Abstract

This work presents, to our knowledge, the first demonstration of the Laser Speckle Contrast Imaging (LSCI) technique with extended depth of field (DOF). We employ wavefront coding on the detected beam to gain quantitative information on flow speeds through a DOF extended two-fold compared to the traditional system. We characterize the system in-vitro using controlled microfluidic experiments, and apply it in-vivo to imaging the somatosensory cortex of a rat, showing improved ability to image flow in a larger number of vessels simultaneously.

© 2013 Optical Society of America

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  1. A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
    [CrossRef]
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    [CrossRef] [PubMed]
  3. J. D. Briers and S. Webster, “Laser speckle contrast analysis (lasca): a nonscanning, full-field technique for monitoring capillary blood flow,” J. Biomed. Opt.1, 174–179 (1996).
    [CrossRef] [PubMed]
  4. M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
    [CrossRef]
  5. A. Fercher and J. Briers, “Flow visualization by means of single-exposure speckle photography,” Opt. Comm.37, 326–330 (1981).
    [CrossRef]
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    [CrossRef] [PubMed]
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    [CrossRef]
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    [CrossRef] [PubMed]
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2013 (2)

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Rapid multiexposure in vivo brain imaging system using vertical cavity surface emitting lasers as a light source,” Appl. Opt.52, C64–C71 (2013).
[CrossRef] [PubMed]

S. Quirin and R. Piestun, “Depth estimation and image recovery using broadband, incoherent illumination with engineered point spread functions,” Appl. Opt.52, A367–A376 (2013).
[CrossRef] [PubMed]

2012 (3)

2011 (3)

2010 (1)

D. A. Boas and A. K. Dunn, “Laser speckle contrast imaging in biomedical optics,” J. Biomed. Opt.15, 011109 (2010).
[CrossRef] [PubMed]

2009 (1)

M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
[CrossRef]

2008 (2)

2007 (1)

Q. Yang, L. Liu, and J. Sun, “Optimized phase pupil masks for extended depth of field,” Opt. Comm.272, 56–66 (2007).
[CrossRef]

2005 (1)

2004 (1)

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

2002 (1)

2001 (1)

A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
[CrossRef]

1996 (2)

E. R. Dowski, W. T. Cathey, and S. C. Bradburn, “Aberration invariant optical/digital incoherent systems,” Optical Review3, A429–A432 (1996).
[CrossRef]

J. D. Briers and S. Webster, “Laser speckle contrast analysis (lasca): a nonscanning, full-field technique for monitoring capillary blood flow,” J. Biomed. Opt.1, 174–179 (1996).
[CrossRef] [PubMed]

1981 (1)

A. Fercher and J. Briers, “Flow visualization by means of single-exposure speckle photography,” Opt. Comm.37, 326–330 (1981).
[CrossRef]

1976 (1)

Atchia, Y.

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Rapid multiexposure in vivo brain imaging system using vertical cavity surface emitting lasers as a light source,” Appl. Opt.52, C64–C71 (2013).
[CrossRef] [PubMed]

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Speckle contrast at deviations from best focus in microfluidic and in vivo,” in Biomedical Optics and 3-D Imaging, (Optical Society of America, 2012), p. BTu3A.49.

Berent, J.

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

Boas, D. A.

D. A. Boas and A. K. Dunn, “Laser speckle contrast imaging in biomedical optics,” J. Biomed. Opt.15, 011109 (2010).
[CrossRef] [PubMed]

S. Yuan, A. Devor, D. A. Boas, and A. K. Dunn, “Determination of optimal exposure time for imaging of blood flow changes with laser speckle contrast imaging,” Appl. Opt.44, 1823–1830 (2005).
[CrossRef] [PubMed]

A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
[CrossRef]

Bolay, H.

A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
[CrossRef]

Bradburn, S. C.

E. R. Dowski, W. T. Cathey, and S. C. Bradburn, “Aberration invariant optical/digital incoherent systems,” Optical Review3, A429–A432 (1996).
[CrossRef]

Breckinridge, J.

J. Breckinridge, D. Voelz, and J. B. Breckinridge, Computational Fourier Optics: A MATLAB Tutorial, Tutorial Text Series (SPIE Press, 2011).

Breckinridge, J. B.

J. Breckinridge, D. Voelz, and J. B. Breckinridge, Computational Fourier Optics: A MATLAB Tutorial, Tutorial Text Series (SPIE Press, 2011).

Briers, J.

A. Fercher and J. Briers, “Flow visualization by means of single-exposure speckle photography,” Opt. Comm.37, 326–330 (1981).
[CrossRef]

Briers, J. D.

J. D. Briers and S. Webster, “Laser speckle contrast analysis (lasca): a nonscanning, full-field technique for monitoring capillary blood flow,” J. Biomed. Opt.1, 174–179 (1996).
[CrossRef] [PubMed]

Caravaca-Aguirre, A. M.

