Microscopes

Question 1

Define magnification?

Answer 1

The degree to which the size of an image is larger than the object itself

Question 2

Define resolution?

Answer 2

The minimum distance apart hat two objects can be in order for them to appear as separate items

Question 3

What is used to measure the size of cells (in a microscope)

Answer 3

Eyepiece graticule

Question 4

What scale are cells/objects measured using a microscope?

Answer 4

Eyepiece units (EpU)

Question 5

What are the rules when completing a biological drawing

Answer 5

• use a sharp pencil
• should take up at least half a page
• lines need to be clear and continuous (no shading or sketchy lines)
• ensure proportions are correct
• label all features that you have shown.
• Rule label lines
• don’t cross label lines or use arrows
• don’t write on label lines
• ensure label lines touch the part you are labelling

Question 6

What is the equation for magnification

Answer 6

Image size / actual size = mag

Question 7

Describe how you could make a temporary mount of a piece of plant tissue to observe the position of starch granules.

Answer 7

• add a drop of water to the glass slide.
• obtain a thin section of plant tissue (potato) to ensure light can pass through
• stain with iodine in a potassium iodide solution
• lower cover slip on slide using a mounted needle (ensures no air bubbles)

Question 8

How do you calibrate a microscope to work out the mean diameter of starch grains?

Answer 8

Measure the diameter of the grain using the eyepiece graticule .
Calibrate the eyepiece graticule using a stage micrometer at the same magnification.
Randomly sample 10+ grains and calculate the mean

Question 9

What is cell fractionation?

Answer 9

The process in which cells are broken up (lysis) and different organelles within the cells are separated out so they can be studied in detail.

Question 10

Before cell fractionation can begin what type of solution must the the tissue be placed in?

Answer 10

A cold, buffered, isotonic (same water potential) solution

Question 11

What does isotonic mean?

Answer 11

The solutions have the same water potential.

Question 12

Why should the solution be cold?

Answer 12

To reduce enzyme activity to prevent organelles being broken down ( by digestion)

Question 13

Why should the solution be buffered?

Answer 13

So the PH doesn’t fluctuate so proteins don’t denature

Question 14

What should the solution be isotonic (have the same water potential)

Answer 14

To prevent organelles shrinking or bursting as a result of osmotic gain/loss of water

Question 15

Describe the process of cell fractionation

Answer 15

• homogenisation - break open cells by vibrating or grinding them up (in a a homogeniser - blender) to release organelles from cells.
• filtration- the resultant fluid (homogenate) is filtered to remove any debris (whole cells/bits of tissue)
• ultracentrifugation - the fragments in the filtered homogenate are separated in a centrifuge at increasing speeds

Question 16

Describe the process of ultracentrifugation

Answer 16

The first centrifuge spin is low-speed and separates the heaviest organelles (nuclei) out first. They are forced to the bottom of the tube where a thin sediment/pellet forms while the less dense remain in suspension above (supernatant)
The supernatant (is removed leaving just the sediment ) is then transferred to the next tube and spun at a faster speed. (This next pellet contains the next most heavy/dense organelle - mitochondria)

Question 17

What are the 3 types of microscopes?

Answer 17

Light/optical
Transmission electron microscope (TEMs)
Scanning electron microscopes (SEMs)

Question 18

What type of electromagnetic radiation do optical microscopes use?

Answer 18

Beams of Visible light (has a long wave length which reduces resolving power)

Question 19

What type of electromagnetic radiation do TEMs and SEMs use?

Answer 19

Beams of high energy electrons which have a shorter wavelength leading to a higher resolving power.

Question 20

Compare the size of magnification between optical microscopes and TEMs/SEMs?

Answer 20

Optical microscopes have a lower magnification (x400)
SEMs and TEMs have a higher magnification (x 1,000,000)

Question 21

Which microscope has the highest resolving power?

Answer 21

TEMs

Question 22

Compare the resolving power for all 3 microscopes.

Answer 22

• optical = lowest (0.2 um)
• SEMs = higher (5 - 20 nm)
• TEMs = highest (0.2 nm)

Question 23

Which is the only microscope to produce a 3D image

Answer 23

SEMs
Scanning electron microscope

Question 24

How do you focus an optical microscope?

Answer 24

Use a pair of lenses

Question 25

How do you focus an electron microscope?

Answer 25

Use electron magnets.

Question 26

Which microscope enables you to view a specimen in colour?

Answer 26

Optical microscope
The TEMs and SEMs only view in black and white.

Question 27

Which microscope can you view live specimens?

Answer 27

Optical microscopes
The electron microscopes take place in a vacuum so only dead / dehydrated specimens can be viewed.

Question 28

Which microscopes require a thin specimens?

Answer 28

Light/optical microscopes and TEMs
SEMs can view thicker specimens as they produce a 3D image

Question 29

Which is the only portable microscope?

Answer 29

Optical microscopes

Question 30

Compare the value of microscopes

Answer 30

• optical microscopes are cheap
• Electron microscopes are expensive.

Question 31

Which microscope requires the most complex staining process?

Answer 31

TEMs
- requires use of heavy metals

SEMs and Optical microscopes don’t need staining.

Question 32

What is an artefact?

Answer 32

Something you see under a microscope (when looking at a prepared sample ) that isn’t actually part of the specimen. (E,g dust)

Question 33

Which microscope suffers mostly from the presence of artifices.

Answer 33

TEMs due to a more complex staining process

Question 34

What are examples of artefacts?

Answer 34

Dust, bubbles and finger prints.