Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

A guide to light-sheet fluorescence microscopy for multiscale imaging

Nature Methods volume 14, pages 360–373 (2017)Cite this article

Subjects

Abstract

The impact of light-sheet fluorescence microscopy (LSFM) is visible in fields as diverse as developmental and cell biology, anatomical science, biophysics and neuroscience. Although adoption among biologists has been steady, LSFM has not displaced more traditional imaging methods despite its often-superior performance. One reason for this is that the field has largely conformed to a do-it-yourself ethic, although the challenges of big image data cannot be overstated. With the most powerful implementations of LSFM available to only a few groups worldwide, the scope of this technique is unnecessarily limited. Here we elucidate the key developments and define a simple set of underlying principles governing LSFM. In doing so, we aim to clarify the decisions to be made for those who wish to develop and use bespoke light-sheet systems and to assist in identifying the best approaches to apply this powerful technique to myriad biological questions.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Learn more

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Light-sheet fluorescence microscopy.
Figure 2: Parallelization of light-sheet generation.
Figure 3: Spatial resolution in light-sheet fluorescence microscopy.
Figure 4: Reflected light-sheet geometries.
Figure 5: Gaussian and Bessel beams for light-sheet generation.
Figure 6: Axially swept light-sheet geometries.
Figure 7: Light-sheet penetration.
Figure 8: Multiview imaging.
Figure 9: Ultrafast volumetric imaging.
Figure 10: Hyperspectral light-sheet microscopy.

Similar content being viewed by others

References

  1. Chhetri, R.K. et al. Whole-animal functional and developmental imaging with isotropic spatial resolution. Nat. Methods 12, 1171–1178 (2015).

    Article  CAS  PubMed  Google Scholar 

  2. Arrenberg, A.B., Stainier, D.Y.R., Baier, H. & Huisken, J. Optogenetic control of cardiac function. Science 330, 971–974 (2010).

    Article  CAS  PubMed  Google Scholar 

  3. Mickoleit, M. et al. High-resolution reconstruction of the beating zebrafish heart. Nat. Methods 11, 919–922 (2014).

    Article  CAS  PubMed  Google Scholar 

  4. Rauzi, M. et al. Embryo-scale tissue mechanics during Drosophila gastrulation movements. Nat. Commun. 6, 8677 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Schmid, B. et al. High-speed panoramic light-sheet microscopy reveals global endodermal cell dynamics. Nat. Commun. 4, 2207 (2013).

    Article  PubMed  CAS  Google Scholar 

  6. Welf, E.S. et al. Quantitative multiscale cell imaging in controlled 3D microenvironments. Dev. Cell 36, 462–475 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Strnad, P. et al. Inverted light-sheet microscope for imaging mouse pre-implantation development. Nat. Methods 13, 139–142 (2016).

    Article  CAS  PubMed  Google Scholar 

  8. Wu, Y. et al. Inverted selective-plane illumination microscopy (iSPIM) enables coupled cell identity lineaging and neurodevelopmental imaging in Caenorhabditis elegans. Proc. Natl. Acad. Sci. USA 108, 17708–17713 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Wu, Y. et al. Spatially isotropic four-dimensional imaging with dual-view-plane illumination microscopy. Nat. Biotechnol. 31, 1032–1038 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Kaufmann, A., Mickoleit, M., Weber, M. & Huisken, J. Multilayer mounting enables long-term imaging of zebrafish development in a light-sheet microscope. Development 139, 3242–3247 (2012).

    Article  CAS  PubMed  Google Scholar 

  11. Vladimirov, N. et al. Light-sheet functional imaging in fictively behaving zebrafish. Nat. Methods 11, 883–884 (2014).

    Article  CAS  PubMed  Google Scholar 

  12. Wolf, S. et al. Whole-brain functional imaging with two-photon light-sheet microscopy. Nat. Methods 12, 379–380 (2015).

    Article  CAS  PubMed  Google Scholar 

  13. Pitrone, P.G. et al. OpenSPIM: an open-access light-sheet microscopy platform. Nat. Methods 10, 598–599 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Gualda, E.J. et al. OpenSpinMicroscopy: an open-source integrated microscopy platform. Nat. Methods 10, 599–600 (2013).

