State Key Laboratory Base of Novel Functional Materials and Preparation Science, Key Laboratory of Advanced Mass Spectrometry and Molecular Analysis of Zhejiang Province, School of Materials Science and Chemical Engineering, Ningbo University, Ningbo, Zhejiang 315211, China.
Analyst. 2021 Jul 26;146(15):4945-4953. doi: 10.1039/d1an00419k.
Two-photon fluorescence imaging is one of the most attractive imaging techniques for monitoring important biomolecules in the biomedical field due to its advantages of low light scattering, high penetration depth, and suppressed photodamage/phototoxicity under near-infrared excitation. However, in actual biological imaging, organic two-photon fluorescent dyes have disadvantages such as high biological toxicity and their fluorescence efficiency is easily affected by the complex environment in organisms. In this study, a novel nanoprobe platform with two-photon dye-doped silica nanoparticles was developed for FRET-based ratiometric biosensing and bioimaging, with endogenous ATP chosen as the target for detection. The nanoprobe has three components: (1) a two-photon dye-doped silica nanoparticle core, which serves as an energy donor for FRET; (2) amino-modified hairpin primers with carboxy fluorescein as an energy acceptor for FRET; (3) an aptamer acting as a recognition unit to realize the probing function. The nanoprobe showed ratiometric fluorescence responses for ATP detection with high sensitivity and high selectivity in vivo. Moreover, the nanoprobe showed satisfactory ratiometric two-photon fluorescence imaging of endogenous ATP in living cells and tissues (penetration depth of 190 nm). These results indicated that novel two-photon silica nanoparticles can be constructed by doping a two-photon fluorescent dye into silica nanoparticles, and they can effectively solve the disadvantages of two-photon fluorescent dyes. These excellent performances indicate that this novel nanoprobe platform will become a very valuable molecular imaging tool, which can be widely used in the biomedical field for drug screening and disease diagnosis and other related research.
双光子荧光成像是生物医学领域中监测重要生物分子最具吸引力的成像技术之一，因为它具有低光散射、高穿透深度和在近红外激发下抑制光损伤/光毒性的优点。然而，在实际的生物成像中，有机双光子荧光染料存在生物毒性高和荧光效率容易受到生物体内复杂环境影响等缺点。在本研究中，开发了一种具有双光子染料掺杂硅纳米粒子的新型纳米探针平台，用于基于 FRET 的比率型生物传感和生物成像，以内源性 ATP 作为检测目标。该纳米探针具有三个组成部分：(1) 双光子染料掺杂硅纳米粒子核，作为 FRET 的能量供体；(2) 带有羧基荧光素的氨基修饰发夹引物，作为 FRET 的能量受体；(3) 作为识别单元的适体，实现探测功能。该纳米探针对 ATP 的检测具有高灵敏度和高选择性的比率荧光响应，体内。此外，该纳米探针在活细胞和组织中对内源性 ATP 进行了令人满意的比率双光子荧光成像（穿透深度为 190nm）。这些结果表明，通过将双光子荧光染料掺杂到硅纳米粒子中可以构建新型双光子硅纳米粒子，并能有效解决双光子荧光染料的缺点。这些优异的性能表明，这种新型纳米探针平台将成为一种非常有价值的分子成像工具，可广泛应用于药物筛选和疾病诊断等相关研究的生物医学领域。
