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J Biomed Opt. 2009 Jan-Feb;14(1):014002. doi: 10.1117/1.3059629.

Two-photon imaging using adaptive phase compensated ultrashort laser pulses.

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  • 1Shanghai Jiao Tong University, Department of Biomedical Engineering, Institute for Laser Medicine and Biophotonics, Shanghai 200240 China.


An adaptive pulse shaper controlled by multiphoton intrapulse interference phase scanning (MIIPS) was used, together with a prism-pair, to measure and cancel high-order phase distortions introduced by a high-numerical-aperture objective and other dispersive elements of a two-photon laser-scanning microscope. The delivery of broad-bandwidth (approximately 100 nm), sub-12-fs pulses was confirmed by interferometric autocorrelation measurements at the focal plane. A comparison of two-photon imaging with transform-limited and second-order-dispersion compensated laser pulses of the same energy showed a 6-to-11-fold improvement in the two-photon excitation fluorescence signal when applied to cells and tissue, and up to a 19-fold improvement in the second harmonic generation signal from a rat tendon specimen.

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