    Article  CAS  PubMed  Google Scholar 

  15. Huisken, J., Swoger, J., Del Bene, F., Wittbrodt, J. & Stelzer, E.H.K. Optical sectioning deep inside live embryos by selective-plane illumination microscopy. Science 305, 1007–1009 (2004).

    Article  CAS  PubMed  Google Scholar 

  16. Huisken, J. & Stainier, D.Y.R. Even fluorescence excitation by multidirectional selective-plane illumination microscopy (mSPIM). Opt. Lett. 32, 2608–2610 (2007).

    Article  PubMed  Google Scholar 

  17. Keller, P.J., Schmidt, A.D., Wittbrodt, J. & Stelzer, E.H.K. Reconstruction of zebrafish early embryonic development by scanned light-sheet microscopy. Science 322, 1065–1069 (2008).

    Article  CAS  PubMed  Google Scholar 

  18. Chen, B.-C. et al. Lattice light-sheet microscopy: imaging molecules to embryos at high spatiotemporal resolution. Science 346, 1257998 (2014).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  19. Cella Zanacchi, F. et al. Live-cell 3D super-resolution imaging in thick biological samples. Nat. Methods 8, 1047–1049 (2011).

    Article  PubMed  CAS  Google Scholar 

  20. Huang, B., Jones, S.A., Brandenburg, B. & Zhuang, X. Whole-cell 3D STORM reveals interactions between cellular structures with nanometer-scale resolution. Nat. Methods 5, 1047–1052 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Gebhardt, J.C.M. et al. Single-molecule imaging of transcription factor binding to DNA in live mammalian cells. Nat. Methods 10, 421–426 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Galland, R. et al. 3D high- and super-resolution imaging using single-objective SPIM. Nat. Methods 12, 641–644 (2015).

    Article  CAS  PubMed  Google Scholar 

  23. Li, T. et al. Axial-plane optical microscopy. Sci. Rep. 4, 7253 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Theer, P., Dragneva, D. & Knop, M. πSPIM: high-NA high-resolution isotropic light-sheet imaging in cell culture dishes. Sci. Rep. 6, 32880 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Tokunaga, M., Imamoto, N. & Sakata-Sogawa, K. Highly inclined thin illumination enables clear single-molecule imaging in cells. Nat. Methods 5, 159–161 (2008).

    Article  CAS  PubMed  Google Scholar 

  26. Gao, L. Optimization of the excitation light sheet in selective-plane illumination microscopy. Biomed. Opt. Express 6, 881–890 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  27. Planchon, T.A. et al. Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination. Nat. Methods 8, 417–423 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Gao, L. et al. Non-invasive imaging beyond the diffraction limit of 3D dynamics in thickly fluorescent specimens. Cell 151, 1370–1385 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Manton, J.D. & Rees, E.J. triSPIM: light-sheet microscopy with isotropic super-resolution. Opt. Lett. 41, 4170–4173 (2016).

    Article  PubMed  Google Scholar 

  30. Zhao, T. et al. Multicolor 4D fluorescence microscopy using ultrathin Bessel light sheets. Sci. Rep. 6, 26159 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Vettenburg, T. et al. Light-sheet microscopy using an Airy beam. Nat. Methods 11, 541–544 (2014).

    Article  CAS  PubMed  Google Scholar 

  32. Zong, W. et al. Large-field high-resolution two-photon digital-scanned light-sheet microscopy. Cell Res. 25, 254–257 (2015).

    Article  CAS  PubMed  Google Scholar 

  33. Dean, K.M. & Fiolka, R. Uniform and scalable light-sheets generated by extended focusing. Opt. Express 22, 26141–26152 (2014).

    Article  PubMed  Google Scholar 

  34. Dean, K.M., Roudot, P., Welf, E.S., Danuser, G. & Fiolka, R. Deconvolution-free subcellular imaging with axially swept light-sheet microscopy. Biophys. J. 108, 2807–2815 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Buytaert, J.A. & Dirckx, J.J. Design and quantitative resolution measurements of an optical virtual sectioning three-dimensional imaging technique for biomedical specimens, featuring two-micrometer slicing resolution. J. Biomed. Opt. 12, 14039 (2011).

