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Oblique-plane single-molecule localization microscopy for tissues and small intact animals

A Publisher Correction to this article was published on 09 September 2019

This article has been updated


Single-molecule localization microscopy (SMLM), while well established for cultured cells, is not yet fully compatible with tissue-scale samples. We introduce single-molecule oblique-plane microscopy (obSTORM), which by directly imaging oblique sections of samples with oblique light-sheet illumination offers a deep and volumetric SMLM platform that is convenient for standard tissue samples and small intact animals. We demonstrate super-resolution imaging at depths of up to 66 µm for cells, Caenorhabditis elegans gonads, Drosophila melanogaster larval brain, mouse retina and brain sections, and whole stickleback fish.

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Fig. 1: Imaging principle and cell-scale demonstration of obSTORM.
Fig. 2: Tissue-scale super-resolution imaging enabled by 45° obSTORM.
Fig. 3: Volumetric super-resolution imaging of PKCα in 60-μm-thick mouse retina sections.

Data availability

The data that support the findings of this study are available from the corresponding authors upon reasonable request.

Code availability

The custom MATLAB codes for the localization analysis used in this study are available as Supplementary Software.

Change history

  • 09 September 2019

    An amendment to this paper has been published and can be accessed via a link at the top of the paper.


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The authors thank L. Li (X. Zhang Lab at the University of California, Berkeley) for providing gold-evaporated silicon wafer mirrors, S. Köhler (A.F. Dernburg Lab at the University of California, Berkeley) for help with C. elegans samples and A. Bormann and T. Square (C.T. Miller Lab at the University of California, Berkeley) for help with stickleback samples. We thank C.T. Miller, J.W. de Jong and H. Adesnik for discussions. X.Z. acknowledges support from the Gordon and Betty Moore Foundation and the Office of Naval Research Multidisciplinary University Research Initiative program (N00014-17-1-2588). K.X. is a Chan Zuckerberg Biohub investigator and acknowledges support from the Bakar Fellows Award, and STROBE, an NSF Science and Technology Center (DMR 1548924). M.W. acknowledges an NSF Graduate Research Fellowship (DGE-1106400).

Author information




J.K. designed and built the microscopy system, calculated theoretical PSFs, prepared fluorescent bead samples, calibrated the optical system and wrote software code for localization analysis. M.W. and S.M. prepared cell samples. S.M. labeled mouse brain tissues. E.A.Z. and J.G.F. prepared retina samples and provided fixed brain sections. N.M. and Z.L.N. prepared Drosophila samples. J.K. and M.W. carried out imaging experiments. J.K. and Y.W. analyzed single-molecule data. X.Z. and K.X. guided the research. J.K., M.W., Y.W., K.X. and X.Z. contributed to writing the manuscript.

Corresponding authors

Correspondence to Ke Xu or Xiang Zhang.

Ethics declarations

Competing interests

J.K., Y.W. and X.Z. have filed a provisional patent application on the microscopy system and method.

Additional information

Peer review information: Rita Strack was the primary editor on this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Supplementary Information

Supplementary Figs. 1–31, Supplementary Table 1 and Supplementary Note

Reporting Summary

Supplementary Video 1

Representative single-molecule raw images by obSTORM (α = 45°) for AF647-labeled microtubules in an A549 cell (Fig. 1d). The video shows 300 frames of n = 60,000 frames at 50 frames per second.

Supplementary Software

MATLAB codes used for single-molecule localization.

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Kim, J., Wojcik, M., Wang, Y. et al. Oblique-plane single-molecule localization microscopy for tissues and small intact animals. Nat Methods 16, 853–857 (2019).

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