3.2.1.3 (methods of studying cells)

Question 1

Define magnification

Answer 1

Increasing the visual size of an object

Question 2

Define resolution

Answer 2

The ability to distinguish two close objects as being separate entities

Question 3

Magnification equation

Answer 3

size of image / size of real object

Question 4

Describe how light microscopes work

Answer 4

Magnify objects using light foccused into glass lenses

Question 5

Magnification of a light microscope

Answer 5

x1500

Question 6

Resolution of a light microscope

Answer 6

200nm

Question 7

Give advantages and disadvantages of light microscopes

Answer 7

Advantages- cheap to buy and run, simple to prepare specimens, living and dead tissue can be observed, colour images obtained
Disadvantages- limited resolution due to wavelength of light, limited magnification (x1500), restricted depth of field

Question 8

What are the two types of electron microscope?

Answer 8

TEM- transmission electron microscope
SEM- scanning electron microscope

Question 9

Describe how TEMs work

Answer 9

Beam of electrons passes through specimen and dispersed by structures in it
Scattered electrons captured on photographic plate
Electromagnets focus beam of electrons
In a vacuum so beam isn’t distorted
2D image

Question 10

Magnification of TEMs

Answer 10

x250,000

Question 11

Resolution of TEMs

Answer 11

2nm

Question 12

Describe how SEMs work

Answer 12

Specimen coated in very thin layer of metal
Beam of electrons bounce off surface onto photographic plate
Allows 3D images to be formed

Question 13

Magnification of SEMs

Answer 13

x100,000

Question 14

Resolution of SEMs

Answer 14

3-20nm

Question 15

Give limitations of SEM and TEM

Answer 15

Both- must be in a vacuum so living specimens can’t be observed, complex staining process required but image still black and white, image may contain artefacts
TEM- specimen must be very thin to allow electrons to penetrate, flat 2D image
SEM- lower resolving power than TEM

Question 16

What is cell fractionation?

Answer 16

Process where cells are broken up and the different organelles they contain separated out

Question 17

What is cell fractionation used for?

Answer 17

To study cell structure and function

Question 18

What are the stages of cell fractionation?

Answer 18

Homogenisation
Ultracentrifugation

Question 19

What type of solution is the tissue put in before cell fractionation and why?

Answer 19

Cold isotonic buffer solution
Cold- reduces enzyme activity which may break down organelles
Isotonic (same water potential as tissue)- prevents organelles bursting/ shrinking due to osmotic gain/loss of water
Buffer- to maintain constant pH to prevent proteins denaturing

Question 20

Describe the process of cell fractionation

Answer 20

1. tissue cut into pieces and put in cold isotonic buffered solution
2. tissue homogenised eg in blender
3. homogenised suspension filtered
4. filtrate centrifuged at low speed
5. supernatent decanted and recentrifuged at higher speeds until desired organelle seperated
6. supernatent analysed for content

Question 21

What order are the organelle pellets formed during cell fractionation?

Answer 21

1. nucleus
2. chloroplasts
3. mitochondria
4. lysosomes
5. endoplasmic reticulum
6. ribosomes