Abstract

Electrically tunable lenses are becoming a widely used optical tool, and have brought significant innovation to microscopy methods. One current limitation of such systems is the difficulty of directly monitor the focal change in real time. Affordable and reliable feedback for such lenses, compatible with any microscopy setup, represents a much-needed improvement that is still not widely available. We discuss here the implementation and technical performance of an optical device to measure with a high frequency response the displacement of the focal offset of a commercial tunable lens with a precision in the range of the axial Point Spread Function (PSF) of the microscope. The technology presented is cost effective and can be employed on any microscopy setup.

© 2016 Optical Society of America

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References

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  1. B. F. Grewe, F. F. Voigt, M. van ’t Hoff, and F. Helmchen, “Fast two-layer two-photon imaging of neuronal cell populations using an electrically tunable lens,” Biomed. Opt. Express 2(7), 2035–2046 (2011).
    [Crossref] [PubMed]
  2. P. Annibale, A. Dvornikov, and E. Gratton, “Electrically tunable lens speeds up 3D orbital tracking,” Biomed. Opt. Express 6(6), 2181–2190 (2015).
    [Crossref] [PubMed]
  3. M. Duocastella, G. Vicidomini, and A. Diaspro, “Simultaneous multiplane confocal microscopy using acoustic tunable lenses,” Opt. Express 22(16), 19293–19301 (2014).
    [Crossref] [PubMed]
  4. Optotune Focus Tunable Lenses, http://www.optotune.com/products/focus-tunable-lenses
  5. S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).
  6. R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
    [Crossref] [PubMed]
  7. V. Crosignani, A. S. Dvornikov, and E. Gratton, “Enhancement of imaging depth in turbid media using a wide area detector,” J. Biophotonics 4(9), 592–599 (2011).
    [PubMed]
  8. F. O. Fahrbach, F. F. Voigt, B. Schmid, F. Helmchen, and J. Huisken, “Rapid 3D light-sheet microscopy with a tunable lens,” Opt. Express 21(18), 21010–21026 (2013).
    [Crossref] [PubMed]

2015 (2)

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

P. Annibale, A. Dvornikov, and E. Gratton, “Electrically tunable lens speeds up 3D orbital tracking,” Biomed. Opt. Express 6(6), 2181–2190 (2015).
[Crossref] [PubMed]

2014 (2)

M. Duocastella, G. Vicidomini, and A. Diaspro, “Simultaneous multiplane confocal microscopy using acoustic tunable lenses,” Opt. Express 22(16), 19293–19301 (2014).
[Crossref] [PubMed]

S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).

2013 (1)

2011 (2)

V. Crosignani, A. S. Dvornikov, and E. Gratton, “Enhancement of imaging depth in turbid media using a wide area detector,” J. Biophotonics 4(9), 592–599 (2011).
[PubMed]

B. F. Grewe, F. F. Voigt, M. van ’t Hoff, and F. Helmchen, “Fast two-layer two-photon imaging of neuronal cell populations using an electrically tunable lens,” Biomed. Opt. Express 2(7), 2035–2046 (2011).
[Crossref] [PubMed]

Annibale, P.

Aravind, A.

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

Aschwanden, M.

S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).

Blum, M.

S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).

Bueeler, M.

S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).

Casutt, S.

S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).

Crosignani, V.

V. Crosignani, A. S. Dvornikov, and E. Gratton, “Enhancement of imaging depth in turbid media using a wide area detector,” J. Biophotonics 4(9), 592–599 (2011).
[PubMed]

Diaspro, A.

Duocastella, M.

Dvornikov, A.

Dvornikov, A. S.

V. Crosignani, A. S. Dvornikov, and E. Gratton, “Enhancement of imaging depth in turbid media using a wide area detector,” J. Biophotonics 4(9), 592–599 (2011).
[PubMed]

Fahrbach, F. O.

Galland, R.

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

Gratton, E.

P. Annibale, A. Dvornikov, and E. Gratton, “Electrically tunable lens speeds up 3D orbital tracking,” Biomed. Opt. Express 6(6), 2181–2190 (2015).
[Crossref] [PubMed]

V. Crosignani, A. S. Dvornikov, and E. Gratton, “Enhancement of imaging depth in turbid media using a wide area detector,” J. Biophotonics 4(9), 592–599 (2011).
[PubMed]

Grenci, G.

