Membranes are protective barriers surrounding all cells, providing structure and shape, and controlling what enters or leaves a cell. These membranes comprise lipids and proteins that work together to allow the cells of complex higher organisms to communicate (e.g. by transfer of vesicles - small membrane 'bags' carrying essential molecular cargo) and function (e.g. by protein-protein and protein-lipid interactions within and between membranes). There is much to learn about how these membrane components work together to support these essential functions. In physiology, in response to stress, and across the life course, membranes and their components undergo constant change. These seemingly small chemical modifications to proteins and lipids profoundly alter cellular shape and activity. However, these changes remain poorly understood and must be defined to enhance our understanding of fundamental physiology in health and disease. We are a vibrant, multidisciplinary team of Midlands leaders in membrane biosciences, based at Aston University, in the heart of the Midlands, within the new £10M Research England-funded Aston Institute for Membrane Excellence (AIME). This proposal seeks funds for the latest generation of mass spectrometer, the Orbitrap Ascend Tribrid ('Ascend') that, based on our preliminary data collected in collaboration with the manufacturer, will revolutionize our ability to detect proteins, lipids and their modifications in biological membranes with the utmost sensitivity, resolution and throughput. This investment will provide the first Ascend in England outside of the London-Cambridge-Oxford "golden triangle", with capability to support not only Aston's researchers, but also further partnerships across the Midlands region and beyond. We have carefully selected the Ascend platform to analyse membranes with the highest sensitivity and resolution, ensuring we can confidently identify the rarest (often previously undetectable) proteins and lipids, including their subtle but critical modifications, which are technically challenging to measure, yet fundamental to fully deciphering the importance, mechanisms and consequences of intercellular signalling. Ascend will enable information-driven discovery and translational research at a level not previously possible across four main research objectives: Defining membrane microenvironments (protein-lipid interactions) that control membrane protein function in health and disease (e.g. Aquaporins and ABC transporters in physiology, inflammation and cancer) offers transformation in advancing biomedical research, drug development, and therapeutic strategies to design selective drugs and innovative treatments, including for traumatic brain injury and neuroinflammation; Defining protein changes under stress and ageing (e.g. platelet receptor changes in wound healing and vascular disease) will help us understand key regulatory signalling processes which, when altered, drive disease; Defining functionally significant vesicle cargo mediating intercellular communication in inflammation and repair (e.g. vanishingly rare enzyme components of extracellular vesicles), enabling new therapeutic approaches for inflammatory conditions, such as non-resolving wounds; Membrane engineering to produce active proteins, improve stress tolerance of industrially-important microbes, and inform design of novel precision-controlled drug delivery systems for cancer. This investment will have significant impact on UK membrane biology, allowing us to understand the details of protein and lipid interactions and modifications that underpin essential cellular responses, in order to develop new therapeutics for diseases and age-associated conditions with unmet clinical needs.