Abstract

We theoretically demonstrate that the spatial resolution of stimulated emission depletion (STED) microscopy can be substantially enhanced without increasing the intensity of the STED beam. In our scheme, tiny nanobeads codoped with donor and acceptor molecules are used as fluorescent probes, in which Förster resonance energy transfer (FRET) can occur with an 100% efficiency between the donors and acceptors. Enhancement of the depletion of acceptors in the nanobeads with the doughnut-shaped depletion beam can lead to an increase of FRET efficiency in the outer area of the excitation spot, which itself is used for deexciting donor molecules and, consequently, enhancing the optical resolution.

© 2010 Optical Society of America

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    [CrossRef] [PubMed]
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2010

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

2009

2008

B. Huang, S. A. Jones, B. Brandenburg, and X. W. Zhuang, Nat. Meth. 5, 1047 (2008).
[CrossRef]

2007

S. W. Hell, Science 316, 1153 (2007).
[CrossRef] [PubMed]

K. I. Willig, B. Harke, R. Medda, and S. W. Hell, Nat. Meth. 4, 915 (2007).
[CrossRef]

2006

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

W. E. Moerner, Nat. Meth. 3, 781 (2006).
[CrossRef]

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, Science 313, 1642 (2006).
[CrossRef] [PubMed]

S. T. Hess, T. P. K. Girirajan, and M. D. Mason, Biophys. J. 91, 4258 (2006).
[CrossRef] [PubMed]

M. J. Rust, M. Bates, and X. W. Zhuang, Nat. Meth. 3, 793(2006).
[CrossRef]

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

2005

M. G. L. Gustafsson, Proc. Natl. Acad. Sci. USA 102, 13081 (2005).
[CrossRef] [PubMed]

1994

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, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

Bates, M.

M. J. Rust, M. Bates, and X. W. Zhuang, Nat. Meth. 3, 793(2006).
[CrossRef]

Betzig, E.

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, Science 313, 1642 (2006).
[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, Science 313, 1642 (2006).
[CrossRef] [PubMed]

Brand, L.

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

Brandenburg, B.

B. Huang, S. A. Jones, B. Brandenburg, and X. W. Zhuang, Nat. Meth. 5, 1047 (2008).
[CrossRef]

Chen, J.

Cheng, Y.

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, Science 313, 1642 (2006).
[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, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[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, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

Felekyan, S.

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

Forthmann, C.

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

Girirajan, T. P. K.

S. T. Hess, T. P. K. Girirajan, and M. D. Mason, Biophys. J. 91, 4258 (2006).
[CrossRef] [PubMed]

Gustafsson, M. G. L.

M. G. L. Gustafsson, Proc. Natl. Acad. Sci. USA 102, 13081 (2005).
[CrossRef] [PubMed]

Harke, B.

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

K. I. Willig, B. Harke, R. Medda, and S. W. Hell, Nat. Meth. 4, 915 (2007).
[CrossRef]

Hell, S. W.

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

S. W. Hell, Science 316, 1153 (2007).
[CrossRef] [PubMed]

K. I. Willig, B. Harke, R. Medda, and S. W. Hell, Nat. Meth. 4, 915 (2007).
[CrossRef]

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

S. W. Hell and J. Wichmann, Opt. Lett. 19, 780 (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, Science 313, 1642 (2006).
[CrossRef] [PubMed]

Hess, S. T.

S. T. Hess, T. P. K. Girirajan, and M. D. Mason, Biophys. J. 91, 4258 (2006).
[CrossRef] [PubMed]

Huang, B.

B. Huang, S. A. Jones, B. Brandenburg, and X. W. Zhuang, Nat. Meth. 5, 1047 (2008).
[CrossRef]

Jahn, R.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

Jones, S. A.

B. Huang, S. A. Jones, B. Brandenburg, and X. W. Zhuang, Nat. Meth. 5, 1047 (2008).
[CrossRef]

Kasper, R.

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[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, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

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, Science 313, 1642 (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, Science 313, 1642 (2006).
[CrossRef] [PubMed]

Lührmann, R.

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

Mason, M. D.

S. T. Hess, T. P. K. Girirajan, and M. D. Mason, Biophys. J. 91, 4258 (2006).
[CrossRef] [PubMed]

Medda, R.

K. I. Willig, B. Harke, R. Medda, and S. W. Hell, Nat. Meth. 4, 915 (2007).
[CrossRef]

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

Moerner, W. E.

W. E. Moerner, Nat. Meth. 3, 781 (2006).
[CrossRef]

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, Science 313, 1642 (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, Science 313, 1642 (2006).
[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, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

Rust, M. J.

M. J. Rust, M. Bates, and X. W. Zhuang, Nat. Meth. 3, 793(2006).
[CrossRef]

Sauer, M.

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

Schaffer, J.

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

Seidel, C. A. M.

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

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, Science 313, 1642 (2006).
[CrossRef] [PubMed]

Tinnefeld, P.

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

Wichmann, J.

Widengren, J.

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

Willig, K. I.

K. I. Willig, B. Harke, R. Medda, and S. W. Hell, Nat. Meth. 4, 915 (2007).
[CrossRef]

Zhuang, X. W.

B. Huang, S. A. Jones, B. Brandenburg, and X. W. Zhuang, Nat. Meth. 5, 1047 (2008).
[CrossRef]

M. J. Rust, M. Bates, and X. W. Zhuang, Nat. Meth. 3, 793(2006).
[CrossRef]

Biophys. J.

S. T. Hess, T. P. K. Girirajan, and M. D. Mason, Biophys. J. 91, 4258 (2006).
[CrossRef] [PubMed]

J. Phys. Chem. A.

C. Eggeling, J. Widengren, L. Brand, J. Schaffer, S. Felekyan, and C. A. M. Seidel, J. Phys. Chem. A. 110, 2979 (2006).
[CrossRef] [PubMed]

Nat. Meth.

K. I. Willig, B. Harke, R. Medda, and S. W. Hell, Nat. Meth. 4, 915 (2007).
[CrossRef]

M. J. Rust, M. Bates, and X. W. Zhuang, Nat. Meth. 3, 793(2006).
[CrossRef]

B. Huang, S. A. Jones, B. Brandenburg, and X. W. Zhuang, Nat. Meth. 5, 1047 (2008).
[CrossRef]

W. E. Moerner, Nat. Meth. 3, 781 (2006).
[CrossRef]

Opt. Lett.

Proc. Natl. Acad. Sci. USA

M. G. L. Gustafsson, Proc. Natl. Acad. Sci. USA 102, 13081 (2005).
[CrossRef] [PubMed]

G. Donnert, J. Keller, R. Medda, M. A. Andrei, S. O. Rizzoli, R. Lührmann, R. Jahn, C. Eggeling, and S. W. Hell, Proc. Natl. Acad. Sci. USA 103, 11440 (2006).
[CrossRef] [PubMed]

Science

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, Science 313, 1642 (2006).
[CrossRef] [PubMed]

S. W. Hell, Science 316, 1153 (2007).
[CrossRef] [PubMed]

Small

R. Kasper, B. Harke, C. Forthmann, P. Tinnefeld, S. W. Hell, and M. Sauer, Small 6, 1379 (2010).
[CrossRef] [PubMed]

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Figures (3)

Fig. 1
Fig. 1

(a) Energy diagram illustrating the concept of FASTED. The excited donor molecules are switched off by the resonance energy transfer between the donor and acceptor molecules. (b) Excitation and depletion wavelengths in FASTED microscopy together with the excitation (solid curve) and emission (dashed curve) spectra of the Cy3–Cy5 FRET pair. The wavelength range for the fluorescence detection in FASTED microscopy is indicated by the gray-filled region.

Fig. 2
Fig. 2

Normalized PSFs on the focal plane for the (a) conventional confocal, (b) SFM, (c) CW-STED, and (d) FASTED microscopes. The reduction factor f max , which is obtained by dividing the fluorescence intensity at the center of each PSF by that at the center of the PSF of the confocal microscope, is presented in each subfigure. The x axis unit is the excitation wavelength λ exc = 510 nm .

Fig. 3
Fig. 3

(a) FWHMs of FASTED (hollow red circles) and CW-STED microscopes (filled blue squares) as functions of the saturation factor, illustrating a significant resolution gain in FASTED over CW-STED microscopy. The reduction factor for FASTED is f max = 0.48 . (b) The reduction factor f max versus the FWHMs of the PSFs of the FASTED microscope at the saturation factors of ζ = 50 (filled blue squares) and ζ = 100 (hollow red circles).

Equations (2)

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{ d n D E d t = n D G k exc n D E k D min ( n A G , n D E ) k T d n A E d t = min ( n A G , n D E ) k T n A E ( k A + k s ) ,
{ n D E = k exc k D + k exc ( k A + k s ) k T ( k D + k exc ) ( k A + k s + k T ) , k exc ( k A + k s ) ( k D + k T ) k T n D E = k exc k D + k exc + k T , k exc ( k A + k s ) ( k D + k T ) k T .

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