Abstract

We demonstrate how optical tweezers can be incorporated into a multiphoton microscope to achieve three-dimensional imaging of trapped cells. The optical tweezers, formed by a cw 1064 nm Nd:YVO4 laser, were used to trap live yeast cells in suspension while the 4′,6-diamidino-2-phenylindole-stained nucleus was imaged in three dimensions by use of a pulsed femtosecond laser. The trapped cell was moved in the axial direction by changing the position of an external lens, which was used to control the divergence of the trapping laser beam. This gives us a simple method to use optical tweezers in the laser scanning of confocal and multiphoton microscopes. It is further shown that the same femtosecond laser as used for the multiphoton imaging could also be used as laser scissors, allowing us to drill holes in the membrane of trapped spermatozoa.

© 2004 Optical Society of America

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    [CrossRef]
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  3. G. J. Brakenhoff, “Imaging modes in confocal scanning light microscopy (cslm),” J. Microsc. (Oxford) 117, 233–242 (1979).
    [CrossRef]
  4. S. Speiser, S. Kimel, “On the possibility of observing photochemical reactions induced by multiphoton absorption,” Chem. Phys. Lett. 7, 19–22 (1970).
    [CrossRef]
  5. W. Denk, J. H. Strickler, W. W. Webb, “2-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
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  6. J. H. Strickler, W. Denk, W. W. Webb, “2-photon excitation in laser scanning microscopy,” Biophys. J. 57, A374 (1990).
  7. W. Denk, “Two-photon excitation in functional biological imaging,” J. Biomed. Opt. 1, 296–304 (1996).
    [CrossRef] [PubMed]
  8. W. Denk, “Multiphoton microscopy: imaging with nonlinear optics,” Photonics Spectra 31, 125–126 (1997).
  9. W. Denk, K. Svoboda, “Photon upmanship: why multiphoton imaging is more than a gimmick,” Neuron 18, 351–357 (1997).
    [CrossRef] [PubMed]
  10. M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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  13. A. Ashkin, J. M. Dziedzic, J. E. Bjorkholm, S. Chu, “Observation of a single-beam gradient force optical trap for dielectric particles,” Opt. Lett. 11, 288–290 (1986).
    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
  15. A. Ashkin, J. M. Dziedzic, “Optical trapping and manipulation of viruses and bacteria,” Science 235, 1517–1520 (1987).
    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
  17. K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).
  18. M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
    [CrossRef] [PubMed]
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  20. K. Svoboda, S. M. Block, “Biological applications of optical forces,” Ann. Rev. Biophys. Biomol. Struct. 23, 247–285 (1994).
    [CrossRef]
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  23. K. Visscher, G. J. Brakenhoff, “Single beam optical trapping integrated in a confocal microscope for biological applications,” Cytometry 12, 486–491 (1991).
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  24. K. Visscher, G. J. Brakenhoff, J. J. Krol, “Micromanipulation by multiple optical traps created by a single fast scanning trap integrated with the bilateral confocal scanning laser microscope,” Cytometry 14, 105–114 (1993).
    [CrossRef]
  25. A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
    [CrossRef]
  26. D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
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    [CrossRef] [PubMed]
  29. U. K. Tirlapur, K. König, “Femtosecond near-infrared laser pulses as a versatile non-invasive tool for intra-tissue nanoprocessing in plants without compromising viability,” Plant J. 31, 365–374 (2002).
    [CrossRef] [PubMed]
  30. E. Fällman, O. Axner, “Design for fully steerable dual-trap optical tweezers,” Appl. Opt. 36, 2107–2113 (1997).
    [CrossRef] [PubMed]
  31. M. Goksör, J. Enger, K. Ramser, D. Hanstorp, “An experimental setup for combining optical tweezers and laser scalpels with advanced imaging techniques,” in Microarrays and Combinatorial Technologies for Biomedical Applications: Design, Fabrication and Analysis, D. V. Nicolau, R. Raghavachari, eds., Proc. SPIE4966, 50–57 (2003).
    [CrossRef]
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    [CrossRef]
  34. U. K. Tirlapur, K. König, “Near-infrared femtosecond laser pulses as a novel non-invasive means for dye-permeation and 3D imaging of localised dye-coupling in the arabidopsis root meristem,” Plant J. 20, 363–370 (1999).
    [CrossRef] [PubMed]
  35. K. König, “Multiphoton microscopy in life sciences,” J. Microsc. (Oxford) 200, 83–104 (2000).
    [CrossRef]
  36. K. König, H. Liang, M. W. Berns, B. J. Tromberg, “Cell damage in near-infrared multimode optical traps as a result of multiphoton absorption,” Opt. Lett. 21, 1090–1092 (1996).
    [CrossRef] [PubMed]
  37. K. König, “Laser tweezers are sources of two-photon excitation,” Cellul. Mol. Biol. 44, 721–733 (1998).
  38. Z. X. Zhang, G. J. Sonek, H. Liang, M. W. Berns, B. J. Tromberg, “Multiphoton fluorescence excitation in continuous-wave infrared optical traps,” Appl. Opt. 37, 2766–2773 (1998).
    [CrossRef]
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    [CrossRef]
  40. Y. Liu, G. J. Sonek, M. W. Berns, K. König, B. J. Tromberg, “2-photon fluorescence excitation in continuous-wave infrared optical tweezers,” Opt. Lett. 20, 2246–2248 (1995).
    [CrossRef]

