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Biology OCR B > Microscopy > Flashcards

Microscopy Flashcards

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1
Q

Define magnification

A

Number of times an image is enlarged compared to the actual object

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2
Q

Define resolution

A

The ability to distinguish between two separate points

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3
Q

How do you improve resolution?

A

Use radiation with a shorter wavelength e.g. beam of electrons.

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4
Q

What glass do light microscopes have?

A

Refractive glass to focus light into the eye

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5
Q

What is the maximum magnification of a light microscope?

A

1500x

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6
Q

What is the maximum resolution of a light microscope?

A

200nm

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7
Q

Give the magnifications of the objective lenses.

A

x4, x10, x40

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8
Q

Describe the pathway of light in a light microscope.

A

From a bulb, through a condenser lens, up through the specimen, through the objective lens and through the eyepiece lens.

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9
Q

Total magnification (formula 1) =

A

eyepiece magnification x objective magnification

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10
Q

Give six advantages of light microscopes.

A

Observe living cells, can use smear preparations with thin specimens, easy to use, inexpensive, don’t require specialist training, transported easily.

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11
Q

Give a disadvantage of light microscopes.

A

Limited resolution meaning most internal cellular structures cannot be seen.

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12
Q

What are stains used for?

A

Make different parts of the specimen stand out more clearly, many specimens are colourless and transparent.

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13
Q

Give examples of stains that can be applied directly to living cells.

A

Methylene blue and iodine solution

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14
Q

What does fixing mean?

A

Fixing a specimen in a position

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15
Q

How can you fix a specimen?

A

Adding acetic acid or alcohol to make the proteins and nucleic acids insoluble, so they are fixed in position

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16
Q

What does fixing do to cells?

A

Kills cells

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17
Q

True or false: stains can be added before or after the fixing process.

A

True

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18
Q

Magnification (formula 2) =

A

Size of image/size of object

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19
Q

mm -> micrometers

A

x1000

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20
Q

micrometers -> mm

A

/1000

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21
Q

Who discovered microscopes and when?

A

Robert Hooke, 1665

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22
Q

Where did the term ‘cell’ originate from?

A

Resembled where monks lived: cells.

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23
Q

Who developed cell theory?

A

Schleiden, Schwann and Virchow

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24
Q

Give the 6 terms of cell theory.

A

The cell is the basic unit of all life forms.
All organisms are made up of one cell (unicellular) or more than one cell (multicellular)
Metabolic processes take place inside cells
New cells are derived from existing cells
Cells contain genetic material of an organism which can be passed from parent cells to new daughter cells.
A cell is the smallest unit of an organism capable of surviving independently.

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25
Q

What does EM stand for?

A

Electron microscope

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26
Q

What is an EM’s source of radiation?

A

Beam of electrons

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27
Q

Who created the electron microscope?

A

Max Knoll and Ernst Ruska

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28
Q

What was the resolution of the basic prototype of the electron microscope?

A

50nm

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29
Q

Shorter wavelength =

A

greater resolving power

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30
Q

What is the resolving power of electron microscopes?

A

0.1nm

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31
Q

Give an advantage of electron microscopes.

A

Higher magnifications can be achieved, enabling smaller objects and finer detail to be seen.

32
Q

Give 5 disadvantages of electron microscopes.

A

Large, expensive, require trained personnel to use them, require a specially designed room to be used in, specimens must be placed in a near vacuum.

33
Q

What is the purpose of electrons being negatively charged in EM?

A

So the beam of electrons can be focused using electromagnets, making up the condenser.

34
Q

Why are specimens placed in a near vacuum with electron microscopes?

A

Air molecules absorb electrons - specimens must be dehydrated and dead

35
Q

Why are electrons focused onto a fluorescent screen?

A

Human eye can’t detect electrons, so a visible image is generated on a screen instead.

36
Q

Define: visible image on a screen

A

Photomicrograph

37
Q

What is an artefact?

A

A dark area, showing structures which aren’t actually part of the natural specimen.

38
Q

What colour are the final images of electron microscopes?

A

Black, white and grey.

39
Q

How can colour be added to photomicrographs?

