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A-Level Biology - Two > 2.1 Microscopes > Flashcards

2.1 Microscopes Flashcards

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1
Q

What was the first microscope to be developed and when?

A
  • light microscope
  • 16th and 17th century
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2
Q

What became accessible in the mid 19th century?

A
  • microscopes with a high enough level of magnification to allow them to see individual cells
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3
Q

What does the cell theory state?

A
  • both plant and animal tissue is composed of cells
  • cells are the basic unit of all life
  • cells only develop from existing cells
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4
Q

How does a compound light microscope work?

A
  • using a visual light source beneath the specimen
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5
Q

What are the four different sample preparation methods?

A

dry mount, squash slides, smear slides, wet mount

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6
Q

How is the dry mount sample preparation done?

A
  • solid samples are either viewed whole or into very thin slices with a sharp blade
  • specimen placed on top of the slide and cover slip cover it
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7
Q

How is the wet mount sample preparation done?

A
  • specimens are suspended in a liquid such as water or an immersion oil
  • a cover slip is placed on an angle
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8
Q

How is the squash slides preparation done?

A
  • a wet mount is first prepared, then a lens tissue is used to gently press down the cover slip
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9
Q

How is the smear slide sample preparation done?

A
  • the edge of a slide is used to smear the sample
  • a cover slip is then placed over the same sample
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10
Q

What happens in basic light microscopy in terms of light?

A
  • sample is illuminated from below with bright white light and observed from above
  • images tend to have low contrast as most cells do not absorb a lot of light
  • resolution is limited of the wavelength of light and diffraction of light as it passes through a sample
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11
Q

What is wide-field microscopy?

A
  • the whole sample is illuminated at once
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12
Q

What is crystal violet and methylene blue staining?

A
  • positively charged dyes
  • attracted to negatively charged materials in cytoplasm
  • leading to staining of cell components
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13
Q

What is nigrosine and congo red staining?

A
  • negatively charged dyes
  • repelled by the negatively charged cytosol
  • these dyes stay outside the cells, leaving the cells unstained, to stand out against the stained background
  • this a negative stain technique
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14
Q

What is differential staining?

A
  • it can distinguish between two types of organisms
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15
Q

What is the gram stain technique?

A
  • used to separate bacteria into two groups, gram-positive/ gram-negative bacteria
  • crystal violet is first applied to a bacterial specimen then iodine, which fixes the dye
  • the slide is then washed with alcohol
  • then stained with safranin dye called a counterstain
  • bacteria will then appear red
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16
Q

What happens to the gram stains with crystal violet staining?

A
  • the gram-positive bacteria retain the crystal-violet stain and will appear blue or purple under a microscope
  • gram-negative bacteria have thinner cell walls and therefore lose the stain
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17
Q

What happens to gram stains with the counterstain?

A
  • gram-positive bacteria are susceptible to the antibiotic penicillin, which inhibits the formation of cell walls
  • gram-negative bacteria have much thinner cell wall that are not susceptible to penicillin
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18
Q

What are the stage involved in the production of slides?

A

Fixing - chemicals like formaldehyde are used to preserve specimens
Sectioning - specimens are dehydrated with alcohols and then are placed in a mould with wax or resin to form a hard block
Staining - specimens are often treated with multiple stains to show different structures
Mounting - the specimens are then secured to a microscope slide and a cover slip placed on top

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19
Q

Magnification is …

A
  • the degree to which the size of an image is larger than the viewed object (specimen) itself
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20
Q

Resolution is …

A
  • the degree to which it is possible to distinguish between two objects (adjacent points) that are very close together
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21
Q

What is the equation for magnification?

A

magnification = size of image/ actual size of object

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22
Q

What is calibrating a microscope mean?

A
  • used when measuring the size of a sample under a microscope by using the eye piece graticule
  • true magnification of the lenses of a microscope changes dependent on the microscope itself
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23
Q

What is an eye piece graticule?

A
  • a glass disc marked with a fine scale from 1 to 100
  • the relative size of the divisions increases with each increase in magnification
  • the scale on the graticule at each magnification is calibrated using a stage micrometer
24
Q

What is a stage micrometer?

A
  • a microscope slide with a very accurate scale in micrometers engraved on it
  • the scale marked on the micrometer slide is usually 100 divisions = 1 mm so 1 division = 1um
25
Q
A
26
Q

What is laser scanning confocal microscopy?

A
  • uses laser beams to scan a specimen tagged with dyes
  • it picks up fluorescence, out of focus light is blocked
  • objects can be seen at different depths
  • a 3D image is generated
  • it is still a light microscope so the resolution is similar to the light microscope
27
Q

Advantages of light microscopes.

A
  • easy to use
  • cheap to purchase (less than £1K)
  • true to colour but sometimes require staining
  • could use live specimens
28
Q

Disadvantages of light microscopy.

