation [1 mark]
When cell plasma MEMBRANES are broken up in a sample to release organelles into a solution.
What is a homogeniser? Give an example. [1 mark]
Equipment that breaks down and separates organelles within a cell.
E.G.
VIBRATING the cells
A blender
Pestle & mortar
What is a centrifuge? [1 mark]
Equipment that spins around a solution to separate organelles by their densities.
What is ultracentrifugation? [1 mark]
The repeated centrifugation of solutions at higher speeds to create organelle-rich pellets.
What is a supernatant liquid [1 mark]
The liquid left on top of the pellet after centrifugation. Has suspended cell organelles with a lighter density.
How is cell fractionation carried out in simple steps?
1) Homogenisation - break cells using blender.
2) Filtration - remove debris.
3) Ultracentrifugation - separate organelles based on density at high speeds.
What are the properties of the solution used during cell fractionation? Explain why these properties are necessary. [4 marks]
> ICE-COLD =
stops enzyme activity - prevents breakdown of organelles.
> BUFFERED =
maintains pH so enzymes/proteins aren’t denatured.
> ISOTONIC =
concentration is kept the same inside & outside the cell to prevent osmosis.
Prevents osmosis so organelles don’t burst or shrink from gain/loss of water.
Describe how a sample of chloroplasts could be isolated from leaves. [4 marks]
1) Homogenise plant cell walls/membrane with blender/pestle & mortar and add a cold, buffered, isotonic solution.
2) Filter sample to remove debris & collect organelle-rich solution.
3) Centrifuge sample at low speed to separate the dense organelle first (nucleus).
4) Ultracentrifuge by repeating spinning at higher speeds. Chloroplast forms a pellet.
5) Syringe supernatant liquid out of tube to separate from pellet.
Describe the process of homogenisation [2 marks]
1) Vibrate the cells of the sample, or the sample is grinded in a blender or pestle & mortar.
This breaks down plasma MEMBRANE of cell, and releases organelles into the cold, isotonic, buffered solution.
Describe the process of filtration [1 mark]
1) Sample solution is filtered to remove large debris.
Describe the process of centrifugation [2 marks]
1) Spinning solution at high speeds to separate them by density.
The lower the density of the organelle, the higher speed is required to separate it.
The most dense organelle forms a pellet.
Then supernatant is removed from liquid, and centrifuged again.
Describe the process of ultracentrifugation [2 marks]
1) Spinning organelles at higher speeds each time to remove the next densest organelle to form a pellet.
State the order of density of organelles from most dense to least dense. [2 marks]
MOST DENSE
> nucleus
chloroplasts
mitochondria
lysosomes
membranes / RER / SER
ribosomes
LEAST DENSE