    Article  Google Scholar 

  36. Gao, L. Extend the field of view of selective-plane illumination microscopy by tiling the excitation light sheet. Opt. Express 23, 6102–6111 (2015).

    Article  PubMed  Google Scholar 

  37. Fu, Q., Martin, B.L., Matus, D.Q. & Gao, L. Imaging multicellular specimens with real-time optimized tiling light-sheet selective-plane illumination microscopy. Nat. Commun. 7, 11088 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Friedrich, M., Gan, Q., Ermolayev, V. & Harms, G.S. STED-SPIM: stimulated emission depletion improves sheet-illumination microscopy resolution. Biophys. J. 100, L43–L45 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Hoyer, P. et al. Breaking the diffraction limit of light-sheet fluorescence microscopy by RESOLFT. Proc. Natl. Acad. Sci. USA 113, 3442–3446 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Palero, J., Santos, S.I.C.O., Artigas, D. & Loza-Alvarez, P. A simple scanless two-photon fluorescence microscope using selective plane illumination. Opt. Express 18, 8491–8498 (2010).

    Article  CAS  PubMed  Google Scholar 

  41. Truong, T.V., Supatto, W., Koos, D.S., Choi, J.M. & Fraser, S.E. Deep and fast live imaging with two-photon scanned light-sheet microscopy. Nat. Methods 8, 757–760 (2011).

    Article  CAS  PubMed  Google Scholar 

  42. Tomer, R., Khairy, K., Amat, F. & Keller, P.J. Quantitative high-speed imaging of entire developing embryos with simultaneous multiview light-sheet microscopy. Nat. Methods 9, 755–763 (2012).

    Article  CAS  PubMed  Google Scholar 

  43. Lemon, W.C. et al. Whole-central nervous system functional imaging in larval Drosophila. Nat. Commun. 6, 7924 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Supatto, W., Truong, T.V., Débarre, D. & Beaurepaire, E. Advances in multiphoton microscopy for imaging embryos. Curr. Opin. Genet. Dev. 21, 538–548 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Ji, N., Magee, J.C. & Betzig, E. High-speed, low-photodamage nonlinear imaging using passive pulse splitters. Nat. Methods 5, 197–202 (2008).

    Article  CAS  PubMed  Google Scholar 

  46. Mahou, P., Vermot, J., Beaurepaire, E. & Supatto, W. Multicolor two-photon light-sheet microscopy. Nat. Methods 11, 600–601 (2014).

    Article  CAS  PubMed  Google Scholar 

  47. Fahrbach, F., Simon, P. & Rohrbach, A. Microscopy with self-reconstructing beams. Nat. Photonics 4, 780–785 (2010).

    Article  CAS  Google Scholar 

  48. Fahrbach, F.O., Gurchenkov, V., Alessandri, K., Nassoy, P. & Rohrbach, A. Self-reconstructing sectioned Bessel beams offer submicron optical sectioning for large fields of view in light-sheet microscopy. Opt. Express 21, 11425–11440 (2013).

    Article  PubMed  Google Scholar 

  49. Fahrbach, F.O., Gurchenkov, V., Alessandri, K., Nassoy, P. & Rohrbach, A. Light-sheet microscopy in thick media using scanned Bessel beams and two-photon fluorescence excitation. Opt. Express 21, 13824–13839 (2013).

    Article  CAS  PubMed  Google Scholar 

  50. Keller, P.J. et al. Fast, high-contrast imaging of animal development with scanned light-sheet-based structured-illumination microscopy. Nat. Methods 7, 637–642 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Fahrbach, F.O. & Rohrbach, A. Propagation stability of self-reconstructing Bessel beams enables contrast-enhanced imaging in thick media. Nat. Commun. 3, 632 (2012).

    Article  PubMed  CAS  Google Scholar 

  52. Silvestri, L., Bria, A., Sacconi, L., Iannello, G. & Pavone, F.S. Confocal light-sheet microscopy: micron-scale neuroanatomy of the entire mouse brain. Opt. Express 20, 20582–20598 (2012).

