Raman probes to determine cellular redox potential

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Understanding the exquisite complexity of a cell, the fundamental unit of all known organisms, provides unrivalled opportunity for innovation for research. In order to investigate cellular function, experimental biology has typically restricted analysis to cellular 'snapshots'. This ignores both the time-dependent changes occurring in each cell and its organelles, and also the dynamic flux of its macromolecules through biochemical pathways. This 'snapshot' approach to biology is unlikely to provide a truly deep understanding of cell biology as the cell involves an integrated and temporal network. As a result, existing reductionist approaches may exacerbate stochastic variability within modern experimental biology that can lead to major 'failure-to-replicate' problems. The generation of a holistic understanding of cellular biology by establishing advanced tools and technologies in order to understand how a cell's molecular circuitry responds and adapts to environmental change in a spatiotemporal manner therefore provides significant opportunity.Redox potential within a cell is essential to cellular health and is intimately involved in many important processes including apoptosis, differentiation, cell cycle and inflammation. The progression of many diseases such as cancer, cardiovascular disease and neurodegenerative disorders have been implicated in aberrant cellular redox potential. Therefore, tools and techniques with which to monitor and determine redox potential in both normal cell function and that associated with disease is critical in the validation of new targets for therapeutic intervention and the development of medicines of the future.Within this project a suite of alkyne probes to determine redox potential, glutathione levels and the concentration of reactive oxygen species will be designed and synthesised. These will be targeted to specific organelles using standard techniques and calibrated within a cellular context using Raman microscopy. This analytical technique offers additional rich, label-free information on the distribution of endogenous biomolecules such as lipids, nucleic acids, and proteins within different cellular compartments (e.g., mitochondria, lysosome, nucleus). Monitoring spatiotemporal changes in response to external signals and stresses within different types of cell will provide spatiotemporal insight into the internal localised environment within organelles and provide effective tools for a cellular analytical approach to understanding and monitoring the cellular redox environment.Successful outcome of this project will deliver a new suite of molecules able to quantitatively measure the redox environment within a cell and inform of the global cellular effect on the perturbation of cellular pathways to external stimuli and disease which will find broad application to those working within the life sciences and chemical biology communities.