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Abstract

Molecular excitation by the simultaneous absorption of two photons provides intrinsic three-dimensional resolution in laser scanning fluorescence microscopy. The excitation of fluorophores having single-photon absorption in the ultraviolet with a stream of strongly focused subpicosecond pulses of red laser light has made possible fluorescence images of living cells and other microscopic objects. The fluorescence emission increased quadratically with the excitation intensity so that fluorescence and photobleaching were confined to the vicinity of the focal plane as expected for cooperative two-photon excitation. This technique also provides unprecedented capabilities for three-dimensional, spatially resolved photochemistry, particularly photolytic release of caged effector molecules.

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Science
Volume 248Issue 49516 April 1990
Pages: 73 - 76

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Published online: 6 April 1990

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Winifried Denk
School of Applied and Engineering Physics, Department of Physics, Cornell University, Ithaca, NY 14853.
James H. Strickler
School of Applied and Engineering Physics, Department of Physics, Cornell University, Ithaca, NY 14853.
Watt W. Webb
School of Applied and Engineering Physics, Department of Physics, Cornell University, Ithaca, NY 14853.

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Volume 248|Issue 4951
6 April 1990
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