    Article  CAS  PubMed  Google Scholar 

  53. Baumgart, E. & Kubitscheck, U. Scanned light-sheet microscopy with confocal slit detection. Opt. Express 20, 21805–21814 (2012).

    Article  PubMed  Google Scholar 

  54. Yang, Z. et al. Dual-slit confocal light-sheet microscopy for in vivo whole-brain imaging of zebrafish. Biomed. Opt. Express 6, 1797–1811 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  55. de Medeiros, G. et al. Confocal multiview light-sheet microscopy. Nat. Commun. 6, 8881 (2015).

    Article  PubMed  CAS  Google Scholar 

  56. Verveer, P.J. et al. High-resolution three-dimensional imaging of large specimens with light-sheet-based microscopy. Nat. Methods 4, 311–313 (2007).

    Article  CAS  PubMed  Google Scholar 

  57. Preibisch, S., Saalfeld, S., Schindelin, J. & Tomancak, P. Software for bead-based registration of selective-plane illumination microscopy data. Nat. Methods 7, 418–419 (2010).

    Article  CAS  PubMed  Google Scholar 

  58. Schmid, B. & Huisken, J. Real-time multiview deconvolution. Bioinformatics 31, 3398–3400 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Preibisch, S. et al. Efficient Bayesian-based multiview deconvolution. Nat. Methods 11, 645–648 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Swoger, J., Verveer, P., Greger, K., Huisken, J. & Stelzer, E.H.K. Multiview image fusion improves resolution in three-dimensional microscopy. Opt. Express 15, 8029–8042 (2007).

    Article  PubMed  Google Scholar 

  61. Krzic, U., Gunther, S., Saunders, T.E., Streichan, S.J. & Hufnagel, L. Multiview light-sheet microscope for rapid in toto imaging. Nat. Methods 9, 730–733 (2012).

    Article  CAS  PubMed  Google Scholar 

  62. Reynaud, E.G., Peychl, J., Huisken, J. & Tomancak, P. Guide to light-sheet microscopy for adventurous biologists. Nat. Methods 12, 30–34 (2015).

    Article  CAS  PubMed  Google Scholar 

  63. Dodt, H.-U. et al. Ultramicroscopy: three-dimensional visualization of neuronal networks in the whole mouse brain. Nat. Methods 4, 331–336 (2007).

    Article  CAS  PubMed  Google Scholar 

  64. Saghafi, S., Becker, K., Hahn, C. & Dodt, H.U. 3D-ultramicroscopy utilizing aspheric optics. J. Biophotonics 7, 117–125 (2014).

    Article  PubMed  Google Scholar 

  65. Golub, I., Chebbi, B. & Golub, J. Toward the optical 'magic carpet': reducing the divergence of a light sheet below the diffraction limit. Opt. Lett. 40, 5121–5124 (2015).

    Article  PubMed  Google Scholar 

  66. Wilding, D. et al. Pupil filters for extending the field of view in light-sheet microscopy. Opt. Lett. 41, 1205–1208 (2016).

    Article  CAS  PubMed  Google Scholar 

  67. Tomer, R., Ye, L., Hsueh, B. & Deisseroth, K. Advanced CLARITY for rapid and high-resolution imaging of intact tissues. Nat. Protoc. 9, 1682–1697 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  68. Fahrbach, F.O., Voigt, F.F., Schmid, B., Helmchen, F. & Huisken, J. Rapid 3D light-sheet microscopy with a tunable lens. Opt. Express 21, 21010–21026 (2013).

    Article  PubMed  Google Scholar 

  69. Ahrens, M.B., Orger, M.B., Robson, D.N., Li, J.M. & Keller, P.J. Whole-brain functional imaging at cellular resolution using light-sheet microscopy. Nat. Methods 10, 413–420 (2013).

    Article  CAS  PubMed  Google Scholar 

  70. Olarte, O.E., Andilla, J., Artigas, D. & Loza-Alvarez, P. Decoupled illumination detection in light-sheet microscopy for fast volumetric imaging. Optica 2, 702 (2015).

