Nanoscopic subcellular imaging enabled by ion beam tomography

• Type of Material: Article
• Authors: Ahmet F Coskun, Stanford University; Guojun Han, Stanford University; Shambavi Ganesh, Georgia Institute of Technology; Shih-Yu Chen, Stanford University; Xavier Rovira Clave, Stanford University; Stefan Harmsen, Stanford University; Sizun Jiang, Stanford University; Christian M Schuerch, Stanford University; Yunhao Bai, Stanford University; Chuck Hitzman, Stanford University; Garry P Nolan, Stanford University
• Language: English
• Date: 2021-02-04
• Publisher: NATURE PORTFOLIO
• Publication Version: Final Published Version
• Copyright Statement: © The Author(s) 2021
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1038/s41467-020-20753-5
• Title of Journal or Parent Work: NATURE COMMUNICATIONS
• Volume: 12
• Issue: 1
• Start Page: 789
• End Page: 789
• Grant/Funding Information: C.M.S. was supported by the Swiss National Science Foundation (P300PB_171189, P400PM_183915).; S.J. is supported by a Stanford Dean’s Fellowship and the Leukemia & Lymphoma Society Career Development Program.; This work was supported by NIH 5R01NS08953304, NIH 5U54CA14914505, Juno Therapeutics, Bill & Melinda Gates Foundation, Array BioPharma, NIH 5UH2AR06767603, NIH 5R25CA18099304, NIH 5R01GM10983604, Department of the Army W81XWH-12-1-0591, W81XWH-14-1-0180, NIH 5R01CA18496804, NIH 5R01GM10983604, and the Rachford and Carlota A. Harris Endowed Professorship to G.P.N.; X.R.-C. is supported by a long-term EMBO fellowship (ALTF 300-2017).; A. F. C. was supported by start-up funds from the Georgia Institute of Technology and Emory University.
• Keywords: HETEROGENEITY; TRANSCRIPTION; Science & Technology - Other Topics; Multidisciplinary Sciences; FLUORESCENCE; Science & Technology; CELLS; LATERAL RESOLUTION; REPLICATION; MESSENGER-RNA
• Research Categories: Health Sciences, Radiology; Engineering, Biomedical

Abstract

Multiplexed ion beam imaging (MIBI) has been previously used to profile multiple parameters in two dimensions in single cells within tissue slices. Here, a mathematical and technical framework for three-dimensional (3D) subcellular MIBI is presented. Ion-beam tomography (IBT) compiles ion beam images that are acquired iteratively across successive, multiple scans, and later assembled into a 3D format without loss of depth resolution. Algorithmic deconvolution, tailored for ion beams, is then applied to the transformed ion image series, yielding 4-fold enhanced ion beam data cubes. To further generate 3D sub-ion-beam-width precision visuals, isolated ion molecules are localized in the raw ion beam images, creating an approach coined as SILM, secondary ion beam localization microscopy, providing sub-25 nm accuracy in original ion images. Using deep learning, a parameter-free reconstruction method for ion beam tomograms with high accuracy is developed for low-density targets. In cultured cancer cells and tissues, IBT enables accessible visualization of 3D volumetric distributions of genomic regions, RNA transcripts, and protein factors with 5 nm axial resolution using isotope-enrichments and label-free elemental analyses. Multiparameter imaging of subcellular features at near macromolecular resolution is implemented by the IBT tools as a general biocomputation pipeline for imaging mass spectrometry.