Topics and Objectives

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During this three-day course, the participants will be taught the theoretical principles, potential applications, advantages and disadvantages of different advanced imaging methods. Lectures will be provided by experts from the participating institutes, product specialists of microscope manufacturers and international speakers. In the afternoon, each technique will be illustrated during on-site and remote demonstration sessions. These demonstration sessions naturally complement the theoretical lectures and are intended to provide insight into the theoretical principles (participants will be encouraged to discuss their own data or research projects). Together they form a solid basis for further individual hands-on training at - or collaboration with the organizing core facilities. 

The first day covers a set of microscopy methods to measure molecular dynamics, i.e. the mobility, stability, interactions and conformational changes of molecules or particles. An introduction to Fluorescence Recovery After Photobleaching (FRAP), Single Particle Tracking (SPT), single molecule Fluorescence Resonance Energy Transfer (smFRET), Fluorescence Correlation Spectroscopy (FCS) and Raster Imaging Correlation Spectroscopy (RICS) will be given followed by demonstration sessions of each technique/experimental set-up. 

During the second day we will focus on mesoscale imaging and non-linear optical (NLO) imaging. Mesoscale imaging is a general term referring to various imaging methods that allow the visualization of larger (up to cm) samples while maintaining subcellular resolution. Two specific techniques will be demonstrated: light sheet microscopy and optical projection tomography. Non-linear imaging involves the visualization of (label-free) contrast mechanisms in which at least 2 photons are recombined into a single photon. Due to inherent optical sectioning, greater optical range and the less invasive nature of red-shifted photons employed in NLO imaging this enables live, three dimensional imaging of larger volumes over extended periods of time.

On the third day, we will zoom in on super-resolution microscopy. Super-resolution microscopy refers to a collection of techniques that are able to circumvent the fundamental resolution limit imposed by diffraction through various means. The techniques covered include: Airyscan, SIM (SIM-TIRF), SML and STED. Lectures will explain the theory behind the different principles to break the diffraction barrier employed in each technique. Practical sessions will illustrate the possibilities, advantages and disadvantages of enhanced resolution imaging.