Cathey, W. T.

W. T. Cathey and E. R. Dowski, “New paradigm for imaging systems,” Appl. Opt.41, 6080–6092 (2002).
[CrossRef] [PubMed]

E. R. Dowski, W. T. Cathey, and S. C. Bradburn, “Aberration invariant optical/digital incoherent systems,” Optical Review3, A429–A432 (1996).
[CrossRef]

Choi, B.

Conkey, D. B.

Cuccia, D. J.

Devor, A.

Dowski, E. R.

W. T. Cathey and E. R. Dowski, “New paradigm for imaging systems,” Appl. Opt.41, 6080–6092 (2002).
[CrossRef] [PubMed]

E. R. Dowski, W. T. Cathey, and S. C. Bradburn, “Aberration invariant optical/digital incoherent systems,” Optical Review3, A429–A432 (1996).
[CrossRef]

Draijer, M.

M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
[CrossRef]

Dufour, S.

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Rapid multiexposure in vivo brain imaging system using vertical cavity surface emitting lasers as a light source,” Appl. Opt.52, C64–C71 (2013).
[CrossRef] [PubMed]

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Speckle contrast at deviations from best focus in microfluidic and in vivo,” in Biomedical Optics and 3-D Imaging, (Optical Society of America, 2012), p. BTu3A.49.

Duncan, D. D.

Dunn, A. K.

D. A. Boas and A. K. Dunn, “Laser speckle contrast imaging in biomedical optics,” J. Biomed. Opt.15, 011109 (2010).
[CrossRef] [PubMed]

S. Yuan, A. Devor, D. A. Boas, and A. K. Dunn, “Determination of optimal exposure time for imaging of blood flow changes with laser speckle contrast imaging,” Appl. Opt.44, 1823–1830 (2005).
[CrossRef] [PubMed]

A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
[CrossRef]

Durkin, A. J.

Fercher, A.

A. Fercher and J. Briers, “Flow visualization by means of single-exposure speckle photography,” Opt. Comm.37, 326–330 (1981).
[CrossRef]

Fiedler, C.

Forster, B.

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

Gonzalez, R. C.

R. C. Gonzalez and R. E. Woods, Digital Image Processing, 3rd ed. (Prentice-Hall, Inc., Upper Saddle River, NJ, USA, 2006).

Goodman, J. W.

Grover, G.

Hondebrink, E.

M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
[CrossRef]

Kirkpatrick, S. J.

Komatsu, S.

Konecky, S. D.

Leeuwen, T.

M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
[CrossRef]

Levi, O.

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Rapid multiexposure in vivo brain imaging system using vertical cavity surface emitting lasers as a light source,” Appl. Opt.52, C64–C71 (2013).
[CrossRef] [PubMed]

H. Levy, D. Ringuette, and O. Levi, “Rapid monitoring of cerebral ischemia dynamics using laser-based optical imaging of blood oxygenation and flow,” Biomed. Opt. Express3, 777–791 (2012).
[CrossRef] [PubMed]

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Speckle contrast at deviations from best focus in microfluidic and in vivo,” in Biomedical Optics and 3-D Imaging, (Optical Society of America, 2012), p. BTu3A.49.

Levy, H.

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Rapid multiexposure in vivo brain imaging system using vertical cavity surface emitting lasers as a light source,” Appl. Opt.52, C64–C71 (2013).
[CrossRef] [PubMed]

H. Levy, D. Ringuette, and O. Levi, “Rapid monitoring of cerebral ischemia dynamics using laser-based optical imaging of blood oxygenation and flow,” Biomed. Opt. Express3, 777–791 (2012).
[CrossRef] [PubMed]

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Speckle contrast at deviations from best focus in microfluidic and in vivo,” in Biomedical Optics and 3-D Imaging, (Optical Society of America, 2012), p. BTu3A.49.

Li, Y.

P. Miao, H. Lu, Q. Liu, Y. Li, and S. Tong, “Laser speckle contrast imaging of cerebral blood flow in freely moving animals,” J. Biomed. Opt.16, 090502 (2011).
[CrossRef] [PubMed]

Liu, L.

Q. Yang, L. Liu, and J. Sun, “Optimized phase pupil masks for extended depth of field,” Opt. Comm.272, 56–66 (2007).
[CrossRef]

Liu, Q.

P. Miao, H. Lu, Q. Liu, Y. Li, and S. Tong, “Laser speckle contrast imaging of cerebral blood flow in freely moving animals,” J. Biomed. Opt.16, 090502 (2011).
[CrossRef] [PubMed]

Lu, H.

P. Miao, H. Lu, Q. Liu, Y. Li, and S. Tong, “Laser speckle contrast imaging of cerebral blood flow in freely moving animals,” J. Biomed. Opt.16, 090502 (2011).
[CrossRef] [PubMed]

Mazhar, A.

