Cell Fractionation

Question 1

Cell Fractionation

Answer 1

used to extract and isolate large numbers of organelles so we can study them

Question 2

Differential Centrifugation

Answer 2

organelles separated accordingly to weight

Question 3

Density Separation

Answer 3

organelles separated according to density

Question 4

Simple Sample Preparation

Answer 4

add a drop of water on to the slide,
obtain a thin section and place on the slide, 
stain with iodine in potassium iodide,
avoid trapping air bubbles,
lower coverslip using a mounted needle

Question 5

Conditions

Answer 5

chilled
isotonic
buffered
homogenised

Question 6

Chilled Solution

Answer 6

low temperature to slow down the rate of enzyme-controlled reactions

Question 7

Isotonic Solution

Answer 7

organelles do not take on or lose water due to osmosis

Question 8

Buffered Solution

Answer 8

prevents denaturing the enzymes inside the organelles

Question 9

Complex Sample Preparation

Answer 9

Cut up tissue sample
-tissue chilled and cut into pieces in cold, isotonic and buffered solution
Homogenise
-tissue homogenized in a pestle homogenizer or a blender to break open cells and release organelles
Filter
-homogenized suspension filtered
Spin
-filtrate centrifuged at low speed to remove debris and partly opened cells

Question 10

Homogenized

Answer 10

break open cells in order to release organelles

Question 11

Supernatant

Answer 11

the liquid in the tube above the sediment,
usually decanted (poured) off carefully

Question 12

Products

Answer 12

nuclei
chloroplasts
mitochondria
membranes

Question 13

Nuclei

Answer 13

centrifuge at low speed for 10 minutes

Question 14

Mitochondria (or chloroplast)

Answer 14

centrifuge at moderate speed for 15 minutes

Question 15

Membranes

Answer 15

centrifuge at high speed for 30 minutes

Question 16

Density Gradient Centrifugation

Answer 16

formed by different concentrations of sucrose solution, 
least dense (most dilute) = top
most dense (most concentrated) = bottom

Question 17

how they separate

Answer 17

accoridng to their density

Question 18

order of gradient

Answer 18

top
-ribosomes
-mitochondria
-chloroplast
-nuclei
-cell debris
bottom

Question 19

how to release organelles

Answer 19

pierce a hole in the bottom of the tube

meaning different organelles can be collected in separate tubes

Question 20

Ultra Centrifugation

Answer 20

special type of centrifugation
samples are spun at high speeds to separate the materials
spins many thousands of times per minute producing up to 100,000g