Abstract

Three-dimensional fluorescence spatial distributions under single-photon and two-photon excitation within a turbid medium are studied with Monte Carlo simulation. It is demonstrated that two-photon excitation has an advantage of producing much less fluorescence light outside the focal region compared with single-photon excitation. With the increase of the concentration of scattering particles in a turbid medium, the position of the maximum fluorescence intensity point shifts from the geometric focal region toward the medium surface. Further studies show that the optical sectioning property of two-photon fluorescence microscopy is degraded in thick turbid media or when the numerical aperture of an objective becomes low.

© 2000 Optical Society of America

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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef]
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    [CrossRef] [PubMed]
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1999 (2)

1998 (3)

1997 (2)

X. Gan, S. Schilders, M. Gu, “Combination of annular aperture and polarization gating methods for efficient microscopic imaging through a turbid medium: theoretical analysis,” Microsc. Microanal. 3, 495–503 (1997).

Y. Guo, Q. Z. Wang, N. Zhadin, F. Liu, S. Demos, D. Calistru, A. Tirksliunas, A. Katz, Y. Budansky, P. P. Ho, R. R. Alfano, “Two-photon excitation of fluorescence from chicken tissue,” Appl. Opt. 36, 968–970 (1997).
[CrossRef] [PubMed]

1996 (1)

1995 (2)

D. W. Piston, B. R. Masters, W. W. Webb, “Three-dimensionally resolved NAD(P)H cellular metabolic redox imaging of the in-situ cornea with two-photon excitation laser scanning microscopy,” J. Microsc. 178, 20–27 (1995).
[CrossRef] [PubMed]

A. Yodh, B. Chance, “Spectroscopy and imaging with diffusing light,” Phys. Today 48(3), 34–40 (1995).
[CrossRef]

1990 (2)

C. J. R. Sheppard, M. Gu, “Image formation in two-photon fluorescence microscopy,” Optik 86, 104–106 (1990).

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

1987 (1)

S. T. Flock, B. C. Wilson, M. S. Patterson, “Total attenuation coefficients and scattering phase functions of tissues and phantom materials at 633 nm,” Med. Phys. 14, 835–841 (1987).
[CrossRef] [PubMed]

Alfano, R. R.

Ben-Letaief, K.

Budansky, Y.

Calistru, D.

Chance, B.

A. Yodh, B. Chance, “Spectroscopy and imaging with diffusing light,” Phys. Today 48(3), 34–40 (1995).
[CrossRef]

Daria, V.

Demos, S.

Denk, W.

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

Flannery, B.

W. Press, W. Vetterling, S. Teukolsky, B. Flannery, Numerical Recipes in Fortran (Cambridge U Press, New York, 1992).

Flock, S. T.

S. T. Flock, B. C. Wilson, M. S. Patterson, “Total attenuation coefficients and scattering phase functions of tissues and phantom materials at 633 nm,” Med. Phys. 14, 835–841 (1987).
[CrossRef] [PubMed]

Gan, X.

X. Gan, S. P. Schilders, M. Gu, “Image formation in turbid media under a microscope,” J. Opt. Soc. Am. A 15, 2052–2058 (1998).
[CrossRef]

X. Gan, S. Schilders, M. Gu, “Combination of annular aperture and polarization gating methods for efficient microscopic imaging through a turbid medium: theoretical analysis,” Microsc. Microanal. 3, 495–503 (1997).

Gu, M.

S. P. Schilders, M. Gu, “Three-dimensional autofluorescence spectroscopy of rat skeletal muscle tissue under two-photon excitation,” Appl. Opt. 38, 720–722 (1999).
[CrossRef]

X. Gan, S. P. Schilders, M. Gu, “Image formation in turbid media under a microscope,” J. Opt. Soc. Am. A 15, 2052–2058 (1998).
[CrossRef]

X. Gan, S. Schilders, M. Gu, “Combination of annular aperture and polarization gating methods for efficient microscopic imaging through a turbid medium: theoretical analysis,” Microsc. Microanal. 3, 495–503 (1997).

C. J. R. Sheppard, M. Gu, “Image formation in two-photon fluorescence microscopy,” Optik 86, 104–106 (1990).

M. Gu, Principles of Three-Dimensional Imaging in Confocal Microscopes (World Scientific, Singapore, 1996).

Guo, Y.

Ho, P. P.

Katz, A.

Kawata, S.

Liu, F.

Mar Blanca, C.

Masters, B. R.

D. W. Piston, B. R. Masters, W. W. Webb, “Three-dimensionally resolved NAD(P)H cellular metabolic redox imaging of the in-situ cornea with two-photon excitation laser scanning microscopy,” J. Microsc. 178, 20–27 (1995).
[CrossRef] [PubMed]

Nakamura, O.

Patterson, M. S.

S. T. Flock, B. C. Wilson, M. S. Patterson, “Total attenuation coefficients and scattering phase functions of tissues and phantom materials at 633 nm,” Med. Phys. 14, 835–841 (1987).
[CrossRef] [PubMed]

Piston, D. W.

D. W. Piston, B. R. Masters, W. W. Webb, “Three-dimensionally resolved NAD(P)H cellular metabolic redox imaging of the in-situ cornea with two-photon excitation laser scanning microscopy,” J. Microsc. 178, 20–27 (1995).
[CrossRef] [PubMed]

Press, W.

