Rapid tool for cell nanoarchitecture integrity assessment

• Published in: Journal of structural biology 2021-12, Vol.213 (4), p.107801-107801, Article 107801
• Main Authors: Gaietta, Guido, Swift, Mark F., Volkmann, Niels, Hanein, Dorit
• Format: Article
• Language: English
• Subjects: Actin Cytoskeleton - chemistry; Actin Cytoskeleton - ultrastructure; Animals; Cells, Cultured; CLEM; Cryo-electron tomography; Cryoelectron Microscopy - methods; Electron Microscope Tomography - methods; Eukaryotic Cells - chemistry; Eukaryotic Cells - classification; Eukaryotic Cells - ultrastructure; HeLa Cells; Humans; Imaging, Three-Dimensional - methods; In-situ cellular methods; Intracellular Membranes - chemistry; Intracellular Membranes - ultrastructure; Life Sciences; Mice; Microscopy, Fluorescence - methods; Mitochondria - chemistry; Mitochondria - ultrastructure; Nanostructures - chemistry; Nanostructures - ultrastructure; NIH 3T3 Cells; Reproducibility of Results; THP-1 Cells; Virus inactivation
• ISSN: 1047-8477; 1095-8657
• DOI: 10.1016/j.jsb.2021.107801

[Display omitted] •Rapid screening assay for high-resolution structural integrity of cells (RCIA).•RCIA benefits all studies relying on 3D integrity of cellular nano-environments.•RCIA can test effects on structural integrity for any type of perturbation.•RCIA reporting of high-resolution integrity was validated by cellular cryo-ET. With the rapid increase and accessibility of high-resolution imaging technologies of cells, the interpretation of results relies more and more on the assumption that the three-dimensional integrity of the surrounding cellular landscape is not compromised by the experimental setup. However, the only available technology for directly probing the structural integrity of whole-cell preparations at the nanoscale is electron cryo-tomography, which is time-consuming, costly, and complex. We devised an accessible, inexpensive and reliable screening assay to quickly report on the compatibility of experimental protocols with preserving the structural integrity of whole-cell preparations at the nanoscale. Our Rapid Cell Integrity Assessment (RCIA) assay is executed at room temperature and relies solely on light microscopy imaging. Using cellular electron cryo-tomography as a benchmark, we verify that RCIA accurately unveils the adverse impact of reagents and/or protocols such as those used for virus inactivation or to arrest dynamic processes on the cellular nanoarchitecture.