Abstract

Stimulated emission depletion (STED) microscopy allows fluorescence far-field imaging with diffraction-unlimited resolution. Unfortunately, extending this technique to three-dimensional (3D) imaging of thick specimens has been inhibited by sample-induced aberrations. Here we present the first implementation of adaptive optics in STED microscopy to allow 3D super-resolution imaging in strongly aberrated imaging conditions, such as those introduced by thick biological tissue.

© 2012 OSA

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  1. D. Toomre and J. Bewersdorf, “A new wave of cellular imaging,” Annu. Rev. Cell Dev. Biol.26(1), 285–314 (2010).
    [CrossRef] [PubMed]
  2. S. W. Hell, “Microscopy and its focal switch,” Nat. Methods6(1), 24–32 (2009).
    [CrossRef] [PubMed]
  3. S. W. Hell and J. Wichmann, “Breaking the diffraction resolution limit by stimulated emission: stimulated-emission-depletion fluorescence microscopy,” Opt. Lett.19(11), 780–782 (1994).
    [CrossRef] [PubMed]
  4. E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
    [CrossRef] [PubMed]
  5. S. T. Hess, T. P. Girirajan, and M. D. Mason, “Ultra-high resolution imaging by fluorescence photoactivation localization microscopy,” Biophys. J.91(11), 4258–4272 (2006).
    [CrossRef] [PubMed]
  6. M. J. Rust, M. Bates, and X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods3(10), 793–796 (2006).
    [CrossRef] [PubMed]
  7. J. B. Ding, K. T. Takasaki, and B. L. Sabatini, “Supraresolution imaging in brain slices using stimulated-emission depletion two-photon laser scanning microscopy,” Neuron63(4), 429–437 (2009).
    [CrossRef] [PubMed]
  8. N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
    [CrossRef] [PubMed]
  9. B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
    [CrossRef] [PubMed]
  10. S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
    [CrossRef] [PubMed]
  11. S. Deng, L. Liu, Y. Cheng, R. Li, and Z. Xu, “Investigation of the influence of the aberration induced by a plane interface on STED microscopy,” Opt. Express17(3), 1714–1725 (2009).
    [CrossRef] [PubMed]
  12. R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
    [CrossRef] [PubMed]
  13. T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
    [CrossRef] [PubMed]
  14. M. J. Booth, M. A. Neil, R. Juskaitis, and T. Wilson, “Adaptive aberration correction in a confocal microscope,” Proc. Natl. Acad. Sci. U.S.A.99(9), 5788–5792 (2002).
    [CrossRef] [PubMed]
  15. N. Ji, D. E. Milkie, and E. Betzig, “Adaptive optics via pupil segmentation for high-resolution imaging in biological tissues,” Nat. Methods7(2), 141–147 (2010).
    [CrossRef] [PubMed]
  16. M. Rueckel, J. A. Mack-Bucher, and W. Denk, “Adaptive wavefront correction in two-photon microscopy using coherence-gated wavefront sensing,” Proc. Natl. Acad. Sci. U.S.A.103(46), 17137–17142 (2006).
    [CrossRef] [PubMed]
  17. X. D. Tao, O. Azucena, M. Fu, Y. Zuo, D. C. Chen, and J. Kubby, “Adaptive optics microscopy with direct wavefront sensing using fluorescent protein guide stars,” Opt. Lett.36(17), 3389–3391 (2011).
    [CrossRef] [PubMed]
  18. M. Schwertner, M. J. Booth, and T. Wilson, “Characterizing specimen induced aberrations for high NA adaptive optical microscopy,” Opt. Express12(26), 6540–6552 (2004).
    [CrossRef] [PubMed]
  19. T. J. Gould, J. R. Myers, and J. Bewersdorf, “Total internal reflection STED microscopy,” Opt. Express19(14), 13351–13357 (2011).
    [CrossRef] [PubMed]
  20. G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
    [CrossRef] [PubMed]
  21. K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
    [CrossRef] [PubMed]
  22. E. Auksorius, B. R. Boruah, C. Dunsby, P. M. Lanigan, G. Kennedy, M. A. Neil, and P. M. French, “Stimulated emission depletion microscopy with a supercontinuum source and fluorescence lifetime imaging,” Opt. Lett.33(2), 113–115 (2008).
    [CrossRef] [PubMed]
  23. M. Booth, “Wave front sensor-less adaptive optics: a model-based approach using sphere packings,” Opt. Express14(4), 1339–1352 (2006).
    [CrossRef] [PubMed]
  24. D. Débarre, E. J. Botcherby, T. Watanabe, S. Srinivas, M. J. Booth, and T. Wilson, “Image-based adaptive optics for two-photon microscopy,” Opt. Lett.34(16), 2495–2497 (2009).
    [CrossRef] [PubMed]
  25. A. Facomprez, E. Beaurepaire, and D. Débarre, “Accuracy of correction in modal sensorless adaptive optics,” Opt. Express20(3), 2598–2612 (2012).
    [CrossRef] [PubMed]
  26. A. Thayil and M. J. Booth, “Self calibration of sensorless adaptive optical microscopes,” J. Eur. Opt. Soc.6, 11045 (2011).
    [CrossRef]
  27. M. A. Neil, M. J. Booth, and T. Wilson, “New modal wave-front sensor: a theoretical analysis,” J. Opt. Soc. Am. A17(6), 1098–1107 (2000).
    [CrossRef] [PubMed]
  28. F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
    [CrossRef] [PubMed]
  29. I. Izeddin, M. El Beheiry, J. Andilla, D. Ciepielewski, X. Darzacq, and M. Dahan, “PSF shaping using adaptive optics for three-dimensional single-molecule super-resolution imaging and tracking,” Opt. Express20(5), 4957–4967 (2012).
    [CrossRef] [PubMed]

2012 (3)

2011 (6)

A. Thayil and M. J. Booth, “Self calibration of sensorless adaptive optical microscopes,” J. Eur. Opt. Soc.6, 11045 (2011).
[CrossRef]

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

T. J. Gould, J. R. Myers, and J. Bewersdorf, “Total internal reflection STED microscopy,” Opt. Express19(14), 13351–13357 (2011).
[CrossRef] [PubMed]

N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
[CrossRef] [PubMed]

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

X. D. Tao, O. Azucena, M. Fu, Y. Zuo, D. C. Chen, and J. Kubby, “Adaptive optics microscopy with direct wavefront sensing using fluorescent protein guide stars,” Opt. Lett.36(17), 3389–3391 (2011).
[CrossRef] [PubMed]

2010 (2)

N. Ji, D. E. Milkie, and E. Betzig, “Adaptive optics via pupil segmentation for high-resolution imaging in biological tissues,” Nat. Methods7(2), 141–147 (2010).
[CrossRef] [PubMed]

D. Toomre and J. Bewersdorf, “A new wave of cellular imaging,” Annu. Rev. Cell Dev. Biol.26(1), 285–314 (2010).
[CrossRef] [PubMed]

2009 (4)

2008 (2)

2006 (7)

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

S. T. Hess, T. P. Girirajan, and M. D. Mason, “Ultra-high resolution imaging by fluorescence photoactivation localization microscopy,” Biophys. J.91(11), 4258–4272 (2006).
[CrossRef] [PubMed]

M. J. Rust, M. Bates, and X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods3(10), 793–796 (2006).
[CrossRef] [PubMed]

M. Rueckel, J. A. Mack-Bucher, and W. Denk, “Adaptive wavefront correction in two-photon microscopy using coherence-gated wavefront sensing,” Proc. Natl. Acad. Sci. U.S.A.103(46), 17137–17142 (2006).
[CrossRef] [PubMed]

M. Booth, “Wave front sensor-less adaptive optics: a model-based approach using sphere packings,” Opt. Express14(4), 1339–1352 (2006).
[CrossRef] [PubMed]

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

2004 (1)

2002 (1)

M. J. Booth, M. A. Neil, R. Juskaitis, and T. Wilson, “Adaptive aberration correction in a confocal microscope,” Proc. Natl. Acad. Sci. U.S.A.99(9), 5788–5792 (2002).
[CrossRef] [PubMed]

2000 (2)

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

M. A. Neil, M. J. Booth, and T. Wilson, “New modal wave-front sensor: a theoretical analysis,” J. Opt. Soc. Am. A17(6), 1098–1107 (2000).
[CrossRef] [PubMed]

1994 (1)

Andilla, J.

Andrei, M. A.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Auksorius, E.

Azucena, O.

Bates, M.

M. J. Rust, M. Bates, and X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods3(10), 793–796 (2006).
[CrossRef] [PubMed]

Beaurepaire, E.

Berning, S.

S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
[CrossRef] [PubMed]

Betzig, E.

N. Ji, D. E. Milkie, and E. Betzig, “Adaptive optics via pupil segmentation for high-resolution imaging in biological tissues,” Nat. Methods7(2), 141–147 (2010).
[CrossRef] [PubMed]

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Bewersdorf, J.

T. J. Gould, J. R. Myers, and J. Bewersdorf, “Total internal reflection STED microscopy,” Opt. Express19(14), 13351–13357 (2011).
[CrossRef] [PubMed]

D. Toomre and J. Bewersdorf, “A new wave of cellular imaging,” Annu. Rev. Cell Dev. Biol.26(1), 285–314 (2010).
[CrossRef] [PubMed]

Bonifacino, J. S.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Booth, M.

Booth, M. J.

Boruah, B. R.

Botcherby, E. J.

Cella Zanacchi, F.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

Chen, D. C.

Cheng, Y.

Ciepielewski, D.

Colón-Ramos, D. A.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Dahan, M.

Darzacq, X.

Davidson, M. W.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Débarre, D.

Del Bue, A.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

Deng, S.

Denk, W.

M. Rueckel, J. A. Mack-Bucher, and W. Denk, “Adaptive wavefront correction in two-photon microscopy using coherence-gated wavefront sensing,” Proc. Natl. Acad. Sci. U.S.A.103(46), 17137–17142 (2006).
[CrossRef] [PubMed]

Diaspro, A.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

Dibaj, P.

S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
[CrossRef] [PubMed]

Ding, J. B.

J. B. Ding, K. T. Takasaki, and B. L. Sabatini, “Supraresolution imaging in brain slices using stimulated-emission depletion two-photon laser scanning microscopy,” Neuron63(4), 429–437 (2009).
[CrossRef] [PubMed]

Donnert, G.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Dunsby, C.

Dyba, M.

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

Eggeling, C.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Egner, A.

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

El Beheiry, M.

Engelhardt, J.

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

Facomprez, A.

Faretta, M.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

French, P. M.

Fu, M.

Furia, L.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

Girirajan, T. P.

S. T. Hess, T. P. Girirajan, and M. D. Mason, “Ultra-high resolution imaging by fluorescence photoactivation localization microscopy,” Biophys. J.91(11), 4258–4272 (2006).
[CrossRef] [PubMed]

Gould, T. J.

Hein, B.

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

Hell, S. W.

S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
[CrossRef] [PubMed]

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
[CrossRef] [PubMed]

S. W. Hell, “Microscopy and its focal switch,” Nat. Methods6(1), 24–32 (2009).
[CrossRef] [PubMed]

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

S. W. Hell and J. Wichmann, “Breaking the diffraction resolution limit by stimulated emission: stimulated-emission-depletion fluorescence microscopy,” Opt. Lett.19(11), 780–782 (1994).
[CrossRef] [PubMed]

Hess, H. F.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Hess, S. T.

S. T. Hess, T. P. Girirajan, and M. D. Mason, “Ultra-high resolution imaging by fluorescence photoactivation localization microscopy,” Biophys. J.91(11), 4258–4272 (2006).
[CrossRef] [PubMed]

Izeddin, I.

Jahn, R.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Jakobs, S.

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

Ji, N.

N. Ji, D. E. Milkie, and E. Betzig, “Adaptive optics via pupil segmentation for high-resolution imaging in biological tissues,” Nat. Methods7(2), 141–147 (2010).
[CrossRef] [PubMed]

Juskaitis, R.

M. J. Booth, M. A. Neil, R. Juskaitis, and T. Wilson, “Adaptive aberration correction in a confocal microscope,” Proc. Natl. Acad. Sci. U.S.A.99(9), 5788–5792 (2002).
[CrossRef] [PubMed]

Keller, J.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Kellner, R. R.

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

Kennedy, G.

Klar, T. A.

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

Kubby, J.

Lanigan, P. M.

Lavagnino, Z.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

Li, R.

Lindwasser, O. W.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Lippincott-Schwartz, J.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Liu, L.

Lührmann, R.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Mack-Bucher, J. A.

M. Rueckel, J. A. Mack-Bucher, and W. Denk, “Adaptive wavefront correction in two-photon microscopy using coherence-gated wavefront sensing,” Proc. Natl. Acad. Sci. U.S.A.103(46), 17137–17142 (2006).
[CrossRef] [PubMed]

Mason, M. D.

S. T. Hess, T. P. Girirajan, and M. D. Mason, “Ultra-high resolution imaging by fluorescence photoactivation localization microscopy,” Biophys. J.91(11), 4258–4272 (2006).
[CrossRef] [PubMed]

Medda, R.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

Milkie, D. E.

N. Ji, D. E. Milkie, and E. Betzig, “Adaptive optics via pupil segmentation for high-resolution imaging in biological tissues,” Nat. Methods7(2), 141–147 (2010).
[CrossRef] [PubMed]

Moneron, G.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Myers, J. R.

Nägerl, U. V.

N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
[CrossRef] [PubMed]

Neil, M. A.

Nelson, J. C.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Olenych, S.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Patterson, G. H.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Perrone Donnorso, M.

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

Rankin, B. R.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Rizzoli, S. O.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

Rueckel, M.

M. Rueckel, J. A. Mack-Bucher, and W. Denk, “Adaptive wavefront correction in two-photon microscopy using coherence-gated wavefront sensing,” Proc. Natl. Acad. Sci. U.S.A.103(46), 17137–17142 (2006).
[CrossRef] [PubMed]

Rust, M. J.

M. J. Rust, M. Bates, and X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods3(10), 793–796 (2006).
[CrossRef] [PubMed]

Sabatini, B. L.

J. B. Ding, K. T. Takasaki, and B. L. Sabatini, “Supraresolution imaging in brain slices using stimulated-emission depletion two-photon laser scanning microscopy,” Neuron63(4), 429–437 (2009).
[CrossRef] [PubMed]

Schmidt, R.

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

Schroeder, J.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Schwarzer, D.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Schwertner, M.

Sougrat, R.

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

Srinivas, S.

Steffens, H.

S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
[CrossRef] [PubMed]

Takasaki, K. T.

J. B. Ding, K. T. Takasaki, and B. L. Sabatini, “Supraresolution imaging in brain slices using stimulated-emission depletion two-photon laser scanning microscopy,” Neuron63(4), 429–437 (2009).
[CrossRef] [PubMed]

Tao, X. D.

Thayil, A.

A. Thayil and M. J. Booth, “Self calibration of sensorless adaptive optical microscopes,” J. Eur. Opt. Soc.6, 11045 (2011).
[CrossRef]

Toomre, D.

D. Toomre and J. Bewersdorf, “A new wave of cellular imaging,” Annu. Rev. Cell Dev. Biol.26(1), 285–314 (2010).
[CrossRef] [PubMed]

Urban, N. T.

N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
[CrossRef] [PubMed]

Walter, A.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

Watanabe, T.

Wichmann, J.

Willig, K. I.

S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
[CrossRef] [PubMed]

N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
[CrossRef] [PubMed]

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

Wilson, T.

Wurm, C. A.

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

Xu, Z.

Zhuang, X.

M. J. Rust, M. Bates, and X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods3(10), 793–796 (2006).
[CrossRef] [PubMed]

Zuo, Y.

