Integrated multi-dimensional molecular organ imaging

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The development of innovative optical imaging technologies has led to a revolution in light microscopy heralding the development of a generation of microscopes that promise powerful new applications for biomedical research. We believe our plans for refinement of commercially available instrumentation, development of novel probes and comprehensive testing in complex biological applications will ensure our advanced light microscopy 'toolkit' can be exploited to deliver innovative research and improved health outcomes.To maximise our contribution to the innovation and application of advanced imaging we focus on intra-vital imaging as this platform allows detailed investigation of biological events at sub-cellular resolution in living specimens at the greatest possible penetration depth. We will deploy a state-of-the-art intra-vital light microscope to acquire images of specifically labelled or label-free molecules up to one millimetre into tissue(s) of live animals typically mouse or zebrafish. Our innovative approach includes complementing these images with 3D mapping in order to provide a much more complete view of organ function in health and disease.Effective, informative, methods for labelling cells prior to intra-vital imaging remains a significant challenge in the wider application of these methods to non-transgenic models. In order to increase the reach of our intra-vital imaging methods we will employ and evaluate novel fluorescent 'Smartprobes'. These Smartprobes are already being used in Edinburgh to image key biological and pathological events in human lungs through cutting-edge fibre-optic microscopy, thus providing a direct translational pathway from planned studies in zebrafish and mice to human diseased organs in life (the technology is readily applicable to the digestive tract and the reproductive system). Our group has already established a strong track record in the use of a wide range of model animals that express fluorescently labelled molecules of interest in key target cells/organs. This expertise will guarantee we develop robust methodology focussed on scientifically challenging questions, such as: What is the involvement of macrophages in the development of breast cancer? How do neurons and glial cells interact in the development, function and repair of the nervous system? How do sex steroid hormones alter cell behaviour in the uterus and how can this be modulated? What is the distribution of central neuronal proteins in the brains of normal compared to model mice suffering from brain disorders?Excitement surrounding the application of Intra-vital imaging to biomedical research is based on its capacity to track cells and their contents within live tissues offering a novel platform for the study of dynamic changes in protein distribution, molecular interactions and tissue composition. Currently, such observations are only possible at single locations within a tissue - our goal is to expand data-acquisition to whole organs, e.g. to interrogate the distribution of synaptic proteins in a mouse brain. To achieve this, we will integrate our intra-vital techniques with systematic 3D image acquisition on a fast confocal microscope, capable of scanning whole sectioned organs. The resulting series of 3D-images will be archived to ensure data preservation and will be made available to the wider research community via searchable web-based organ libraries. These molecular 'organ maps' detailing the location of specific labelled proteins will allow us to refine subsequent 'focussed' functional intra-vital imaging and provide high-content 3D organ maps with functional measurements from live specimens.These innovative studies will provide data for our translational research pipeline and inform development of improved diagnostics and therapies for health problems including cancer, multiple sclerosis, infertility, cardiovascular and neurodegenerative diseases.