W. Press, W. Vetterling, S. Teukolsky, B. Flannery, Numerical Recipes in Fortran (Cambridge U Press, New York, 1992).

Saloma, C.

Schilders, S.

X. Gan, S. Schilders, M. Gu, “Combination of annular aperture and polarization gating methods for efficient microscopic imaging through a turbid medium: theoretical analysis,” Microsc. Microanal. 3, 495–503 (1997).

Schilders, S. P.

Schmitt, J. M.

Sheppard, C. J. R.

C. J. R. Sheppard, M. Gu, “Image formation in two-photon fluorescence microscopy,” Optik 86, 104–106 (1990).

Strickler, J. H.

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

Teukolsky, S.

W. Press, W. Vetterling, S. Teukolsky, B. Flannery, Numerical Recipes in Fortran (Cambridge U Press, New York, 1992).

Tirksliunas, A.

Vetterling, W.

W. Press, W. Vetterling, S. Teukolsky, B. Flannery, Numerical Recipes in Fortran (Cambridge U Press, New York, 1992).

Wang, Q. Z.

Webb, W. W.

D. W. Piston, B. R. Masters, W. W. Webb, “Three-dimensionally resolved NAD(P)H cellular metabolic redox imaging of the in-situ cornea with two-photon excitation laser scanning microscopy,” J. Microsc. 178, 20–27 (1995).
[CrossRef] [PubMed]

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

Wilson, B. C.

S. T. Flock, B. C. Wilson, M. S. Patterson, “Total attenuation coefficients and scattering phase functions of tissues and phantom materials at 633 nm,” Med. Phys. 14, 835–841 (1987).
[CrossRef] [PubMed]

Ying, J.

Yodh, A.

A. Yodh, B. Chance, “Spectroscopy and imaging with diffusing light,” Phys. Today 48(3), 34–40 (1995).
[CrossRef]

Zhadin, N.

Appl. Opt. (5)

J. Microsc. (1)

D. W. Piston, B. R. Masters, W. W. Webb, “Three-dimensionally resolved NAD(P)H cellular metabolic redox imaging of the in-situ cornea with two-photon excitation laser scanning microscopy,” J. Microsc. 178, 20–27 (1995).
[CrossRef] [PubMed]

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

Med. Phys. (1)

S. T. Flock, B. C. Wilson, M. S. Patterson, “Total attenuation coefficients and scattering phase functions of tissues and phantom materials at 633 nm,” Med. Phys. 14, 835–841 (1987).
[CrossRef] [PubMed]

Microsc. Microanal. (1)

X. Gan, S. Schilders, M. Gu, “Combination of annular aperture and polarization gating methods for efficient microscopic imaging through a turbid medium: theoretical analysis,” Microsc. Microanal. 3, 495–503 (1997).

Optik (1)

C. J. R. Sheppard, M. Gu, “Image formation in two-photon fluorescence microscopy,” Optik 86, 104–106 (1990).

Phys. Today (1)

A. Yodh, B. Chance, “Spectroscopy and imaging with diffusing light,” Phys. Today 48(3), 34–40 (1995).
[CrossRef]

Science (1)

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

Other (2)

M. Gu, Principles of Three-Dimensional Imaging in Confocal Microscopes (World Scientific, Singapore, 1996).

W. Press, W. Vetterling, S. Teukolsky, B. Flannery, Numerical Recipes in Fortran (Cambridge U Press, New York, 1992).

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Figures (8)

Fig. 1
Fig. 1

Three-dimensional fluorescence intensity distribution I 1p(x, y, z) for 1-p excitation (NA = 0.25): (a) n = 2, (b) n = 4, (c) n = 6, (d) n = 8.

Fig. 2
Fig. 2

Three-dimensional fluorescence intensity distribution I 2p(x, y, z) for 2-p excitation (NA = 0.25): (a) n = 2, (b) n = 4, (c) n = 6, (d) n = 8.

Fig. 3
Fig. 3

Two-dimensional xz intensity cross section V 1p(x, z) for 1-p excitation (NA = 0.25): (a) n = 2, (b) n = 4, (c) n = 6, (d) n = 8.

Fig. 4
Fig. 4

Two-dimensional xz intensity cross section V 2p(x, z) for 2-p excitation (NA = 0.25): (a) n = 2, (b) n = 4, (c) n = 6, (d) n = 8.

Fig. 5
Fig. 5

Axial intensity distribution S(z) for a 1-p fluorescence microscope (NA = 0.25).

Fig. 6
Fig. 6

Axial intensity distribution S(z) for a 2-p fluorescence microscope (NA = 0.25).

Fig. 7
Fig. 7

Axial intensity distribution S(z) for different NA values under 1-p excitation (n = 4).

Fig. 8
Fig. 8

Axial intensity distribution S(z) for different NA values under 2-p excitation (n = 4).

Tables (1)

Tables Icon

Table 1 Turbid-Medium-Associated Parameters That Are Comparable with Those Used in Experimentsa

Equations (3)

Equations on this page are rendered with MathJax. Learn more.

I1px, y, z  Iexx, y, z, I2px, y, z  Iex2x, y, z.
Vx, z  -Ix, y, zdy.
Sz  -Ix, y, zdxdy.

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