Methods of Studying Cells

Question 1

What is magnification?

Answer 1

The number of times greater the image is than the specimen.

Question 2

How can magnification can be calculated?

Answer 2

magnification = image size/actual size

Question 3

What is resolution?

Answer 3

The ability/how well a microscope distinguishes between two points that are close together.

Question 4

What are the limitations of an optical microscope?

Answer 4

• Limited by wavelength of visible light.
• Impossible to resolve 2 objects that are closer than half a wavelength of visible light.
• Visible light wavelength is 500-650nm.
• Wavelength of light not short enough / too long to see smaller organelles.

Question 5

What are the steps for preparing a cheek cell slide?

Answer 5

Making a cheek cell slide:
1). Scrape inside of cheek with cotton bud.
2). Smear it on slide.
3). Add 1 drop of methylene blue stain.
4). Add a coverslip.

Question 6

What are the steps for preparing an onion cell slide?

Answer 6

1). Using tweezers peel off a thin layer of onion skin.
2). Lay this carefully onto the slide.
3). Add 1 drop of iodine.
4). Add a coverslip.
5). Remove any excess iodine with absorbing with a tissue.

Question 7

What are the features of electron microscopes?

Answer 7

• Very short wavelength, therefore high resolving power.
• Electrons negatively charged so beams can be focused using electromagnets.
• Electrons absorbed or deflected by molecules in the air so vacuum needed.

Question 8

What are the principles of a TEM?

Answer 8

• Beams of electrons pass through thin sections of specimen.
• Some parts absorb electrons (appear darker) or allow electrons to pass through (lighter).
• Forms flat 2d image.

Question 9

What are the advantages of a TEM?

Answer 9

+ High resolution (0.1nm)
+ Magnifications up to 10,000x

Question 10

What are the disadvantages of a TEM?

Answer 10

• Vacuum, water must be removed, specimens are dead.
• Lengthy preservation and staining.
• Foreign artefacts can be accidentally introduced.

Question 11

What are the principles of an SEM?

Answer 11

• Electron beams bounce off surfaces of objects.
• 3D structure, e.g. viruses can be seen.
• 20nm resolving power (lower than TEM).

Question 12

What are the steps of cell fractionation?

Answer 12

• Placing tissue in solution (cold, buffered, isotonic)
• Homogenation
• Ultracentrifugation

Question 13

The solution that the tissue is placed in is what 3 things? Why is it these 3 things?

Answer 13

• Cold: to reduce enzyme activity that might break down the organelle.
• Buffered: so pH doesn’t fluctuate as change in pH could alter the structure of organelle/affect functioning enzymes.
• Isotonic (same water potential): prevent organelle shrinking or bursting as a result of osmotic loss or gain of water.

Question 14

Describe homogenation.

Answer 14

• Cells broken up by homogenizer (blender), releasing organelles.
• Resultant fluid (homogenate) is filtered to remove complete cells and large debris.

Question 15

Describe ultracentrifugation.

Answer 15

• Fragments in filtered homogenate are separated by a centrifuge.
• Centrifuge spins tubes of homogenate at high speeds to create centrifugal force.
• Tube of filtrate placed into centrifuge and spun at a lower speed.
• Heaviest organelles (nuclei) are forced to the bottom forming a pellet.
• Fluid at top (supernatant) removed and spun at a higher speed.
• Next heaviest pellet forms.

Question 16

Order of size of pellet formed?

Answer 16

Nuclei (largest), chloroplast (if plant cell), mitochondria, lysosomes, endoplasmic reticulum, ribosomes.