2003 (2)

D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
[CrossRef]

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

2002 (3)

U. K. Tirlapur, K. König, “Femtosecond near-infrared laser pulses as a versatile non-invasive tool for intra-tissue nanoprocessing in plants without compromising viability,” Plant J. 31, 365–374 (2002).
[CrossRef] [PubMed]

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

F. Helmchen, W. Denk, “New developments in multiphoton microscopy,” Curr. Opin. Neurobiol. 12, 593–601 (2002).
[CrossRef] [PubMed]

2000 (3)

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

K. König, “Multiphoton microscopy in life sciences,” J. Microsc. (Oxford) 200, 83–104 (2000).
[CrossRef]

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

1999 (1)

U. K. Tirlapur, K. König, “Near-infrared femtosecond laser pulses as a novel non-invasive means for dye-permeation and 3D imaging of localised dye-coupling in the arabidopsis root meristem,” Plant J. 20, 363–370 (1999).
[CrossRef] [PubMed]

1998 (4)

1997 (3)

W. Denk, “Multiphoton microscopy: imaging with nonlinear optics,” Photonics Spectra 31, 125–126 (1997).

W. Denk, K. Svoboda, “Photon upmanship: why multiphoton imaging is more than a gimmick,” Neuron 18, 351–357 (1997).
[CrossRef] [PubMed]

E. Fällman, O. Axner, “Design for fully steerable dual-trap optical tweezers,” Appl. Opt. 36, 2107–2113 (1997).
[CrossRef] [PubMed]

1996 (4)

K. König, H. Liang, M. W. Berns, B. J. Tromberg, “Cell damage in near-infrared multimode optical traps as a result of multiphoton absorption,” Opt. Lett. 21, 1090–1092 (1996).
[CrossRef] [PubMed]

W. Denk, “Two-photon excitation in functional biological imaging,” J. Biomed. Opt. 1, 296–304 (1996).
[CrossRef] [PubMed]

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

1995 (1)

1994 (1)

K. Svoboda, S. M. Block, “Biological applications of optical forces,” Ann. Rev. Biophys. Biomol. Struct. 23, 247–285 (1994).
[CrossRef]

1993 (1)

K. Visscher, G. J. Brakenhoff, J. J. Krol, “Micromanipulation by multiple optical traps created by a single fast scanning trap integrated with the bilateral confocal scanning laser microscope,” Cytometry 14, 105–114 (1993).
[CrossRef]

1991 (1)

K. Visscher, G. J. Brakenhoff, “Single beam optical trapping integrated in a confocal microscope for biological applications,” Cytometry 12, 486–491 (1991).
[CrossRef] [PubMed]

1990 (3)

W. Denk, J. H. Strickler, W. W. Webb, “2-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

J. H. Strickler, W. Denk, W. W. Webb, “2-photon excitation in laser scanning microscopy,” Biophys. J. 57, A374 (1990).

R. M. Tyrrell, S. M. Keyse, “New trends in photobiology—the interaction of uva radiation with cultured cells,” J. Photochem. Photobiol. B 4, 349–361 (1990).
[CrossRef] [PubMed]

1987 (2)

A. Ashkin, J. M. Dziedzic, T. Yamane, “Optical trapping and manipulation of single cells using infrared-laser beams,” Nature 330, 769–771 (1987).
[CrossRef] [PubMed]

A. Ashkin, J. M. Dziedzic, “Optical trapping and manipulation of viruses and bacteria,” Science 235, 1517–1520 (1987).
[CrossRef] [PubMed]

1986 (1)

1985 (1)

M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
[CrossRef] [PubMed]

1980 (1)

