Abstract

Adaptive optics has been used to compensate the detrimental effects of aberrations in a range of high-resolution microscopes. We investigate how backscattered laser illumination can be used as the source for direct wavefront sensing using a pinhole-filtered Shack–Hartmann wavefront sensor. It is found that the sensor produces linear response to input aberrations for a given specimen. The gradient of this response is dependent upon experimental configuration and specimen structure. Cross sensitivity between modes is also observed. The double pass nature of the microscope system leads in general to lower sensitivity to odd-symmetry aberration modes. The results show that there is potential for use of this type of wavefront sensing in microscopes.

© 2013 Optical Society of America

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References

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2012 (2)

S. A. Rahman and M. J. Booth, “Adaptive optics for high-resolution microscopy: wave front sensing using back scattered light,” Proc. SPIE 8253, 82530I (2012).
[CrossRef]

X. Tao, J. Crest, S. Kotadia, O. Azucena, D. C. Chen, W. Sullivan, and J. Kubby, “Live imaging using adaptive optics with fluorescent protein guide-stars,” Opt. Express 20, 15969–15982 (2012).
[CrossRef]

2011 (4)

2010 (1)

J. W. Cha, J. Ballesta, and P. T. C. So, “Shack-Hartmann wavefront-sensor-based adaptive optics system for multiphoton microscopy,” J. Biomed. Opt. 15, 046022 (2010).
[CrossRef]

2009 (2)

2007 (1)

M. J. Booth, “Adaptive optics in microscopy,” Phil. Trans. R. Soc. A 365, 2829–2843 (2007).
[CrossRef]

2006 (1)

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. USA 103, 17137–17142 (2006).
[CrossRef]

2005 (1)

2001 (1)

1997 (1)

T. Wilson, R. Juškaitis, and P. Higdon, “The imaging of dielectric point scatterers in conventional and confocal polarization microscopes,” Opt. Commun. 141, 298–313 (1997).
[CrossRef]

1995 (3)

1980 (1)

Andilla, J.

Artal, P.

Artigas, D.

Aviles-Espinosa, R.

Azucena, O.

Ballesta, J.

J. W. Cha, J. Ballesta, and P. T. C. So, “Shack-Hartmann wavefront-sensor-based adaptive optics system for multiphoton microscopy,” J. Biomed. Opt. 15, 046022 (2010).
[CrossRef]

Booth, M.

Booth, M. J.

S. A. Rahman and M. J. Booth, “Adaptive optics for high-resolution microscopy: wave front sensing using back scattered light,” Proc. SPIE 8253, 82530I (2012).
[CrossRef]

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, 2495–2497 (2009).
[CrossRef]

M. J. Booth, “Adaptive optics in microscopy,” Phil. Trans. R. Soc. A 365, 2829–2843 (2007).
[CrossRef]

M. J. Booth, T. Wilson, H. Sun, T. Ota, and S. Kawata, “Methods for the characterization of deformable membrane mirrors,” Appl. Opt. 44, 5131–5139 (2005).
[CrossRef]

Botcherby, E. J.

Cha, J. W.

J. W. Cha, J. Ballesta, and P. T. C. So, “Shack-Hartmann wavefront-sensor-based adaptive optics system for multiphoton microscopy,” J. Biomed. Opt. 15, 046022 (2010).
[CrossRef]

Chen, D. C.

Crest, J.

Dainty, J. C.

Débarre, D.

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. USA 103, 17137–17142 (2006).
[CrossRef]

Diaz-Santana, L.

Fernandez, B.

Fragola, A.

P. Vermeulen, E. Muro, T. Pons, V. Loriette, and A. Fragola, “Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers,” J. Biomed. Opt. 16, 076019 (2011).
[CrossRef]

Fu, M.

Garcia, D.

Green, D. G.

Grieve, K.

Gu, M.

M. Gu, Advanced Optical Imaging Theory (Springer, 2000).

Higdon, P.

T. Wilson, R. Juškaitis, and P. Higdon, “The imaging of dielectric point scatterers in conventional and confocal polarization microscopes,” Opt. Commun. 141, 298–313 (1997).
[CrossRef]

Iglesias, I.

