microscopy and magnification

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

minimum distance apart 2 objects can be distinguished as separate objects

Question 3

what is the magnification calculation?

Answer 3

Question 4

what are the principles of using an optical (light) microscope?

Answer 4

-light focused using glass lenses
-light passes through specimen, different structures absorb different amounts and wavelengths
-generates a 2D image of a cross-section
-specimen must be thin

Question 5

what are the advantages of using an optical microscope?

Answer 5

-can show colour
-simple preparation
-can view living organisms

Question 6

what are the limitations of using an optical microscope?

Answer 6

-low resolution due to long wavelength of light
-cant see internal structure of organelles or ribosomes
-low magnification (x1,500)

Question 7

what are the principles of using a transmission electron microscope (TEM)?

Answer 7

-electrons focused using electromagnets
-electrons pass through specimen, denser parts absorb more and appear darker
-generates a 2D image of a cross section
-specimen must be thinner
-can only view dead/dehydrated specimens as uses a vacuum

Question 8

what are the advantages of using TEM?

Answer 8

-very high resolution due to short wavelength of electrons
-Can see internal structures of
organelles and ribosomes
-higher magnification than SEM
(x1,000,000)

Question 9

what are the limitations of using TEM?

Answer 9

-complex preparation so artefacts often present
-does not show colour

Question 10

what are the principles of using a scanning electron microscope (SEM)?

Answer 10

-electrons focused using electromagnets
-electrons deflected/bounced off specimen surface
-generates a 3D image of surface
-specimen does not need to be thin
-can only view dead/dehydrated specimens as uses a vacuum

Question 11

what are the advantages of using SEM?

Answer 11

high resolution due to short wavelength of electrons
-high magnification(x1,000,000)

Question 12

what are the limitation of using SEM?

Answer 12

-cant see internal structures
-complex preparation so artefacts often present
-does not show colour

Question 13

suggest how the scientific community distinguished between artefacts(e.g. dust, air bubbles occurring during preparation) and cell organelles

Answer 13

-scientists prepared specimens in different ways
-if an object was seen with one technique but not another, it was more likely to be an artefact than an organelle

Question 14

unit conversions

Answer 14

cm → mm = x10

Question 15

describe how the size of an object viewed with an optical microscope can be measured

Answer 15

1. Line up (scale of) eyepiece graticule with (scale of) stage micrometre
2. Calibrate eyepiece graticule - use stage micrometre to calculate size of divisions on eyepiece graticule
3. Take micrometre away and use graticule to measure how many divisions make up the object
4. Calculate size of object by multiplying number of divisions by size of division
5. Recalibrate eyepiece graticule at different magnifications

Question 16

describe how the student could use an eye piece to determine the mean diameter of the organelle

Answer 16

1.calibrate the eye piece graticule
2. using the stage micrometre
3.meaure the organelle/cell length
4.repeat the measurement and calculate a mean

Question 17

how could you determine the mean length of a cell using an eye piece graticule?

Answer 17

• calibrate the graticule using a stage micrometre
• measure the length pf a number of cells at random using the graticule
• calculate a mean

Question 18

when preparing a slide for viewing why must you press down hard on the coverslip?

Answer 18

to squash the tissue (don’t push side ways to avoid cells rolling together)

Question 19

why must the specimen be thin?

Answer 19

-single layer of cells
-to allow light pass through

Question 20

how do you prepare a temporary mount?

Answer 20

• add drop of water to slide
• take a thin section of tissue (1 cell thick)
  -place on glass slide (float on water)
  -add stain
  -place on coverslip
  -press down firmly