Abstract

We characterize experimentally the influence of sample structure and beam focusing on signal level in third-harmonic generation (THG) microscopy. In the case of a homogeneous spherical sample, the dependence of the signal on the size of the sphere can be controlled by modifying the Rayleigh length of the excitation beam. More generally, the influence of excitation focusing on the signal depends on sample geometry, allowing one to highlight certain structures within a complex system. We illustrate this point by focusing-based contrast modulation in THG images of Drosophila embryos.

© 2005 Optical Society of America

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    [CrossRef]

2005 (1)

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

2004 (1)

2002 (1)

2000 (1)

1999 (1)

A. Zumbusch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

1998 (1)

1997 (1)

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, Appl. Phys. Lett. 70, 922 (1997).
[CrossRef]

1990 (1)

W. Denk, J. H. Strickler, and W. W. Webb, Science 248, 73 (1990).
[CrossRef] [PubMed]

Barad, Y.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, Appl. Phys. Lett. 70, 922 (1997).
[CrossRef]

Beaurepaire, E.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

D. Débarre, W. Supatto, E. Farge, B. Moulia, M.-C. Schanne-Klein, and E. Beaurepaire, Opt. Lett. 29, 2881 (2004).
[CrossRef]

Blanchard-Desce, M.

Boyd, R. W.

R. W. Boyd, Nonlinear Optics, 2nd ed. (Academic, 2003).

Brakenhoff, G. J.

Brouzés, E.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

Cheng, J.-X.

Débarre, D.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

D. Débarre, W. Supatto, E. Farge, B. Moulia, M.-C. Schanne-Klein, and E. Beaurepaire, Opt. Lett. 29, 2881 (2004).
[CrossRef]

Denk, W.

W. Denk, J. H. Strickler, and W. W. Webb, Science 248, 73 (1990).
[CrossRef] [PubMed]

Eisenberg, H.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, Appl. Phys. Lett. 70, 922 (1997).
[CrossRef]

Farge, E.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

D. Débarre, W. Supatto, E. Farge, B. Moulia, M.-C. Schanne-Klein, and E. Beaurepaire, Opt. Lett. 29, 2881 (2004).
[CrossRef]

Holtom, G. R.

A. Zumbusch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Horowitz, M.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, Appl. Phys. Lett. 70, 922 (1997).
[CrossRef]

Martin, J.-L.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

Mertz, J.

Moreaux, L.

Moulia, B.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

D. Débarre, W. Supatto, E. Farge, B. Moulia, M.-C. Schanne-Klein, and E. Beaurepaire, Opt. Lett. 29, 2881 (2004).
[CrossRef]

Müller, M.

Sandre, O.

Schanne-Klein, M.-C.

Silberberg, Y.

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, Appl. Phys. Lett. 70, 922 (1997).
[CrossRef]

Squier, J. A.

Strickler, J. H.

W. Denk, J. H. Strickler, and W. W. Webb, Science 248, 73 (1990).
[CrossRef] [PubMed]

Supatto, W.

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

D. Débarre, W. Supatto, E. Farge, B. Moulia, M.-C. Schanne-Klein, and E. Beaurepaire, Opt. Lett. 29, 2881 (2004).
[CrossRef]

Webb, W. W.

W. Denk, J. H. Strickler, and W. W. Webb, Science 248, 73 (1990).
[CrossRef] [PubMed]

Wilson, K. R.

Xie, X. S.

J.-X. Cheng and X. S. Xie, J. Opt. Soc. Am. B 19, 1604 (2002).
[CrossRef]

A. Zumbusch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Zumbusch, A.

A. Zumbusch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Appl. Phys. Lett. (1)

Y. Barad, H. Eisenberg, M. Horowitz, and Y. Silberberg, Appl. Phys. Lett. 70, 922 (1997).
[CrossRef]

J. Opt. Soc. Am. B (1)

Opt. Express (1)

Opt. Lett. (2)

Phys. Rev. Lett. (1)

A. Zumbusch, G. R. Holtom, and X. S. Xie, Phys. Rev. Lett. 82, 4142 (1999).
[CrossRef]

Proc. Natl. Acad. Sci. U.S.A. (1)

W. Supatto, D. Débarre, B. Moulia, E. Brouzés, J.-L. Martin, E. Farge, and E. Beaurepaire, Proc. Natl. Acad. Sci. U.S.A. 102, 1047 (2005).
[CrossRef]

Science (1)

W. Denk, J. H. Strickler, and W. W. Webb, Science 248, 73 (1990).
[CrossRef] [PubMed]

Other (1)

R. W. Boyd, Nonlinear Optics, 2nd ed. (Academic, 2003).

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Figures (3)

Fig. 1
Fig. 1

(a) Measured THG power from a polystyrene bead centered at the beam focus as a function of bead diameter for three focusing conditions corresponding to δ z = 2 , 2.5, and 3.5 μ m (respectively represented by white, gray, and black squares. Solid curves are a guide for the eye). (b) Theoretical THG power from a bead at the beam focus, assuming a refractive index of 1.57 at 1180 nm and 1.61 at 393 nm for polystyrene. (c) Total THG power from a bead during a volume scan, normalized to the bead volume. (d), (e) Axial THG profile through a (d) 0.6 and a (e) 3.0 μ m bead, with δ z = 2 μ m .

Fig. 2
Fig. 2

THG power as a function of sample structure and focusing conditions. (a) Excitation beam scanned along an interface. (b) Excitation beam scanned in a gel of beads. (c) Total THG power obtained from a 30 μ m × 30 μ m area in the geometry in (a). (d) Same measurement in the geometry in (b) for 0.6 (circles) and 3 μ m (squares) beads. Solid curves are theoretical calculations.

Fig. 3
Fig. 3

Focusing-based contrast enhancement in THG images. (a) Total THG power obtained from the outer membrane of a Drosophila embryo [black arrowhead in (c)] as a function of δ z . (b) THG power obtained from internal organelles under the same conditions [white arrowhead in (d)]. (c), (d) Images recorded 50 μ m above the embryo equator for δ z = 2.5 and 6 μ m , normalized to the cubed excitation intensity and displayed with the same color code. Inverted contrast. Image acquisition time of 2.5 s.

Equations (1)

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E THG ( R ) V d V ( 1 + k 3 2 ) exp ( i k 3 R r ) 4 π R r P THG ( r ) ,

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