Abstract

Fourier multiplexed FLIM (FmFLIM) tomography enables multiplexed 3D lifetime imaging of whole embryos. In our previous FmFLIM system, the spatial resolution was limited to 25 μm because of the trade-off between the spatial resolution and the imaging depth. In order to achieve cellular resolution imaging of thick specimens, we built a tomography system with dual-color Bessel beam. In combination with FmFLIM, the Bessel FmFLIM tomography system can perform parallel 3D lifetime imaging on multiple excitation-emission channels at a cellular resolution of 2.8 μm. The image capability of the Bessel FmFLIM tomography system was demonstrated by 3D lifetime imaging of dual-labeled transgenic zebrafish embryos.

© 2017 Optical Society of America

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References

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    [Crossref] [PubMed]
  4. J. Sharpe, “Optical Projection Tomography,” Annu. Rev. Biomed. Eng. 6(1), 209–228 (2004).
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    [Crossref] [PubMed]
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    [Crossref] [PubMed]
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  18. J. Leach, G. M. Gibson, M. J. Padgett, E. Esposito, G. McConnell, A. J. Wright, and J. M. Girkin, “Generation of achromatic Bessel beams using a compensated spatial light modulator,” Opt. Express 14(12), 5581–5587 (2006).
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2015 (1)

M. Zhao, X. Wan, Y. Li, W. Zhou, and L. Peng, “Multiplexed 3D FRET imaging in deep tissue of live embryos,” Sci. Rep. 5, 13991 (2015).
[Crossref] [PubMed]

2014 (3)

2012 (1)

F. O. Fahrbach and A. Rohrbach, “Propagation stability of self-reconstructing Bessel beams enables contrast-enhanced imaging in thick media,” Nat. Commun. 3, 632 (2012).
[Crossref] [PubMed]

2011 (3)

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

R.-A. Lorbeer, M. Heidrich, C. Lorbeer, D. F. Ramírez Ojeda, G. Bicker, H. Meyer, and A. Heisterkamp, “Highly efficient 3D fluorescence microscopy with a scanning laser optical tomograph,” Opt. Express 19(6), 5419–5430 (2011).
[Crossref] [PubMed]

2010 (3)

2007 (1)

J. R. Walls, J. G. Sled, J. Sharpe, and R. M. Henkelman, “Resolution improvement in emission optical projection tomography,” Phys. Med. Biol. 52(10), 2775–2790 (2007).
[Crossref] [PubMed]

2006 (2)

2005 (2)

J.-A. Conchello and J. W. Lichtman, “Optical sectioning microscopy,” Nat. Methods 2(12), 920–931 (2005).
[Crossref] [PubMed]

F. Helmchen and W. Denk, “Deep tissue two-photon microscopy,” Nat. Methods 2(12), 932–940 (2005).
[Crossref] [PubMed]

2004 (1)

J. Sharpe, “Optical Projection Tomography,” Annu. Rev. Biomed. Eng. 6(1), 209–228 (2004).
[Crossref] [PubMed]

2002 (2)

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Z. Ding, H. Ren, Y. Zhao, J. S. Nelson, and Z. Chen, “High-resolution optical coherence tomography over a large depth range with an axicon lens,” Opt. Lett. 27(4), 243–245 (2002).
[Crossref] [PubMed]

Ahlgren, U.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Ashok, A.

Bachmann, A. H.

Baldock, R.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Betzig, E.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Bicker, G.

Bouma, B. E.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Chen, Z.

Conchello, J.-A.

J.-A. Conchello and J. W. Lichtman, “Optical sectioning microscopy,” Nat. Methods 2(12), 920–931 (2005).
[Crossref] [PubMed]

Davidson, D.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Davidson, M. W.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Denk, W.

F. Helmchen and W. Denk, “Deep tissue two-photon microscopy,” Nat. Methods 2(12), 932–940 (2005).
[Crossref] [PubMed]

Ding, Z.

Esposito, E.

Fahrbach, F. O.

F. O. Fahrbach and A. Rohrbach, “Propagation stability of self-reconstructing Bessel beams enables contrast-enhanced imaging in thick media,” Nat. Commun. 3, 632 (2012).
[Crossref] [PubMed]

F. O. Fahrbach, P. Simon, and A. Rohrbach, “Microscopy with self-reconstructing beams,” Nat. Photonics 4(11), 780–785 (2010).
[Crossref]

F. O. Fahrbach and A. Rohrbach, “A line scanned light-sheet microscope with phase shaped self-reconstructing beams,” Opt. Express 18(23), 24229–24244 (2010).
[Crossref] [PubMed]

Galbraith, C. G.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Galbraith, J. A.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Gao, L.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Gardecki, J. A.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Gibson, G. M.

Girkin, J. M.

Hecksher-Sørensen, J.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Heidrich, M.

