基于脱氧核酶的双光子内标型纳米荧光探针的构建及阿兹海默症脑组织成像应用研究

It is of great significance to accurately monitor abnormal concentration of metal ions in brain tissues for studying the mechanism and early intervention of Alzheimer’s disease. DNAzymes-based fluorescent nanoprobes with advantages of high affinity and high specificity have become a powerful tool in the field of biosensing and bioimaging. However, there are still some shortcomings of reported DNAzymes-based fluorescent nanoprobes, such as the shallow tissue depth, large fluorescence background, difficulty to quantitatively detect in living cells, limiting the application of sensing and imaging in brain tissues of Alzheimer’s disease. This project aims to realize high-sensitivity detection and high-resolution imaging metal ions in brain tissues of Alzheimer’s disease by DNAzymes fluorescent nanoprobes. Combining high-specificity DNAzymes, nanotechnology and two-photon fluorescent imaging technology, we would develop series of DNAzymes-based two-photon fluorescent nanoprobes with high-sensitivity, high-selectivity, high-resolution and deep tissue depth. We try to design two-photon fluorophores as signaling molecules and internal references. Using ratios of fluorescent intensity of signaling molecules and internal references as detection signals, it could effectively avoid detection inaccuracy coming from different concentration of probes diffused in cells, intensity changes of excited light sources, and so on. With the introduction of internal references, it is expected to improve detection accuracy of DNAzymes-based fluorescent nanoprobes and achieve quantitative detection in living cells. This project could broaden the biosensing and bioimaging application of DNAzymes-based fluorescent nanoprobes in brain tissues of Alzheimer’s disease, further providing new methods to study the mechanism of Alzheimer’s disease induced by overexpressed metal ions.

准确监测脑组织中金属离子的浓度异常变化对阿兹海默症发病机理的研究及早期干预具有重要意义。高亲和力、高特异性的脱氧核酶纳米荧光探针已成为生物检测与成像的强有力工具。然而穿透深度有限、背景荧光干扰大、难以用于细胞内定量检测等问题限制了脱氧核酶纳米探针应用于阿兹海默症脑组织成像检测，本项目拟结合高特异性的脱氧核酶识别单元、纳米技术及双光子荧光成像技术，通过引入内标型策略，构筑双光子荧光信号分子和内标分子对，构建灵敏度高、选择性好、成像分辨率高及组织穿透深度深的双光子内标型纳米荧光探针，以信号分子和内标分子的荧光强度比值作为输出信号，消除荧光探针进入细胞浓度以及环境因素的干扰，实现活细胞内金属离子的定量检测，拓宽脱氧核酶纳米荧光探针在阿兹海默症脑组织中的高灵敏、高特异性检测与成像应用，为过量金属离子诱发阿兹海默症这一发病机制的研究提供新的分析手段。

细胞中活性小分子浓度的异常变化与疾病的发生发展密切相关。针对目前活性小分子成像分析中存在的特异性不够、穿透深度有限、背景荧光干扰大等关键科学问题，本项目利用功能核酸作为特异性分子识别单元，结合纳米技术、双光子荧光成像技术、近红外二区荧光/光声成像技术，构建了系列高灵敏、高特异性的成像探针，实现了深层组织及活体中活性小分子的实时、原位、动态成像以及成像指导下的肿瘤治疗，在细胞及活体水平上为肿瘤等疾病的发生发展机制研究提供分析方法及工具。在项目的资助下，共发表SCI论文6篇，其中以一作或通讯作者发表的SCI论文4篇，包括Chem. Soc. Rev. 1篇，Chem. Sci. 1篇，Anal. Chem. 1篇，Sci. China Chem. 1篇；参加国内学术会议5次；培养硕士生3名。