High-throughput digital microplate microscopy reader for the study of cellular responses to infection and stress

Viruses require living cells to multiply. When doing so, viruses cause a complete re-organisation of the infected cell, overall allowing the production of thousands of new viral particles but also suppressing the ability of the host to fight infection. Crucial insights into this complex interplay can be obtained imaging living cells and viruses, which is only possible using microscopes. Most microscopes are, however, able to image a few cells at a time making it very lengthy and laborious to obtain information from a representative number of cells. In addition, cells undergo their own life cycle and variations exist between them even in the same petri dish. A way to overcome this problem is the use of high-throughput microscopes able to image thousands of cells and analyse their properties on an individual basis. This proposal concerns the acquisition of one of such imaging systems for the Section of Virology at the University of Surrey, a unit of research formed by >20 investigators from 6 different groups studying human viruses and their interplay with human cells. The instrument we propose to acquire will enable us to track where viruses go inside a cell and how they manipulate the intracellular environment. We will be able to visualise viral features such as gene expression and replication factories, as well as the cell's response to infection in the form of stress granules, mitochondrial reorganisation and cell and nucleus morphology. More importantly, we will be able to measure these events qualitatively and quantitatively in hundreds of individual cells in specified conditions, generating high quality reproducible data in a short period of time. As examples, we have recently discovered a viral protein expressed in poxviruses (including the emerging monkeypox virus) that induces the clustering of mitochondria (the energy factories of the cell). The kinetics of assembly and disassembly as well as the size and morphology of these clusters in different conditions remains unknown but can be elucidated with a high-throughput multimode microscopy unit. Similarly, we have recently discovered that cells infected with herpes simplex virus fail to export their mRNAs from the nucleus, causing a blockade of cellular functions that allows efficient virus infection. The mechanisms behind this process are unclear but can be researched with an imaging system equipped with a spot counting module. Altogether the requested instrument will not only increase our capacity to deliver ground-breaking research, but also provide novel perspectives on viral infection that are at present not possible with other equipment.

This proposal concerns the acquisition of an Agilent Biotek Cytation 5 Cell Imaging Multimode Reader for the Section of Virology at the University of Surrey, a unit of research formed by >20 investigators from 6 different groups studying human viruses and their interplay with human cells. The instrument will complement our existing imaging facilities, which include a Nikon A1 inverted confocal microscope, a Zeiss LSM 980 confocal microscope and an ONI Nanoimager for super-resolution microscopy. The Cytation 5 combines automated digital microscopy with multi-mode detection using monochromators and filter-based fluorescence optical systems for 4 independent channels. It is compatible with 6- to 1536-well microplates, microscope slides, cell culture dishes and T25 cell culture flasks; offers both atmosphere and temperature controls; and detects absorbance and luminescence. The modular nature of the software makes it the most versatile on the market, and it is also equipped with high-accuracy injectors. The sophisticated image analysis software performs stitching and Z-projection, as well as quantitative and qualitative analysis and spot counting. The proposal is spearheaded by the Elliott Lab and Maluquer Lab, which will use the instrument to provide novel insights into the biology of alpha-herpesviruses and orthopoxviruses, respectively. In addition, the instrument will be used by 4 more research groups studying noroviruses, flaviviruses and respiratory viruses such as SARS-CoV-2, RSV and Influenza virus. The equipment will therefore be instrumental in our ability to discover novel virus-host interactions and the mechanisms that govern them in different cells. This knowledge will eventually provide novel therapeutic avenues for a collection of established and emergent viruses with enormous burden on human health worldwide.