Abstract

In this work, we investigate the non-ablative, non-thermal photo-modification of collagen fibers by femtosecond Ti:Sa laser. The effect was induced and simultaneously registered during the repetitive laser scanning of type I collagen (rat tail and bovine Achilles’ tendon), and bovine cornea. An irreversible increase in two-photon autofluorescence and a decrease in second harmonic generation intensities were associated with the collagen femtosecond laser photo-modification. Confocal spectral imaging revealed the formation of new fluorescent species. Controllable nonlinear photo-modification of collagen fibers and bovine cornea with ~2 µm spatial resolution was demonstrated.

© 2008 Optical Society of America

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    [CrossRef]
  2. W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
    [CrossRef]
  3. R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).
  4. T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
    [CrossRef]
  5. K. König, I. Riemann, and W. Fritzsche, "Nanodissection of human chromosomes with near-infrared femtosecond laser pulses," Opt. Lett. 26, 819-821 (2001).
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  6. C. Schaffer, N. Nishimura, E. Glezer, A. Kim, and E. Mazur, "Dynamics of femtosecond laser-induced breakdown in water from femtoseconds to microseconds," Opt. Express 10, 196-203 (2002).
    [PubMed]
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  9. M. Han, G. Giese, L. Zickler, H. Sun, and J. F. Bille, "Mini-invasive corneal surgery and imaging with femtosecond lasers," Opt. Express 12, 4275-4281 (2004).
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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    [CrossRef] [PubMed]
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2007 (5)

B. G. Wang, I. Riemann, H. Schubert, K. J. Halbhuber, and K. Koenig, "In-vivo intratissue ablation by nanojoule near-infrared femtosecond laser pulses," Cell Tissue Res. 328, 515-520 (2007).
[CrossRef] [PubMed]

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

2006 (2)

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

T. A. Theodossiou, C. Thrasivoulou, C. Ekwobi, and D. L. Becker, "Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections," Biophys. J. 91, 4665-4677 (2006).
[CrossRef] [PubMed]

2005 (3)

A. Vogel, J. Noack, G. Huttman, and G. Paltauf, "Mechanisms of femtosecond laser nanosurgery of cells and tissues," Appl. Phys. B 81, 1015-1047 (2005).
[CrossRef]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

2004 (2)

M. F. Yanik, H. Cinar, N. Cinar, A. Chisholm, Y. Jin, and A. B. Yakar, "Neurosurgery: Functional regeneration after laser axotomy," Nature 432, 882 (2004).
[CrossRef]

M. Han, G. Giese, L. Zickler, H. Sun, and J. F. Bille, "Mini-invasive corneal surgery and imaging with femtosecond lasers," Opt. Express 12, 4275-4281 (2004).
[CrossRef] [PubMed]

2002 (3)

C. Schaffer, N. Nishimura, E. Glezer, A. Kim, and E. Mazur, "Dynamics of femtosecond laser-induced breakdown in water from femtoseconds to microseconds," Opt. Express 10, 196-203 (2002).
[PubMed]

K. König, O. Krauss, and I. Riemann, "Intratissue surgery with 80 MHz nanojoule femtosecond laser pulses in the near infrared," Opt. Express. 10, 171-176 (2002).

T. Theodossiou, G. S.  Rapti, V.  Hovhannisyan, E.  Georgiou, K.  Politopoulos, and D.  Yova, "Thermally induced irreversible conformational changes in collagen probed by optical second harmonic generation and laser-induced fluorescence," Lasers Med. Sci. 17, 34-41 (2002).
[CrossRef] [PubMed]

2001 (2)

A. I. Rem, J. A. Oosterhuis, H. G. Journée-de Korver, T. J van den Berg, and J. E. Keunen, "Temperature dependence of thermal damage to the sclera: exploring the heat tolerance of the sclera for transscleral thermotherapy," Exp. Eye Res. 72, 153-62 (2001).
[CrossRef] [PubMed]

K. König, I. Riemann, and W. Fritzsche, "Nanodissection of human chromosomes with near-infrared femtosecond laser pulses," Opt. Lett. 26, 819-821 (2001).
[CrossRef]

