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Improved methods for spatial-temporal fluorescence imaging of cell activity Supervisors Cyril Ruckebusch (LASIR CNRS U. de Lille) Peter Dedecker (Department of Chemistry, KU Leuven)

Keywords Super-resolution, Fluorescence, Imaging, Living cell, Chemometrics, Data analysis, Image processing

Summary The PhD projects aim to develop a new methodology -from cell preparation to data analysis- to extract superresolution spatial-temporal information on cell activity in response to some external stress. Ultimately this work aims to provide new insights into how biological systems are structured at the nanoscale, and how this structures provides e.g. signaling specificity, using the latest methodologies in imaging and data analysis.

Context Fluorescence microscopy is about the only technique that can provide structural and dynamic information on selected components in live cells while only minimally interfering with the chemical or biological system under study. Previously limited to a spatial resolution of a few hundred nanometers due to diffraction, the recent development of super-resolution fluorescence imaging, which is one of the major technological advances of the last decade, has allowed direct access to the tens of nanometers length scale. To achieve such super-resolution in wide-field the key is the use of smart fluorophores that display fluorescence dynamics in combination with tailored data analysis and image processing techniques. However, simply visualizing the spatial distribution of molecules is not sufficient to understand biological issues occurring in living cells. In particular, improved super-resolution fluorescence imaging techniques are required to follow and investigate biological dynamics.

Research project One of the most important issues in biology is understanding signaling activities in living cells. How does the cell transfer specific signals through a complex interaction network in order to trigger an action in response to a stressing event ? To achieve this transduction, the cell must regulate its response by managing the spatial temporal organization of its constituents. This means that a given molecule not only has to be present in the cell but has to be there in the right place at a particular time. Advances in biosensor technology (chemically smart labels) have made possible to visualize and quantify the cellular activation dynamics in living cells. Information about when and where activity arises is assessed following how the fluorescence emission of the probe changes conditional on some aspects of the environment. One approach consists of mapping Forster Resonant Energy Transfer activity at an interesting spatial and temporal resolution. However, the construction of diffraction unlimited and high-sensitivity superresolution FRET activity maps requires acquiring and interpreted multicolor donor acceptor data, as well as developing specific data analysis methods. On top of that, several side issues complicating the development of robust models have to be solved such as handling stationary signals, dealing with strong photobleaching and low S/N to noise ratio or unraveling signal multiplexing when several biosensors are present in the same cell.

Application The candidate should be highly motivated, enthusiastic and opened to interdisciplinary science and international collaborations. The position is administratively based in Lille but work will be equally shared between U. de Lille and KU Leuven both offering an ideal environment and top-level equipments. The PhD applicant will join a 36 months bi-national (“cotutelle internationale”) PhD Program and will get diploma from both universities. Candidates should hold a master in physics, (bio-)chemistry or equivalent, and ideally have experience or have shown a strong interest in optical microscopy and data/image analysis. To apply, please send a CV and the name of 2 references to Cyril Ruckebusch (Cyril.ruckebusch chez univ-lille1.fr) and Peter Dedecker (peter.dedecker chez chem.kuleuven.be).