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Turning single-molecule localization microscopy into a quantitative bioanalytical tool

Nature Protocols volume 12, pages 453–460 (2017)Cite this article

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Abstract

Single-molecule localization microscopy (SMLM) generates super-resolution images by serially detecting individual fluorescent molecules. The power of SMLM, however, goes beyond images: biologically relevant information can be extracted from the mathematical relationships between the positions of the fluorophores in space and time. Here we review the history of SMLM and how recent progress in methods for spatial point analysis has enabled quantitative measurement of SMLM data, providing insights into biomolecule patterning, clustering and oligomerization in biological systems.

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Figure 1: Timeline of SMLM developments, seen as a recapitulation of the SMLM workflow.
Figure 2: Spatial analysis approaches applied to identical SMLM data.
Figure 3: Scheme for kinetics-based molecular counting in SMLM data.

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Authors and Affiliations

  1. ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, Australia

    Philip R Nicovich & Katharina Gaus

  2. EMBL Australia Node in Single Molecule Science, School of Medical Sciences, University of New South Wales, Sydney, Australia

    Philip R Nicovich & Katharina Gaus

  3. Department of Physics and Randall Division of Cell and Molecular Biophysics, King's College London, London, UK

    Dylan M Owen

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  1. Philip R Nicovich
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P.R.N., D.M.O. and K.G. wrote the manuscript. P.R.N. prepared the figures.

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Correspondence to Katharina Gaus.

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Nicovich, P., Owen, D. & Gaus, K. Turning single-molecule localization microscopy into a quantitative bioanalytical tool. Nat Protoc 12, 453–460 (2017). https://doi.org/10.1038/nprot.2016.166

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