[OLD] Cells - Cell Fractionation Flashcards
Why is the centrifugal solution isotonic?
So osmosis won’t occur so the organelles won’t shrivel or burst
Why is the centrifugal solution ice-cold?
To prevent enzymes self-digesting
Which is the densest organelle?
Nucleus
Name 4 methods of homogenisation
Osmotic shock
Ultrasonic vibration
Force through a grid
Very close fitting plunger in a tube
what is homogenisation?
breaking up the cells
what does homogenisation do?
breaks up the plasma membrane and releases the organelles into solution
What are the conditions needed for homogenisation?
solution must be ice cold to reduce activity of enzymes that break down organelles.
solution should be isotonic to prevent damage to organelles through osmosis.
a buffer solution should be added to maintain the pH.
steps of cell fractionation
homogenisation
filtration
ultracentrifugation
what happens during filtration?
the homogenised cell solution is filtered through a gauze to separate any large cell debris or tissue debris, like connective tissue, from the organelles. The organelles are much smaller than the debris, so they pass through the gauze.
what happens during ultracentrifugation?
cell fragments centrifuged in a tube at a low speed. heaviest organelles form a thick sediment at the bottom of the tube called the pellet. the rest of the organelles stay suspended in the supernatant. the supernatant is drained off and centrifuged in another tube at a higher speed. this process is repeated at higher and higher speeds until all the organelles are separated out.
what is the supernatant?
the fluid above the sediment/pellet
order of mass of organelles?
nuclei, mitochondria, lysosomes, endoplasmic reticulum, ribosomes
(in plants it’s nuclei, chloroplasts, mitochondria)
