Electron Microscopy Flashcards
Compare light and electron microscopes
What is the Rayleigh criteria
What is the relationship between the resolution limit and wavelength
What is meant by resolution
What are the properties of electrons
What could happen when an electron encounters an atom
What different sources can be used in an electron microscope
Thermionic emission source
Field emission gun
Electrons emitted by source
What is a thermionic source
What is a field emission gun
What does the electromagnetic lens do in an electron microscope
What three types of lenses are there in an electron microscope
What are apertures
How were images recorded in the past
What is a charged coupled device
What is a direct electron detector
Label the different parts of an electron microscope
Describe the pathway of the e- beam in EM to generate an image
What are the 2 types of contrast
Amplitude contrast
Phase contrast
Due to electron wave-particle duality
What is amplitude contrast
What are the 3 factors that contribute to amplitude contrast
What is phase contrast
How does a complex wave propagate
Why is the amplitude of the scattered complex waves small
What happens when the scattered and in scattered waves encounter the objective lens of an EM
What are the threes ways in which the small amplitude of the scattered wave can contribute to the unscattered wave
Maximally in the opposite direction of the unscattered wave
Minimally to the unscattered wave
Maximally in the same direction of the scattered wave
How can the small amplitude of the scattered wave can contribute to the unscattered wave maximally in the opposite direction
(Detector sees sum of scattered and unscattered wave)
How can the small amplitude of the scattered wave can contribute to the unscattered wave minimally to the scattered wave (edit)
How can the small amplitude of the scattered wave can contribute to the unscattered wave maximally in the same direction
How can defocusing the objective lens create contrast
What are spherical aberrations
What is the CTF
Why does the CTF need to be corrected
How can a phase plate be used for defocus imaging
What happens once images are obtained from EM
What happens after negative staining imaging to obtain a model
What happens in cryo-EM to obtain accurate models
What is defocus estimation
Why are particles selected in cryo EM
How can selected particles be classified using 2D classification
What 2D classification be used to do
What is projection theorem used for
How can orientation parameters be defined
What can projection matching be used for
What is meant by iterative refinement
What is 3D classification
What can classification be used for
What are the downfalls of EM
Why can noise in EM images result in inaccurate models
Why my inaccurate structures still be obtained even after improving the signal to noise ratio
What is model bias
What could over refinement of particles lead to
Give an overview of cryo-EM
What does EM allow for in imageing of a sample
Size
Range
Quantity
Environment
Complexity and heterogeneity
What does it mean when EM allows for imaging of sample in terms of size
What does it mean when EM allows for imaging of sample in terms of range
What does it mean when EM allows for imaging of sample in terms of quantity
What does it mean when EM allows for imaging of sample in terms of environment
What does it mean when EM allows for imaging of sample in terms of complexity and heterogeneity
What is the goal of sample preparation
What are the two sample prep techniques for the 2 types of EM
What is the characteristics of the EM grid
Explain the negative staining drop to drop method
How is the sample loaded into the electron microscope in negative staining
What are the advantages of negative staining
What are the disadvantages of negative staining
What is the first step of cryo-EM
What is the second step of cryo-EM
What are the different types of ice
What is the major challenge for cryo-EM
Driving force for this difficulty is the air-water interface (where surface of suspended liquid meets air )
What could happen at the air-water interface
How can the effect of the air-water interface be reduced
How is the sample loaded in cryo-EM + what factors need to be controlled
Why is low dose imaging used to image the sample in cryo-EM
What three modes does low-dose imaging cycle through in cryo-EM
How do you calculate the amount of in elastic scattering through an electron microscope
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
What factors can influence the orientation of particles in ice for cryo EM
The speed at which the sample is frozen
pi of the protein
