Advanced Optical Imaging & Single-Molecule Biophysics

In this intensive course, students will learn and apply the working principles and concepts behind various advanced imaging and single-molecule techniques that have revolutionized the way we can monitor inter- and intra-molecular interactions in the life-, materials-, and food sciences. In the first part of the course, the theoretical and experimental background will be taught and practiced during lectures and tutorials. The second part of the course focuses on practical work and applications, offering the student hands-on experience with single-molecule experiments: The students will (1) build and design optical microscopes with the open UC2 microscopy platform, (2) track fluorescently labelled proteins (CRISPR-Cas) in live bacteria, (3) read and distinguish the four DNA bases label-free using nanopore technology, and (4) use FRET as a molecular ruler to accurately determine nanometer distances within single DNA strands. Regarding experimental data analysis, the principles of image processing, interpretation of multi-dimensional datasets, and time trace analysis including machine learning approaches will be treated. All of this prepares the students for the third part of the course, where they will study, discuss, and present some of the latest literature of this dynamic research field. The course is concluded with a written exam.

After successful completion of this course students are expected to be able to:

• Understand and apply state-of-the-art methodologies in (fluorescence) microscopy and single-molecule biophysics
• Demonstrate critical thinking and problem-solving capabilities that are required for research in the academic as well as industrial environment (R&D)
• Be capable of understanding, explaining, and applying the principles of advanced imaging and single-molecule biophysics methods, including (1) optical phenomena and advanced microscopy (UC2), (2) nanopores for label free detection of biomolecules (nanopores), (3) single-molecule Foerster resonance energy transfer (smFRET), and (4) single-molecule localisation microscopy and single-particle tracking (SMLM)
• Understand the limitations and strengths of these techniques (achievable spatial and temporal resolution, contrast, information content and possible applications)
• Understand and apply suitable image and data processing techniques
• Interpret and understand experimental results in terms of nanoscale information with a space and time component

Assumed Knowledge:
BIP24306Spectroscopy and Imaging;
BIP32803 Biophysical Imaging.
Analytical chemistry or similar courses that provide a basic understanding of molecular life sciences / biophysical chemistry topics