Abstract

We demonstrate a widefield multiphoton microscope and a temporally decorrelated, multifocal, multiphoton microscope that is based on a high-efficiency array of cascaded beamsplitters. Because these microscopes use ultrashort pulse excitation over large areas of the sample, they allow efficient use of the high-average power available from modern ultrashort pulse lasers.

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References

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  1. W. Denk, J.H. Strickler, and W.W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990).
  2. Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, "Nonlinear scanning laser microscopy by third harmonic generation," Appl. Phys. Lett. 70, 922-4 (1997).
  3. M. Sonnleitner, G. J. Schutz, and Th Schmidt, "Imaging individual molecules by two-photon excitation," Chem. Phys. Lett. 300, 221-6 (1999).
  4. E.H.K. Stelzer, S. Hell, S. Lindek, R. Stricker, R. Pick, C. Storz, G. Ritter, and N. Salmon, "Nonlinear absorption extends confocal fluorescence microscopy into the ultra-violet regime and confines the illumination volume.," Opt. Commun. 104, 223-228 (1994).
  5. K. Konig, P. T. C. So, W. W. Mantulin, and E. Gratton, "Cellular response to near-infrared femtosecond laser pulses in two-photon microscopes," Opt. Lett. 22, 135-6 (1997).
  6. J. Bewersdorf, R. Pick, and S. W. Hell, "Multifocal multiphoton microscopy," Opt. Lett. 23, 655-7 (1998).
  7. A. H. Buist, M. Muller, J. Squier, and G. J. Brakenhoff, "Real time two-photon absorption microscopy using multi point excitation," J. Microscopy 192, 217-26 (1998).
  8. K. Fujita, O. Nakamura, T. Kaneko, M. Oyamada, T. Takamatsu, and S. Kawata, "Confocal multipoint multiphoton excitation microscope with microlens and pinhole arrays," Opt. Comm. 174, 7-12 (2000).
  9. D. N. Fittinghoff and J. A. Squier, "Time-decorrelated multifocal array for multiphoton microscopy and micromachining," Opt. Lett. 25, 1213-1215 (2000).
  10. A. Egner and S. W. Hell, "Time multiplexing and parallelization in multifocal multiphoton microscopy," J. Opt. Soc. Am. A (Optics, Image Science and Vision) 17, 1192-201 (2000).
  11. Heidi Dobson, "Pollen and pollen-coat lipids: chemical survey and role in pollen selection by solitary bees (Pollenkitt, Oligolecty)," PhD Dissertation, University of California, Berkeley (1985).
  12. M. G. L. Gustafsson, D. A. Agard, and J. W. Sedat, "I/sup 5/M: 3D widefield light microscopy with better than 100 nm axial resolution," J. Microscopy 195, 10-16 (1999).

Other (12)

W. Denk, J.H. Strickler, and W.W. Webb, "Two-photon laser scanning fluorescence microscopy," Science 248, 73-76 (1990).

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, "Nonlinear scanning laser microscopy by third harmonic generation," Appl. Phys. Lett. 70, 922-4 (1997).

M. Sonnleitner, G. J. Schutz, and Th Schmidt, "Imaging individual molecules by two-photon excitation," Chem. Phys. Lett. 300, 221-6 (1999).

E.H.K. Stelzer, S. Hell, S. Lindek, R. Stricker, R. Pick, C. Storz, G. Ritter, and N. Salmon, "Nonlinear absorption extends confocal fluorescence microscopy into the ultra-violet regime and confines the illumination volume.," Opt. Commun. 104, 223-228 (1994).

K. Konig, P. T. C. So, W. W. Mantulin, and E. Gratton, "Cellular response to near-infrared femtosecond laser pulses in two-photon microscopes," Opt. Lett. 22, 135-6 (1997).

J. Bewersdorf, R. Pick, and S. W. Hell, "Multifocal multiphoton microscopy," Opt. Lett. 23, 655-7 (1998).

A. H. Buist, M. Muller, J. Squier, and G. J. Brakenhoff, "Real time two-photon absorption microscopy using multi point excitation," J. Microscopy 192, 217-26 (1998).

K. Fujita, O. Nakamura, T. Kaneko, M. Oyamada, T. Takamatsu, and S. Kawata, "Confocal multipoint multiphoton excitation microscope with microlens and pinhole arrays," Opt. Comm. 174, 7-12 (2000).

D. N. Fittinghoff and J. A. Squier, "Time-decorrelated multifocal array for multiphoton microscopy and micromachining," Opt. Lett. 25, 1213-1215 (2000).

A. Egner and S. W. Hell, "Time multiplexing and parallelization in multifocal multiphoton microscopy," J. Opt. Soc. Am. A (Optics, Image Science and Vision) 17, 1192-201 (2000).

Heidi Dobson, "Pollen and pollen-coat lipids: chemical survey and role in pollen selection by solitary bees (Pollenkitt, Oligolecty)," PhD Dissertation, University of California, Berkeley (1985).

M. G. L. Gustafsson, D. A. Agard, and J. W. Sedat, "I/sup 5/M: 3D widefield light microscopy with better than 100 nm axial resolution," J. Microscopy 195, 10-16 (1999).

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