2000 (1)

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, "Collagen structure and nonlinear susceptibility: effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity," Lasers Surg. Med. 27, 329-335 (2000).
[CrossRef] [PubMed]

1999 (1)

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

1997 (2)

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

R. I. Price, S. Lees, and D. A. Kirschner, "X-ray diffraction analysis of tendon collagen at ambient and cryogenic temperatures: role of hydration," Int. J. Biol. Macromol. 20, 23-33 (1997).
[CrossRef] [PubMed]

1994 (1)

W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
[CrossRef]

1989 (1)

D. Stern, C. A. Puliafito, E. T. Dobei, and W. T. Reidy, "Corneal ablation by nanosecond, picosecond and femtosecond laser pulses at 532 nm and 625 nm," Arch. Ophthalmol. 107, 587-592 (1989).
[CrossRef]

1979 (1)

D. J. S. Hulmes and A. Miller, "Quasi-hexagonal molecular packing in collagen fibrils," Nature 282, 878-880 (1979).
[CrossRef] [PubMed]

1969 (1)

E. D. Eanes and E. J. Miller, "Effect of covalent cross-linking on the X-ray diffraction properties of chick bone and rat tail tendon collagens," Arch. Biochem. Biophys. 129, 769-771 (1969).
[CrossRef] [PubMed]

Abrusci, C.

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

Akiyama, S.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Andreeva, I. V.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Averkiev, S. V.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Bartels, A.

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

Becker, D. L.

T. A. Theodossiou, C. Thrasivoulou, C. Ekwobi, and D. L. Becker, "Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections," Biophys. J. 91, 4665-4677 (2006).
[CrossRef] [PubMed]

Bille, J. F.

M. Han, G. Giese, L. Zickler, H. Sun, and J. F. Bille, "Mini-invasive corneal surgery and imaging with femtosecond lasers," Opt. Express 12, 4275-4281 (2004).
[CrossRef] [PubMed]

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

Castillejo, M.

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

Catalina, F.

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

Chen, W. L.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

Chen, Y. F.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

Chisholm, A.

M. F. Yanik, H. Cinar, N. Cinar, A. Chisholm, Y. Jin, and A. B. Yakar, "Neurosurgery: Functional regeneration after laser axotomy," Nature 432, 882 (2004).
[CrossRef]

Cinar, H.

M. F. Yanik, H. Cinar, N. Cinar, A. Chisholm, Y. Jin, and A. B. Yakar, "Neurosurgery: Functional regeneration after laser axotomy," Nature 432, 882 (2004).
[CrossRef]

Cinar, N.

M. F. Yanik, H. Cinar, N. Cinar, A. Chisholm, Y. Jin, and A. B. Yakar, "Neurosurgery: Functional regeneration after laser axotomy," Nature 432, 882 (2004).
[CrossRef]

Da Silva, L. B.

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, "Collagen structure and nonlinear susceptibility: effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity," Lasers Surg. Med. 27, 329-335 (2000).
[CrossRef] [PubMed]

Del Amo, A.

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

Dobei, E. T.

D. Stern, C. A. Puliafito, E. T. Dobei, and W. T. Reidy, "Corneal ablation by nanosecond, picosecond and femtosecond laser pulses at 532 nm and 625 nm," Arch. Ophthalmol. 107, 587-592 (1989).
[CrossRef]

Dong, C. Y.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

Du, D.

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

Eanes, E. D.

E. D. Eanes and E. J. Miller, "Effect of covalent cross-linking on the X-ray diffraction properties of chick bone and rat tail tendon collagens," Arch. Biochem. Biophys. 129, 769-771 (1969).
[CrossRef] [PubMed]

Ehlers, A.

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

Eichler, J.

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, "Collagen structure and nonlinear susceptibility: effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity," Lasers Surg. Med. 27, 329-335 (2000).
[CrossRef] [PubMed]

Ekwobi, C.

T. A. Theodossiou, C. Thrasivoulou, C. Ekwobi, and D. L. Becker, "Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections," Biophys. J. 91, 4665-4677 (2006).
[CrossRef] [PubMed]

Elner, V. M.

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

Fritzsche, W.

Fukusaki, E.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Georgiou, E.

T. Theodossiou, G. S.  Rapti, V.  Hovhannisyan, E.  Georgiou, K.  Politopoulos, and D.  Yova, "Thermally induced irreversible conformational changes in collagen probed by optical second harmonic generation and laser-induced fluorescence," Lasers Med. Sci. 17, 34-41 (2002).
[CrossRef] [PubMed]

Giese, G.

Glezer, E.

Grabner, G.

W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
[CrossRef]

Halbhuber, K. J.

B. G. Wang, I. Riemann, H. Schubert, K. J. Halbhuber, and K. Koenig, "In-vivo intratissue ablation by nanojoule near-infrared femtosecond laser pulses," Cell Tissue Res. 328, 515-520 (2007).
[CrossRef] [PubMed]

Han, M.

Hirao, K.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Horvath, C.

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

Hovhannisyan, V.

T. Theodossiou, G. S.  Rapti, V.  Hovhannisyan, E.  Georgiou, K.  Politopoulos, and D.  Yova, "Thermally induced irreversible conformational changes in collagen probed by optical second harmonic generation and laser-induced fluorescence," Lasers Med. Sci. 17, 34-41 (2002).
[CrossRef] [PubMed]

Hsiao, C. Y.

Hulmes, D. J. S.

D. J. S. Hulmes and A. Miller, "Quasi-hexagonal molecular packing in collagen fibrils," Nature 282, 878-880 (1979).
[CrossRef] [PubMed]

Husinsky, W.

W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
[CrossRef]

Huttman, G.

A. Vogel, J. Noack, G. Huttman, and G. Paltauf, "Mechanisms of femtosecond laser nanosurgery of cells and tissues," Appl. Phys. B 81, 1015-1047 (2005).
[CrossRef]

Ignatieva, N. Yu.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Jan, G. J.

Janke, C.

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

Jee, S. H.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

Jin, Y.

M. F. Yanik, H. Cinar, N. Cinar, A. Chisholm, Y. Jin, and A. B. Yakar, "Neurosurgery: Functional regeneration after laser axotomy," Nature 432, 882 (2004).
[CrossRef]

Journée-de Korver, H. G.

A. I. Rem, J. A. Oosterhuis, H. G. Journée-de Korver, T. J van den Berg, and J. E. Keunen, "Temperature dependence of thermal damage to the sclera: exploring the heat tolerance of the sclera for transscleral thermotherapy," Exp. Eye Res. 72, 153-62 (2001).
[CrossRef] [PubMed]

Juhasz, T.

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

Kajiyama, S.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Kamensky, V. A.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Kautek, W.

W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
[CrossRef]

Kim, A.

Kim, B. M.

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, "Collagen structure and nonlinear susceptibility: effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity," Lasers Surg. Med. 27, 329-335 (2000).
[CrossRef] [PubMed]

Kirschner, D. A.

R. I. Price, S. Lees, and D. A. Kirschner, "X-ray diffraction analysis of tendon collagen at ambient and cryogenic temperatures: role of hydration," Int. J. Biol. Macromol. 20, 23-33 (1997).
[CrossRef] [PubMed]

Koenig, K.

B. G. Wang, I. Riemann, H. Schubert, K. J. Halbhuber, and K. Koenig, "In-vivo intratissue ablation by nanojoule near-infrared femtosecond laser pulses," Cell Tissue Res. 328, 515-520 (2007).
[CrossRef] [PubMed]

König, K.

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

K. König, O. Krauss, and I. Riemann, "Intratissue surgery with 80 MHz nanojoule femtosecond laser pulses in the near infrared," Opt. Express. 10, 171-176 (2002).

K. König, I. Riemann, and W. Fritzsche, "Nanodissection of human chromosomes with near-infrared femtosecond laser pulses," Opt. Lett. 26, 819-821 (2001).
[CrossRef]

Krauss, O.

K. König, O. Krauss, and I. Riemann, "Intratissue surgery with 80 MHz nanojoule femtosecond laser pulses in the near infrared," Opt. Express. 10, 171-176 (2002).

Kruger, J.

W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
[CrossRef]

Kurtz, R. M.

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

Le Harzic, R.

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

Lees, S.

R. I. Price, S. Lees, and D. A. Kirschner, "X-ray diffraction analysis of tendon collagen at ambient and cryogenic temperatures: role of hydration," Int. J. Biol. Macromol. 20, 23-33 (1997).
[CrossRef] [PubMed]

Lin, L. C.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

Lin, S. J.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

Lin, W. C.

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

Liu, X.

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

Lo, W.

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

Loesel, F. H.

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

Lunin, V. V.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Martin, S.

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

Mazur, E.

Miller, A.

D. J. S. Hulmes and A. Miller, "Quasi-hexagonal molecular packing in collagen fibrils," Nature 282, 878-880 (1979).
[CrossRef] [PubMed]

Miller, E. J.

E. D. Eanes and E. J. Miller, "Effect of covalent cross-linking on the X-ray diffraction properties of chick bone and rat tail tendon collagens," Arch. Biochem. Biophys. 129, 769-771 (1969).
[CrossRef] [PubMed]

Mitterer, S.

W. Kautek, S. Mitterer, J. Kruger, W. Husinsky, and G. Grabner, "Femtosecond-pulse laser ablation of human corneas," Appl. Phys. A 58, 513-518 (1994).
[CrossRef]

Miura, K.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Mouro, G.

T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
[CrossRef]

Mourou, G. A.

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

Myakov, A. V.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Nishimura, N.

Noack, J.

A. Vogel, J. Noack, G. Huttman, and G. Paltauf, "Mechanisms of femtosecond laser nanosurgery of cells and tissues," Appl. Phys. B 81, 1015-1047 (2005).
[CrossRef]

Oosterhuis, J. A.

A. I. Rem, J. A. Oosterhuis, H. G. Journée-de Korver, T. J van den Berg, and J. E. Keunen, "Temperature dependence of thermal damage to the sclera: exploring the heat tolerance of the sclera for transscleral thermotherapy," Exp. Eye Res. 72, 153-62 (2001).
[CrossRef] [PubMed]

Oujja, M.

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

Paltauf, G.

A. Vogel, J. Noack, G. Huttman, and G. Paltauf, "Mechanisms of femtosecond laser nanosurgery of cells and tissues," Appl. Phys. B 81, 1015-1047 (2005).
[CrossRef]

Politopoulos, K.

T. Theodossiou, G. S.  Rapti, V.  Hovhannisyan, E.  Georgiou, K.  Politopoulos, and D.  Yova, "Thermally induced irreversible conformational changes in collagen probed by optical second harmonic generation and laser-induced fluorescence," Lasers Med. Sci. 17, 34-41 (2002).
[CrossRef] [PubMed]

Price, R. I.

R. I. Price, S. Lees, and D. A. Kirschner, "X-ray diffraction analysis of tendon collagen at ambient and cryogenic temperatures: role of hydration," Int. J. Biol. Macromol. 20, 23-33 (1997).
[CrossRef] [PubMed]

Puliafito, C. A.

D. Stern, C. A. Puliafito, E. T. Dobei, and W. T. Reidy, "Corneal ablation by nanosecond, picosecond and femtosecond laser pulses at 532 nm and 625 nm," Arch. Ophthalmol. 107, 587-592 (1989).
[CrossRef]

Rapti, G. S.

T. Theodossiou, G. S.  Rapti, V.  Hovhannisyan, E.  Georgiou, K.  Politopoulos, and D.  Yova, "Thermally induced irreversible conformational changes in collagen probed by optical second harmonic generation and laser-induced fluorescence," Lasers Med. Sci. 17, 34-41 (2002).
[CrossRef] [PubMed]

Rebollar, E.

M. Oujja, E. Rebollar, C. Abrusci, A. Del Amo, F. Catalina, and M. Castillejo, "UV, visible and IR laser interaction with gelatine," J. Phys.: Conf. Ser. 59, 571-574 (2007).
[CrossRef]

Reidy, W. T.

D. Stern, C. A. Puliafito, E. T. Dobei, and W. T. Reidy, "Corneal ablation by nanosecond, picosecond and femtosecond laser pulses at 532 nm and 625 nm," Arch. Ophthalmol. 107, 587-592 (1989).
[CrossRef]

Reiser, K. M.

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, "Collagen structure and nonlinear susceptibility: effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity," Lasers Surg. Med. 27, 329-335 (2000).
[CrossRef] [PubMed]

Rem, A. I.

A. I. Rem, J. A. Oosterhuis, H. G. Journée-de Korver, T. J van den Berg, and J. E. Keunen, "Temperature dependence of thermal damage to the sclera: exploring the heat tolerance of the sclera for transscleral thermotherapy," Exp. Eye Res. 72, 153-62 (2001).
[CrossRef] [PubMed]

Riemann, I.

B. G. Wang, I. Riemann, H. Schubert, K. J. Halbhuber, and K. Koenig, "In-vivo intratissue ablation by nanojoule near-infrared femtosecond laser pulses," Cell Tissue Res. 328, 515-520 (2007).
[CrossRef] [PubMed]

A. Ehlers, I. Riemann, S. Martin, R. Le Harzic, A. Bartels, C. Janke, and K. König, "High (1 GHz) repetition rate compact femtosecond laser: A powerful multiphoton tool for nanomedicine and nanobiotechnology," J. Appl. Phys. 102, 014701-6 (2007).
[CrossRef]

K. König, O. Krauss, and I. Riemann, "Intratissue surgery with 80 MHz nanojoule femtosecond laser pulses in the near infrared," Opt. Express. 10, 171-176 (2002).

K. König, I. Riemann, and W. Fritzsche, "Nanodissection of human chromosomes with near-infrared femtosecond laser pulses," Opt. Lett. 26, 819-821 (2001).
[CrossRef]

Rubenchik, A. M.

B. M. Kim, J. Eichler, K. M. Reiser, A. M. Rubenchik, and L. B. Da Silva, "Collagen structure and nonlinear susceptibility: effects of heat, glycation, and enzymatic cleavage on second harmonic signal intensity," Lasers Surg. Med. 27, 329-335 (2000).
[CrossRef] [PubMed]

Sakakura, M.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Schaffer, C.

Schubert, H.

B. G. Wang, I. Riemann, H. Schubert, K. J. Halbhuber, and K. Koenig, "In-vivo intratissue ablation by nanojoule near-infrared femtosecond laser pulses," Cell Tissue Res. 328, 515-520 (2007).
[CrossRef] [PubMed]

Si, J.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

So, P. T.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

Sobol, E. N.

N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
[CrossRef] [PubMed]

Squier, J. A.

R. M. Kurtz, X. Liu, V. M. Elner, J. A. Squier, D. Du, and G. A. Mourou, "Photodisruption in the human cornea as a function of laser pulse width," J. Refract. Surg. 13, 653-658 (1997).

Stern, D.

D. Stern, C. A. Puliafito, E. T. Dobei, and W. T. Reidy, "Corneal ablation by nanosecond, picosecond and femtosecond laser pulses at 532 nm and 625 nm," Arch. Ophthalmol. 107, 587-592 (1989).
[CrossRef]

Sun, H.

Sun, Y.

Y. Sun, W. L. Chen, S. J. Lin, S. H. Jee, Y. F. Chen, L. C. Lin, P. T. So, and C. Y. Dong, "Investigating mechanisms of collagen thermal denaturation by high resolution second-harmonic generation imaging," Biophys J. 91, 2620-2625 (2006).
[CrossRef] [PubMed]

S. J. Lin, C. Y. Hsiao, Y. Sun, W. Lo, W. C. Lin, G. J. Jan, S. H. Jee, and C. Y. Dong, "Monitoring the thermally induced structural transitions of collagen using second harmonic generation microscopy," Opt. Lett. 30, 622-624 (2005).
[CrossRef] [PubMed]

Tamaru, Y.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Tan, H. Y.

