2.1.1 Microscopy Flashcards

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1
Q

What are the different types of microscopy?

A
  • Light microscopy
  • Transmission Electron microscope
  • Scanning Electron Microscope
  • Laser Confocal microscope
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2
Q

What are the pros and cons of using a light microscope?

A

Pros : Easy to see large cellular structures, eg cell walls, chloroplasts, nuclei. Samples can be alive or dead, Images in colour

Cons : Low magnification and resolution, ultra-cellular structure cannot be seen

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3
Q

What is the radiation source and radiation used for a light microscope? How is this radiation focused?

A

The radiation source is light and radiation used is light. This radiation is focused using glass lenses.

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4
Q

How are cell structures seen with light microscope images?

A

By using staining dyes

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5
Q

What is the highest magnification and resolution in a light microscope

A

M : x1500
R : 200nm

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6
Q

How does a TEM work

A

U

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7
Q

How does a TEM work

A

A beam of electrons is fired through a thin section of a specimen. Denser areas of the cell absorb more electrons making them appear darker on the image

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8
Q

Max Magnification and resolution of TEM?

A

M : x500,000
R : 0.2 nm

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9
Q

Pros and cons of TEM?

A

Pros: highest magnification and resolution,

Cons: Always a 2D greyscale image, colour can be added digitally, specimen always needs to be dead due to the vacuum used

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10
Q

How does a SEM work

A

A beam of electrons is fired at the surface of specimen, knocks electrons off and these reflected electrons are collected t form an image.

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11
Q

Max Magnification and resolution of SEM?

A

M : x100,000

R : 10nm

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12
Q

Pros and cons of SEM

A

Pros: Can produce a 3d image of high magnification and resolution

Cons: always a greyscale image although colour can be added digitally, Specimen has to be dead since a vacuum is used

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13
Q

What is the radiation source, radiation used and method of focusing radiation for electron microscopes?

A

The radiation source is an electron gun, electrons are the radiation used and electromagnets are used to focus the radiation

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14
Q

Why do you need a vacuum for electron microscopes?

A

Scattering of electrons by air molecules interferes with image production

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15
Q

How does a Laser scanning confocal microscope work?

A

Cells are stained with fluorescent dye and a thick section of tissue is scanned with a laser beam,
Light is focused through a pinhole on a detector - the pinhole means any out of focus light is blocked so clearer images

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16
Q

Pros and Cons of a Laser Confocal Microscope?

A

Pros : Clearer images than light microscopes, laser bean can be focused at a specific depth eliminating the blur in optical scopes caused by out of focus lenses so clearer images, 3D images can be reconstructed as images taken can be fed into a computer
Cons:

17
Q

When were cells first discovered and by who?

A

1660s by Robert Hooke

18
Q

What is magnification?

A

The ability of a lens to enlarge the size of any optical image

19
Q

Magnification equation

A

Image / Actual size

20
Q

What is resolution

A

The ability to distinguish between two points as separate to see detail

21
Q

Why are stains used?

A

Stains are taken up better by different parts of the cell, the contrast is what makes the different parts show up better

22
Q

Preparing samples for staining - light microscope

A

Sample left to air dry, heat fixed, can then take up stains

23
Q

Preparing samples for staining - Electron microscopes

A

Samples dipped in solution of heavy metals, metal ions scatter creating contrast and some areas show up darker.

24
Q

Preparing a sample

A
  • fixation - prevent decay - formaldehyde added in a form of phosphate buffered solution
  • embedment - infiltrated with paraffin wax
  • sectioning - a single layer of cells cut 3-5micrometers with a microtome
  • staining
  • covered with coverslip
25
Q

What’s the differences between a wet and dry mount

A

Wet slides you add a drop of water and place the coverslip at an angle to avoid air bubbles unlike a dry mount which involves no water so the cover slide doesn’t need to be placed at an angle