    Article  Google Scholar 

  71. Quirin, S. et al. Calcium imaging of neural circuits with extended depth-of-field light-sheet microscopy. Opt. Lett. 41, 855–858 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. Tomer, R. et al. SPED light-sheet microscopy: fast mapping of biological system structure and function. Cell 163, 1796–1806 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  73. Dunsby, C. Optically sectioned imaging by oblique-plane microscopy. Opt. Express 16, 20306–20316 (2008).

    Article  CAS  PubMed  Google Scholar 

  74. Bouchard, M.B. et al. Swept confocally aligned planar excitation (SCAPE) microscopy for high-speed volumetric imaging of behaving organisms. Nat. Photonics 9, 113–119 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  75. Jahr, W., Schmid, B., Schmied, C., Fahrbach, F.O. & Huisken, J. Hyperspectral light-sheet microscopy. Nat. Commun. 6, 7990 (2015).

    Article  PubMed  Google Scholar 

  76. Cutrale, F. et al. Hyperspectral phasor analysis enables multiplexed 5D in vivo imaging. Nat. Methods 14, 149–152 (2017).

    Article  CAS  PubMed  Google Scholar 

  77. Ji, N. Adaptive optical-fluorescence microscopy for biological imaging. Nat. Methods http://dx.doi.org/10.1038/nmeth.4218 (2017).

  78. Masson, A. et al. High-resolution in-depth imaging of optically cleared thick samples using an adaptive SPIM. Sci. Rep. 5, 16898 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  79. Wilding, D., Pozzi, P., Soloviev, O., Vdovin, G. & Verhaegen, M. Adaptive illumination based on direct wavefront sensing in a light-sheet fluorescence microscope. Opt. Express 24, 24896–24906 (2016).

    Article  PubMed  Google Scholar 

  80. Simmonds, R.D. & Booth, M.J. Modelling of multiconjugate adaptive optics for spatially variant aberrations in microscopy. J. Opt. 15, 094010 (2013).

    Article  Google Scholar 

  81. Royer, L.A. et al. Adaptive light-sheet microscopy for long-term, high-resolution imaging in living organisms. Nat. Biotechnol. 34, 1267–1278 (2016).

    Article  CAS  PubMed  Google Scholar 

  82. Scherf, N. & Huisken, J. The smart and gentle microscope. Nat. Biotechnol. 33, 815–818 (2015).

    Article  CAS  PubMed  Google Scholar 

  83. Hoebe, R.A. et al. Controlled light-exposure microscopy reduces photobleaching and phototoxicity in fluorescence live-cell imaging. Nat. Biotechnol. 25, 249–253 (2007).

    Article  CAS  PubMed  Google Scholar 

  84. Conrad, C. et al. Micropilot: automation of fluorescence-microscopy-based imaging for systems biology. Nat. Methods 8, 246–249 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  85. Chmielewski, A.K. et al. Fast imaging of live organisms with sculpted light sheets. Sci. Rep. 5, 9385 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  86. Prevedel, R. et al. Simultaneous whole-animal 3D imaging of neuronal activity using light-field microscopy. Nat. Methods 11, 727–730 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  87. McGorty, R. et al. Open-top selective-plane illumination microscope for conventionally mounted specimens. Opt. Express 23, 16142–16153 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Chung, K., Crane, M.M. & Lu, H. Automated on-chip rapid microscopy, phenotyping and sorting of C. elegans. Nat. Methods 5, 637–643 (2008).

    Article  CAS  PubMed  Google Scholar 

  89. Pardo-Martin, C. et al. High-throughput hyperdimensional vertebrate phenotyping. Nat. Commun. 4, 1467 (2013).

    Article  PubMed  CAS  Google Scholar 

  90. Pardo-Martin, C. et al. High-throughput in vivo vertebrate screening. Nat. Methods 7, 634–636 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Gualda, E.J. et al. SPIM-fluid: open source light-sheet-based platform for high-throughput imaging. Biomed. Opt. Express 6, 4447–4456 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  92. Heemskerk, I. & Streichan, S.J. Tissue cartography: compressing bio-image data by dimensional reduction. Nat. Methods 12, 1139–1142 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  93. Guan, Z. et al. Compact plane illumination plugin device to enable light-sheet fluorescence imaging of multicellular organisms on an inverted wide-field microscope. Biomed. Opt. Express 7, 194–208 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  94. Paiè, P., Bragheri, F., Bassi, A. & Osellame, R. Selective-plane illumination microscopy on a chip. Lab Chip 16, 1556–1560 (2016).