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

Teng, S. W.

H. Y. Tan, S. W. Teng, W. Lo, W. C. Lin, S. J. Lin, S. H. Jee, and C. Y. Dong, "Characterizing the thermally induced structural changes to intact porcine eye, part 1: second harmonic generation imaging of cornea stroma," J. Biomed. Opt. 10, 054019-5 (2005).
[CrossRef] [PubMed]

Theodossiou, T.

T. Theodossiou, G. S.  Rapti, V.  Hovhannisyan, E.  Georgiou, K.  Politopoulos, and D.  Yova, "Thermally induced irreversible conformational changes in collagen probed by optical second harmonic generation and laser-induced fluorescence," Lasers Med. Sci. 17, 34-41 (2002).
[CrossRef] [PubMed]

Theodossiou, T. A.

T. A. Theodossiou, C. Thrasivoulou, C. Ekwobi, and D. L. Becker, "Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections," Biophys. J. 91, 4665-4677 (2006).
[CrossRef] [PubMed]

Thrasivoulou, C.

T. A. Theodossiou, C. Thrasivoulou, C. Ekwobi, and D. L. Becker, "Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections," Biophys. J. 91, 4665-4677 (2006).
[CrossRef] [PubMed]

Tsutsumi, M.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Ueda, M.

M. Sakakura, S. Kajiyama, M. Tsutsumi, J. Si, E. Fukusaki, Y. Tamaru, S. Akiyama, K. Miura, K. Hirao, and M. Ueda, "Femtosecond pulsed laser as a microscalpel for microdissection and isolation of specific sections from biological samples," Jpn. J. Appl. Phys. 46, 5859-5864 (2007).
[CrossRef]

Vogel, A.

A. Vogel, J. Noack, G. Huttman, and G. Paltauf, "Mechanisms of femtosecond laser nanosurgery of cells and tissues," Appl. Phys. B 81, 1015-1047 (2005).
[CrossRef]

Wang, B. G.

B. G. Wang, I. Riemann, H. Schubert, K. J. Halbhuber, and K. Koenig, "In-vivo intratissue ablation by nanojoule near-infrared femtosecond laser pulses," Cell Tissue Res. 328, 515-520 (2007).
[CrossRef] [PubMed]

Yakar, A. B.

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N. Yu. Ignatieva, O. L. Zakharkina, I. V. Andreeva, E. N. Sobol, V. A. Kamensky, A. V. Myakov, S. V. Averkiev, and V. V. Lunin, "IR Laser and heat-induced changes in annulus fibrosus collagen structure," Photochem. Photobiol. 83, 675-685 (2007).
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T. Juhasz, F. H. Loesel, R. M. Kurtz, C. Horvath, J. F. Bille, and G. Mouro, "Corneal refractive surgery with femtosecond lasers," IEEE J. Quantum Electron. 5, 902-909 (1999).
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M. F. Yanik, H. Cinar, N. Cinar, A. Chisholm, Y. Jin, and A. B. Yakar, "Neurosurgery: Functional regeneration after laser axotomy," Nature 432, 882 (2004).
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Supplementary Material (2)

» Media 1: AVI (2437 KB)     
» Media 2: AVI (2245 KB)     

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Figures (12)

Fig. 1.
Fig. 1.

(Movie 1.avi, 2.4 Mb). Dynamics of photo-modification of collagen fibers from rat tail during fs illumination. Temporal dependences of TPAF and SHG intensities in 2 regions of interest (ROIs, white oval regions in the image II) are plotted on the left. Images in frames I, II and III were acquired respectively at 17, 28, and 37 seconds of the illumination process. An estimated speed of the photo-modification extension is ~0.7 µm/sec. Pseudo-colors of red and green represent SHG and TPAF respectively. Laser power was 10 mW and the frame height is 33 µm (Red: SHG; green: TPAF). [Media 1]

Fig. 2.
Fig. 2.

Temporal dependence of TPAF and SHG intensities in a large ROI (white circle, radius=46 µm) within a collagen fiber network from bovine Achilles’ tendon. The laser power was 29 mW. (Red: SHG; green: TPAF).