C. J. R. Sheppard, T. Wilson, “Image formation in confocal scanning microscopes,” Optik 55, 331–342 (1980).

1979 (2)

G. J. Brakenhoff, “Imaging modes in confocal scanning light microscopy (cslm),” J. Microsc. (Oxford) 117, 233–242 (1979).
[CrossRef]

G. J. Brakenhoff, P. Blom, P. Barends, “Confocal scanning light microscopy with high aperture immersion lenses,” J. Microsc. (Oxford) 117, 219–232 (1979).
[CrossRef]

1970 (1)

S. Speiser, S. Kimel, “On the possibility of observing photochemical reactions induced by multiphoton absorption,” Chem. Phys. Lett. 7, 19–22 (1970).
[CrossRef]

Acker, H.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Arndt-Jovin, D. J.

Ashkin, A.

A. Ashkin, J. M. Dziedzic, T. Yamane, “Optical trapping and manipulation of single cells using infrared-laser beams,” Nature 330, 769–771 (1987).
[CrossRef] [PubMed]

A. Ashkin, J. M. Dziedzic, “Optical trapping and manipulation of viruses and bacteria,” Science 235, 1517–1520 (1987).
[CrossRef] [PubMed]

A. Ashkin, J. M. Dziedzic, J. E. Bjorkholm, S. Chu, “Observation of a single-beam gradient force optical trap for dielectric particles,” Opt. Lett. 11, 288–290 (1986).
[CrossRef] [PubMed]

Axner, O.

Barends, P.

G. J. Brakenhoff, P. Blom, P. Barends, “Confocal scanning light microscopy with high aperture immersion lenses,” J. Microsc. (Oxford) 117, 219–232 (1979).
[CrossRef]

Berchner-Pfannschmidt, U.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Berns, M. W.

Z. X. Zhang, G. J. Sonek, H. Liang, M. W. Berns, B. J. Tromberg, “Multiphoton fluorescence excitation in continuous-wave infrared optical traps,” Appl. Opt. 37, 2766–2773 (1998).
[CrossRef]

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

K. König, H. Liang, M. W. Berns, B. J. Tromberg, “Cell damage in near-infrared multimode optical traps as a result of multiphoton absorption,” Opt. Lett. 21, 1090–1092 (1996).
[CrossRef] [PubMed]

Y. Liu, G. J. Sonek, M. W. Berns, K. König, B. J. Tromberg, “2-photon fluorescence excitation in continuous-wave infrared optical tweezers,” Opt. Lett. 20, 2246–2248 (1995).
[CrossRef]

M. W. Berns, Y. Tadir, H. Liang, B. J. Tromberg, “Laser scissors and tweezers,” in Laser Tweezers in Cell Biology, M. P. Sheetz, ed. (Academic, San Diego, Calif., 1998), pp. 71–98.

Bestvater, F.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Bjorkholm, J. E.

Block, S. M.

K. Svoboda, S. M. Block, “Biological applications of optical forces,” Ann. Rev. Biophys. Biomol. Struct. 23, 247–285 (1994).
[CrossRef]

S. M. Block, “Optical tweezers: a new tool for biophysics,” in Noninvasive Techniques in Cell Biology, J. K. Foskett, S. Grinstein, eds. (Wiley-Liss, New York, 1990), pp. 372–402.

Blom, P.

G. J. Brakenhoff, P. Blom, P. Barends, “Confocal scanning light microscopy with high aperture immersion lenses,” J. Microsc. (Oxford) 117, 219–232 (1979).
[CrossRef]

Booth, M.

Brakenhoff, G. J.

K. Visscher, G. J. Brakenhoff, J. J. Krol, “Micromanipulation by multiple optical traps created by a single fast scanning trap integrated with the bilateral confocal scanning laser microscope,” Cytometry 14, 105–114 (1993).
[CrossRef]

K. Visscher, G. J. Brakenhoff, “Single beam optical trapping integrated in a confocal microscope for biological applications,” Cytometry 12, 486–491 (1991).
[CrossRef] [PubMed]

G. J. Brakenhoff, P. Blom, P. Barends, “Confocal scanning light microscopy with high aperture immersion lenses,” J. Microsc. (Oxford) 117, 219–232 (1979).
[CrossRef]

G. J. Brakenhoff, “Imaging modes in confocal scanning light microscopy (cslm),” J. Microsc. (Oxford) 117, 233–242 (1979).
[CrossRef]

Chu, S.

Cunningham, M. L.

M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
[CrossRef] [PubMed]

Denk, W.

