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A Level Biology > Microscopy > Flashcards

Microscopy Flashcards

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1
Q

What is magnification

A

How many times bigger the image produced by the microscope is than the real life object

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2
Q

What is resolution

A

The ability to distinguish between 2 objects that are close together (seeing two structures that are very close as two separate objects)

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3
Q

How to calculate magnification of a light microscope

A

Total magnification = Eyepiece lens magnification (10) x Objective lens magnification

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4
Q

Magnification equation

A

IAM triangle

           Image size

Actual size Magnification

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5
Q

See images for labelled diagram of microscope

A
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6
Q

Description of Light microscope

A
  • Use light to form an image
  • Using light, it is difficult to distinguish between (resolve) two objects that are closer than the wavelength of light 500-650nm
  • up to x2000 magnification, maximum resolution is 0.2pm
  • Can be used to observe eukaryotic cells, nuclei & possibly mitochondria & chloroplasts. Cannot observe ribosomes, ER or lysosomes
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7
Q

Description of Electron microscope

A
  • Use electrons to form an image
  • This increases the resolution, giving more detail bc a beam of electrons has a much smaller wavelength than light.
  • Can resolve 2 objects that are extremely close tg. Maximum resolution of 0.2nm (1000x greater than optical microscopes)
  • Can observe small organelles sa ribosomes, ER & lysosomes
  • 2 types of e microscopes: Transmission electron microscope, Scanning electron microscope
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8
Q

Advantages of Light microscope

A
  • inexpensive to buy & operate
  • small & portable
  • simple sample preparation
  • vacuum is not required
  • natural colour of sample is seen (or stains are used)
  • specimens can be living or dead
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9
Q

Disadvantages of Electron microscope

A
  • expensive to buy & operate
  • large & needs to be installed
  • complex sample preparation which often distorts material
  • vacuum is required
  • black & white images produced (can only be coloured digitally)
  • specimens must be dead
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10
Q

What are Scanning electron microscopes

A
  • SEMs scan a beam of electrons across the surface of the specimen. This bounces off the surface & knocks off electrons from the specimen, which are detected in a cathode ray tube forming an image
  • SEMs therefore form 3D images that show the surface of a specimen
  • lower res images than TEMs. Magnification x500,000 or less
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11
Q

Advantages of Scanning electron microscope

A
  • can be used on thick or 3D specimens
  • allow external 3D structure to be observed
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12
Q

Disadvantages of Scanning electron microscope

A
  • lower resolution than TEMs
  • cannot be used on live specimen (unlike optical microscopes)
  • do not produce a colour image (unlike optical microscopes)
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13
Q

What are Transmission electron microscopes

A
  • TEMs use electromagnets to focus a beam of electrons, which is transmitted through the specimen
  • Denser parts of the specimen absorb more electrons, making them appear darker on the final image
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14
Q

Advantages of Transmission electron microscope

A
  • high resolution images = more detail
  • internal structures can be seen
  • magnification x1,000,000 or more
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15
Q

Disadvantages of Transmission electron microscope

A
  • only used w very thin specimens or thin sections of an object
  • cannot be used to observe live specimens bc of the vacuum inside the TEM plus all the water must be removed from the specimen
  • lengthy treatment required to prepare specimens. Artefacts could be introduced - these look like real structures but are acc the results of preserving & staining
  • do not produce a colour image & are in 2D
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16
Q

What are Laser scanning confocal microscopes

A
  • relatively new tech
  • cells are viewed and stained w fluorescent dyes
  • a thick section of tissue or small living organisms are scanned w a laser beam (focused light)
  • the laser beam is reflected by the fluorescent dyes
  • multiple depths of tissue section/organisms are scanned to produce an image. As if the laser beam is building the image layer by layer
17
Q

Advantages of Laser scanning confocal microscopes

A
  • used on thick or 3D structures
  • external 3D structure is observed
  • very clear, high resolution bc the laser beam can be focused at a very specific depth. Cytoskeleton can be observed
18
Q

Disadvantages of Laser scanning confocal microscopes

A
  • slow process & takes a long time to obtain an image
  • laser could cause photodamage to the cells
19
Q

What is significant about tissues used in microscopy being transparent

A

Many tissues used in microscopy are transparent, letting light & electrons through them
- this makes it difficult to see details
- stains are often used to colour the tissues

20
Q

Staining for light microscopy

A

Coloured dyes are used when staining specimens
- the absorb specific colours of light whilst reflecting others, making structures visible
- certain tissues absorb certain dyes, depending on their chemical nature
- sometimes specimens are stained w more than one dye to ensure diff tissues who up (differential staining)
- chloroplasts do not need staining
- most colours are NOT natural

common stains: toluidine blue, methylene blue, phloroglucinal and eosin

21
Q

Staining for electron microscopy

A

When using TEMs, the specimen must be stained to absorb/scatter the electrons
- electrons have no colour so dyes used cause the tissues to show up black or shades of grey
- heavy metal compounds are commonly used as stains bc they provide a gd contrast by absorbing/scattering electrons. Eg, osmium tetroxide & ruthenium tetroxide
- any of the colour present in electron micrographs is NOT natural & it is NOT as a result of staining
- colours are often added to image using image processing software

22
Q

Converting units

A

mm
/1000
μm
/1000
mm
/10
cm
/100
m
/1000
km

23
Q

What is the magnification of the eyepiece lens

A

x10

24
Q

How to deal with scale bars (microscopes calc)

A
  1. measure actual length of scale bar w ruler (convert from cm to mm and then to μm)
  2. divide by length it represents
  3. this gives the magnification

A Level Biology (8 decks)

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