    Article  PubMed  CAS  Google Scholar 

  95. Engelbrecht, C.J., Voigt, F. & Helmchen, F. Miniaturized selective-plane illumination microscopy for high-contrast in vivo fluorescence imaging. Opt. Lett. 35, 1413–1415 (2010).

    Article  PubMed  Google Scholar 

  96. Plöschner, M. et al. Multimode fiber: light-sheet microscopy at the tip of a needle. Sci. Rep. 5, 18050 (2015).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  97. Oshima, Y. et al. Light-sheet-excited spontaneous Raman imaging of a living fish by optical sectioning in a wide-field Raman microscope. Opt. Express 20, 16195–16204 (2012).

    Article  CAS  Google Scholar 

  98. Rocha-Mendoza, I. et al. Rapid spontaneous Raman light-sheet microscopy using cw lasers and tunable filters. Biomed. Opt. Express 6, 3449–3461 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  99. Yang, Z., Downie, H., Rozbicki, E., Dupuy, L.X. & MacDonald, M.P. Light sheet tomography (LST) for in situ imaging of plant roots. Opt. Express 21, 16239–16247 (2013).

    Article  PubMed  CAS  Google Scholar 

  100. Mayer, J. et al. OPTiSPIM: integrating optical projection tomography in light-sheet microscopy extends specimen characterization to nonfluorescent contrasts. Opt. Lett. 39, 1053–1056 (2014).

    Article  CAS  PubMed  Google Scholar 

  101. Bassi, A., Schmid, B. & Huisken, J. Optical tomography complements light-sheet microscopy for in toto imaging of zebrafish development. Development 142, 1016–1020 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Doerr, J. et al. Whole-brain 3D mapping of human neural transplant innervation. Nat. Commun. 8, 14162 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  103. Engelbrecht, C.J. et al. Three-dimensional laser microsurgery in light-sheet-based microscopy (SPIM). Opt. Express 15, 6420–6430 (2007).

    Article  PubMed  Google Scholar 

  104. Yang, Z., Piksarv, P., Ferrier, D.E.K., Gunn-Moore, F.J. & Dholakia, K. Macro-optical trapping for sample confinement in light-sheet microscopy. Biomed. Opt. Express 6, 2778–2785 (2015).

    Article  PubMed  PubMed Central  Google Scholar 

  105. Fahrbach, F.O. & Rohrbach, A. A line-scanned light-sheet microscope with phase-shaped self-reconstructing beams. Opt. Express 18, 24229–24244 (2010).

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by the Max Planck Society (R.M.P. and J.H.), the European Research Council (ERC Consolidator grant 647885; J.H.) and a fellowship from the Human Frontier Science Program (HFSP) (LT000321/2015-C; R.M.P.). We thank members of the Huisken lab for discussions and critical comments on the manuscript.

Author information

Authors and Affiliations

  1. Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany

    Rory M Power & Jan Huisken

  2. Department of Medical Engineering, Morgridge Institute for Research, Madison, Wisconsin, USA

    Rory M Power & Jan Huisken

  3. Department of Biomedical Engineering, University of Wisconsin, Madison, Wisconsin, USA

    Rory M Power & Jan Huisken

Authors
  1. Rory M Power
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jan Huisken
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Jan Huisken.

Ethics declarations

Competing interests

J.H. is a co-inventor on patent US 20060033987 and an inventor on patent US 20110115895, which are related to light-sheet microscopy.

Supplementary information

Supplementary Text and Figures

Supplementary Figure 1, Supplementary Tables 1–5 and Supplementary Notes 1–8 (PDF 1760 kb)

Rights and permissions

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Power, R., Huisken, J. A guide to light-sheet fluorescence microscopy for multiscale imaging. Nat Methods 14, 360–373 (2017). https://doi.org/10.1038/nmeth.4224

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nmeth.4224

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing