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Using Multiphoton Microscopy for the Study of Embryogenesis

Published online by Cambridge University Press:  02 July 2020

John White ,
Victoria Centonze ,
David Wokosin  and
William Mohler
John White
Affiliation:
Integrated Microscopy Resource University of Wisconsin, Madison53706
Victoria Centonze
Affiliation:
Integrated Microscopy Resource University of Wisconsin, Madison53706
David Wokosin
Affiliation:
Integrated Microscopy Resource University of Wisconsin, Madison53706
William Mohler
Affiliation:
Laboratory of Molecular Biology, University of Wisconsin, Madison53706
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Extract

Multiphoton fluorescence excitation imaging is an optical sectioning technique for fluorescence microscopy. At very high photon densities, two or more photons may coordinately excite an energy transition in a fluorophore that corresponds to the sum of the energies of the individual photons. by this means, a fluorophore may be excited by a wavelength that is considerably longer than its single photon excitation wavelength. Ultra-fast pulsed (femtosecond) lasers can produce the peak power densities in the focal volume of an objective lens needed to provide sufficient 2- or 3- photon excitation events for imaging. The use of short-pulse lasers provides the high peak powers necessary for imaging yet with modest mean power levels that do not thermally damage biological specimens. Production of multiphoton events depends on the square of photon density for 2-photon excitation and the cube of photon density for 3-photon excitation. The power density therefore rapidly falls off away from the focal volume of an objective lens, thereby confining fluorescence excitation to the focal volume.

Type
Biological Applications of Multi-photon Excitation Fluorescence Imaging
Information
Microscopy and Microanalysis , Volume 3 , Issue S2: Proceedings: Microscopy & Microanalysis '97, Microscopy Society of America 55th Annual Meeting, Microbeam Analysis Society 31st Annual Meeting, Histochemical Society 48th Annual Meeting, Cleveland, Ohio, August 10-14, 1997 , August 1997 , pp. 307 - 308
Copyright
Copyright © Microscopy Society of America 1997

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References

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4. This work was supported by NIH Grants P41-RR00570 and RO1 GM52454-02Google Scholar