Heisterkamp, A.

Helmchen, F.

F. Helmchen and W. Denk, “Deep tissue two-photon microscopy,” Nat. Methods 2(12), 932–940 (2005).
[Crossref] [PubMed]

Henkelman, R. M.

J. R. Walls, J. G. Sled, J. Sharpe, and R. M. Henkelman, “Resolution improvement in emission optical projection tomography,” Phys. Med. Biol. 52(10), 2775–2790 (2007).
[Crossref] [PubMed]

Hill, B.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Lasser, T.

Leach, J.

Leitgeb, R. A.

Li, Y.

Liang, R.

Lichtman, J. W.

J.-A. Conchello and J. W. Lichtman, “Optical sectioning microscopy,” Nat. Methods 2(12), 920–931 (2005).
[Crossref] [PubMed]

Liu, L.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Lorbeer, C.

Lorbeer, R.-A.

McConnell, G.

Meyer, H.

Milkie, D. E.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Nadkarni, S. K.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Nelson, J. S.

Padgett, M. J.

Peng, L.

Perry, P.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Planchon, T. A.

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

Ramírez Ojeda, D. F.

Ren, H.

Rohrbach, A.

F. O. Fahrbach and A. Rohrbach, “Propagation stability of self-reconstructing Bessel beams enables contrast-enhanced imaging in thick media,” Nat. Commun. 3, 632 (2012).
[Crossref] [PubMed]

F. O. Fahrbach, P. Simon, and A. Rohrbach, “Microscopy with self-reconstructing beams,” Nat. Photonics 4(11), 780–785 (2010).
[Crossref]

F. O. Fahrbach and A. Rohrbach, “A line scanned light-sheet microscope with phase shaped self-reconstructing beams,” Opt. Express 18(23), 24229–24244 (2010).
[Crossref] [PubMed]

Ross, A.

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Sharpe, J.

J. R. Walls, J. G. Sled, J. Sharpe, and R. M. Henkelman, “Resolution improvement in emission optical projection tomography,” Phys. Med. Biol. 52(10), 2775–2790 (2007).
[Crossref] [PubMed]

J. Sharpe, “Optical Projection Tomography,” Annu. Rev. Biomed. Eng. 6(1), 209–228 (2004).
[Crossref] [PubMed]

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Simon, P.

F. O. Fahrbach, P. Simon, and A. Rohrbach, “Microscopy with self-reconstructing beams,” Nat. Photonics 4(11), 780–785 (2010).
[Crossref]

Sled, J. G.

J. R. Walls, J. G. Sled, J. Sharpe, and R. M. Henkelman, “Resolution improvement in emission optical projection tomography,” Phys. Med. Biol. 52(10), 2775–2790 (2007).
[Crossref] [PubMed]

Steinmann, L.

Tearney, G. J.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Toussaint, J. D.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Villiger, M.

Walls, J. R.

J. R. Walls, J. G. Sled, J. Sharpe, and R. M. Henkelman, “Resolution improvement in emission optical projection tomography,” Phys. Med. Biol. 52(10), 2775–2790 (2007).
[Crossref] [PubMed]

Wan, X.

M. Zhao, X. Wan, Y. Li, W. Zhou, and L. Peng, “Multiplexed 3D FRET imaging in deep tissue of live embryos,” Sci. Rep. 5, 13991 (2015).
[Crossref] [PubMed]

Wright, A. J.

Yagi, Y.

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Zhang, H.

Zhao, M.

Zhao, Y.

Zhou, W.

Annu. Rev. Biomed. Eng. (1)

J. Sharpe, “Optical Projection Tomography,” Annu. Rev. Biomed. Eng. 6(1), 209–228 (2004).
[Crossref] [PubMed]

Biomed. Opt. Express (1)

Nat. Commun. (1)

F. O. Fahrbach and A. Rohrbach, “Propagation stability of self-reconstructing Bessel beams enables contrast-enhanced imaging in thick media,” Nat. Commun. 3, 632 (2012).
[Crossref] [PubMed]

Nat. Med. (1)

L. Liu, J. A. Gardecki, S. K. Nadkarni, J. D. Toussaint, Y. Yagi, B. E. Bouma, and G. J. Tearney, “Imaging the subcellular structure of human coronary atherosclerosis using micro-optical coherence tomography,” Nat. Med. 17(8), 1010–1014 (2011).
[Crossref] [PubMed]

Nat. Methods (3)

T. A. Planchon, L. Gao, D. E. Milkie, M. W. Davidson, J. A. Galbraith, C. G. Galbraith, and E. Betzig, “Rapid three-dimensional isotropic imaging of living cells using Bessel beam plane illumination,” Nat. Methods 8(5), 417–423 (2011).
[Crossref] [PubMed]

