Cryo-Electron Microscopy

What it Is? Principle? What are Its Pros and cons?

Q: What is Cry o- El ectron Microscopy?

.

Ans: Essentially, Crye-electron microscopy (Crye-EM) is a type of transmission

electron microscopy that allows specimen of interest to be viewed at cryogenic
temperatures. They are very valuable tool for viewing and studying t he
structures of various biological molecules.

Q: What is Principle of electron microscopy?

Ans: For this we need to first understand the Significance of Electrons (Electrons
vs. Photons):
Photons are packets of energy/basic particles of light, hence, everything that ~e
sees with our eyes is due to the fact that these particles reflect those physical

objects.
th
· However, some of the objects/specimen are too small compared to e
wavelength of photons, they are unable to interact making it impossible to view

them.
When it comes to the wavelength of electrons, it is small enough to interact with
the objects making it possible to observe them. Therefore, electrons become
more suitable for the purposes of observing the small components of cells as
well as a variety of molecular structures.

e.g. TEM and SEM

Q: How the Cryo-EM Works?

Ans:
1) Rather than using the limited wavelength of light, electrons with much lower
wavelengths are used as the source of light.

2) In Crye-Electron Microscopy the sample under observation is usuaHy frozen

(frozen-hydrated) for preservation purposes.
 tis very important that the process of freezing the sampl~ is very quick. This
vents the frozen water around the specimen sample from forming cubic ice.
~he event that ice is formed, it readily absorbs the electron beam, which in
rn .obscures the sample. For this reason, it is essential that the sample be
unged in to the cooling liquid rapidly so that the water only freezes around the
pecimen for clear images.
) * While liquid nitrogen can also be used for the freezing process, ethane is
sed instead given that it has higher heat capacity and is also liquid at
mperatures slightly above th.at of liquid nitrogen. As such, it is sufficiently cold
o freeze water rapidly and appropriately without boiling _off.
) To observe macromolecules in their native hydrated state, the sample is
·rnply embedded in vitrified water (formation of an aqueous solid without
crystalline ice through ultra-rapid cooling at temperatures below - 130 °C.,)
·frozen hydrated as directly visualized on the microscope. Here, no stains are
. used given that the surrounding buffer allows for enough contrast to observe
the specimen.
6) Importantly, different samples have different protocols. For this reason, it is
important to confirm the right procedure for specific specimen.
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in Uquid ethane and stored in iquid nitrogen. freezing must be

aystals. Ice lattices will not only absorb the electron beam
sample's structures. If !he sample is lroz,n quiddy enough,
does not aystamse. ec--c,;.'.,

· such as viruses) suspended in random orientations in

matter with a beam of electrons. Newly developed •a~ect
er digital camera•style imaging.

different views of the molerule into a 3D model. The

s or hundreds of thousands of particle images.
o help traas the movement of particles and so reduce

that the molea,le of Interest does not have to

simply can not be ~tamsed; others have tJ,eir

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In theirri'ative environment, and there

can 1Ufflr as a n,sult of electron i,racf~tlon.

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droph'l t1 Iravel and s pread (blua atrDW!i) onln the EM grid at. Iha grtd Is moved Inwa rd the ayogtWl (hlaclt a rrow). c, Scribing-based deposltlon hwofw5 a
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