Wang Weichao, Liu Yujia, Xi Peng, Ren Qiushi
Department of Biomedical Engineering, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, No. 800 Dong-Chuan Road, Shanghai 200240, China.
Appl Opt. 2010 Dec 10;49(35):6703-9. doi: 10.1364/AO.49.006703.
Short pulses can induce high nonlinear excitation, and thus they should be favorable for use in multiphoton microscopy. However, the large spectral dispersion can easily destroy the advantages of the ultrashort pulse if there is no compensation. The group delay dispersion (GDD), third-order dispersion, and their effects on the intensity and bandwidth of second-harmonic generation (SHG) signal were analyzed. We found that the prism pair used for compensating the GDD of the two-photon microscope actually introduces significant negative high-order dispersion (HOD), which dramatically narrowed down the two-photon absorption probability for ultrashort pulses. We also investigated the SHG signal after GDD and HOD compensation for different pulse durations. Without HOD compensation, the SHG efficiency dropped significantly for a pulse duration below 20 fs. We experimentally compared the SHG and two-photon excited fluorescence (TPEF) signal intensity for 11 fs versus 50 fs pulses, a pulse duration close to that commonly used in conventional multiphoton microscopy. The result suggested that after adaptive phase compensation, the 11fs pulse can yield a 3.2- to 6.0-fold TPEF intensity and a 5.1-fold SHG intensity, compared to 50 fs pulses.
短脉冲能够诱导高非线性激发，因此它们在多光子显微镜中应用应具有优势。然而，如果没有补偿，大的光谱色散很容易破坏超短脉冲的优势。分析了群延迟色散（GDD）、三阶色散及其对二次谐波产生（SHG）信号强度和带宽的影响。我们发现，用于补偿双光子显微镜GDD的棱镜对实际上引入了显著的负高阶色散（HOD），这极大地降低了超短脉冲的双光子吸收概率。我们还研究了不同脉冲持续时间在GDD和HOD补偿后的SHG信号。在没有HOD补偿的情况下，当脉冲持续时间低于20 fs时，SHG效率显著下降。我们通过实验比较了11 fs和50 fs脉冲的SHG和双光子激发荧光（TPEF）信号强度，50 fs脉冲的持续时间接近传统多光子显微镜常用的持续时间。结果表明，经过自适应相位补偿后，与50 fs脉冲相比，11 fs脉冲可产生3.2至6.0倍的TPEF强度和5.1倍的SHG强度。
