Protocol for live-cell fluorescence-guided cryoFIB-milling and electron cryo-tomography of virus-infected cells

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-...

Descripción completa

Detalles Bibliográficos
Autores principales: Franken, Linda E., Rosch, Rene, Laugks, Ulrike, Grünewald, Kay
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Elsevier 2022
Materias:
Protocol
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9508610/
https://www.ncbi.nlm.nih.gov/pubmed/36149798
http://dx.doi.org/10.1016/j.xpro.2022.101696
Descripción
Sumario: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).

Ejemplares similares