Correlative montage parallel array cryo-tomography for in situ structural cell biology
J.E. Yang et al. "Correlative montage parallel array cryo-tomography for in situ structural cell biology". Nature Methods 20: 1537-1543 (2023) [DOI:10.1038/s41592-023-01999-5]
Imaging large fields of view while preserving high-resolution structural information remains a challenge in low-dose cryo-electron tomography. Here we present robust tools for montage parallel array cryo-tomography (MPACT) tailored for vitrified specimens. The combination of correlative cryo-fluorescence microscopy, focused-ion-beam milling, substrate micropatterning, and MPACT supports studies that contextually define the three-dimensional architecture of cells. To further extend the flexibility of MPACT, tilt series may be processed in their entirety or as individual tiles suitable for sub-tomogram averaging, enabling efficient data processing and analysis.
All relevant data are available from the corresponding author upon reasonable request. A set of raw frames (Extended Data Fig. 9a; a representative 3 × 3 montage cryo-tilt series) is provided as a demo dataset and available to download at https://github.com/wright-cemrc-projects/cryoet-montage/tree/main/Tutor… for demonstration of pre-processing steps including montage tilt-series generation and stitching. Sub-tomogram averages have been deposited in the Electron Microscopy Data Bank (https://www.emdataresource.org/) under accession numbers EMD-40308 (Extended Data Fig. 10; sub-tomogram average of total RSV F-pair particles picked from individual tile tilt/sub-tilt montage cryo-ET tomogram tiles) and EMD-40307 (Extended Data Fig. 10; sub-tomogram average of overdose removed RSV F-pair particles picked from individual tile tilt/sub-tilt montage cryo-ET tomogram tiles).