Fig. 3.
Fig. 3.

(Movie 3.avi, 2.2 Mb). Dynamics of NIR, fs laser induced photo-modification in bovine cornea during the continuous scanning. Multiphoton images were acquired at various illumination and scan times (4, 14, 24 and 50 seconds). Laser power was 21 mW and the frame size is 16×16 µm2. (Red : SHG; green: TPAF). [Media 2]

Fig. 4.
Fig. 4.

Dependence of TPAF (1,2) and SHG (3, 4) signals on scan number n during illumination of the bovine cornea within a selected circular region (radius 6 µm). Two pixel residence times were selected. Data sets 1 and 3 correspond to the pixel residence time of 6.4 µs. Data sets 2 and 4 were acquired at the pixel residence time of 3.2 µs. The solid curves represent quadratic polynomial fits to data sets 1 and 2 and n is the scan number.

Fig. 5.
Fig. 5.

Dependence of photo-modification efficiency ( R ) on the pixel residence time. R was determined from TPAF curves (Fig. 4 (1) and (2)) using equation R=√r/n.

Fig. 6.
Fig. 6.

Temporal (1, 3) and dosage (2, 4) dependence of TPAF (1, 2) and SHG (3, 4) intensities during multiple scans of bovine cornea samples. (A): Power was 24 mW and the scan frequency was changed from 5 Hz to 0.5 Hz. (B): Power was 28.5 mW and the scan frequency was changed from 0.5 Hz to 5 Hz (B). The frame scan time was 0.2 sec and image size is 46 µm.

Fig. 7.
Fig. 7.

Multiphoton imaging of collagen fibers from rat tail at various illumination times (1, 22, 37, 38 and 45 seconds, green: TPAF, red: SHG). A pause of 15 seconds was introduced between the scans at 22 and 37 seconds. The power was 38 mW and the frame size is 20 µm.

Fig. 8.
Fig. 8.

Reflected (frame 1), fluorescence (frames 2–8) and combined (frame 9) confocal images of bovine cornea treated with 780 nm. Illumination time was 30 sec. For comparison, the multiphoton image is shown in (frame 10) (green: TPAF; red: SHG). Boundaries of the photo-modified region are clearly visible in the TPAF image. The power was 23 mW and the images were acquired the day following NIR fs laser illumination. The confocal spectral imaging bandwidth for 1–-7 images was 21 nm. (Frame size: 100 µm).

Fig. 9.
Fig. 9.

Bovine cornea fluorescence spectra in the non-illuminated region (curve 1) and fs laser illuminated region (2). Fluorescence experimental data points are connected by smoothed lines.

Fig. 10.
Fig. 10.

Collagen fiber cutting. TPAF (green) and SHG (red) images of a collagen fiber from rat tail before (A) and after (B) excision by fs laser radiation with power of 45 mW. An increase in TPAF was observed at the cutting site (blue line). The experimental parameters are: cutting time - 6 sec, pixel residence time - 26 µs, and frame rate - 10 Hz. Scale bar: 20 µm.

Fig. 11.
Fig. 11.

(A) Fs, NIR irradiation induced bending of collagen fiber from bovine Achilles’ tendon. The laser power was 45 mW. The scale bar is 10 µm. (Red: SHG; green: TPAF). (B) Observed collagen fiber shrinkage. Equatorial profiles of the collagen SHG averaged intensity in the selected rectangular region at different times of irradiation. (C) Temporal dependence of mean SHG intensity in the selected circular region shown in (A). It was observed that SHG intensity increased concurrently with shrinking of the collagen fiber.

Fig. 12.
Fig. 12.

Femtosecond laser lithography in bovine corneal stroma. The letters “V” ((A) and (C)) and “I” (B) were sculptured with 20 mW of fs laser. Image (A) was acquired using fs laser with power of 5 mW (red: SHG; green: TPAF). (B) and (C) are confocal images (blue: reflected confocal signal at 458nm; green: autofluorescence between 505–550 nm; red: autofluorescence between 550–670 nm). (Frame height: 17 µm).

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