Academic Courses

Advanced optical methods provide tools for observing objects with high spatial resolution, high spectral resolution and high sensitivity. These methods are based on pure optical effects as well as effects that involve photon-matter interaction.

The course will cover a significant number of the most important methods in optical imaging and microscopy that are used in the life sciences and biophysics. The course will start with a review of the fundamental physics required for the understanding of imaging and microscopy methods and will continue with a study of the more important methods.

The aim of the course is to provide on one hand a solid physical background to these optical microscopy methods, and on the other hand to understand the importance, capabilities and limitations of these methods.

The course will take place for one semester (two quarters), 2 hours a week (14 weeks). The course will start with review lectures and will follow with seminars on various topics prepared and presented by the students (complemented by a review on the subject). Along the course, there will be few literature study sessions. In each study, a paper will be studied and discussed. This will require preparation of the literature in advance.

Evaluation method: Home projects: 40%, Seminar presentation and preparation: 20%, Final exam: 40% (may be modified).

Topics:

1. Electric fields in matter [overview]
2. Maxwell’s equations in vacuum & matter
3. Diffraction theory of light
4. Point spread function & Optical Transfer function
5. Filtering methods (phase-contrast, Nomarski, etc.)
6. Optical detection systems, CCD’s, PMT
7. Introduction to fluorescence microscopy
8. Fluorescence Resonance Energy Transfer
9. Surface Plasmon Resonance imaging
10. Confocal Microscopy
11. 4pi microscopy
12. n-photon microscopy
13. Near-field microscopy
14. Fluorescence Correlation Spectroscopy
15. Advanced fluorescent molecules: quantum-dots
16. Fluorescent Proteins, molecular beacons
17. Spectral Imaging & Multicolor FISH
18. I5M microscopy
19. Saturation Depletion Microscopy

1. Introduction to Solid state
2. Diffraction
3. Electron Microscopy
1. Transmission electron microscopy: Introduction and principles
1. Electron sources and electron guns
2. Electromagnetic lenses
3. Resolution and limiting factors
4. Imaging and contrast formation mechanisms
5. The contrast transfer function
6. Sample preparation, Vacuum systems, CCD cameras
2. Scanning transmission electron microscopy (STEM)
3. Analysis (STEM/TEM) – DF/BF
4. Scanning electron microscope (SEM)
4. Advanced topics.

1. define advantages and disadvantages of transmission electron microscopy (TEM) and scanning electron microscopy (SEM) methods;
2. list known artifacts which appear due to unappropriate sample preparation;
3. choose appropriate SEM/TEM sample preparation technique;
4. summarise principles of SEM, TEM and scanning –transmission (STEM) set-ups: vacuum system, detectors, scanning coils, electro-magnetic lenses, apertures, stages/holders and electron guns;
5. define terms such as resolution of these microscopes, magnification, depth of field and depth of focus and relate influence of various factors on these terms;
6. conlude contrast mechanisms in studied electron microscopes;
7. describe special techniques such as HRTEM, HRSEM, cryo TEM and ESEM;
8. differentiate among different electorn diffraction methods;
9. index polycrystalline and single crystalline electron diffraction patterns;
10. explain EDS and WDS methods, choose appropriate one for solution of analytical problems in material science

Topics
Introduction to Transmission Electron Microscopy (TEM)
Lenses
Aberrations
Electron Guns

Scattering – Elastic and inelastic
Contrast transfer function
Resolution contrast and magnification

Image formation – contrast mechanisms
Electron crystallography (Ext.)
Tomography (Ext.)
Analytical methods (Ext.)
Cryogenic TEM techniques
CTF, real and reverse space

Image analysis
Microscopic imaging of colloidal particles and liquid crystalline phases.

Demonstrations

Room temperature TEM
Imaging – CCD basics
Cryo-TEM preparation
Cryo-TEM imaging

Main topics covered in the course:
• Bioimage informatics
• High content single cell phenotypic profiling
• Quantifying cell motility: from the intraceullular to the multicellular scales
• Classifying cell state with deep learning
• Generative models for cell structure with deep learning
• Enhancing cell image quality with deep learning
• Quantifying causality with fluctuations analysis (without perturbations)
• Molecular and cellular heterogeneity
• Public data repositories, data harmonization, integration and fusion
• Importing ideas from systems biology