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Updated: Nov 18, 2025

Applying Live Cell Imaging and Cryo-Electron Tomography to Resolve Spatiotemporal Features of the Legionella pneumophila Dot/Icm Secretion System
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Nanoscopic subcellular imaging enabled by ion beam tomography.

Ahmet F Coskun1,2,3, Guojun Han4, Shambavi Ganesh5,6

  • 1Baxter Laboratory, Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, CA, USA. ahmet.coskun@bme.gatech.edu.

Nature Communications
|February 5, 2021
PubMed
Summary
This summary is machine-generated.

This study introduces 3D subcellular Multiplexed Ion Beam Imaging (MIBI) using Ion-Beam Tomography (IBT) and Secondary Ion Beam Localization Microscopy (SILM). This advanced technique visualizes genomic regions, RNA, and proteins in 3D with nanoscale precision.

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Area of Science:

  • Biophysics
  • Cell Biology
  • Biotechnology

Background:

  • Multiplexed Ion Beam Imaging (MIBI) has enabled 2D cellular profiling in tissues.
  • A need exists for 3D subcellular resolution imaging to understand complex biological structures.

Purpose of the Study:

  • To present a mathematical and technical framework for 3D subcellular MIBI.
  • To develop novel computational methods for enhanced 3D ion beam imaging.
  • To enable high-resolution 3D visualization of molecular distributions within cells and tissues.

Main Methods:

  • Ion-Beam Tomography (IBT) for iterative image acquisition and 3D assembly.
  • Algorithmic deconvolution for enhanced ion beam data cubes.
  • Secondary Ion Beam Localization Microscopy (SILM) for sub-25 nm localization.
  • Deep learning for parameter-free tomogram reconstruction.
  • Isotope-enrichment and label-free elemental analyses.

Main Results:

  • Achieved 5 nm axial resolution in 3D volumetric imaging.
  • Successfully visualized 3D distributions of genomic regions, RNA transcripts, and protein factors.
  • Demonstrated near-macromolecular resolution for multiparameter imaging of subcellular features.
  • Validated the approach in cultured cancer cells and tissues.

Conclusions:

  • IBT provides an accessible pipeline for 3D subcellular imaging mass spectrometry.
  • The developed framework significantly advances the capabilities of MIBI for biological research.
  • This technology enables unprecedented visualization of molecular organization in 3D at the nanoscale.