Ultracentrifugation

Question 1

What is cell fragmentation?

Answer 1

How we separate organelle out on a cell

Question 2

Explain steps of cell fractionation and ultracentrifugation?

Answer 2

-cell is cut into pieces and placed in a cold isotonic buffer solution and organelles are suspended in a solution
Homogenetion
-placed in a homogeniser (scientific blender) and cell wall and membrane be is broken
-it is then filtered out to then collect any unwanted solid particles like cell debris or nuclei
-the resulting cell homogenate is spun in centrifuge (less it is spun the larger the pellets in centrifuge and the more it is spun the smaller the pellets produced) creating supernatant (top liquid) and nuclei bottom,liquid is removed leaving sediment of nuclei and the supernatant is then transferred to another tube and spun
(The heaviest is made 1st,the lightest 2nd)
-then spun again until small organelles aren’t made.

Question 3

Why is cell put into a cold isotonic buffer solution?

Answer 3

-Cold to it reduces enzymes activity
-isotonic:same water potential in and out of cell so then it doesn’t shrinking nor bursting.
-buffer:keeps ph same.

Question 4

Why is the cell homogenise?

Answer 4

To break down cell wall and membrane and now only organelles are left.

Question 5

Why filter the resolution suspension?

Answer 5

To separate organelle from the unwanted solid particle e.g cell debris and nuclei

Question 6

What is ultracentrifugation?

Answer 6

Is a powerful laboratory technique used to separate and analyse particles

Question 7

Give me the order of fractionation from large to small

Answer 7

-Nuclei
-Chloroplast
-Mitochondria
-Lysosomes
-ER
-ribosome