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26 February 2010 Pulse shaping for reducing photodamage in multiphoton microscopy
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We formulate a simple strategy for mitigation of laser-induced damage through pulse shaping and demonstrate experimentally the effect of laser pulse duration on the degree of optically induced damage for two-photon microscopy imaging. We use a broadband Ti:Sapphire laser source, aided with a shaper, and adjust both the laser pulse duration and energy to maintain constant two-photon excitation efficiency. The damage is assessed by the dynamics of two-photon excited autofluorescence intensity and sample morphology during prolonged laser exposure. We observe that for a 5-μm layer of skin tissue the damage rate is independent of the pulse shape, which suggests that the primary damage (bleaching) mechanism stems from the two-photon excitation itself. For optically thick dried blood samples, taken as another example, the data suggests that the damage is driven by one-photon absorption. In the later case, it is favorable to use shorter laser pulses to mitigate photodamage while maintaining adequate intensity of two-photon excited autofluorescence.
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Dmitry Pestov, Yair Andegeko, Vadim V. Lozovoy, and Marcos Dantus "Pulse shaping for reducing photodamage in multiphoton microscopy", Proc. SPIE 7569, Multiphoton Microscopy in the Biomedical Sciences X, 756926 (26 February 2010); doi: 10.1117/12.852289;

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