The iBiOs platform for advanced live-cell imaging, ultrastructural analyses and correlative light and electron microscopy in SPP 2225

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The proposed research project aims to develop advanced correlative light and electron microscopy (CLEM) imaging techniques for the analysis of host-pathogen interactions. Current limitations of conventional imaging methods include inadequate spatio-temporal resolution, difficulties in interpreting structurally complex host-pathogen interactions, and lack of 3D information needed to understand microbial pathogen exit strategies. Recent technological advances, such as high pressure freezing/freeze substitution (HPF-FS) and super-resolution (SR) CLEM, have enabled high-resolution visualisation of host-pathogen interactions. The iBiOs platform provides a range of advanced fluorescence and electron microscopy systems, i.e. combining lattice light-sheet microscopy (LLSM) imaging with 2D/3D electron microscopy, on-section CLEM and near-infrared branding (NIRB) to generate landmarks in polarised confluent cells and tissues. The facility is unique in its ability to combine live cell imaging with subsequent cryo-fixation for EM of pathogenic samples under BL2/S2 conditions and will be used to develop and optimise imaging workflows to analyse pathogen exit strategies within the SPP 2225 consortium. A large number of project leaders studying pathogens within SPP 2225 have already expressed interest in the iBiOs imaging platform. The resulting data will be useful for comparative analyses of pathogen exit strategies, addressing the need for tailored methods and innovative workflows to analyse pathogen escape from host compartments or host cells.