This proposal seeks to acquire a Bruker Opterra II swept-field confocal microscope with photoactivation module for live cell imaging for the Center for Cell and Molecular Imaging (CCMI) at Yale University School of Medicine. The ability to dynamically monitor cells in tissues, and organelles and proteins within cells has transformed our understanding of cell and developmental biology. Also, new fluorescent sensors enable the measurement of multiple physiological parameters including calcium levels and pH (as examples). Importantly, to fully take advantage of live cell imaging in order to probe cellular behaviors, we need microscopy that is highly sensitive to low light emissions from fluorophores, can acquire images quickly to capture rapid and dynamic behaviors, and has sufficient spatial resolution to detect cellular components. The combination of these three criteria poses a significant challenge for live cell imaging. Our preliminary results suggest that the unique imaging architecture of the Bruker Opterra II with its one dimensional array of pinholes/slits that creates a ?swept field confocal? performs optimally for our needs compared to other confocal technologies. Specific projects that would immediately take advantage of this proposal are diverse and numerous: 1) analysis of beating cilia during ciliogenesis and cilia regeneration in various organs and tissues 2) calcium fluxes in cardiac cilia 3) monitoring cytokinesis and proteins essential for the process 3) protein tracking in cells 4) axon regeneration 4) synaptic signaling and development in neurons 5) visualizing gene networks during the maternal to zygotic transition. By providing access to a live cell imaging microscope that optimally balances speed, sensitivity and resolution, the requested instrument will dramatic enhance the productivity and discovery in these NIH sponsored projects.
The requested microscope would permit visualization of cells and the components of cells while they are alive and dynamically so that these processes can be watched and understood. This in turn will help us to better understand cell function in normal and disease states. The machine would be located in a core microscopy facility that would allow investigators from throughout the medical school to have access to it.
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