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Cited by 722Advanced Electron Microscopy Techniques and ApplicationsAdvanced Fluorescence Microscopy TechniquesBiotin and Related Studies

Engineered ascorbate peroxidase as a genetically encoded reporter for electron microscopy

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Martell, Jeffrey D, Deerinck, Thomas J, Sancak, Yasemin, Poulos, Thomas L, Mootha, Vamsi K, Sosinsky, Gina E, Ellisman, Mark H, Ting, Alice Y·2012·Nature Biotechnology
DOI 10.1038/nbt.2375PMC PMC3699407
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Abstract

Electron microscopy (EM) is the standard method for imaging cellular structures with nanometer resolution, but existing genetic tags are inactive in most cellular compartments or require light and can be difficult to use. Here we report the development of 'APEX', a genetically encodable EM tag that is active in all cellular compartments and does not require light. APEX is a monomeric 28-kDa peroxidase that withstands strong EM fixation to give excellent ultrastructural preservation. We demonstrate the utility of APEX for high-resolution EM imaging of a variety of mammalian organelles and specific proteins using a simple and robust labeling procedure. We also fused APEX to the N or C terminus of the mitochondrial calcium uniporter (MCU), a recently identified channel whose topology is disputed. These fusions give EM contrast exclusively in the mitochondrial matrix, suggesting that both the N and C termini of MCU face the matrix. Because APEX staining is not dependent on light activation, APEX should make EM imaging of any cellular protein straightforward, regardless of the size or thickness of the specimen.

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  • The APEX system with DAB as substrate was adapted for use in diplomonads to reveal the ultrastructural localization of a fusion partner.

    “We have adapted the APEX system (25) for use in diplomonads with DAB as a substrate to reveal the ultrastructural localization of a fusion partner.”

    From Comparative Cell Biology and Evolution of Annexins in Diplomonads (2016) · cites this paper as [25]

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Research works printed but not linked: 33

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Authors
Martell, J. D., Deerinck, T. J., Sancak, Y., Poulos, T. L., Mootha, V. K., Sosinsky, G. E., Ellisman, M. H., & Ting, A. Y.
Affiliations
  • Massachusetts Institute of Technology (US)
  • University of California San Diego (US)
  • Broad Institute (US)
  • University of California, Irvine (US)
Corresponding author
Ting, Alice Y
ORCID
Venue
Nature Biotechnology
Publisher
Springer Science and Business Media LLC
Published
2012-10-21
Volume
30
Issue
11
Pages
1143-1148
ISSN
  • 0733-222X (ISSN-L)
  • 1087-0156
  • 1546-1696
DOI
10.1038/nbt.2375
PMC
PMC3699407
OpenAlex
W2160737695
Cited by
722
References
0
Citations by year
2026: 54 · 2025: 51 · 2024: 63 · 2023: 49 · 2022: 37 · 2021: 68 · 2020: 70 · 2019: 75 · 2018: 38 · 2017: 61 · 2016: 52 · 2015: 46 · 2014: 37 · 2013: 18 · 2012: 2
FWCI
14.46
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Biotechnology (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
  • Digital Access to Scholarship at Harvard (DASH) (Harvard University) (submittedVersion, cc-by)
  • DSpace@MIT (Massachusetts Institute of Technology) (submittedVersion, cc-by-nc)
  • eScholarship (California Digital Library) (submittedVersion, cc-by)
  • PMC
Funders
  • National Institutes of Health (DP1OD003961)
  • National Institute of General Medical Sciences (GM077465, R01GM072881, GM072881, P41GM103412, GM-065937, R01GM042614, GM42614, R01GM065937, R01GM077465)
  • National Center for Research Resources (P41RR004050, P41RR002250)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Advanced Electron Microscopy Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Biophysics › Advanced Fluorescence Microscopy Techniques
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Biotin and Related Studies
Keywords
APEX, ultrastructure preservation, mitochondrial calcium uniporter, protein localization, organelle imaging
MeSH terms
Microscopy, Electron, Protein Engineering, Genes, Reporter, Molecular Imaging, Ascorbate Peroxidases

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