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

Grewe, B. F.

Helmchen, F.

Huisken, J.

Schmid, B.

Sibarita, J. B.

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

Studer, V.

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

van ’t Hoff, M.

Viasnoff, V.

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

Vicidomini, G.

Voigt, F. F.

Biomed. Opt. Express (2)

J. Biophotonics (1)

V. Crosignani, A. S. Dvornikov, and E. Gratton, “Enhancement of imaging depth in turbid media using a wide area detector,” J. Biophotonics 4(9), 592–599 (2011).
[PubMed]

Nat. Methods (1)

R. Galland, G. Grenci, A. Aravind, V. Viasnoff, V. Studer, and J. B. Sibarita, “3D high- and super-resolution imaging using single-objective SPIM,” Nat. Methods 12(7), 641–644 (2015).
[Crossref] [PubMed]

Opt. Express (2)

Proc. SPIE 8982, Optical Components and Materials (1)

S. Casutt, M. Bueeler, M. Blum, and M. Aschwanden, “Fast and precise continuous focusing with focus tunable lenses,” Proc. SPIE 8982, Optical Components and Materials XI, 89820Y (2014).

Other (1)

Optotune Focus Tunable Lenses, http://www.optotune.com/products/focus-tunable-lenses

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Figures (4)

Fig. 1
Fig. 1 Diagram of the configurations tested to implement a feedback module fitting a commercial ETL a) One fiber is used to provide excitation light on the surface of the ETL. Two stacked fibers are used in detection, to achieve a differential measurement. Orange: plastic ring fitting an ETL custom holder on the side where it threads into any RMS microscope turret. b) Retro-reflected configuration: one excitation and one detection fibers are used, on the same side. The collection fiber measures changes in intensity due to defocus. c) (top) Picture of the electrical lens setup with two optical fibers reaching into the case. (bottom) Plastic ring fitting an ETL custom holder on the side where it threads into a RMS microscope turret with two fibers entering at 45deg. e) (top) Picture of the arrangement illustrated in b. The reflecting surface (aluminum foil here) on the lens side, opposite to the fibers, is clearly visible (bottom) picture of a 500 µm fiber cut at 45deg and polished.
Fig. 2
Fig. 2 Feedback signal measured for two of the configurations discussed in Fig. 1. a) Change in focal distance vs Voltage applied to the ETL for three different objectives. b) (top) Representation of the PSF of the microscope using a 20x 0.4NA objective. The approximate axial waist is 1 μm. (bottom) The graph shows the changes in the measured signal using the differential configuration described in Fig. 2(a). Each point is obtained by averaging the signal over an acquisition of 20s (0.05 Hz). c) Signal measured in reflected transmission geometry while changing the focal distance of the ETL in steps covering over 400 μm (using a 20x, 0.4 NA objective). The linear fit has a slope of −0.5 mV/mA. d) Enlarged view of the linear region of detector output vs lens current. Steps in lens current of 2 mA (corresponding to about 7 μm axial displacement) are clearly resolvable.
Fig. 3
Fig. 3 a) Stability of lens focal offset signal as a function of time upon modulation (8 Hz) of the lens focal offset with a square wave of 2.7 mA amplitude. b) Enlarged view of a portion of the axial modulation as a function of time acquired at a frequency of 60 Hz, and c) 15 Hz.
Fig. 4
Fig. 4 Measurement of the focal plane displacement using the add-on in a retro-reflected configuration in a microscope. a) Signal measured in reflected transmission geometry while changing the focal distance of the ETL in steps covering over 300 μm (using a 20x, 0.4 NA objective). Error bars are given respectively by the standard deviation of the measured PMT signal (x) and by half the axial PSF width (y). As the current to the lens is changed, the sample is refocused using a stepper motor with μm-step sensitivity, providing the focal offset values in the x-axis. Blue solid markers indicate the measured position signal upon re-focusing the sample (image insets) at two selected focal offsets. (A 1mV offset is arbitrarily introduced in the plot to avoid complete overlap). b) Microscope objective (20x, 0.4 NA) mounted on the ETL housing containing the position sensitive device as illustrated in Fig. 1(e). The red arrows indicate the input and output optical fibers respectively.

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