State Key Laboratory of Bioactive Substances and Function of Natural Medicine, Beijing Key Laboratory of Active Substances Discovery and Drugability Evaluation, Institute of Materia Medica, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100050, China.
Flourescence Products, ISS, Inc., 1602 Newton Drive, Champaign, IL 61822, USA.
Adv Healthc Mater. 2024 Jun;13(15):e2304223. doi: 10.1002/adhm.202304223. Epub 2024 Mar 5.
Two-photon excitation (TPE) microscopy with near-infrared (NIR) emission has emerged as a promising technique for deep-tissue optical imaging. Recent developments in fluorescence lifetime imaging with long-lived emission probes have further enhanced the spatial resolution and precision of fluorescence imaging, especially in complex systems with short-lived background signals. In this study, two innovative lysosome-targeting probes, Cz-NA and tCz-NA, are introduced. These probes offer a combination of advantages, including TPE (λ = 880 nm), NIR emission (λ = 650 nm), and thermally activated delayed fluorescence (TADF) with long-lived lifetimes (1.05 and 1.71 µs, respectively). These characteristics significantly improve the resolution and signal-to-noise ratio in deep-tissue imaging. By integrating an acousto-optic modulator (AOM) device with TPE microscopy, the authors successfully applied Cz-NA in two-photon excited delayed fluorescence (TPEDF) imaging to track lysosomal adaptation and immune responses to inflammation in mice. This study sheds light on the relationship between lysosome tubulation, innate immune responses, and inflammation in vivo, providing valuable insights for the development of autofluorescence-free molecular probes in the future.
双光子激发（TPE）显微镜与近红外（NIR）发射结合，已成为一种很有前途的用于深层组织光学成像的技术。最近，具有长寿命发射探针的荧光寿命成像技术的发展进一步提高了荧光成像的空间分辨率和精度，特别是在具有短寿命背景信号的复杂系统中。在这项研究中，引入了两种创新的溶酶体靶向探针 Cz-NA 和 tCz-NA。这些探针具有 TPE（λ=880nm）、NIR 发射（λ=650nm）和热激活延迟荧光（TADF）的优点，寿命长（分别为 1.05 和 1.71µs）。这些特性显著提高了深层组织成像的分辨率和信噪比。通过将声光调制器（AOM）与 TPE 显微镜集成，作者成功地将 Cz-NA 应用于双光子激发延迟荧光（TPEDF）成像中，以追踪溶酶体适应和对炎症的先天免疫反应。这项研究揭示了体内溶酶体管化、先天免疫反应和炎症之间的关系，为未来开发无自发荧光的分子探针提供了有价值的见解。
