unit 2,ultracentrifugation Flashcards
what is cell fractionation
where cells are broken up and the different organelles they contain are separated out , depending on their size and mass using gravity
what are the three stages in the process called
homogenisation, filtration, ultracentrifugation
why is the tissue placed in cold solution
to reduce enzyme activity that may break down organelles , prevent enzyme digestion
why is the tissue placed in an isotonic solution (same water potential as original tissue)
to prevent organelles bursting/ shrinking due to osmotic loss/ gain of water
why is the tissue placed in placed in a buffer solution
to maintain constant ph , so proteins don’t denature
what is homogenising
it is blending , it breaks up the tissue to break open cell membranes and release the organelles
why is the homogenate filtered
to remove large debris that will be heavy and sink to the bottom of the test tube
what is the process of centrifuging
Starts at slow speed to allow the large fragments to collect at the bottom of the test tube and the small ones to stay within the liquid (supernatant).
It is then re spun at a higher speed to collect the smaller fragments at the bottom of the test tube forming a new pellet.
this method is repeated until smaller organelles will be recovered
what is the liquid within centrifuging called
supernatant
what speed and time duration do the nucleus require to be seperatee
x1000 , for 10 minutes
speed and time duration required for extraction of chloroplasts, mitochondria
x3500 , 10 minute duration
speed and time duration required for extraction of lysosomes and endoplasmic reticulum
x16,500 , 20 mins
speed and duration required for extraction of ribosomes
x100,000 , 60 mins