Jesacher, A.

Juškaitis, R.

T. Wilson, R. Juškaitis, and P. Higdon, “The imaging of dielectric point scatterers in conventional and confocal polarization microscopes,” Opt. Commun. 141, 298–313 (1997).
[CrossRef]

Kawata, S.

Kotadia, S.

Kubby, J.

Levecq, X.

López-Gil, N.

Loriette, V.

P. Vermeulen, E. Muro, T. Pons, V. Loriette, and A. Fragola, “Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers,” J. Biomed. Opt. 16, 076019 (2011).
[CrossRef]

Losada, M. A.

Loza-Alvarez, P.

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. USA 103, 17137–17142 (2006).
[CrossRef]

Marcos, S.

Muro, E.

P. Vermeulen, E. Muro, T. Pons, V. Loriette, and A. Fragola, “Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers,” J. Biomed. Opt. 16, 076019 (2011).
[CrossRef]

Navarro, R.

Nieto, M.

Olarte, O. E.

Ota, T.

Pons, T.

P. Vermeulen, E. Muro, T. Pons, V. Loriette, and A. Fragola, “Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers,” J. Biomed. Opt. 16, 076019 (2011).
[CrossRef]

Porcar-Guezenec, R.

Rahman, S. A.

S. A. Rahman and M. J. Booth, “Adaptive optics for high-resolution microscopy: wave front sensing using back scattered light,” Proc. SPIE 8253, 82530I (2012).
[CrossRef]

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. USA 103, 17137–17142 (2006).
[CrossRef]

So, P. T. C.

J. W. Cha, J. Ballesta, and P. T. C. So, “Shack-Hartmann wavefront-sensor-based adaptive optics system for multiphoton microscopy,” J. Biomed. Opt. 15, 046022 (2010).
[CrossRef]

Southwell, W. H.

Srinivas, S.

Sullivan, W.

Sun, H.

Tao, X.

Thayil, A.

Vermeulen, P.

P. Vermeulen, E. Muro, T. Pons, V. Loriette, and A. Fragola, “Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers,” J. Biomed. Opt. 16, 076019 (2011).
[CrossRef]

Watanabe, T.

Williams, D. R.

Wilson, T.

Zuo, Y.

Appl. Opt. (1)

Biomed. Opt. Express (1)

J. Biomed. Opt. (2)

J. W. Cha, J. Ballesta, and P. T. C. So, “Shack-Hartmann wavefront-sensor-based adaptive optics system for multiphoton microscopy,” J. Biomed. Opt. 15, 046022 (2010).
[CrossRef]

P. Vermeulen, E. Muro, T. Pons, V. Loriette, and A. Fragola, “Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers,” J. Biomed. Opt. 16, 076019 (2011).
[CrossRef]

J. Opt. Soc. Am. (1)

J. Opt. Soc. Am. A (4)

Opt. Commun. (1)

T. Wilson, R. Juškaitis, and P. Higdon, “The imaging of dielectric point scatterers in conventional and confocal polarization microscopes,” Opt. Commun. 141, 298–313 (1997).
[CrossRef]

Opt. Express (1)

Opt. Lett. (4)

Phil. Trans. R. Soc. A (1)

M. J. Booth, “Adaptive optics in microscopy,” Phil. Trans. R. Soc. A 365, 2829–2843 (2007).
[CrossRef]

Proc. Natl. Acad. Sci. USA (1)

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. USA 103, 17137–17142 (2006).
[CrossRef]

Proc. SPIE (1)

S. A. Rahman and M. J. Booth, “Adaptive optics for high-resolution microscopy: wave front sensing using back scattered light,” Proc. SPIE 8253, 82530I (2012).
[CrossRef]

Other (2)

T. Wilson, ed. Confocal Microscopy (Academic, 1990).

M. Gu, Advanced Optical Imaging Theory (Springer, 2000).

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Figures (12)

Fig. 1.
Fig. 1.

Schematic of the microscope wavefront sensing system. Several optical elements have been omitted for clarity. Arrows show the light propagation direction in the illumination and detection paths.

Fig. 2.
Fig. 2.