J.-A. Conchello and J. W. Lichtman, “Optical sectioning microscopy,” Nat. Methods 2(12), 920–931 (2005).
[Crossref] [PubMed]

F. Helmchen and W. Denk, “Deep tissue two-photon microscopy,” Nat. Methods 2(12), 932–940 (2005).
[Crossref] [PubMed]

Nat. Photonics (1)

F. O. Fahrbach, P. Simon, and A. Rohrbach, “Microscopy with self-reconstructing beams,” Nat. Photonics 4(11), 780–785 (2010).
[Crossref]

Opt. Express (5)

Opt. Lett. (3)

Phys. Med. Biol. (1)

J. R. Walls, J. G. Sled, J. Sharpe, and R. M. Henkelman, “Resolution improvement in emission optical projection tomography,” Phys. Med. Biol. 52(10), 2775–2790 (2007).
[Crossref] [PubMed]

Sci. Rep. (1)

M. Zhao, X. Wan, Y. Li, W. Zhou, and L. Peng, “Multiplexed 3D FRET imaging in deep tissue of live embryos,” Sci. Rep. 5, 13991 (2015).
[Crossref] [PubMed]

Science (1)

J. Sharpe, U. Ahlgren, P. Perry, B. Hill, A. Ross, J. Hecksher-Sørensen, R. Baldock, and D. Davidson, “Optical projection tomography as a tool for 3D microscopy and gene expression studies,” Science 296(5567), 541–545 (2002).
[Crossref] [PubMed]

Supplementary Material (3)

NameDescription
» Visualization 1: MP4 (2822 KB)      3D intensity and lifetime projection of the tail of a zebrafish embryo.
» Visualization 2: MP4 (7306 KB)      Intensity and lifetime cross-sections of the tail of a zebrafish embryo in z-direction.
» Visualization 3: MP4 (2318 KB)      Intensity and lifetime cross-sections of the tail of a zebrafish embryo in x-direction.

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Figures (5)

Fig. 1
Fig. 1 Schematics of Bessel FmFLIM-SLOT tomography. (A) Schematic of key optical and electronic components of the Bessel FmFLIM-SLOT system. (B) Detailed schematic of the Michelson interferometer, which produces wavelength-dependent frequency sweeping interferometric modulation for all laser lines. f1 = 50 mm, f2 = 50 mm (C) Optical schematic of two-color Bessel beam generation. The modulated Gaussian beam from the Michelson interferometer is expanded and directed to the SLM. An Iris is placed at the Fourier plane of the SLM to select the first order diffracted beams. A 4° prism at the image plane of the SLM is used to compensate the dispersion. f3 = 400 mm, f4 = 250 mm. QWP, quarter-wave plates; PBS, polarizing beamsplitter; SLM, spatial light modulator.

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Fig. 2
Fig. 2 Cross sectional images of Bessel beams taken after the dispersion compensation from a BK7 prism. (A) Beam profiles from different laser lines (405nm, 488nm, 561nm and 640nm), images are captured by a beam profiler (Thorlabs) at 450mm away from the prism. (B) Theoretical and experimental 488 nm Bessel beam intensity cross-sections taken at the dashed line in A. The diameter of the central lobe is 169 μm (zero to zero) (C) Overlaid cross-sections of all wavelengths, showing the overlap of 488 nm and 561 nm Bessel beams and dispersion residues in 405 nm and 640 nm beams.

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Fig. 3
Fig. 3 Characterization of the FOV and resolution. (A) Side projection image of a Bessel beam propagating in a sample tube filled with fluorescent dye. Image is acquired by an EMCCD through a 40x objective (0.8 NA, Olympus) and an 85 mm tube lens. (B) Peak intensity curve along the propagation axis, showing the Bessel beam has a FWHM length of 292 μm. (C) Reconstructed cross-section image of the internal cavity of a FEP tube. (D) Intensity profile across the FEP-water interface (outside circle in C) and water-agar interface (inside circle in C). The profile was calculated by interpolating the image in C along a line (yellow line in C) normal to two interfaces, and averaging all lines around the circular interface. The interpolation step size is 0.9 μm.

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Fig. 4
Fig. 4 3D two-channel lifetime images of the tail of a zebrafish embryo at 40 hpf.

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Fig. 5
Fig. 5 Background in raw projection images due to Bessel side bands and its removal in the tomography reconstruction (A) Intensity projection image of a zebrafish embryo tail, calculated by integrating the 3D reconstructed image. (B) Raw projection image at the same projection angle of A. (C) Intensity profiles along the dashed lines marked in A and B, showing a uniform background in the raw projection image due to Bessel sidebands (orange line), which was removed in the tomographic 3D image (blue line) after the inverse Radon transform.

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Equations (1)

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u( x,y )= ∫ P( θ ′ ,k ) exp[ 2πik( xcosθ+ysinθ ) ]k dkdθ,

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