Abstract

We report on three-dimensional autofluorescence spectroscopy obtained from rat skeletal muscle tissue under two-photon excitation by an ultrashort pulsed-laser beam. It is demonstrated that two types of fluorophores within the skeletal muscle tissue can be simultaneously excited with the laser beam at a wavelength of 800 nm. The two fluorophores exhibited unique fluorescence spectral peaks at wavelengths of 450 and 550 nm. These spectroscopic signals can be used to form a three-dimensional image, giving the information about the biochemical makeup of the skeletal muscle tissue.

© 1999 Optical Society of America

Full Article  |  PDF Article

References

  • View by:
  • |
  • |
  • |

  1. B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
    [CrossRef] [PubMed]
  2. J. Aubin, “Autofluorescence of viable cultured mammalian cells,” J. Histochem. Cytochem. 27, 36–43 (1979).
    [CrossRef] [PubMed]
  3. R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
    [CrossRef] [PubMed]
  4. R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
    [CrossRef]
  5. G. C. Tang, A. Pradhan, R. R. Alfano, “Spectroscopic differences between human cancer and normal lung and breast tissues,” Lasers Surg. Med. 9, 290–295 (1989).
    [CrossRef] [PubMed]
  6. W. Denk, J. H. Strickler, W. W. Webb, “Two photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
    [CrossRef] [PubMed]
  7. Y. Guo, Q. Z. Wang, N. Zhadin, F. Liu, S. Demos, D. Calistru, A. Tirksliunas, A. Katz, Y. Bunansky, P. P. Ho, R. R. Alfano, “Two-photon excitation of fluorescence from chicken tissue,” Appl. Opt. 36, 968–970 (1997).
    [CrossRef] [PubMed]
  8. C. J. R. Sheppard, M. Gu, “Image formation in two-photon fluorescence microscopy,” Optik 86, 104–106 (1990).
  9. M. Gu, Principles of Three-Dimensional Imaging in Confocal Microscopes (World Scientific, Singapore, 1996).
  10. B. R. Masters, P. T. C. So, E. Gratton, “Multiphoton excitation fluorescence microscopy and spectroscopy of in vivo human skin,” Biophys. J. 72, 2405–2412 (1997).
    [CrossRef] [PubMed]
  11. W. J. Krause, Essentials of Human Histology, (Little, Brown and Company, Boston, Mass., 1994).
  12. Y. R. Shen, Principles of Nonlinear Optics (Wiley, New York, 1984).

1997 (2)

1990 (2)

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

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

1989 (1)

G. C. Tang, A. Pradhan, R. R. Alfano, “Spectroscopic differences between human cancer and normal lung and breast tissues,” Lasers Surg. Med. 9, 290–295 (1989).
[CrossRef] [PubMed]

1984 (1)

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

1979 (2)

J. Aubin, “Autofluorescence of viable cultured mammalian cells,” J. Histochem. Cytochem. 27, 36–43 (1979).
[CrossRef] [PubMed]

R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
[CrossRef] [PubMed]

1962 (1)

B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
[CrossRef] [PubMed]

Alfano, M. A.

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

Alfano, R. R.

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

G. C. Tang, A. Pradhan, R. R. Alfano, “Spectroscopic differences between human cancer and normal lung and breast tissues,” Lasers Surg. Med. 9, 290–295 (1989).
[CrossRef] [PubMed]

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

Aubin, J.

J. Aubin, “Autofluorescence of viable cultured mammalian cells,” J. Histochem. Cytochem. 27, 36–43 (1979).
[CrossRef] [PubMed]

Benson, R. C.

R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
[CrossRef] [PubMed]

Bunansky, Y.

Calistru, D.

Chance, B.

B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
[CrossRef] [PubMed]

Cohen, P.

B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
[CrossRef] [PubMed]

Cordero, J.

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

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]

Gratton, E.

B. R. Masters, P. T. C. So, E. Gratton, “Multiphoton excitation fluorescence microscopy and spectroscopy of in vivo human skin,” Biophys. J. 72, 2405–2412 (1997).
[CrossRef] [PubMed]

Gu, M.

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.

Jobsis, F.

B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
[CrossRef] [PubMed]

Katz, A.

Krause, W. J.

W. J. Krause, Essentials of Human Histology, (Little, Brown and Company, Boston, Mass., 1994).

Liu, F.

Longo, F. W.

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

Masters, B. R.

