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Structural Biology > Electron Microscopy > Flashcards

Electron Microscopy Flashcards

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1
Q

Compare light and electron microscopes

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2
Q

What is the Rayleigh criteria

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3
Q

What is the relationship between the resolution limit and wavelength

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4
Q

What is meant by resolution

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5
Q

What are the properties of electrons

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6
Q

What could happen when an electron encounters an atom

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7
Q

What different sources can be used in an electron microscope

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Thermionic emission source
Field emission gun

Electrons emitted by source

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8
Q

What is a thermionic source

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9
Q

What is a field emission gun

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10
Q

What does the electromagnetic lens do in an electron microscope

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11
Q

What three types of lenses are there in an electron microscope

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12
Q

What are apertures

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13
Q

How were images recorded in the past

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14
Q

What is a charged coupled device

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15
Q

What is a direct electron detector

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16
Q

Label the different parts of an electron microscope

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17
Q

Describe the pathway of the e- beam in EM to generate an image

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18
Q

What are the 2 types of contrast

A

Amplitude contrast
Phase contrast

Due to electron wave-particle duality

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19
Q

What is amplitude contrast

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20
Q

What are the 3 factors that contribute to amplitude contrast

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21
Q

What is phase contrast

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22
Q

How does a complex wave propagate

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23
Q

Why is the amplitude of the scattered complex waves small

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24
Q

What happens when the scattered and in scattered waves encounter the objective lens of an EM

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25
Q

What are the threes ways in which the small amplitude of the scattered wave can contribute to the unscattered wave

A

Maximally in the opposite direction of the unscattered wave
Minimally to the unscattered wave
Maximally in the same direction of the scattered wave

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26
Q

How can the small amplitude of the scattered wave can contribute to the unscattered wave maximally in the opposite direction

A

(Detector sees sum of scattered and unscattered wave)

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27
Q

How can the small amplitude of the scattered wave can contribute to the unscattered wave minimally to the scattered wave (edit)

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28
Q

How can the small amplitude of the scattered wave can contribute to the unscattered wave maximally in the same direction

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29
Q

How can defocusing the objective lens create contrast

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30
Q

What are spherical aberrations

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31
Q

What is the CTF

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32
Q

Why does the CTF need to be corrected

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33
Q

How can a phase plate be used for defocus imaging

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34
Q

What happens once images are obtained from EM

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35
Q

What happens after negative staining imaging to obtain a model

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36
Q

What happens in cryo-EM to obtain accurate models

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37
Q

What is defocus estimation

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38
Q

Why are particles selected in cryo EM

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39
Q

How can selected particles be classified using 2D classification

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40
Q

What 2D classification be used to do

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41
Q

What is projection theorem used for

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42
Q

How can orientation parameters be defined

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43
Q

What can projection matching be used for

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44
Q

What is meant by iterative refinement

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45
Q

What is 3D classification
What can classification be used for

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46
Q

What are the downfalls of EM

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47
Q

Why can noise in EM images result in inaccurate models

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48
Q

Why my inaccurate structures still be obtained even after improving the signal to noise ratio
What is model bias

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49
Q

What could over refinement of particles lead to

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50
Q

Give an overview of cryo-EM

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51
Q

What does EM allow for in imageing of a sample

A

Size
Range
Quantity
Environment
Complexity and heterogeneity

52
Q

What does it mean when EM allows for imaging of sample in terms of size

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53
Q

What does it mean when EM allows for imaging of sample in terms of range

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54
Q

What does it mean when EM allows for imaging of sample in terms of quantity

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55
Q

What does it mean when EM allows for imaging of sample in terms of environment

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56
Q

What does it mean when EM allows for imaging of sample in terms of complexity and heterogeneity

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57
Q

What is the goal of sample preparation

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58
Q

What are the two sample prep techniques for the 2 types of EM

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59
Q

What is the characteristics of the EM grid

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60
Q

Explain the negative staining drop to drop method

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61
Q

How is the sample loaded into the electron microscope in negative staining

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62
Q

What are the advantages of negative staining

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63
Q

What are the disadvantages of negative staining

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64
Q

What is the first step of cryo-EM

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65
Q

What is the second step of cryo-EM

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66
Q

What are the different types of ice

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67
Q

What is the major challenge for cryo-EM

A

Driving force for this difficulty is the air-water interface (where surface of suspended liquid meets air )

68
Q

What could happen at the air-water interface

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69
Q

How can the effect of the air-water interface be reduced

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70
Q

How is the sample loaded in cryo-EM + what factors need to be controlled

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71
Q

Why is low dose imaging used to image the sample in cryo-EM

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72
Q

What three modes does low-dose imaging cycle through in cryo-EM

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73
Q

How do you calculate the amount of in elastic scattering through an electron microscope

A

10–20% of electrons undergo inelastic scattering, depending on the material and conditions.
So do percentage of total e- suppled to calculate amount of e- inelastically scattered

74
Q

What factors can influence the orientation of particles in ice for cryo EM

A

The speed at which the sample is frozen
pi of the protein

Structural Biology (8 decks)

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