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Correlative Microscopy for 3D Structural Analysis of Dynamic Interactions
Published on: June 24, 2013
Protocol for live-cell fluorescence-guided cryoFIB-milling and electron cryo-tomography of virus-infected cells
Linda E Franken1, Rene Rosch2, Ulrike Laugks3
1Leibniz Institute of Virology (LIV), Martinistraße 52, 20251 Hamburg, Germany; Centre for Structural Systems Biology, Notkestraße 85, 22607 Hamburg, Germany.
Abstract:
Here, we present a protocol for assessing virus-infected cells using electron cryo-tomography (cryoET). It includes the basic workflows of seeding cells, plunge-freezing, clipping, cryo-focused ion beam milling (cryoFIB-milling), and cryoET, as well as two optional modules: micropatterning and live-cell fluorescence microscopy. We use an A549 human cell line and the virus HAdV5-pIX-mcherry in this protocol, but the comprehensive workflow can be easily transferred to other cell types and different types of virus infection or treatment. For complete details on the use and execution of this protocol, please refer to Pfitzner et al. (2021).
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