Elsevier (Cell Press)
Three-dimensional (3D) visualization of vitrified cells can uncover structures of subcellular complexes without chemical fixation or staining. Here, we present a pipeline integrating three imaging modalities to visualize the same specimen at cryogenic temperature at different scales: cryo-fluorescence confocal microscopy, volume cryo-focused ion beam scanning electron microscopy, and transmission cryo-electron tomography. Our proof-of-concept benchmark revealed the 3D distribution of organelles and subcellular structures in whole heat-shocked yeast cells, including the ultrastructure of protein inclusions that recruit fluorescently-labeled chaperone Hsp104. Since our workflow efficiently integrates imaging at three different scales and can be applied to other types of cells, it could be used for large-scale phenotypic studies of frozen-hydrated specimens in a variety of healthy and diseased conditions with and without treatments.
Wu, Gong-Her; Mitchell, Patrick G.; Galaz-Montoya, Jesus G.; Hecksel, Corey W.; Sontag, Emily M.; Gangadharan, Vimal; Marshman, Jeffrey; Mankus, David; Bisher, Margaret E.; Lytton-Jean, Abigail K.R.; Frydman, Judith; Czymmek, Kirk; and Chiu, Wah, "Multi-scale 3D Cryo-Correlative Microscopy for Vitrified Cells" (2020). Biological Sciences Faculty Research and Publications. 825.
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