Annu. Rev. Cell Dev. Biol. (1)

D. Toomre and J. Bewersdorf, “A new wave of cellular imaging,” Annu. Rev. Cell Dev. Biol.26(1), 285–314 (2010).
[CrossRef] [PubMed]

Biophys. J. (3)

S. T. Hess, T. P. Girirajan, and M. D. Mason, “Ultra-high resolution imaging by fluorescence photoactivation localization microscopy,” Biophys. J.91(11), 4258–4272 (2006).
[CrossRef] [PubMed]

N. T. Urban, K. I. Willig, S. W. Hell, and U. V. Nägerl, “STED Nanoscopy of Actin Dynamics in Synapses Deep Inside Living Brain Slices,” Biophys. J.101(5), 1277–1284 (2011).
[CrossRef] [PubMed]

B. R. Rankin, G. Moneron, C. A. Wurm, J. C. Nelson, A. Walter, D. Schwarzer, J. Schroeder, D. A. Colón-Ramos, and S. W. Hell, “Nanoscopy in a living multicellular organism expressing GFP,” Biophys. J.100(12), L63–L65 (2011).
[CrossRef] [PubMed]

J. Eur. Opt. Soc. (1)

A. Thayil and M. J. Booth, “Self calibration of sensorless adaptive optical microscopes,” J. Eur. Opt. Soc.6, 11045 (2011).
[CrossRef]

J. Opt. Soc. Am. A (1)

Nat. Methods (6)

F. Cella Zanacchi, Z. Lavagnino, M. Perrone Donnorso, A. Del Bue, L. Furia, M. Faretta, and A. Diaspro, “Live-cell 3D super-resolution imaging in thick biological samples,” Nat. Methods8(12), 1047–1049 (2011).
[CrossRef] [PubMed]

K. I. Willig, R. R. Kellner, R. Medda, B. Hein, S. Jakobs, and S. W. Hell, “Nanoscale resolution in GFP-based microscopy,” Nat. Methods3(9), 721–723 (2006).
[CrossRef] [PubMed]

M. J. Rust, M. Bates, and X. Zhuang, “Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM),” Nat. Methods3(10), 793–796 (2006).
[CrossRef] [PubMed]

S. W. Hell, “Microscopy and its focal switch,” Nat. Methods6(1), 24–32 (2009).
[CrossRef] [PubMed]

R. Schmidt, C. A. Wurm, S. Jakobs, J. Engelhardt, A. Egner, and S. W. Hell, “Spherical nanosized focal spot unravels the interior of cells,” Nat. Methods5(6), 539–544 (2008).
[CrossRef] [PubMed]

N. Ji, D. E. Milkie, and E. Betzig, “Adaptive optics via pupil segmentation for high-resolution imaging in biological tissues,” Nat. Methods7(2), 141–147 (2010).
[CrossRef] [PubMed]

Neuron (1)

J. B. Ding, K. T. Takasaki, and B. L. Sabatini, “Supraresolution imaging in brain slices using stimulated-emission depletion two-photon laser scanning microscopy,” Neuron63(4), 429–437 (2009).
[CrossRef] [PubMed]

Opt. Express (6)

Opt. Lett. (4)

Proc. Natl. Acad. Sci. U.S.A. (4)

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, “Macromolecular-scale resolution in biological fluorescence microscopy,” Proc. Natl. Acad. Sci. U.S.A.103(31), 11440–11445 (2006).
[CrossRef] [PubMed]

M. Rueckel, J. A. Mack-Bucher, and W. Denk, “Adaptive wavefront correction in two-photon microscopy using coherence-gated wavefront sensing,” Proc. Natl. Acad. Sci. U.S.A.103(46), 17137–17142 (2006).
[CrossRef] [PubMed]

T. A. Klar, S. Jakobs, M. Dyba, A. Egner, and S. W. Hell, “Fluorescence microscopy with diffraction resolution barrier broken by stimulated emission,” Proc. Natl. Acad. Sci. U.S.A.97(15), 8206–8210 (2000).
[CrossRef] [PubMed]

M. J. Booth, M. A. Neil, R. Juskaitis, and T. Wilson, “Adaptive aberration correction in a confocal microscope,” Proc. Natl. Acad. Sci. U.S.A.99(9), 5788–5792 (2002).
[CrossRef] [PubMed]

Science (2)

E. Betzig, G. H. Patterson, R. Sougrat, O. W. Lindwasser, S. Olenych, J. S. Bonifacino, M. W. Davidson, J. Lippincott-Schwartz, and H. F. Hess, “Imaging intracellular fluorescent proteins at nanometer resolution,” Science313(5793), 1642–1645 (2006).
[CrossRef] [PubMed]