SHWFS measurements of the DM-induced even aberration modes, (a, b) Z5 (astigmatism) and (c, d) Z11 (spherical), for the mirror specimen. (a, c): Measured aberration amplitude as a function of induced aberration amplitude. (b, d): Maximum measured range of all Zernike modes when ±0.6rad (rms) of Z5 or Z11 was induced, showing crosstalk between modes. The inset in (b) shows the definition of the Zernike modes.

Fig. 3.
Fig. 3.

Percentage of missing SHWFS spots for different pinhole sizes for different amplitudes of modes (a) Z5 and (b) Z11. (c) Cross sections through the Zernike modes Z5 and Z11 with RMS amplitude 0.5 rad for θ=π/2. The average gradient of Z11 is higher, leading to more light loss at the pinhole and more missing spots in the SHWFS.

Fig. 4.
Fig. 4.

Sensitivity to the induced Zernike mode of index i (black) and the total rms crosstalk sensitivity measured in other modes (gray) for Zernike modes 5 to 11 using the mirror specimen for (a) 100 μm diameter pinhole, (b) 300 μm pinhole, and (c) no pinhole.

Fig. 5.
Fig. 5.

(a) Confocal reflection microscope image of the 200 nm bead specimen. (b–d) SHWFS spot patterns using light backscattered from the sample 200 nm beads. A subregion of the pupil is shown. (b) Stationary specimen using a 200 μm pinhole, showing significant intensity variation across pupil. (c) Scanning and frame averaging with no pinhole, showing near uniform intensity but with out-of-focus light causing secondary spots within each subaperture domain. (d) Scanning and averaging, using a 200 μm pinhole, showing greater uniformity and no secondary spots.

Fig. 6.
Fig. 6.

SHWFS measurements of the even aberration modes Z5 (astigmatism) and Z11 (spherical) and the the odd aberration modes Z7 (coma) and Z9 (trefoil) for the specimen of 200 nm diameter beads suspended in gelatin. Left: Measured aberration amplitude as a function of induced aberration amplitude. Right: Maximum measured amplitude of all modes, showing modal cross talk.

Fig. 7.
Fig. 7.

Sensitivity (black) and crosstalk (gray) for Zernike modes 5 to 11 using the specimen of 200 nm diameter beads in gelatin with (a) 200 μm diameter pinhole, (b) 300 μm pinhole, and (c) 600 μm pinhole.

Fig. 8.
Fig. 8.

Confocal reflection microscope image of the artificial collagen specimen.

Fig. 9.
Fig. 9.

Sensitivity (black) and crosstalk (gray) for Zernike modes 5 to 11 using the artificial collagen specimen with (a) 200 μm diameter pinhole, (b) 300 μm pinhole, and (c) 600 μm pinhole.

Fig. 10.
Fig. 10.

Sensitivity (black) and crosstalk (gray) for Zernike modes 5 to 11 using the gold bead specimen, with (a) 300 μm and (b) 600 μm pinhole.

Fig. 11.
Fig. 11.

Asymmetric and polarization-filtered detection. SHWFS spot patterns using asymmetric illumination and detection with the specimen of 200 nm diameter beads in gelatin: (a) stationary specimen, (b) scanned specimen with time averaging. The bright region in the center corresponds to the specular reflection component. Sensitivity (black) and crosstalk (gray) using a 300 μm diameter pinhole: (c) using symmetric illumination/detection, (d) using asymmetric illumination/detection. SHWFS spot patterns using polarization filtering: (e) stationary specimen, (f) scanned specimen with time averaging. Sensitivity (black) and crosstalk (gray) for: (g) no polarization filtering, (h) polarization filtering.

Fig. 12.
Fig. 12.

Sensitivity (black) and crosstalk (gray) for Zernike modes 5 to 11 using the C. Elegans specimen with a 600 μm pinhole. Key: s/a, symmetric/asymmetric illumination and detection; p, polarization filtering. Image: Confocal reflection microscope image of the region of the C. Elegans specimen used for the aberration measurements.

Tables (1)

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Table 1. Condition Numbers of Sensitivity Matrices Shown to Two Significant Figuresa

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cactual=S1cmeasured.
κ=SS1,

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