Determining composition of micron-scale protein deposits in neurodegenerative disease by spatially targeted optical microproteomics

  1. Kevin C Hadley
  2. Rishi Rakhit
  3. Hongbo Guo
  4. Yulong Sun
  5. James EN Jonkman
  6. Joanne McLaurin
  7. Lili-Naz Hazrati
  8. Andrew Emili
  9. Avijit Chakrabartty  Is a corresponding author
  1. University of Toronto, Canada
  2. Stanford University, United States
  3. University Health Network, Canada

Abstract

Spatially targeted optical microproteomics (STOMP) is a novel proteomics technique for interrogating micron-scale regions of interest (ROI) in mammalian tissue, with no requirement for genetic manipulation. Methanol or formalin fixed specimens are stained with fluorescent dyes or antibodies to visualize ROIs, then soaked in solutions containing the photo-tag: 4-benzoylbenzyl-glycyl-hexahistidine. Confocal imaging along with two photon excitation are used to covalently couple photo-tags to all proteins within each ROI, to a resolution of 0.67 µm in the xy-plane and 1.48 µm axially. After tissue solubilization, photo-tagged proteins are isolated and identified by mass spectrometry. As a test case, we examined amyloid plaques in an Alzheimer's disease (AD) mouse model and a postmortem AD case, confirming known plaque constituents and discovering new ones. STOMP can be applied to various biological samples including cell lines, primary cell cultures, ex vivo specimens, biopsy samples and fixed postmortem tissue.

Article and author information

Author details

  1. Kevin C Hadley

    Department of Medical Biophysics, Princess Margaret Cancer Centre, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  2. Rishi Rakhit

    Department of Chemical and Systems Biology, Stanford University, Stanford, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Hongbo Guo

    The Banting and Best Department of Medical Research, Terrence Donnelly Centre for Cellular & Biomolecular Research, Department of Molecular Genetics, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  4. Yulong Sun

    Department of Medical Biophysics, Princess Margaret Cancer Centre, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  5. James EN Jonkman

    Advanced Optical Microscopy Facility, Princess Margaret Cancer Centre, University Health Network, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  6. Joanne McLaurin

    Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  7. Lili-Naz Hazrati

    Tanz Centre for Research in Neurodegenerative Diseases, Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  8. Andrew Emili

    The Banting and Best Department of Medical Research, Terrence Donnelly Centre for Cellular & Biomolecular Research, Department of Molecular Genetics, University of Toronto, Toronto, Canada
    Competing interests
    The authors declare that no competing interests exist.
  9. Avijit Chakrabartty

    Departments of Biochemistry and Medical Biophysics, Princess Margaret Cancer Centre, University of Toronto, Toronto, Canada
    For correspondence
    chakrab@uhnresearch.ca
    Competing interests
    The authors declare that no competing interests exist.

Reviewing Editor

  1. Jeffery W Kelly, Scripps Research Institute, United States

Ethics

Animal experimentation: This study was performed in strict accordance with the University of Toronto Animal Care Committee Guidelines.

Human subjects: The work presented was performed in compliance with recognized international standards, including the International Conference on Harmonization (ICH), the Council for International Organizations of Medical Sciences (CIOMS) and the principles of the Declaration of Helsinki. Use of human tissue was in accordance with the University Health Network Research Ethics Board. The Human brain samples were collected as part of the Canadian Brain Tissue Bank (CBTB). At the time of collection, informed consent was obtained.

Version history

  1. Received: June 20, 2015
  2. Accepted: September 15, 2015
  3. Accepted Manuscript published: September 29, 2015 (version 1)
  4. Version of Record published: November 5, 2015 (version 2)

Copyright

© 2015, Hadley et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

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  1. Kevin C Hadley
  2. Rishi Rakhit
  3. Hongbo Guo
  4. Yulong Sun
  5. James EN Jonkman
  6. Joanne McLaurin
  7. Lili-Naz Hazrati
  8. Andrew Emili
  9. Avijit Chakrabartty
(2015)
Determining composition of micron-scale protein deposits in neurodegenerative disease by spatially targeted optical microproteomics
eLife 4:e09579.
https://doi.org/10.7554/eLife.09579

Share this article

https://doi.org/10.7554/eLife.09579

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