Abstract

Focal modulation microscopy (FMM) is an emerging microscopy technique for fluorescence imaging of thick biological tissue in vivo. A spatial phase modulator is a critical component whose characteristics have a significant impact on the performance of a FMM system. We have designed a simple spatial phase modulator based on a tilting glass plate that provides superb modulation stability. Image quality has been improved remarkably after integrating such a modulator into a FMM system.

© 2009 Optical Society of America

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References

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  1. A. Waggoner, “Fluorescent labels for proteomics and genomics,” Curr. Opin. Chem. Biol. 10, 62-66 (2006).
    [CrossRef] [PubMed]
  2. R. Y. Tsien, “The green fluorescent protein,” Annu. Rev. Biochem. 67, 509-544 (1998).
    [CrossRef] [PubMed]
  3. C. Dunsby and P. M. W. French, “Techniques for depth-resolved imaging through turbid media including coherence-gated imaging,” J. Phys. D 36, R207-R227 (2003).
    [CrossRef]
  4. G. E. Anderson, F. Liu, and R. R. Alfano, “Microscope imaging through highly scattering media,” Opt. Lett. 19, 981-983(1994).
    [CrossRef] [PubMed]
  5. V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
    [CrossRef]
  6. M. Minsky, “Microscopy apparatus,” U.S. patent 3,013,467 (19 December 1961).
  7. C. J. R. Sheppard and A. Choudhury, “Image formation in the scanning microscope,” Opt. Acta 24, 1051-1073 (1977).
    [CrossRef]
  8. R. H. Webb, “Confocal optical microscopy,” Rep. Prog. Phys. 59, 427-471 (1996).
    [CrossRef]
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    [CrossRef]
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    [CrossRef] [PubMed]
  11. M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
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    [CrossRef]
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    [CrossRef]
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    [CrossRef] [PubMed]

2008 (1)

2006 (2)

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

A. Waggoner, “Fluorescent labels for proteomics and genomics,” Curr. Opin. Chem. Biol. 10, 62-66 (2006).
[CrossRef] [PubMed]

2003 (4)

2001 (1)

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

1998 (1)

R. Y. Tsien, “The green fluorescent protein,” Annu. Rev. Biochem. 67, 509-544 (1998).
[CrossRef] [PubMed]

1997 (1)

V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
[CrossRef]

1996 (2)

1994 (2)

1991 (1)

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

1990 (1)

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73-76 (1990).
[CrossRef] [PubMed]

1977 (1)

C. J. R. Sheppard and A. Choudhury, “Image formation in the scanning microscope,” Opt. Acta 24, 1051-1073 (1977).
[CrossRef]

Alfano, R. R.

Anderson, G. E.

Beaurepaire, E.

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

Brill, A.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Chaigneau, E.

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

Chang, W.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Charpak, S.

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

Chen, N.

Choudhury, A.

C. J. R. Sheppard and A. Choudhury, “Image formation in the scanning microscope,” Opt. Acta 24, 1051-1073 (1977).
[CrossRef]

Deng, X.

Denk, W.

Dunsby, C.

C. Dunsby and P. M. W. French, “Techniques for depth-resolved imaging through turbid media including coherence-gated imaging,” J. Phys. D 36, R207-R227 (2003).
[CrossRef]

Flotte, T.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

French, P. M. W.

C. Dunsby and P. M. W. French, “Techniques for depth-resolved imaging through turbid media including coherence-gated imaging,” J. Phys. D 36, R207-R227 (2003).
[CrossRef]

Fujimoto, J. G.

J. G. Fujimoto, “Optical coherence tomography for ultrahigh resolution in vivo imaging,” Nat. Biotechnol. 21, 1361-1367(2003).
[CrossRef] [PubMed]

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Gregory, Y.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Gu, M.

Haramati, S.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Harmelin, A.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Hasan, M. T.

Hee, M. R.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Huang, D.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Kalchenko, V.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Kempe, M.

Knüttel, A.

Kon, I. L.

V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
[CrossRef]

Lapid, K.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Lapidot, T.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Lin, C. P.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Liu, F.

Maksimova, I. L.

V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
[CrossRef]

Malina, V.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Mertz, J.

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

Minsky, M.

M. Minsky, “Microscopy apparatus,” U.S. patent 3,013,467 (19 December 1961).

Oheim, M.

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

Puliafito, C. A.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Rudolph, W.

Schmitt, J. M.

Schuman, J. S.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Sheppard, C. J. R.

N. Chen, C. Wong, and C. J. R. Sheppard, “Focal modulation microscopy,” Opt. Express 16, 18764-18769 (2008).
[CrossRef]

C. J. R. Sheppard and A. Choudhury, “Image formation in the scanning microscope,” Opt. Acta 24, 1051-1073 (1977).
[CrossRef]

Shivtiel, S.

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

Stinson, W. G.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Strickler, J. H.

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73-76 (1990).
[CrossRef] [PubMed]

Swanson, E. A.

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Theer, P.

Tsien, R. Y.

R. Y. Tsien, “The green fluorescent protein,” Annu. Rev. Biochem. 67, 509-544 (1998).
[CrossRef] [PubMed]

Tuchin, V. V.

V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
[CrossRef]

Waggoner, A.

