Methods of studying cells

Question 1

What are the benefits of using an electron microscope rather than a light microscope?

Answer 1

• The light microscope has a relatively long wavelength of light rays so they can only distinguish 2 objects that are 0.2um apart.
• An electron microscope uses beams of electrons so it has a shorter wavelength rather than light so it can distinguish 2 objects that are only 0.1um apart.

Question 2

What is the equation for magnification?

Answer 2

Magnification = image / real

MIR

Question 3

What are the units of length?

Answer 3

km = 10^3m
m = 1m
mm = 10^-3m
um = 10^-6m
nm = 10^-9m

Question 4

What is the resolution?

Answer 4

Resolution of a microscope is the minimum distance apart that 2 objects can be in order to appear as 2 separate items.

Question 5

What does resolution depend on?

Answer 5

Wavelength or form of radiation used.
E.g. Light microscopes is about 0.2um so objects which are 0.2um or more apart will appear as one single object. Greater the resolution, better the clarity.

Question 6

What does an increase of magnification do?

Answer 6

It increases the size of an image but it does not always increase the resolution.

Question 7

What is cell fractionation?

Answer 7

The process where cells are been broken up and the different organelles they contain are separated out so we can study the structures and functions at a better detail.

Question 8

Before cell fractionation can begin, it must be placed into a cold, buffered solution which has the same water potential as the tissue, why?

Answer 8

Cold - reduce enzyme activities which might break down the organelles.
Buffered - pH does not change, because any change in the pH can alter the structure of the organelles and therefore the functions of the enzymes.
Same water potential as the tissue - prevents the organelles from bursting or shrinking due to osmotic gain or loss of water.

Question 9

What are the 2 stages of cell fractionation and briefly explain how it works?

Answer 9

1) Homogenation - Cells are broken down by a homogeniser (blender), which allows the organelles to be released. The fluid - homogenate, is filtered to remove any complete cells and large pieces of debris.
2) Ultracentrifugation - Fragments in the filtered homogenates are been separated in a machine called centrifuge and it spins the tube at a very high speed in order to create a centrifugal force.

Question 10

Explain the steps to ultracentrifugation.

Answer 10

• The tubes of filtrates are placed in the centrifuge and spun at a slow speed.
• Heaviest organelles - nuclei would be forced towards the bottom where it forms a thin layer of sediment.
• The fluid at the top of the test tube (supernatant) is removed leaving behind the sediment.
• The supernatant is transferred to another tube and spun in the centrifuge at a faster speed than before and the next heaviest organelle - mitochondria are forced to the bottom.
• The process repeats until everything is separated.