SIGNIFICANCE Stimulated Raman scattering (SRS) and pump-probe microscopy are implementations of multiphoton microscopy that acquire high-resolution, label-free images of live samples encoded with molecular contrast. Most commercial multiphoton microscopes cannot access these techniques since they require sample illumination by two temporally synchronized ultrafast pulse trains. We present a compact and robust way of synchronizing an additional Ti:sapphire laser with a conventional single-beam multiphoton microscope to realize an instrument that can acquire images with enhanced molecular specificity. AIM A passive optical synchronization scheme for a pair of commercially available, unmodified modelocked Ti:sapphire lasers was developed. The suitability of this synchronization scheme for advanced biomedical microscopy was investigated. APPROACH A pair of modelocked Ti:sapphire lasers were aligned in master-slave configuration. Five percent of the master laser output was used to seed the modelocking in the slave laser cavity. The timing jitter of the master and slave pulse trains was characterized using an optical autocorrelator. The synchronized output of both lasers was coupled into a laser scanning microscope and used to acquire spectral focusing SRS and pump-probe microscopy images from biological and nonbiological samples. RESULTS A timing jitter between the modelocked pulse trains of 0.74 fs was recorded. Spectral focusing SRS allowed spectral discrimination of polystyrene and polymethyl methacrylate beads. Pump-probe microscopy was used to record excited state lifetime curves from hemoglobin in intact red blood cells. CONCLUSION Our work demonstrates a simple and robust method of upgrading single-beam multiphoton microscopes with an additional ultrafast laser. The resulting dual-beam instrument can be used to acquire label-free images of sample structure and composition with high biochemical specificity.

中文翻译：

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超低定时抖动，用于受激拉曼散射和泵浦探针显微镜的 Ti:Al2O3 同步。

意义受激拉曼散射 (SRS) 和泵浦探针显微术是多光子显微术的实现，可获取使用分子对比度编码的活样本的高分辨率、无标记图像。大多数商用多光子显微镜无法使用这些技术，因为它们需要通过两个时间同步的超快脉冲序列对样品进行照明。我们提出了一种紧凑而稳健的方式，将额外的钛蓝宝石激光器与传统的单光束多光子显微镜同步，以实现一种能够获取具有增强分子特异性的图像的仪器。目的 为一对市售的、未修改的锁模钛蓝宝石激光器开发了一种无源光同步方案。研究了这种同步方案对先进生物医学显微镜的适用性。方法 一对锁模钛蓝宝石激光器按主从配置排列。主激光输出的 5% 用于在从激光腔中播种锁模。使用光学自相关器表征主从脉冲序列的定时抖动。两种激光器的同步输出耦合到激光扫描显微镜中，用于从生物和非生物样品中获取光谱聚焦 SRS 和泵浦探针显微镜图像。结果 记录了 0.74 fs 的锁模脉冲序列之间的定时抖动。光谱聚焦 SRS 允许对聚苯乙烯和聚甲基丙烯酸甲酯珠进行光谱区分。泵探针显微镜用于记录完整红细胞中血红蛋白的激发态寿命曲线。结论我们的工作展示了一种简单而可靠的方法，可以使用额外的超快激光器升级单光束多光子显微镜。由此产生的双光束仪器可用于获取具有高生化特异性的样品结构和组成的无标记图像。