Microscopes

Question 1

How can we calculate magnification?

Question 2

What does resolution mean?

Answer 2

How much two objects can be distinguished from each other when placed very closely together.

Question 3

What are the stages of cell fractionation?

Answer 3

1. Homogenisation.
2. Filtration.
3. Ultracentrifugation

Question 4

What happens during homogenisation and why is it done?

Answer 4

The tissue is placed in an isotonic, ice-cold, buffered solution and then blended using a homogenizing blender. This is done to break up the cells and release the organelles.

Question 5

Why must the solution the tissue is placed in be ice-cold?

Answer 5

To slow down enzyme action and prevent the digestion of the organelles.

Question 6

Why must the solution the tissue is placed in be buffered?

Answer 6

To maintain the pH and to prevent the breakdown of the proteins that make up the organelles.

Question 7

Why must the solution the tissue is placed in be isotonic?

Answer 7

Prevents osmosis of water out of the organelles. This stops the shrinkage of organelles.

Question 8

What happens during filtration?

Answer 8

The homogenate formed is filtered using a cheese cloth to remove any unbroken cells or cell debris.

Question 9

What happens during ultracentrifugation?

Answer 9

1. The homogenate is placed inside a test tube in the centrifuge. The centrifuge is then spun at low speeds and time to remove denser organelles like the nucleus. The organelles sink to the bottom of the beaker and can be collected as a pellet. The other organelles float at the top and form a solution called the supernatant.
2. The supernatant is spun at increasing speeds and time to remove less dense organelles such as the mitochondria and then the lysosomes. These sink to the bottom of the test tube and can be collected as a pellet.
3. The organelles can then be observed using a microscope.

Question 10

What are the differences between optical microscopes and electron microscopes?

Answer 10

1. Optical microscopes use light to create an image and electron microscopes use electron beams.
2. Electron microscopes have a higher magnification and resolution.
3. Electron microscopes require the use of a vacuum and optical ones don’t. This means that optical microscopes can be used to observe living organisms and electron microscopes can’t.

Question 11

Why do electron microscopes have a higher resolution?

Answer 11

Electron beams have a shorter wavelength compared to light.

Question 12

How does a Transmission Electron Microscope work?

Answer 12

Electrons are fired at this sample using an electron gun. The parts of the sample that absorb those electrons appear darker in the image formed.

Question 13

How does a Scanning Electron Microscope work?

Answer 13

Electrons are scattered across the surface of a sample. The pattern of the scatter is used to create an image.

Question 14

What are the advantages and disadvantages of a TEM over an SEM?

Answer 14

1. TEM microscopes have a higher resolution and magnification compared to SEM microscopes.
2. The sample for SEM microscopes doesn’t have to be very thin, unlike the ones for TEM microscopes.
3. TEM microscopes make a 2D black and white image whereas SEM microscopes for a 3D coloured image.