A
  • low resolution due to wavelength of light (0.2um)
  • low magnification (max x1250)
  • specimens are thin; may not be representative
29
Q

Advantages of scanning electron microscopy (SEM).

A
  • much higher resolution than light microscopy (1nm)
  • provides detailed images of surface structures
  • high magnification (x200000)
  • 3D image
30
Q

Disadvantages of scanning electron microscopy (SEM).

A
  • expensive
  • extensive training required
  • samples must be dead (vacuum, stains)
  • black and white false colour
31
Q
A
32
Q

Advantages of transmition electron microscopes (TEM).

A
  • much higher resolution than light microscopes (1nm)
  • provides detailed images of interior structures
  • high magnification (x500000)
33
Q

Advantages of transmition electron microscopes (TEM).

A
  • much higher resolution than light microscopes (1nm)
  • provides detailed images of interior structures
  • high magnification (x500000)
34
Q
A
35
Q

Disadvantages of transmition electron microscopes (TEM).

A
  • expensive
  • extensive training required
  • samples must be dead (vacuum, stains)
  • black and white false colour
36
Q

What is electron microscopy?

A
  • a beam of electrons with a wavelength of less than 1nm is used to illuminate the specimen
  • more detail of cell ultrastructure can be seen because electrons have a much smaller wavelength than light waves
  • they can produce images with magnifications up to x500,000 and still have clear resolution
37
Q

What is a TEM?

A
  • a beam of electrons is transmitted through a specimen and focused to produce an image
  • similar to light microscopy
  • has the best resolution with a resolving power of 0.5nm
38
Q

What is SEM?

A
  • a beam of electrons in sent across the surface of a specimen and the reflected electrons are collected
  • the resolving power is from 3-10nm, so the resolution is not as good as the TEM but 3D images of surfaces are produced
  • this gives us valuable information about the appearance of different organisms
39
Q

How is a sample prepared for electron microscopes?

A
  • the inside of an electron microscope is a vacuum to ensure the electron beams travel in straight lines and this means samples must be prepared in a specific way
  • preparation involves fixation using chemicals or freezing, staining with heavy metals and dehydration with solvents
  • samples for a TEM will then be set in resin and may be stained again
  • samples for SEM may be fractured to expose the inside and will then need to be coated with heavy metals
40
Q

What is an artefact and how is it used?

A
  • a visible structural detail caused by processing the specimen and not a feature of the specimen
  • the bubbles that get trapped under the cover slip as you prepare a slide for light microscopy are artefacts
41
Q

What does the term ‘mesosome’ mean?

A
  • name given to investigations (inward folding’s) of cell membranes that were observed using an electron microscope after bacterial specimens had been chemically fixed
42
Q

What do conventional optical microscopes use?

A
  • visible light to illuminate specimens and a lens to produce a magnified image
43
Q

In fluorescent microscopes what is used to illuminate a specimen?

A
  • a higher light intensity
  • if treated with with a fluorescent chemical (a fluorescent ‘dye’)
44
Q

What is fluorescence?

A
  • the absorption and re-radiation of light
45
Q

What type of light is used to produce a magnified image?

A
  • light of a longer wavelength and lower energy is emitted
46
Q

What does a laser scanning confocal microscope do?

A
  • moves a single spot of focused light across a specimen (point illumination)
47
Q

Focal plane is …

A
  • the distance that gives the sharpest focus
48
Q

How is a 2D and 3D image produced in LSCM?

A

2D - the spot illuminating the specimen is moved across the specimen
3D - creating images at different focal planes

49
Q

What is a practical application of LSCM?

A
  • it is non-invasive
  • used in the diagnosis of diseases of the eye
  • being developed for use in endoscopic procedures
50
Q

What is a beamsplitter?

A
  • a dichroic mirror, which only reflects one wavelength from the laser, but allows other wavelengths produced by the same to pass through
51
Q

What does atomic force microscopy do?

A
  • gathers information about a specimen by ‘feeling’ its surface with a mechanical probe
52
Q

What is a mechanical probe?

A
  • these are scanning microscopes that generate three-dimensional images of surfaces
53
Q

What are the two principles of super resolved fluorescent microscopy?

A
  • involves building up a very high resolution image by combining many small images
  • involves superimposing many images with normal resolution to create one very high resolution image
54
Q

What are the benefits of being able to follow certain molecules in SRFM?

A
  • cellular processes
  • proteins involved in Parkinson’s and Alzheimer’s disease can be observed interacting
  • fertilsed eggs dividing into embryos can be studied at a molecular level
55
Q

What is a benefit of ATM?

A
  • fixation and staining are not required
  • specimens can be viewed in almost normal cell conditions without damage caused during the preparation of specimens for electron microscopy
  • living systems can be examined
  • very high resolution

A-Level Biology - Two (14 decks)

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