Learn · Fundamentals of Cryo-Electron Microscopy (Cryo-EM) and 3D Image Processing in Structural Biology
Lecture 40 of 57Week 8 Negative Staining & its Applications with Ultramicrotomy in TEM
Ultramicrotomy and Sectioning for Biological Molecules
Prof. Somnath Dutta · IISc Bangalore
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- Fundamentals of Cryo-Electron Microscopy (Cryo-EM) and 3D Image Processing in Structural Biology, IISc Bangalore (at NPTEL)
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- CC BY-NC-SA, as NPTEL states it. Non-commercial use only.
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All the lectures
- Introduction of the microscope
- Image formation by glass lenses and its role in wavelength
- Different components of electron microscope (EM)
- Electron specimen interaction
- Elements of TEM: electron gun - I
- Elements of TEM: electron gun - II
- Comparison between the various types of electron gun
- Elements of TEM: electromagnetic lens (EM) - I
- Trajectory of electrons in EM lens
- Elements of TEM: electromagnetic lens (EM) - II
- Image Formation by EM Lenses: Understanding Ray Diagrams
- Aberrations and Focusing in Cryo-EM imaging
- Understanding Chromatic, Spherical Aberrations, and Astigmatism in Cryo-EM
- Aberrations and Practical Imaging in Cryo-EM
- Electron Dose Control and Beam Optimization in Cryo-EM
- Introduction to TEM Holders and Grids in Cryo-EM
- Overfocused and Underfocused Images
- Elements of TEM: Pumps in EM System
- Elements of TEM: Detectors in EM System
- Imaging and Effect of Thermal Drift
- Wave and its properties
- Addition/superposition of waves - I
- Addition/superposition of waves - II
- Decomposition of complex waves
- wave and their transformation
- Complex 2D Images & Its Diffraction Pattern
- Sample Preparation & Determination for TEM
- Biological Sample Purification - I
- Biological Sample Purification - II
- Different Sample Preparation Techniques for TEM Imaging & Different Types of Grids used for Imaging
- TEM Grids: Types, Handling & Surface Coatings
- Carbon Coaters, Glow Discharge & Grid Surface Treatment
- Negative Staining in TEM: Stains, Grid Preparation & Protocols
- Carbon Sandwich & Metal Shadowing Techniques in TEM
- Staining Quality in TEM: Thick, Thin & Positive Staining
- Negative Staining – Imaging Quality & Artifacts
- Negative Staining – Particle Distribution & Contrast
- Applications of Negative Staining in Structural Biology
- Negative Staining for Complexes and Interactions
- Ultramicrotomy and Sectioning for Biological Molecules
- Cryo-EM Grid Preparation & Vitrification
- Grid Types & Sample Distribution in Cryo-EM
- Blotting Parameters & Ice Conditions
- Motion Correction & Defocus in Cryo-EM
- Microscope Alignment & Imaging Limitations
- CTF Estimation & Defocus in Cryo-EM
- Phase Flip & Motion Correction
- High Pressure & Rapid Freezing Methods
- Cryo-Ultra Microtomy & FIB-SEM
- Graphene Oxide Grids in Cryo-EM
- Motion Correction & 2D Classification Basics
- Concepts & Methods in 2D/3D Classification
- 3D Reconstruction & Resolution Assessment
- Practical: TEM Setup & Negative Staining
- Practical: Cryo-EM Plunge Freezing
- Practical: Cryo-EM Grid Clipping
- Practical: Grid Loading & Imaging Software