A. Waggoner, “Fluorescent labels for proteomics and genomics,” Curr. Opin. Chem. Biol. 10, 62-66 (2006).
[CrossRef] [PubMed]

Webb, R. H.

R. H. Webb, “Confocal optical microscopy,” Rep. Prog. Phys. 59, 427-471 (1996).
[CrossRef]

Webb, W. W.

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73-76 (1990).
[CrossRef] [PubMed]

Welsch, E.

Wong, C.

Yadlowsky, M.

Zimnyakov, D. A.

V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
[CrossRef]

Annu. Rev. Biochem. (1)

R. Y. Tsien, “The green fluorescent protein,” Annu. Rev. Biochem. 67, 509-544 (1998).
[CrossRef] [PubMed]

Appl. Opt. (1)

Curr. Opin. Chem. Biol. (1)

A. Waggoner, “Fluorescent labels for proteomics and genomics,” Curr. Opin. Chem. Biol. 10, 62-66 (2006).
[CrossRef] [PubMed]

J. Biomed. Opt. (2)

V. Kalchenko, S. Shivtiel, V. Malina, K. Lapid, S. Haramati, T. Lapidot, A. Brill, and A. Harmelin, “Use of lipophilic near-infrared dye in whole-body optical imaging of hematopoietic cell homing,” J. Biomed. Opt. 11, 050507 (2006).
[CrossRef] [PubMed]

V. V. Tuchin, I. L. Maksimova, D. A. Zimnyakov, and I. L. Kon, “Light propagation in tissues with controlled optical properties,” J. Biomed. Opt. 2, 401-417 (1997).
[CrossRef]

J. Neurosci. Methods (1)

M. Oheim, E. Beaurepaire, E. Chaigneau, J. Mertz, and S. Charpak, “Two photon microscopy in brain tissue: parameters influencing the imaging depth,” J. Neurosci. Methods 111, 29-37 (2001).
[CrossRef] [PubMed]

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

J. Phys. D (1)

C. Dunsby and P. M. W. French, “Techniques for depth-resolved imaging through turbid media including coherence-gated imaging,” J. Phys. D 36, R207-R227 (2003).
[CrossRef]

Nat. Biotechnol. (1)

J. G. Fujimoto, “Optical coherence tomography for ultrahigh resolution in vivo imaging,” Nat. Biotechnol. 21, 1361-1367(2003).
[CrossRef] [PubMed]

Opt. Acta (1)

C. J. R. Sheppard and A. Choudhury, “Image formation in the scanning microscope,” Opt. Acta 24, 1051-1073 (1977).
[CrossRef]

Opt. Express (1)

Opt. Lett. (2)

Rep. Prog. Phys. (1)

R. H. Webb, “Confocal optical microscopy,” Rep. Prog. Phys. 59, 427-471 (1996).
[CrossRef]

Science (2)

W. Denk, J. H. Strickler, and W. W. Webb, “Two-photon laser scanning fluorescence microscopy,” Science 248, 73-76 (1990).
[CrossRef] [PubMed]

D. Huang, E. A. Swanson, C. P. Lin, J. S. Schuman, W. G. Stinson, W. Chang, M. R. Hee, T. Flotte, Y. Gregory, C. A. Puliafito, and J. G. Fujimoto, “Optical coherence tomography,” Science 254, 1178-1181 (1991).
[CrossRef] [PubMed]

Other (2)

M. Minsky, “Microscopy apparatus,” U.S. patent 3,013,467 (19 December 1961).

MeadowLark Optics spatial light modulator, http://www.meadowlarkoptics.com/products/slmLanding.php.

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Figures (7)

Fig. 1
Fig. 1

(a) Bisected back aperture of the pupil. (b) Excitation light path in FMM with a spatial phase modulator. Half of the beam (shaded areas) is subject to periodic phase modulation.

Fig. 2
Fig. 2

Schematic for the reflection mode phase modulator consisting of two mirrors.

Fig. 3
Fig. 3

(a) Simulated focal excitation waveform and (b) its power spectrum from an ideal two-mirror modulator are compared with the (c) waveform and (d) power spectrum when the optical path length difference drifts away.

Fig. 4
Fig. 4

Fluorescence image of chicken chondrocyte showing the effect of drifting optical path length difference. The scale bar is at 20 μm . The depth is at around the 200 μm region.

Fig. 5
Fig. 5

(a) Glass plate mounted on a galvanometer, intersecting half of the excitation beam whose cross section is indicated by the gray circle. (b) Optical paths of the half-beam propagating through the glass plate (dashed lines) and the half-beam propagating in the air (solid lines).

Fig. 6
Fig. 6

Measured intensity modulation at the focal point from the tilting plate spatial phase modulator.

Fig. 7
Fig. 7

(a) Confocal and (b) FMM images of chondrocytes obtained from chicken cartilage at a depth of 400 μm . (c) and (d) Higher magnification view of the boxed regions in (a) and (b), respectively. Scale bar: 20 μm . The blurred cell outline and out-of-focus chondrocyte cell presence detected in the right portion in (a) and (c) indicate poorer sectioning compared to the FMM techniques illustrated in (b) and (d).

Equations (1)

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φ ( t ) = 2 π λ D ( n 2 sin 2 α ( t ) cos α ( t ) ) ,

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