Underpinning Instruments: Australia’s first cryogenic focused Ion beam-scanning electron microscope

The Cryo-FIB/SEM at the Monash Ramaciotti Electron Microscopy Centre

The Cryo-FIB/SEM at the Monash Ramaciotti Electron Microscopy Centre

Original source: imagingcoe.org/wp-content/ uploads/2018/04/2017_Annual-Report_ARC-CoE-in- Advanced-Molecular-Imaging.pdf

The Monash Ramaciotti Electron Microscopy Centre, in partnership with the Imaging CoE, has installed Australia’s first cryoFIB/SEM. The instrument was purchased in part through the award of an ARC LIEF grant awarded to Dr Georg Ramm Director, Ramaciotti Centre for Cryo-Electron Microscopy, Prof. James Whisstock, Prof Kat Gaus and colleagues. The cryo-FIB/SEM functions as a exquisitely fine molecular milling machine. In the context of biology, cryo-FIB/ SEM’s are used to carve out specific portions (or cryolamella) of cryo-genically preserved cells.

This step is essential, as cells themselves are too thick to image at high resolution using transmission electron microscopy. The advantage of using a cryo-FIB/ SEM over simpler approaches (for example cryo-sectioning) is that users can very precisely target and prepare the region of the cell that they are interested in studying. Once suitably prepared cryolamella derived from cells are transferred to a Titan Krios TEM, and detailed images are obtained using an approach called tomography. Here, the cryo-lamella are imaged at a sweep of different angles, and a 3-dimensional picture (or tomogram) is produced. MASSIVE is used extensively for CryoEM tomography and will be ab important tool for structural biologists using the cryo-FIB/ SEM.

The availability of this instrument sets the scene for Australian scientists being able to conduct “in situ” structural biology experi- ments. Here, through averaging of different tomograms, large protein complexes are imaged in the context of their native, cellular environment. The cryo-FIB/SEM thus moves us one step closer to one of the great ambi- tions of modern biology – the visualization of cells at near molecular resolution. In the context of the Imaging Centre, we anticipate using the cryo-FIB/ SEM in a wide variety of experiments, for example in the context of understanding how immune effectors interact with target cells.

The Imaging CoE is an affiliate partner of MASSIVE.