Cryogenic Electron Microscopy (SEM and FIB-SEM)
Contacts:
Kristian Mølhave, DTU-Nanotech, 2512 6672 (kristian.molhave@nanotech.dtu.dk)
Anne Marie Rasmussen, CEN.
http://www.nanotech.dtu.dk/Medarbejdere/Staff/Molhave_Kristian.aspx
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| Image showing how a cryogenic plant cell sample inside a FIB-SEM can be cut open with the FIB and imaged with the SEM. |
Introduction: At the Center for Electron Nanoscopy, some of the microscopes have the ability to image samples at cryogenic temperatures (at -196C). We need to gain experience in using these systems for imaging liquid and biological samples – optimizing ways to prepare samples, handle them, image them in SEM, and hopefully perform both focused ion beam milling (FIB) and SEM imaging on the frozen samples.
Project goal: The overall goal is develop protocols for sample treatment to image the freshly cut surfaces of cryogenically frozen samples. This can be done bu fracturing the sample, cutting with cryo cooled knives (cryo-ultramicrotome) or FIB-SEM. With well controlled processes like microtomes and FIB-SEM, 3D images can be made of the sample by repeating the cutting process.
Description: You will use the SEM with attached cryo sample-preparation system to learn to handle and image cryo samples. You will hence learn and experiment with various types of samples. We would like to use this to image cells and nanostructured solutions/mixtures such as plants, tissues, and food products. There is a lot of interesting physics in the process, scince freezing must be done very rapidly (~20ms to -196 C) to avoid large ice crystal destroying the sample. Then test the different ways to cut the samples: freeze fracture and cryo-microtomy to compare how they perform. If everything works well, we will move to the more complicated FIB-SEM process for cutting.
Perspectives: Cryogenic electron microscopy has a promising future because it allows to image samples without drying them out or in other ways try to prepare them for the vacuum environment in the microscope. Once we learn to handle this equipment properly, we expect that it will become a standard method at CEN and with time increasingly also for TEM.