F. Helmchen, W. Denk, “New developments in multiphoton microscopy,” Curr. Opin. Neurobiol. 12, 593–601 (2002).
[CrossRef] [PubMed]

W. Denk, “Two-photon fluorescence microscopy: high resolution imaging in scattering tissue,” Eur. J. Neurosci. 10, 10602 (1998).

W. Denk, “Multiphoton microscopy: imaging with nonlinear optics,” Photonics Spectra 31, 125–126 (1997).

W. Denk, K. Svoboda, “Photon upmanship: why multiphoton imaging is more than a gimmick,” Neuron 18, 351–357 (1997).
[CrossRef] [PubMed]

W. Denk, “Two-photon excitation in functional biological imaging,” J. Biomed. Opt. 1, 296–304 (1996).
[CrossRef] [PubMed]

W. Denk, J. H. Strickler, W. W. Webb, “2-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

J. H. Strickler, W. Denk, W. W. Webb, “2-photon excitation in laser scanning microscopy,” Biophys. J. 57, A374 (1990).

Diez, A.

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

Dziedzic, J. M.

A. Ashkin, J. M. Dziedzic, “Optical trapping and manipulation of viruses and bacteria,” Science 235, 1517–1520 (1987).
[CrossRef] [PubMed]

A. Ashkin, J. M. Dziedzic, T. Yamane, “Optical trapping and manipulation of single cells using infrared-laser beams,” Nature 330, 769–771 (1987).
[CrossRef] [PubMed]

A. Ashkin, J. M. Dziedzic, J. E. Bjorkholm, S. Chu, “Observation of a single-beam gradient force optical trap for dielectric particles,” Opt. Lett. 11, 288–290 (1986).
[CrossRef] [PubMed]

Eliceiri, K. W.

D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
[CrossRef]

Enger, J.

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

M. Goksör, J. Enger, K. Ramser, D. Hanstorp, “An experimental setup for combining optical tweezers and laser scalpels with advanced imaging techniques,” in Microarrays and Combinatorial Technologies for Biomedical Applications: Design, Fabrication and Analysis, D. V. Nicolau, R. Raghavachari, eds., Proc. SPIE4966, 50–57 (2003).
[CrossRef]

Ericsson, M.

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

Fällman, E.

Feurer, T.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Giovanazzi, S. M.

M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
[CrossRef] [PubMed]

Goksör, M.

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

M. Goksör, J. Enger, K. Ramser, D. Hanstorp, “An experimental setup for combining optical tweezers and laser scalpels with advanced imaging techniques,” in Microarrays and Combinatorial Technologies for Biomedical Applications: Design, Fabrication and Analysis, D. V. Nicolau, R. Raghavachari, eds., Proc. SPIE4966, 50–57 (2003).
[CrossRef]

Greulich, K. O.

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

K. O. Greulich, Micromanipulation by Light in Biology and Medicine (Birkhäuser, Basel, Switzerland, 1999).

Hacker, M.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Hagberg, P.

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

Hanstorp, D.

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

M. Goksör, J. Enger, K. Ramser, D. Hanstorp, “An experimental setup for combining optical tweezers and laser scalpels with advanced imaging techniques,” in Microarrays and Combinatorial Technologies for Biomedical Applications: Design, Fabrication and Analysis, D. V. Nicolau, R. Raghavachari, eds., Proc. SPIE4966, 50–57 (2003).
[CrossRef]

Haugland, R. P.

R. P. Haugland, Handbook of Fluorescent Probes and Research Products (Molecular Probes, Leiden, The Netherlands, 2002).

Hell, S. W.

Helmchen, F.

F. Helmchen, W. Denk, “New developments in multiphoton microscopy,” Curr. Opin. Neurobiol. 12, 593–601 (2002).
[CrossRef] [PubMed]

Hoffmann, A.

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

Horste, G. M. Z.

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

Johnson, J. S.

M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
[CrossRef] [PubMed]

Jovin, T. M.

Keyse, S. M.

R. M. Tyrrell, S. M. Keyse, “New trends in photobiology—the interaction of uva radiation with cultured cells,” J. Photochem. Photobiol. B 4, 349–361 (1990).
[CrossRef] [PubMed]

Kimel, S.

S. Speiser, S. Kimel, “On the possibility of observing photochemical reactions induced by multiphoton absorption,” Chem. Phys. Lett. 7, 19–22 (1970).
[CrossRef]

Kirsch, A. K.

König, K.