Miao, P.

P. Miao, H. Lu, Q. Liu, Y. Li, and S. Tong, “Laser speckle contrast imaging of cerebral blood flow in freely moving animals,” J. Biomed. Opt.16, 090502 (2011).
[CrossRef] [PubMed]

Moskowitz, M. A.

A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
[CrossRef]

Owen, C.

Piestun, R.

Quirin, S.

Rice, T. B.

Ringuette, D.

Sage, D.

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

Steenbergen, W.

M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
[CrossRef]

Sun, J.

Q. Yang, L. Liu, and J. Sun, “Optimized phase pupil masks for extended depth of field,” Opt. Comm.272, 56–66 (2007).
[CrossRef]

Takahashi, Y.

Tong, S.

P. Miao, H. Lu, Q. Liu, Y. Li, and S. Tong, “Laser speckle contrast imaging of cerebral blood flow in freely moving animals,” J. Biomed. Opt.16, 090502 (2011).
[CrossRef] [PubMed]

Tromberg, B. J.

Unser, M.

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

Van De Ville, D.

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

Voelz, D.

J. Breckinridge, D. Voelz, and J. B. Breckinridge, Computational Fourier Optics: A MATLAB Tutorial, Tutorial Text Series (SPIE Press, 2011).

Webster, S.

J. D. Briers and S. Webster, “Laser speckle contrast analysis (lasca): a nonscanning, full-field technique for monitoring capillary blood flow,” J. Biomed. Opt.1, 174–179 (1996).
[CrossRef] [PubMed]

Woods, R. E.

R. C. Gonzalez and R. E. Woods, Digital Image Processing, 3rd ed. (Prentice-Hall, Inc., Upper Saddle River, NJ, USA, 2006).

Yang, Q.

Q. Yang, L. Liu, and J. Sun, “Optimized phase pupil masks for extended depth of field,” Opt. Comm.272, 56–66 (2007).
[CrossRef]

Yuan, S.

Appl. Opt. (1)

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Rapid multiexposure in vivo brain imaging system using vertical cavity surface emitting lasers as a light source,” Appl. Opt.52, C64–C71 (2013).
[CrossRef] [PubMed]

Appl. Opt. (3)

Biomed. Opt. Express (3)

J. Biomed. Opt. (3)

P. Miao, H. Lu, Q. Liu, Y. Li, and S. Tong, “Laser speckle contrast imaging of cerebral blood flow in freely moving animals,” J. Biomed. Opt.16, 090502 (2011).
[CrossRef] [PubMed]

D. A. Boas and A. K. Dunn, “Laser speckle contrast imaging in biomedical optics,” J. Biomed. Opt.15, 011109 (2010).
[CrossRef] [PubMed]

J. D. Briers and S. Webster, “Laser speckle contrast analysis (lasca): a nonscanning, full-field technique for monitoring capillary blood flow,” J. Biomed. Opt.1, 174–179 (1996).
[CrossRef] [PubMed]

J. Cerebral Blood Flow Metab. (1)

A. K. Dunn, H. Bolay, M. A. Moskowitz, and D. A. Boas, “Dynamic imaging of cerebral blood flow using laser speckle,” J. Cerebral Blood Flow Metab.21, 195–201 (2001).
[CrossRef]

J. Opt. Soc. Am. (1)

J. Opt. Soc. Am. A (2)

Lasers in Medical Science (1)

M. Draijer, E. Hondebrink, T. Leeuwen, and W. Steenbergen, “Review of laser speckle contrast techniques for visualizing tissue perfusion,”Lasers in Medical Science24, 639–651 (2009).
[CrossRef]

Microscopy Res. Technique (1)

B. Forster, D. Van De Ville, J. Berent, D. Sage, and M. Unser, “Complex wavelets for extended depth-of-field: A new method for the fusion of multichannel microscopy images,” Microscopy Res. Technique65, 33–42 (2004).
[CrossRef]

Opt. Comm. (2)

A. Fercher and J. Briers, “Flow visualization by means of single-exposure speckle photography,” Opt. Comm.37, 326–330 (1981).
[CrossRef]

Q. Yang, L. Liu, and J. Sun, “Optimized phase pupil masks for extended depth of field,” Opt. Comm.272, 56–66 (2007).
[CrossRef]

Opt. Express (1)

Opt. Lett. (1)

Optical Review (1)

E. R. Dowski, W. T. Cathey, and S. C. Bradburn, “Aberration invariant optical/digital incoherent systems,” Optical Review3, A429–A432 (1996).
[CrossRef]

Other (3)

Y. Atchia, H. Levy, S. Dufour, and O. Levi, “Speckle contrast at deviations from best focus in microfluidic and in vivo,” in Biomedical Optics and 3-D Imaging, (Optical Society of America, 2012), p. BTu3A.49.

