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Two-Photon Laser Scanning Fluorescence Microscopy
Two-photon excitation microscopy
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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“Two-Photon Laser Scanning Fluorescence Microscopy” is a paper by Winifried Denk James H. Strickler Watt W. Webb published in 1990. It has an Open Access status of “closed”. You can read and download a PDF Full Text of this paper here.