Effect of pulse duration on two-photon excited fluorescence and second harmonic generation in nonlinear optical microscopy.

Publication Type:

Journal Article


Journal of biomedical optics, Volume 11, Issue 2, p.020501 (2006)


Algorithms, Equipment Design, Equipment Failure Analysis, Humans, Image Enhancement, Microscopy, Fluorescence, Multiphoton, Nonlinear Dynamics, Reproducibility of Results, Sensitivity and Specificity, Signal Processing, Computer-Assisted, Skin


We have developed a multiphoton microscopy (MPM) system using a 12-fs Ti:sapphire laser with adjustable dispersion precompensation in order to examine the impact of pulse duration on nonlinear optical signals. The efficiencies of two-photon-excited fluorescence (TPEF) and second harmonic generation (SHG) were studied for various pulse durations, measured at the sample, ranging from approximately 400 fs to sub-20 fs. Both TPEF and SHG increased proportionally to the inverse of the pulse duration for the entire tested range. Because of improved signal-to-noise ratio, sub-20-fs pulses were used to enhance MPM imaging depth by approximately 160%, compared to 120-fs pulses, in human skin.

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