R. C. Gonzalez and R. E. Woods, Digital Image Processing, 3rd ed. (Prentice-Hall, Inc., Upper Saddle River, NJ, USA, 2006).

J. Breckinridge, D. Voelz, and J. B. Breckinridge, Computational Fourier Optics: A MATLAB Tutorial, Tutorial Text Series (SPIE Press, 2011).

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Figures (8)

Fig. 1
Fig. 1

Schematic of the experimental setup. The setup consists of two stages: a relay stage and a 4-f imaging stage. The relay stage is used to set the magnification of the system and translate the object plane that is parallel to the optical table to an intermediate image plane that is parallel to the SLM surface. The objective is mounted on a translation stage with μm-resolution. The unit-magnification 4-f imaging stage contains an SLM placed in the Fourier plane. The SLM is used to impose a cubic phase mask on the incoming beam. Sample phase masks are shown in the right-hand side of the figure: i) α = 23 m−2, ii) α =35 m−2, iii) α = 47 m−2, iv) α = 58 m−2

Fig. 2
Fig. 2

Point spread function (PSF) change with misfocus for the traditional (left) and extended DOF (right) systems, illustrating the behaviour of the system with misfocus. Images taken were of a 10μm-diameter pinhole illuminated from the back with a red LED, at 5× magnification.

Fig. 3
Fig. 3

Flow diagram for the procedure to simulate speckle propagation in our optical system. Free space propagation is performed using the paraxial Fresnel transfer function of free space, similar to a procedure in [16]. The capillary strip is re-randomized N times, and resulting raw speckle intensity images are averaged. Misfocus is simulated by changing the distance between the object plane and the first lens to be different than the focal length of the lens.

Fig. 4
Fig. 4

Results of the numerical simulations. The speckle pattern in the “capillary” was re-randomized N = 15 times. Simulations were performed for N = 5 and 10 to confirm the linear relation between speckle contrast reduction and N, showing almost identical results. a) Relative change in the flow index with misfocus. α values are in units of m−2 b) The DOF and velocity range as a function of α m−2.

Fig. 5
Fig. 5

a) in-vitro characterization device: a rectangular tub contains scattering phantom, which can be filled to desired depth, with a holes that fix hollow glass capillaries. Hollow glass tubes with an outer diameter of 360 μm and inner diameters varying from 75 μm to 150 μm were placed in the holes. b) Cross section of the device, showing the size and relative placement of the holes. c) A schematic of the experimental setup. A perfusion pump is used to control the flow rate in a glass capillary, while a HeNe laser is used as a light source.

Fig. 6
Fig. 6

Maximum flow index vs. average flow speed in a 75 μm-wide capillary, for different values of α. The horizontal axis shows the average flow velocity in the capillary that is set using the perfusion pump. The vertical axis shows the corresponding velocity as measured using LSCI. For traditional LSCI (black squares), the linear relation between the two holds. For extended-DOF LSCI, the linear relation breaks down for higher values of α and higher flow velocities.

Fig. 7
Fig. 7

DOF of the LSCI technique. a) Relative flow speed deviation from its maximum value with misfocus inside a 75 μm-wide capillary, suspended in air. b) DOF, defined as the range of misfocus where the relative flow speed is within 20% of its maximum value, as a function of α, for a capillary immersed in air and in phantom.

Fig. 8
Fig. 8

in-vivo imaging results. a) A flow index map of the vasculature, produced using the complex wavelet transform method [18], for illustrative purposes. The vasculature map is an interpolation of traditional LSCI maps taken at different positions of the microscope objective with respect to the brain surface. Scale bar is 400 μm. b) a selection of two regions of vasculature illustrating the effect of the extended-DOF LSCI technique: columns 1 and 3 from the left illustrate the vessels flow index profiles as a function of misfocus for conventional LSCI, while columns 2 and 4 from the left illustrate the vessel flow profiles for extended-DOF LSCI. Consecutive frame are 150 μm apart in misfocus. c) Transverse flow velocity profile for selected vessels in a). The vertical axis is the transverse direction across a vessel, in μm. The horizontal axis is the position of the microscope objective against an arbitrary height above the brain surface (z = 0), in μm. The left-hand set of plots pertain to the conventional LSCI technique, while the right-hand set of plots pertain to the extended-DOF LSCI technique where α = 47 m−2. These plots were normalized by the maximum velocity in every vessel, where the color map shows the normalized speed. The vertical dashed line indicates a position of the objective where accurate flow information can be read from all selected vessels using the extended-DOF LSCI technique.

Equations (2)

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K = σ ( I ) I = τ c 2 T [ 1 exp ( 2 T τ c ) ] .
ϕ ( x , y ) = 2 π λ α ( x 3 + y 3 ) ,

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