CONTEXT: At the forefront of biological research is understanding how cells interact with their environment. Selective interaction with the extracellular milieu and the recognition of molecular signatures presented on a cell surface enables organisms to respond to environmental changes, which is often altered in disease states. Many cell targeting molecules are advancing our understanding of how these interactions influence different cell phenotypes. However, distinct challenges in the development of these targeting molecules (e.g., small molecules, antibodies, aptamers) necessitates the need for innovative bioanalytical infrastructure to dissect and characterise their binding profile in live cells. AIMS & OBJECTIVES: The purpose of this equipment grant is to establish the UK's first Real-Time Interaction Cytometry (RT-IC) facility, which will enable users to measure the binding properties of molecules (e.g., kinetic rates; affinity, avidity) directly on live cells. The Glas-cyto facility will be part of Glasgow's wider biophysical centre of excellence formed between the Universities of Strathclyde (UoS) and Glasgow (UoG), enabling the UK userbase integrated access to a breadth of bioanalytical and biophysical facilities to examine cellular binding interactions. The heliXcyto equipment will be suitable for a wide range of analyses for eukaryotic cell types and will be underpinned by support from dedicated research technical professionals (RTPs) within the Strathclyde Centre for Molecular Bioscience (UoS) and the Integrated Protein Analysis Facility (UoG). THE RESEARCH THAT THE EQUIPMENT WILL ENABLE: Our dedicated facility will enable users the unique opportunity to culture live cells alongside instrumentation that will quantify binding interactions in less than 30 min. Access to such a facility will enable our user base to develop better cell-targeting biologics (e.g., antibodies, aptamers, Theme 1), deliver a new design proxy for the development of cell targeting molecules for the development of diagnostic platforms (Theme 2), and further our understanding of protein trafficking and cell-selective recognition of G-protein coupled receptors by small molecules (Theme 3). POTENTIAL APPLICATIONS & BENEFITS: The current state-of-the-art in the biophysical analysis of extracellular interactions have predominantly focused on low throughput assays (e.g., CETSA, In Cell Pulse DiscoverX, SPR), which require the downstream isolation of analytes, or qualitative analysis of interactions by flow cytometry. The major drawback of these techniques is that binding is inferred rather than quantified. Detailed quantitative knowledge of spatial patterns of receptor expression, and reconciling these data with binding interactions can accelerate the translational potential of novel targeting molecules in early discovery. Establishing this dedicated facility within Glasgow will provide, for the first time, the ability to quantify binding interactions on live cells, under conditions more closely mimicking their native environment. The Glas-cyto facility will enable new training opportunities across the breadth of the UK's biological user base, and infrastructure to enhance interactions with industrial partners.

Viral infections pose one of the biggest global threats to human populations and agriculture. Successful prevention, monitoring and treatment of viral infections requires the availability of fast and reliable diagnostic methods which can not only sensitively, but rapidly detect a viral infection of interest and differentiate between viral infections. This is particularly important in the winter months where rapid diagnosis of viral infections emerging from SARS-Cov-2 relative to influenza strains is essential in order to assist medical practitioners to suggest the most appropriate interventions and treatment. At present, methods do not exist which can rapidly detect viral infections in a low-cost, point-of-care device. We propose to develop a biosensing technology which can not only detect viral components, but also has the potential for the platform to be reusable and regeneratable. Central to these developments is the use of fluorous technology as a tool to immobilise elements which detect viral components. Much akin to Teflon, fluorous technology has the dual advantage as a method which can immobilise molecular components which have a complementary fluorous tag, and reduces non-specific binding to non-fluorous biomolecules, thus improving the sensitivity of the approach. Furthermore, the fluorous-directed immobilisation event is inherently reversible by a simple washing step with organic solvent. In this proposal, we will demonstrate the modularity of the strategy to detect viral RNA (by RT-PCR) or protein (by direct detection of intact viral particles). This will provide a powerful new tool for the biosciences which has the potential to be used for any application which requires rapid detection of pathogenic infections.