This paper presents a two-photon phase-resolved fluorescence-lifetime measurement method based on the use of an ultrashort pulse laser. The proposed method also involves the use of a lock-in amplifier to control the phase difference between the reference and fluorescence signals, thereby facilitating the use of an alternative method for determining fluorescence lifetimes. Verification of the fluorescence lifetimes as measured in this study was performed using rhodamine B and a cellular thermoprobe as samples. In this study, we assume that the fluorescence decay was monoexponential in all cases. Rhodamine B was observed to exhibit an average fluorescence lifetime of 2.15 ns, whereas a temperature sensitivity of 1.39 ns C-1 over a temperature range of 33.79-37.2 °C was demonstrated for the cellular thermoprobe. These results validate the feasibility of the proposed method for accurate measurement of fluorescence lifetimes using a simple laser configuration.

中文翻译：

---

基于超短激光的两光子相分辨荧光寿命测量方法。

本文提出了一种基于超短脉冲激光的双光子相分辨荧光寿命测量方法。所提出的方法还涉及使用锁定放大器来控制参考信号和荧光信号之间的相位差，从而有助于使用另一种方法来确定荧光寿命。使用若丹明B和细胞热探针作为样品，对本研究中测量的荧光寿命进行了验证。在这项研究中，我们假设在所有情况下荧光衰减都是单指数的。观察到若丹明B的平均荧光寿命为2.15 ns，而对于细胞热探针，在33.79-37.2°C的温度范围内，其温度敏感性为1.39 ns C-1。