Soft X-Ray Tomography: Filling the Gap Between Light and Electrons for Imaging Hydrated Biological Cells

Electron microscopy (EM) overcomes the resolution limitation inherent in light microscopy and can reveal the ultrastructure of cells and tissues. EM can be performed at high resolution and across significant volumes, using a range of systems, from transmission EM (TEM) to serial block face scanning EM (SBF SEM) and focused ion beam scanning EM (FIB SEM) [1]. However, chemically-fixed and resin-embedded samples suffer from a range of artifacts, particularly related to the dehydration process, such as shrinkage and extraction of molecules. To avoid dehydration, samples can be vitrified by plunge freezing and high pressure freezing, which preserves cells and tissues in their fully hydrated state. These vitrified samples can be imaged in TEMs and FIB SEMs equipped with cooled stages and using low dose imaging regimes. Whilst ultrastructure is improved, cryo-techniques come with their own set of technical challenges. Contamination-free transfer between cryo-preparation and cryo-imaging systems is non-trivial, as is imaging samples thicker than ~1 μm.