Over the last decade, cryo-electron microscopy (cryo-EM) has stepped up as the mainstream technology for studying the structure of cells, viruses and protein complexes at molecular resolution. This “resolution revolution” was made possible by recent advances in microscope design and imaging hardware, paired with enhanced image processing and automation capabilities. Cryo-EM involves flash-freezing of a thin layer of sample on a support, which is then visualised in its frozen hydrated state by transmission electron microscopy. This can be achieved with very low quantity of protein and in the buffer of choice, without the use of any stain, which is very useful to determine structure-function correlations of macromolecules. Subsequently, data from a large number of projection images, featuring identical copies of a protein complex in different orientations, are combined to generate a 3D reconstruction of the structure.

In this talk, I will introduce Cryo-EM with a particular focus on image formation, data acquisition and analysis.  I will also introduce the newly established collaborative research centre for Cryo-Electron Microscopy based in Milan (C-EMi) at Dept. of BioSciences. The facility, first of its kind in Italy, owns a FEI Talos Arctica 200 keV Transmission and Scanning Electron Microscope (STEM) equipped with FEI Falcon 3 EC, a state-of-the art direct electron detector. C-EMi is set up as a powerful, versatile laboratory for high-resolution structural characterization of biological samples.