Multicolour multilevel STED nanoscopy of actin/spectrin organization at synapses

S. Sidenstein, E. D’Este, M. Böhm, J.G. Danzl, V. Belov, S. Hell, Scientific Reports 6 (2016) 1–8.

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Journal Article | Published | English
Author
Sidenstein, Sven; D'Este, Elisa; Böhm, Marvin; Danzl, Johann GIST Austria ; Belov, Vladimir; Hell, Stefan
Abstract
Superresolution fluorescence microscopy of multiple fluorophores still requires development. Here we present simultaneous three-colour stimulated emission depletion (STED) nanoscopy relying on a single STED beam at 620 nm. Toggling the STED beam between two or more power levels ("multilevelSTEDv) optimizes resolution and contrast in all colour channels, which are intrinsically co-aligned and well separated. Three-colour recording is demonstrated by imaging the nanoscale cytoskeletal organization in cultured hippocampal neurons. The down to ∼35 nm resolution identified periodic actin/betaII spectrin lattices along dendrites and spines; however, at presynaptic and postsynaptic sites, these patterns were found to be absent. Both our multicolour scheme and the 620 nm STED line should be attractive for routine STED microscopy applications.
Publishing Year
Date Published
2016-05-25
Journal Title
Scientific Reports
Acknowledgement
We acknowledge the assistance of I. Herfort with neuron preparation, and of J. Bienert and K. Müller with analyses of the dye 540R derivatives. We thank T. Gilat and E. Rothermel for sample preparation as well as J. Keller, F. Winter (all MPI-BPC) and C.A. Wurm (Abberior Instruments) for helpful discussion, and S.J. Sahl (MPI-BPC) for a critical reading of the manuscript.
Volume
6
Page
1 - 8
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Sidenstein S, D’Este E, Böhm M, Danzl JG, Belov V, Hell S. Multicolour multilevel STED nanoscopy of actin/spectrin organization at synapses. Scientific Reports. 2016;6:1-8. doi:10.1038/srep26725
Sidenstein, S., D’Este, E., Böhm, M., Danzl, J. G., Belov, V., & Hell, S. (2016). Multicolour multilevel STED nanoscopy of actin/spectrin organization at synapses. Scientific Reports, 6, 1–8. https://doi.org/10.1038/srep26725
Sidenstein, Sven, Elisa D’Este, Marvin Böhm, Johann G Danzl, Vladimir Belov, and Stefan Hell. “Multicolour Multilevel STED Nanoscopy of Actin/Spectrin Organization at Synapses.” Scientific Reports 6 (2016): 1–8. https://doi.org/10.1038/srep26725.
S. Sidenstein, E. D’Este, M. Böhm, J. G. Danzl, V. Belov, and S. Hell, “Multicolour multilevel STED nanoscopy of actin/spectrin organization at synapses,” Scientific Reports, vol. 6, pp. 1–8, 2016.
Sidenstein S, D’Este E, Böhm M, Danzl JG, Belov V, Hell S. 2016. Multicolour multilevel STED nanoscopy of actin/spectrin organization at synapses. Scientific Reports. 6, 1–8.
Sidenstein, Sven, et al. “Multicolour Multilevel STED Nanoscopy of Actin/Spectrin Organization at Synapses.” Scientific Reports, vol. 6, Nature Publishing Group, 2016, pp. 1–8, doi:10.1038/srep26725.

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