U. K. Tirlapur, K. König, “Femtosecond near-infrared laser pulses as a versatile non-invasive tool for intra-tissue nanoprocessing in plants without compromising viability,” Plant J. 31, 365–374 (2002).
[CrossRef] [PubMed]

K. König, “Multiphoton microscopy in life sciences,” J. Microsc. (Oxford) 200, 83–104 (2000).
[CrossRef]

U. K. Tirlapur, K. König, “Near-infrared femtosecond laser pulses as a novel non-invasive means for dye-permeation and 3D imaging of localised dye-coupling in the arabidopsis root meristem,” Plant J. 20, 363–370 (1999).
[CrossRef] [PubMed]

K. König, “Laser tweezers are sources of two-photon excitation,” Cellul. Mol. Biol. 44, 721–733 (1998).

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

K. König, H. Liang, M. W. Berns, B. J. Tromberg, “Cell damage in near-infrared multimode optical traps as a result of multiphoton absorption,” Opt. Lett. 21, 1090–1092 (1996).
[CrossRef] [PubMed]

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

Y. Liu, G. J. Sonek, M. W. Berns, K. König, B. J. Tromberg, “2-photon fluorescence excitation in continuous-wave infrared optical tweezers,” Opt. Lett. 20, 2246–2248 (1995).
[CrossRef]

Krol, J. J.

K. Visscher, G. J. Brakenhoff, J. J. Krol, “Micromanipulation by multiple optical traps created by a single fast scanning trap integrated with the bilateral confocal scanning laser microscope,” Cytometry 14, 105–114 (1993).
[CrossRef]

Liang, H.

Liu, Y.

Liu, Y. G.

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

Monajembashi, S.

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

Nyström, T.

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

Patrizio, P.

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

Peak, M. J.

M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
[CrossRef] [PubMed]

Pilarczyk, G.

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

Porwol, T.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Ramser, K.

M. Goksör, J. Enger, K. Ramser, D. Hanstorp, “An experimental setup for combining optical tweezers and laser scalpels with advanced imaging techniques,” in Microarrays and Combinatorial Technologies for Biomedical Applications: Design, Fabrication and Analysis, D. V. Nicolau, R. Raghavachari, eds., Proc. SPIE4966, 50–57 (2003).
[CrossRef]

Schnetter, C. M.

Sheppard, C. J. R.

C. J. R. Sheppard, T. Wilson, “Image formation in confocal scanning microscopes,” Optik 55, 331–342 (1980).

Sonek, G.

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

Sonek, G. J.

Speiser, S.

S. Speiser, S. Kimel, “On the possibility of observing photochemical reactions induced by multiphoton absorption,” Chem. Phys. Lett. 7, 19–22 (1970).
[CrossRef]

Spiess, E.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Squirrell, J. M.

D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
[CrossRef]

Stobrawa, G.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Strickler, J. H.

W. Denk, J. H. Strickler, W. W. Webb, “2-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

J. H. Strickler, W. Denk, W. W. Webb, “2-photon excitation in laser scanning microscopy,” Biophys. J. 57, A374 (1990).

Svaasand, L.

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

Svoboda, K.

W. Denk, K. Svoboda, “Photon upmanship: why multiphoton imaging is more than a gimmick,” Neuron 18, 351–357 (1997).
[CrossRef] [PubMed]

K. Svoboda, S. M. Block, “Biological applications of optical forces,” Ann. Rev. Biophys. Biomol. Struct. 23, 247–285 (1994).
[CrossRef]

Tadir, Y.

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

M. W. Berns, Y. Tadir, H. Liang, B. J. Tromberg, “Laser scissors and tweezers,” in Laser Tweezers in Cell Biology, M. P. Sheetz, ed. (Academic, San Diego, Calif., 1998), pp. 71–98.

Tirlapur, U. K.

U. K. Tirlapur, K. König, “Femtosecond near-infrared laser pulses as a versatile non-invasive tool for intra-tissue nanoprocessing in plants without compromising viability,” Plant J. 31, 365–374 (2002).
[CrossRef] [PubMed]

U. K. Tirlapur, K. König, “Near-infrared femtosecond laser pulses as a novel non-invasive means for dye-permeation and 3D imaging of localised dye-coupling in the arabidopsis root meristem,” Plant J. 20, 363–370 (1999).
[CrossRef] [PubMed]

Tromberg, B. J.

