About the course
Electron microscopy in combination with image analysis is increasingly powerful in producing 3D structures of individual molecules and large macromolecular complexes that are unapproachable by other methods. This course is focused on the concepts and theories behind electron microscopy and will be taught in a reverse classroom format with pre-recorded lectures. Students will be responsible for watching these online lectures prior to class, which will be live roundtable discussions led by guest lecturers and SEMC staff on the practice of solving molecular structures by electron microscopy.
Class structure (1.5 hr class)
The course will be offered by the New York Structural Biology Center but given the COVID-19 health situation will be held virtually. Classes will be held by Zoom (Mondays 3:30-5pm and select Wednesdays 3:30-5pm). Times may be subject to change.
- Pre-recorded lectures and other online content from previous years will be used.
- Weekly roundtable discussions to supplement the online content. Open-ended discussions on the topic will be lead by course instructors after a brief introduction to the topic. Conversation starters may come from concept check questions from Grant Jensen’s online course (Getting Started in Cryo-EM from Caltech) or presentation on newer developments in the field.
- The weekly meetings will be supplemented by Journal Clubs and remote practicals.
EM fundamentals section (month of January)
Lecture2 : Jan 13 – Challenges in biological EM & Sample Prep and practical
Lecturer: SEMC staff
[Video] [PDF of slides]
[Roundtable 2 & T12 room practical: January 13, 2021] [F20 room practical: January 13, 2021]
Lecture3 : Jan 20 – Support films and practical
Lecturer: SEMC staff
[Video] [PDF of slides]
[Roundtable 3 & Plungefreezer practical: January 20, 2021]
EM crystallography section
Single particle section
EM challenges and new frontiers
1 week in April covering:
- Intro and overview of ET
- CryoCLEM-FIB-ET workflow
- Cryo-Applications and sub-tomogram averaging
- Choosing the right in situ target and model system
- Automated reconstruction, subtomogram averaging methods & data interpretation
- EMAN2 tools, sub-tomogram averaging and annotation
- The future of in situ cellular structural biology
The lecture slides are provided for educational purposes only. All rights are reserved by their original authors. Please contact them if you would like to use or repost their content.
Practicals/Journal club will be regularly scheduled over the semester to supplement the instruction. Typically on Wednesday @3:30pm.
- Practical 1:
Jan 13 [TEM microscope worksheet][defocus series supplement]
- Practical 2:
Jan 20 [Sample preparation worksheet]
- Practical 3:
Feb 3 [Pre-processing worksheet][Pre-processing tutorial][Pre-processing practical: February 3, 2021]
- Feb 10 – A comparison of original and modern plunge freezing techniques
Adrian et al. 1984 & Razinknov et al. 2016
These papers are focused on sample preparation methodology.
- Feb 17 – The best voltage for biological cryo-EM
Peet et al. 2019 & Naydenova et al. 2019
These papers deal with the microscopes and cover details of optics and information loss in EM.
- Feb 24 – The beginning of the resolution revolution
Liao et al. 2013
Seminal work on TRPV1 by Yifan Cheng’s lab. Widely considered to be the “beginning” of routine high resolution cryo-EM. The discussion should focus on the technological and scientific advancements that made this work possible. There are far too many to explicitly assign, so it is left more open-ended.
- Mar 10 – HIV trimer controversy and Einstein from noise
Mao et al. 2013 & Henderson 2013 & van Heel 2013
The first paper is a cryo-EM structure of the HIV trimer, and the second two are criticisms of that paper’s EM data processing. It is a significant case study in what can go wrong in EM.
- Mar 24 – Challenges in and recommendations for validating cryo-EM data
Henderson et al. 2012 & Neumann et al. 2018
The first paper is the set of recommendations from the first Electron Microscopy Task Force Meeting, and the second is a more recent approach to validation in light of the resolution revolution.
For researchers taking the course for credit, the final score will be tabulated as follows:
- Recitation 40%
- Practical Worksheet 10% x 4
- Attendance 20%
- Course coordinator: Ed Eng (NYSBC)
- Teaching assistant: Micah Rapp (Columbia/NYSBC)