S. Berning, K. I. Willig, H. Steffens, P. Dibaj, and S. W. Hell, “Nanoscopy in a living mouse brain,” Science335(6068), 551 (2012).
[CrossRef] [PubMed]

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Figures (5)

Fig. 1
Fig. 1

Schematic of AO STED setup. FI - Faraday isolator; GLP - Glan laser polarizer; GB - glass block; DS - delay stage; AOM - acousto-optical modulator; PMF - polarization-maintaining fiber; SLM1/2 - spatial light modulators; PC - photonic crystal fiber; AOTF - acousto-optical tunable filter; PBS - polarizing beam splitter cube; λ/2 - half-wave plate; λ/4 - quarter-wave plate; DM1/2 - dichroic mirrors; F - bandpass filter; MMF - multimode fiber; APD - avalanche photodiode; OBJ - objective lens; xyz - 3 axes piezo sample stage. Insets show typical phase patterns for each SLM where phase contributions include a baseline flatness correction for the SLM (provided by the manufacturer), a circular blazed grating (for off-axis phase modulation) that defines the active area, correction for system-induced aberration, and a central λ/2 phase mask (SLM1 only).

Fig. 2
Fig. 2

Metric curves as a function of the SLM-applied aberration for correction in the depletion beam path when using a central λ/2 phase mask. Example curves are shown for Zernike modes 5 (astigmatism), 7 (coma), 9 (trefoil), 11 (1st spherical), 22 (2nd spherical), and 37 (3rd spherical) including sharpness (S; green lines), brightness (B; black lines), and combined (M; red lines) metrics. A prime (e.g. Z11) indicates that a particular mode was corrected to be displacement free. Insets show corresponding normalized (-π to + π) phase distributions in the objective back aperture. Data was obtained by imaging 200 nm fluorescent beads in STED mode.

Fig. 3
Fig. 3

Adaptive correction of residual system aberrations. (A, B) STED PSF using the central λ/2 phase mask before (A) and after (B) correction for system aberrations with corresponding phase patterns used on SLM2. (C, D) xz STED images of 100 nm fluorescent bead attached to coverglass and imaged (C) before and (D) after correction of system aberrations. (E) Axial line profiles of pixels summed across horizontal dimension of dashed boxes in (C) and (D). Fitted curves give FWHM of 280 nm (Gaussian fit) for the uncorrected STED image and 140 nm (Lorentzian fit) when adaptive optics is used to compensate for system-induced aberrations in the STED beam path.

Fig. 4
Fig. 4

Images of 100 nm fluorescent beads through ~55 µm layer of glycerol. xz images were acquired in confocal mode (A) before and (B) after correction of sample induced aberrations, and STED mode with (C) correction to the excitation beam only and (D) correction to excitation and depletion beam paths. (E) Axial line profiles of pixels summed across horizontal dimension of dashed boxes in (C) in (D) show improvement in both resolution and signal when aberrations are corrected in the STED beam path.

Fig. 5
Fig. 5

AO STED images of fluorescent beads through zebrafish retina sections. (A-F) Results for beads imaged though ~14 µm of retina. Lateral and axial sections of a single fluorescent bead imaged in (A, D) confocal, (B, E) STED, and (C, F) AO STED show improvement in signal and resolution when adaptive aberration correction is applied to the depletion beam path. (G-L) Similar image sequences for beads imaged through ~25 µm of retina. Axial profiles of beads in AO STED images were ~208 nm and ~249 nm for (F) and (L), respectively. Color bar in (E) also applies to (B), (C), and (F). Color bar in (K) also applies to (H), (I), and (L). (M-O) Volume renderings for data shown in (A-F) for (M) confocal, (N) STED, and (O) AO STED data. (N) and (O) plotted on same color scale for comparison of signal.

Tables (2)

Tables Icon

Table 1 Definition of Zernike Modes, Zi

Tables Icon

Table 2 Couplings Between Displacement and Aberration Zernike Modes

Equations (3)

Equations on this page are rendered with MathJax. Learn more.

M=S+σβB[ 1 1+ e k( S S T ) ]
S= n,m μ n,m I ^ n,m ( n 2 + m 2 ) / n,m I ^ n,m
μ n,m ={ 1 n 2 + m 2 w 0 n 2 + m 2 >w

Metrics