Z. X. Zhang, G. J. Sonek, H. Liang, M. W. Berns, B. J. Tromberg, “Multiphoton fluorescence excitation in continuous-wave infrared optical traps,” Appl. Opt. 37, 2766–2773 (1998).
[CrossRef]

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

K. König, H. Liang, M. W. Berns, B. J. Tromberg, “Cell damage in near-infrared multimode optical traps as a result of multiphoton absorption,” Opt. Lett. 21, 1090–1092 (1996).
[CrossRef] [PubMed]

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

Y. Liu, G. J. Sonek, M. W. Berns, K. König, B. J. Tromberg, “2-photon fluorescence excitation in continuous-wave infrared optical tweezers,” Opt. Lett. 20, 2246–2248 (1995).
[CrossRef]

M. W. Berns, Y. Tadir, H. Liang, B. J. Tromberg, “Laser scissors and tweezers,” in Laser Tweezers in Cell Biology, M. P. Sheetz, ed. (Academic, San Diego, Calif., 1998), pp. 71–98.

Tyrrell, R. M.

R. M. Tyrrell, S. M. Keyse, “New trends in photobiology—the interaction of uva radiation with cultured cells,” J. Photochem. Photobiol. B 4, 349–361 (1990).
[CrossRef] [PubMed]

Uhl, V.

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

Visscher, K.

K. Visscher, G. J. Brakenhoff, J. J. Krol, “Micromanipulation by multiple optical traps created by a single fast scanning trap integrated with the bilateral confocal scanning laser microscope,” Cytometry 14, 105–114 (1993).
[CrossRef]

K. Visscher, G. J. Brakenhoff, “Single beam optical trapping integrated in a confocal microscope for biological applications,” Cytometry 12, 486–491 (1991).
[CrossRef] [PubMed]

Webb, W. W.

W. Denk, J. H. Strickler, W. W. Webb, “2-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

J. H. Strickler, W. Denk, W. W. Webb, “2-photon excitation in laser scanning microscopy,” Biophys. J. 57, A374 (1990).

White, J. G.

D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
[CrossRef]

Wilms, S.

Wilson, T.

C. J. R. Sheppard, T. Wilson, “Image formation in confocal scanning microscopes,” Optik 55, 331–342 (1980).

Wokosin, D. L.

D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
[CrossRef]

Wotzlaw, C.

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

Yamane, T.

A. Ashkin, J. M. Dziedzic, T. Yamane, “Optical trapping and manipulation of single cells using infrared-laser beams,” Nature 330, 769–771 (1987).
[CrossRef] [PubMed]

Zhang, Z. X.

Ann. Rev. Biophys. Biomol. Struct. (1)

K. Svoboda, S. M. Block, “Biological applications of optical forces,” Ann. Rev. Biophys. Biomol. Struct. 23, 247–285 (1994).
[CrossRef]

Appl. Opt. (2)

Appl. Phys. B (1)

A. Hoffmann, G. M. Z. Horste, G. Pilarczyk, S. Monajembashi, V. Uhl, K. O. Greulich, “Optical tweezers for confocal microscopy,” Appl. Phys. B 71, 747–753 (2000).
[CrossRef]

Biophys. J. (1)

J. H. Strickler, W. Denk, W. W. Webb, “2-photon excitation in laser scanning microscopy,” Biophys. J. 57, A374 (1990).

Cellul. Mol. Biol. (2)

K. König, “Laser tweezers are sources of two-photon excitation,” Cellul. Mol. Biol. 44, 721–733 (1998).

K. König, L. Svaasand, Y. G. Liu, G. Sonek, P. Patrizio, Y. Tadir, M. W. Berns, B. J. Tromberg, “Determination of motility forces of human spermatozoa using an 800 nm optical trap,” Cellul. Mol. Biol. 42, 501–509 (1996).

Chem. Phys. Lett. (1)

S. Speiser, S. Kimel, “On the possibility of observing photochemical reactions induced by multiphoton absorption,” Chem. Phys. Lett. 7, 19–22 (1970).
[CrossRef]

Curr. Opin. Neurobiol. (1)

F. Helmchen, W. Denk, “New developments in multiphoton microscopy,” Curr. Opin. Neurobiol. 12, 593–601 (2002).
[CrossRef] [PubMed]

Cytometry (2)

K. Visscher, G. J. Brakenhoff, “Single beam optical trapping integrated in a confocal microscope for biological applications,” Cytometry 12, 486–491 (1991).
[CrossRef] [PubMed]

K. Visscher, G. J. Brakenhoff, J. J. Krol, “Micromanipulation by multiple optical traps created by a single fast scanning trap integrated with the bilateral confocal scanning laser microscope,” Cytometry 14, 105–114 (1993).
[CrossRef]

EMBO Rep. (1)

M. Goksör, A. Diez, J. Enger, D. Hanstorp, T. Nyström, “Analysis of molecular diffusion in ftsK cell division mutants using laser surgery,” EMBO Rep. 4, 867–871 (2003).
[CrossRef] [PubMed]

Eur. J. Neurosci. (1)

W. Denk, “Two-photon fluorescence microscopy: high resolution imaging in scattering tissue,” Eur. J. Neurosci. 10, 10602 (1998).

