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. 2001 Mar;80(3):1568–1574. doi: 10.1016/S0006-3495(01)76129-2

Coherent scattering in multi-harmonic light microscopy.

L Moreaux 1, O Sandre 1, S Charpak 1, M Blanchard-Desce 1, J Mertz 1
PMCID: PMC1301348  PMID: 11222317

Abstract

By focusing a pulsed laser beam into a sample, harmonic up-conversion can be generated as well as multi-photon excited fluorescence. Whereas multi-photon excited fluorescence microscopy is well established, the use of multi-harmonic generation for three-dimensional image contrast is very recent. Both techniques can provide similar resolution and, for adequate radiating source density, comparable signal levels, allowing them to be combined in a single versatile instrument. However, harmonic generation differs fundamentally from fluorescence generation in that it is coherent and produces radiation patterns that are highly sensitive to phase. As such, multi-harmonic generation microscopy provides a unique window into molecular spatial organization that is inaccessible to fluorescence.

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Selected References

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  1. Albota M., Beljonne D., Brédas J. L., Ehrlich J. E., Fu J. Y., Heikal A. A., Hess S. E., Kogej T., Levin M. D., Marder S. R. Design of organic molecules with large two-photon absorption cross sections. Science. 1998 Sep 11;281(5383):1653–1656. doi: 10.1126/science.281.5383.1653. [DOI] [PubMed] [Google Scholar]
  2. Bolz J., Novak N., Staiger V. Formation of specific afferent connections in organotypic slice cultures from rat visual cortex cocultured with lateral geniculate nucleus. J Neurosci. 1992 Aug;12(8):3054–3070. doi: 10.1523/JNEUROSCI.12-08-03054.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bouevitch O., Lewis A., Pinevsky I., Wuskell J. P., Loew L. M. Probing membrane potential with nonlinear optics. Biophys J. 1993 Aug;65(2):672–679. doi: 10.1016/S0006-3495(93)81126-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Campagnola P. J., Wei M. D., Lewis A., Loew L. M. High-resolution nonlinear optical imaging of live cells by second harmonic generation. Biophys J. 1999 Dec;77(6):3341–3349. doi: 10.1016/S0006-3495(99)77165-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Denk W., Strickler J. H., Webb W. W. Two-photon laser scanning fluorescence microscopy. Science. 1990 Apr 6;248(4951):73–76. doi: 10.1126/science.2321027. [DOI] [PubMed] [Google Scholar]
  6. Dufour S., Beauvais-Jouneau A., Delouvée A., Thiery J. P. Differential function of N-cadherin and cadherin-7 in the control of embryonic cell motility. J Cell Biol. 1999 Jul 26;146(2):501–516. doi: 10.1083/jcb.146.2.501. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Fluhler E., Burnham V. G., Loew L. M. Spectra, membrane binding, and potentiometric responses of new charge shift probes. Biochemistry. 1985 Oct 8;24(21):5749–5755. doi: 10.1021/bi00342a010. [DOI] [PubMed] [Google Scholar]
  8. Grinvald A., Hildesheim R., Farber I. C., Anglister L. Improved fluorescent probes for the measurement of rapid changes in membrane potential. Biophys J. 1982 Sep;39(3):301–308. doi: 10.1016/S0006-3495(82)84520-7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gähwiler B. H. Organotypic cultures of neural tissue. Trends Neurosci. 1988 Nov;11(11):484–489. doi: 10.1016/0166-2236(88)90007-0. [DOI] [PubMed] [Google Scholar]
  10. Kuzyk MG, Dirk CW. Effects of centrosymmetry on the nonresonant electronic third-order nonlinear optical susceptibility. Phys Rev A. 1990 May 1;41(9):5098–5109. doi: 10.1103/physreva.41.5098. [DOI] [PubMed] [Google Scholar]
  11. Loew L. M., Simpson L. L. Charge-shift probes of membrane potential: a probable electrochromic mechanism for p-aminostyrylpyridinium probes on a hemispherical lipid bilayer. Biophys J. 1981 Jun;34(3):353–365. doi: 10.1016/S0006-3495(81)84854-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Maiti S., Shear J. B., Williams R. M., Zipfel W. R., Webb W. W. Measuring serotonin distribution in live cells with three-photon excitation. Science. 1997 Jan 24;275(5299):530–532. doi: 10.1126/science.275.5299.530. [DOI] [PubMed] [Google Scholar]
  13. Marder S. R., Beratan D. N., Cheng L. T. Approaches for optimizing the first electronic hyperpolarizability of conjugated organic molecules. Science. 1991 Apr 5;252(5002):103–106. doi: 10.1126/science.252.5002.103. [DOI] [PubMed] [Google Scholar]
  14. Müller M, Squier J, Wilson KR, Brakenhoff GJ. 3D microscopy of transparent objects using third-harmonic generation. J Microsc. 1998 Sep;191(3):266–274. doi: 10.1046/j.1365-2818.1998.00399.x. [DOI] [PubMed] [Google Scholar]
  15. Peleg G., Lewis A., Linial M., Loew L. M. Nonlinear optical measurement of membrane potential around single molecules at selected cellular sites. Proc Natl Acad Sci U S A. 1999 Jun 8;96(12):6700–6704. doi: 10.1073/pnas.96.12.6700. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sandre O., Moreaux L., Brochard-Wyart F. Dynamics of transient pores in stretched vesicles. Proc Natl Acad Sci U S A. 1999 Sep 14;96(19):10591–10596. doi: 10.1073/pnas.96.19.10591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Weiss S. Fluorescence spectroscopy of single biomolecules. Science. 1999 Mar 12;283(5408):1676–1683. doi: 10.1126/science.283.5408.1676. [DOI] [PubMed] [Google Scholar]
  18. Xu C., Zipfel W., Shear J. B., Williams R. M., Webb W. W. Multiphoton fluorescence excitation: new spectral windows for biological nonlinear microscopy. Proc Natl Acad Sci U S A. 1996 Oct 1;93(20):10763–10768. doi: 10.1073/pnas.93.20.10763. [DOI] [PMC free article] [PubMed] [Google Scholar]

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