Cell Fractionation

Centrifuge (2)

A machine that spins containers

The heaviest material in the container will fall to the bottom

Cell fractionation (2)

Uses centrifugation to break uo the components of a cell

Seperates them by size for study

What is an isotonic solution? (2)

A solution which has the same water potential as the substance placed inside it

Prevents organelles from shrinking or bursting by osmosis

C. Fractionation: Step 1 (3)

Cells are placed in a solution

Cold - to reduce enzyme activity that could break the cell

Isotonic - to prevent them bursting or shrinking (osmosis)

Buffered - to maintain pH

C. Fractionation: Step 2 (3)

Homogenization

Cells are broken by a homogenizer (blender) - this releases the organelles from the cell walls / membranes

Fluid gained is called homogenate - a filter is used to remove any whole cells or large debris

C. Fractionation: Step 3 (4)

Ultracentrifugation

Homogenate is spun at a very high speed to create a centrifugal force

Organelles seperate according to mass

Heaviest fall to the bottom at lower speed

C. Fractionation: Step 5 (2)

The supernatant is removed and respun at higher speeds

Step 4 is repeated until all that is left is the lighest eg. ribosomes

What is the order of organelle fractionation?

Nucleus → (chloroplast) → mitochondria → Lysosymes → ER → Ribosomes

What are the heaviest organelles in the centrifuge called? (2)

Pellet

They are forced downwards and collect at the bottom of the tube

What is the supernatant? (2)

Fluid at the top of the tube composed of smaller, leess dense organelles

This is removed and re-spun