Hum. Reprod. (1)

K. König, Y. Tadir, P. Patrizio, M. W. Berns, B. J. Tromberg, “Effects of ultraviolet exposure and near infrared laser tweezers on human spermatozoa,” Hum. Reprod. 11, 2162–2164 (1996).
[CrossRef] [PubMed]

J. Bacteriol. (1)

M. Ericsson, D. Hanstorp, P. Hagberg, J. Enger, T. Nyström, “Sorting out bacterial viability with optical tweezers,” J. Bacteriol. 182, 5551–5555 (2000).
[CrossRef] [PubMed]

J. Biomed. Opt. (1)

W. Denk, “Two-photon excitation in functional biological imaging,” J. Biomed. Opt. 1, 296–304 (1996).
[CrossRef] [PubMed]

J. Microsc. (Oxford) (4)

F. Bestvater, E. Spiess, G. Stobrawa, M. Hacker, T. Feurer, T. Porwol, U. Berchner-Pfannschmidt, C. Wotzlaw, H. Acker, “Two-photon fluorescence absorption and emission spectra of dyes relevant for cell imaging,” J. Microsc. (Oxford) 208, 108–115 (2002).
[CrossRef]

K. König, “Multiphoton microscopy in life sciences,” J. Microsc. (Oxford) 200, 83–104 (2000).
[CrossRef]

G. J. Brakenhoff, “Imaging modes in confocal scanning light microscopy (cslm),” J. Microsc. (Oxford) 117, 233–242 (1979).
[CrossRef]

G. J. Brakenhoff, P. Blom, P. Barends, “Confocal scanning light microscopy with high aperture immersion lenses,” J. Microsc. (Oxford) 117, 219–232 (1979).
[CrossRef]

J. Photochem. Photobiol. B (1)

R. M. Tyrrell, S. M. Keyse, “New trends in photobiology—the interaction of uva radiation with cultured cells,” J. Photochem. Photobiol. B 4, 349–361 (1990).
[CrossRef] [PubMed]

Nature (1)

A. Ashkin, J. M. Dziedzic, T. Yamane, “Optical trapping and manipulation of single cells using infrared-laser beams,” Nature 330, 769–771 (1987).
[CrossRef] [PubMed]

Neuron (1)

W. Denk, K. Svoboda, “Photon upmanship: why multiphoton imaging is more than a gimmick,” Neuron 18, 351–357 (1997).
[CrossRef] [PubMed]

Opt. Lett. (4)

Optik (1)

C. J. R. Sheppard, T. Wilson, “Image formation in confocal scanning microscopes,” Optik 55, 331–342 (1980).

Photochem. Photobiol. (1)

M. L. Cunningham, J. S. Johnson, S. M. Giovanazzi, M. J. Peak, “Photosensitized production of superoxide anion by monochromatic (290–405 nm) ultraviolet-irradiation of nadh and nadph coenzymes,” Photochem. Photobiol. 42, 125–128 (1985).
[CrossRef] [PubMed]

Photonics Spectra (1)

W. Denk, “Multiphoton microscopy: imaging with nonlinear optics,” Photonics Spectra 31, 125–126 (1997).

Plant J. (2)

U. K. Tirlapur, K. König, “Near-infrared femtosecond laser pulses as a novel non-invasive means for dye-permeation and 3D imaging of localised dye-coupling in the arabidopsis root meristem,” Plant J. 20, 363–370 (1999).
[CrossRef] [PubMed]

U. K. Tirlapur, K. König, “Femtosecond near-infrared laser pulses as a versatile non-invasive tool for intra-tissue nanoprocessing in plants without compromising viability,” Plant J. 31, 365–374 (2002).
[CrossRef] [PubMed]

Rev. Sci. Instrum. (1)

D. L. Wokosin, J. M. Squirrell, K. W. Eliceiri, J. G. White, “Optical workstation with concurrent, independent multiphoton imaging and experimental laser microbeam capabilities,” Rev. Sci. Instrum. 74, 193–201 (2003).
[CrossRef]

Science (2)