B. R. Masters, P. T. C. So, E. Gratton, “Multiphoton excitation fluorescence microscopy and spectroscopy of in vivo human skin,” Biophys. J. 72, 2405–2412 (1997).
[CrossRef] [PubMed]

McKhann, G. M.

R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
[CrossRef] [PubMed]

Meyer, R. A.

R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
[CrossRef] [PubMed]

Pradhan, A.

G. C. Tang, A. Pradhan, R. R. Alfano, “Spectroscopic differences between human cancer and normal lung and breast tissues,” Lasers Surg. Med. 9, 290–295 (1989).
[CrossRef] [PubMed]

Schoener, B.

B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
[CrossRef] [PubMed]

Shen, Y. R.

Y. R. Shen, Principles of Nonlinear Optics (Wiley, New York, 1984).

Sheppard, C. J. R.

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

So, P. T. C.

B. R. Masters, P. T. C. So, E. Gratton, “Multiphoton excitation fluorescence microscopy and spectroscopy of in vivo human skin,” Biophys. J. 72, 2405–2412 (1997).
[CrossRef] [PubMed]

Strickler, J. H.

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

Tang, G. C.

G. C. Tang, A. Pradhan, R. R. Alfano, “Spectroscopic differences between human cancer and normal lung and breast tissues,” Lasers Surg. Med. 9, 290–295 (1989).
[CrossRef] [PubMed]

Tata, D. B.

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

Tirksliunas, A.

Tomashefsky, P.

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

Wang, Q. Z.

Webb, W. W.

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

Zauba, M. E.

R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
[CrossRef] [PubMed]

Zhadin, N.

Appl. Opt. (1)

Biophys. J. (1)

B. R. Masters, P. T. C. So, E. Gratton, “Multiphoton excitation fluorescence microscopy and spectroscopy of in vivo human skin,” Biophys. J. 72, 2405–2412 (1997).
[CrossRef] [PubMed]

IEEE J. Quantum Electron. (1)

R. R. Alfano, D. B. Tata, J. Cordero, P. Tomashefsky, F. W. Longo, M. A. Alfano, “Laser induced fluorescence spectroscopy from native cancerous and normal tissue,” IEEE J. Quantum Electron. QE-20, 1507–1511 (1984).
[CrossRef]

J. Histochem. Cytochem. (2)

J. Aubin, “Autofluorescence of viable cultured mammalian cells,” J. Histochem. Cytochem. 27, 36–43 (1979).
[CrossRef] [PubMed]

R. C. Benson, R. A. Meyer, M. E. Zauba, G. M. McKhann, “Cellular autofluorescence—is it due to flavins?” J. Histochem. Cytochem. 27, 44–48 (1979).
[CrossRef] [PubMed]

Lasers Surg. Med. (1)

G. C. Tang, A. Pradhan, R. R. Alfano, “Spectroscopic differences between human cancer and normal lung and breast tissues,” Lasers Surg. Med. 9, 290–295 (1989).
[CrossRef] [PubMed]

Optik (1)

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

Science (2)

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

B. Chance, P. Cohen, F. Jobsis, B. Schoener, “Intracellular oxidation states in vivo,” Science 137, 499–508 (1962).
[CrossRef] [PubMed]

Other (3)

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

W. J. Krause, Essentials of Human Histology, (Little, Brown and Company, Boston, Mass., 1994).

Y. R. Shen, Principles of Nonlinear Optics (Wiley, New York, 1984).

Cited By

OSA participates in CrossRef's Cited-By Linking service. Citing articles from OSA journals and other participating publishers are listed here.

Alert me when this article is cited.


Figures (5)

Fig. 1
Fig. 1

Two-dimensional autofluorescence image sections of the rat skeletal muscle tissue recorded at different depths. The depth spacing is 6 µm.

Fig. 2
Fig. 2

Reconstructed 3-D autofluorescence image of the rat skeletal muscle tissue at different rotation view angles: (a) 0°, (b) 45°, (c) 90°.

Fig. 3
Fig. 3

Autofluorescence spectra from different points within the rat skeletal muscle tissue.

Fig. 4
Fig. 4

Autofluorescence intensity as a function of the excitation laser power.

Fig. 5
Fig. 5

Transverse [(a) and (b)] and vertical [(c) and (d)] image sections of the rat skeletal muscle tissue recorded at autofluorescence wavelengths 450 [(a) and (c)] and 550 nm [(b) and (d)].

Metrics