W10 Applications of Electron Microscopy

Question 1

what is CTF estimation

Answer 1

the process of fixing the imperfections in EM that are affecting the image, via 2 main steps:

phase flipping: flipping of “phase” of image to correct errors caused by defocus or spherical aberration

amplitude correction: fixes parts of image where contrast may be too strong or weak > balance brightness and darkness in image to make it look more accurate

Question 2

steps in cryo-EM processing

Answer 2

motion correction: to fix blurry or distorted images caused by movement of sample

CTF correction: to fix distortions caused by microscope being out of focus of imperfections of lens

particle picking: selecting individual particles in image to find relevant ones to analyse

2D classification: selected particles grouped into categories based on their similar shapes or patterns

initial 3D model: create rough 3D model using different 2D views

refined 3D model: refined using advanced algorithms

Question 3

what is CAR-T cell therapy

Answer 3

type of immunotherapy used to treat cancer

involves modifying a patient’s own T cells to better recognise and attack cancer cells

Question 4

key contributions of cryo-EM

Answer 4

contributes to understanding the function of transmembrane protein and large dynamics complex

molecular details provided by structure are the foundation for drug discovery and optimisation

Question 5

limits of cryo-EM

Answer 5

expensive

throughput: data acquisition takes 1-2 days or even longer

molecule with lower binding affinity may not be visible