A. Ashkin, J. M. Dziedzic, “Optical trapping and manipulation of viruses and bacteria,” Science 235, 1517–1520 (1987).
[CrossRef] [PubMed]

W. Denk, J. H. Strickler, W. W. Webb, “2-photon laser scanning fluorescence microscopy,” Science 248, 73–76 (1990).
[CrossRef] [PubMed]

Other (5)

M. Goksör, J. Enger, K. Ramser, D. Hanstorp, “An experimental setup for combining optical tweezers and laser scalpels with advanced imaging techniques,” in Microarrays and Combinatorial Technologies for Biomedical Applications: Design, Fabrication and Analysis, D. V. Nicolau, R. Raghavachari, eds., Proc. SPIE4966, 50–57 (2003).
[CrossRef]

R. P. Haugland, Handbook of Fluorescent Probes and Research Products (Molecular Probes, Leiden, The Netherlands, 2002).

S. M. Block, “Optical tweezers: a new tool for biophysics,” in Noninvasive Techniques in Cell Biology, J. K. Foskett, S. Grinstein, eds. (Wiley-Liss, New York, 1990), pp. 372–402.

K. O. Greulich, Micromanipulation by Light in Biology and Medicine (Birkhäuser, Basel, Switzerland, 1999).

M. W. Berns, Y. Tadir, H. Liang, B. J. Tromberg, “Laser scissors and tweezers,” in Laser Tweezers in Cell Biology, M. P. Sheetz, ed. (Academic, San Diego, Calif., 1998), pp. 71–98.

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Figures (4)

Fig. 1
Fig. 1

Experimental setup for the optical tweezers in combination with the detectors used for multiphoton and confocal imaging. The laser beam forming the optical tweezers, shown in red, is controlled by a shutter (S) and expanded by a 1:4 beam expander (BE). The beam is guided by a mirror (M) and thereafter split in two arms by use of a beam-splitting cube (BSC). One arm is used for multiphoton imaging. The IR trapping laser is then reflected by a gimbal mounted mirror (GMM) and focused by the external lens L3, which has been mounted on a translator stage. The trapping laser light is transmitted through the first dichroic mirror (DM2) and reflected by the second dichroic mirror (DM1) into the back aperture of the microscope objective (MO). Finally, the optical trap is formed in the focal plane of the objective. For trapping in confocal mode, the trapping beam is instead guided through the fluorescence arm by means of the dichroic mirror DM4. The optical paths for imaging are shown in blue. CL and TD are the condenser lens and the transmission detector, respectively. CH, cassette holder; D1 and D2, direct detectors; AOTF, acousto-optic tunable filter; Hg, mercury lamp; BSU, beam-shaping unit; ICU, instrumental control unit; EP, eye piece; HF, heat filter; EF, excitation filter; ND, neutral density filter; BF, blocking filter; DM3, dichroic mirror.

Fig. 2
Fig. 2

A 3D image of the nucleus of a trapped yeast cell and three optical sections. The cell was trapped by applying optical tweezers, and the DAPI-stained nucleus was excited by use of a pulsed femtosecond laser. The emission was detected by use of the detectors for multiphoton imaging (D1 in Fig. 1). The field view of the 3D image is approximately 5 × 5 μm, whereas the optical sections are approximately 2.5 × 2.5 μm. The full 3D image can be found at www.fy.chalmers.se/atom/research/tweezers/nucleus.

Fig. 3
Fig. 3

(a) Transmission image of a spermatozoa trapped by the optical tweezers. The cell is trapped in a media containing two fluorescent dyes, and their fluorescence response are shown in (b). The first dye, propidium iodide (detector 1 in the ICU), cannot penetrate an intact cell membrane. For SYTO9 (detector 2 in the ICU), on the other hand, the cell membrane is permeable. With maximum output power, the pulsed femtosecond laser is used to weaken the membrane of the spermatozoa until the PI dye can leak through. As the dye starts to diffuse into the cell, the emitted signal shifts from detector 2 to detector 1. The images in (b) were taken 0, 30, and 60 s after incision. The fluorescence was measured by use of the detectors for confocal imaging.

Fig. 4
Fig. 4

The cross section of the trapping laser is high enough to induce two-photon excitation of the trapped object. (a) shows a trapped spermatozoa by use of the transmission detector. The cell has been labeled with a fluorophore that normally is excited around 530 nm, but is now excited with the 1064-nm optical tweezers. (b) shows the fluorescence signal that has been accumulated for 1 min by use of the detectors for confocal imaging.

Equations (1)

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Δz=fobjectiveflens2Δd,

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