A

Through specialist computer programs to produce false-colour electron micrographs.

40
Q

Name the two types of electron microscope.

A

Transmission electron microscope and scanning electron microscope

41
Q

What does TEM stand for?

A

Transmission electron microscope

42
Q

What does SEM stand for?

A

Scanning electron microscope

43
Q

Why do specimens have to be a lot thinner when using a TEM?

A

Electrons cannot penetrate materials as well as light can

44
Q

What is used to stain specimens for the TEM?

A

Heavy metals because they have a large, positively charged nucleus that scatter the electrons

45
Q

Scattered electrons do not hit the fluorescent screen and so

A

leave a dark area in the image.

46
Q

Images produced by a TEM are:

A

2D and black and white.

47
Q

How do SEMs work differently to TEMs?

A

Electrons don’t actually pass through the specimen

48
Q

What happens to electrons in SEMs?

A

They are reflected off the surface of the specimen

49
Q

Describe the pattern of electrons in SEMs

A

Beam of electrons is passed backwards and forwards over the surface of the specimen in a regular pattern

50
Q

What does the pattern of electrons do?

A

Reflects the contours of the specimen

51
Q

Images produced by SEMs are:

A

3D (made through computer analysis) and are black/white.

52
Q

What does CLSM stand for?

A

Confocal laser scanning micrscope

53
Q

When was CLSM developed?

A

1980s

54
Q

What is the main advantage of a CLSM?

A

Ability to produce focused images of thick specimens at various depths

55
Q

What images does a CLSM produce?

A

High resolution images which are 3D.

56
Q

Describe the process of optical sectioning.

A
  1. A point source of light is directed on to the object plane
  2. Fluorescence light is then directed through a detector pinhole, enhanced using a photomultiplier and displayed on a computer screen as a pixel.
  3. Scan the object point by point, recording one pixel at a time
  4. A series of optical sections can be combined to form an image stack, enabling 3D images to be produced.
57
Q

What is the function of fluorescent markers?

A

Detection of biological objects, commonly used in genetics and microbiology.

58
Q

Define the term fluorescence

A

Absorption and re-radiation of light.

59
Q

What is a fluorochrome/fluorescent marker?

A

A chemical dye which is applied to a specimen and then reflects the laser light to produce a pixel on a computer screen.

60
Q

Why is a laser used in CLSM?

A

To obtain higher intensities of light, improving illumination of the specimen.

61
Q

What is the function of the confocal pinhole?

A

Prevents unwanted radiation passing to the detector, ensuring a sharp image is obtained.

62
Q

Explain the purpose of the pinhole aperture in confocal microscopy.

A

Reduces blurring and increases resolution.

63
Q

What can images produced from CLSM be?

A

2D or 3D.

64
Q

What is the best horizontal resolution of a confocal microscope?

A

0.2 micrometers

65
Q

What is the best vertical resolution of a confocal microscope?

A

0.5 micrometers

66
Q

What is the maximum magnification of the confocal microscope?

A

200nm

67
Q

Why is the resolution of the CLSM the same as a light microscope?

A

It is using light as a source of radiation.

68
Q

Give 5 advantages of CLSM.

A

Produces focused images of thigh specimens by optical sectioning, high resolution, 3D images, live specimens can be used, fluorescent markers highlight biological structures.

69
Q

Describe how fluorescent markers can be used in CLSM.

A

Markers can attach to chemicals, antibodies and different biological structures. It has a wavelength of emission, meaning it will find structures with the laser.

70
Q

What is the resolving power of a SEM in nm?

A

20nm

71
Q

How is the beam focused in TEMs and SEMs?

A

Electromanets

72
Q

What is the maximum magnification for a TEM?

A

250,000x

73
Q

What is the maximum magnification for a SEM?

A

100,000x

74
Q

What is the maximum magnification for a CLSM?

A

2000x

75
Q

Maximum resolution for TEMs and SEMs?

A

0.1nm

76
Q

Can a live specimen be used with TEMs and SEMs?

A

No

77
Q

What mirror is present in CLSM?

A

Dichroic mirror

Biology OCR B (8 decks)

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