Li Bingqian, Yu Shengrong, Feng Rong, Qian Zhiling, He Kangdi, Mao Guo-Jiang, Cao Yuting, Tang Keqi, Gan Ning, Wu Yong-Xiang
State Key Laboratory Base of Novel Functional Materials and Preparation Science, Key Laboratory of Advanced Mass Spectrometry and Molecular Analysis of Zhejiang Province, Zhejiang Engineering Research Center of Advanced Mass Spectrometry and Clinical Application, Institute of Mass Spectrometry, School of Materials Science and Chemical Engineering, Ningbo University, Ningbo, Zhejiang 315211, China.
Ningbo Zhenhai Institute of Mass Spectrometry, Ningbo, Zhejiang 315211, China.
Anal Chem. 2023 Oct 10;95(40):14925-14933. doi: 10.1021/acs.analchem.3c02216. Epub 2023 Sep 28.
Bioimaging is widely used in various fields of modern medicine. Fluorescence imaging has the advantages of high sensitivity, high selectivity, noninvasiveness, in situ imaging, and so on. However, one-photon (OP) fluorescence imaging has problems, such as low tissue penetration depth and low spatiotemporal resolution. These disadvantages can be solved by two-photon (TP) fluorescence imaging. However, TP imaging still uses fluorescence intensity as a signal. The complexity of organisms will inevitably affect the change of fluorescence intensity, cause false-positive signals, and affect the accuracy of the results obtained. Fluorescence lifetime imaging (FLIM) is different from other kinds of fluorescence imaging, which is an intrinsic property of the material and independent of the material concentration and fluorescence intensity. FLIM can effectively avoid the fluctuation of TP imaging based on fluorescence intensity and the interference of autofluorescence. Therefore, based on silica-coated gold nanoclusters (AuNCs@SiO) combined with nucleic acid probes, the dual-mode nanoprobe platform was constructed for TP and FLIM imaging of intracellular endogenous miRNA-21 for the first time. First, the dual-mode nanoprobe used a dual fluorescence quencher of BHQ2 and graphene oxide (GO), which has a high signal-to-noise ratio and anti-interference. Second, the dual-mode nanoprobe can detect miR-21 with high sensitivity and selectivity in vitro, with a detection limit of 0.91 nM. Finally, the dual-mode nanoprobes performed satisfactory TP fluorescence imaging (330.0 μm penetration depth) and FLIM (τ = 50.0 ns) of endogenous miR-21 in living cells and tissues. The dual-mode platforms have promising applications in miRNA-based early detection and therapy and hold much promise for improving clinical efficacy.
生物成像在现代医学的各个领域中得到了广泛应用。荧光成像具有高灵敏度、高选择性、非侵入性、原位成像等优点。然而，单光子（OP）荧光成像存在诸如组织穿透深度低和时空分辨率低等问题。这些缺点可以通过双光子（TP）荧光成像来解决。然而，TP成像仍然使用荧光强度作为信号。生物体的复杂性将不可避免地影响荧光强度的变化，产生假阳性信号，并影响所获得结果的准确性。荧光寿命成像（FLIM）与其他类型的荧光成像不同，它是材料的固有属性，与材料浓度和荧光强度无关。FLIM可以有效避免基于荧光强度的TP成像的波动和自发荧光的干扰。因此，首次基于二氧化硅包覆的金纳米团簇（AuNCs@SiO）与核酸探针相结合，构建了用于细胞内源性miRNA-21的TP和FLIM成像的双模式纳米探针平台。首先，双模式纳米探针使用了BHQ2和氧化石墨烯（GO）的双荧光猝灭剂，具有高信噪比和抗干扰能力。其次，双模式纳米探针在体外能够以高灵敏度和选择性检测miR-21，检测限为0.91 nM。最后，双模式纳米探针在活细胞和组织中对内源性miR-21进行了令人满意的TP荧光成像（穿透深度为330.0μm）和FLIM（τ = 50.0 ns）。该双模式平台在基于miRNA的早期检测和治疗中具有广阔的应用前景，有望提高临床疗效。
