System upgrade for a Thermo Fisher GLACIOS 200Kv Cryo Electron Microscope

This application from the University of Manchester is to replace an obsolete camera on a BBSRC-funded Glacios cryo-electron microscope with a Falcon4i camera and Selectris energy filter. This will enhance our capabilities and transform usage with higher quality images and more efficient data collection and will make cryo-electron tomography data collection possible. Cryo-Electron Microscopy (Cryo-EM) provides us with details of tissue and cellular structure, as well as directly imaging proteins or multi-protein complexes (e.g. enzymes, biological motors and viruses) too small to be detected by light microscopes. It was the advent of the direct electron detector technology that provided Cryo-EM with the high sensitivity required to image proteins at atomic resolution. Newer detectors, such as the Falcon4i are even more sensitive, to enable imaging of proteins that were previously too small or flexible. Another enabling technology in Cryo-EM, is the development of imaging filters, such as the Selectris energy filter, which improves contrast in the images. At the University of Manchester, Cryo-EM supports a wide portfolio of BBSRC funded projects in addressing their central research questions. Moreover, to understand the function of proteins, or complexes in their native cellular or tissue environment many of these projects are now extending to cryo-electron tomography (Cryo-ET), where a new system is needed to image these thicker samples. Our current microscope, a Thermo Fisher 200 kV Glacios instrument was purchased in 2020 with BBSRC funding. The instrument has been very productive in terms of usage, publications and supported grant applications. We have also produced 20+ high resolution structures from Manchester research groups including fibres/fibrils, molecular machines, viruses and a range of individual proteins. The instrument was designed to be upgraded in a modular fashion, and we are now at a point where the original Falcon 3 camera is incompatible with the newest versions of the software used to operate the microscope and for tomography and thus the detector will soon be obsolete. The Falcon4i-Selectris would continue to enable high-profile research at Manchester using Cryo-EM including: Studying structural matrix proteins involved in inflammation and tissue integrity, (e.g. defective mucosal surfaces in the gut.) Investigation of how enzymes work and then manipulating them for biotechnology or bioremediation applications. Understanding how proteins are made and processed. Some proteins can ‘mis-fold’ and cause disease (e.g. in Alzheimer’s). Investigating how proteins in membranes can sense mechanical pressure and transport ions. Developing new technology based on virus-like particles for new vaccines, Mass Spectrometry for protein separation and Nanobody-based tools for Cryo-EM. The investment in a modernised instrument will amplify these projects and uplift the initial BBSRC investment from 2020. The University of Manchester recognises this by contributing 20% funding towards the upgrade purchase along with associated estates costs to refurbish the Facility with field cancellation hardware and a new control room. Additionally, the University has already funded a high-pressure freezer for preservation of cell and tissue samples, upgraded the University supercomputer with a dedicated GPU bank and provide an expert Research Technology Professional (RTP) to run projects through the instrument within a core facility platform.