The rational design of a zero cross-talk self-referenced ratiometric fluorescent probe for accurate molecular imaging in living cells is still a major challenge. Herein, we report a zero cross-talk fluorescent polymer nanoprobe (named ZC-FPN) with a self-referenced ratiometric effect for lysosomal hypochlorous acid (HClO) imaging using single-wavelength excitation. The newly designed polymer nanoprobe (ZC-FPN) is prepared by using a simple co-precipitation method through self-assembly of two amphiphilic block copolymers, which are covalently linked with a porphyrin fluorophore (HClO-responsive unit) and a naphthalimide fluorophore (reference unit), respectively. Remarkably, the ZC-FPN displays an unprecedented separation of two emission peaks (∼191 nm), which can avoid the spectral cross-talk problem of the emission-shift type ratiometric probes and is highly favorable for high-resolution bioimaging. In addition, the as-prepared ZC-FPN also shows good water dispersibility, high selectivity, excellent long-term fluorescence stability (>10 weeks) and low cytotoxicity. More importantly, the positive charge of the amino moiety enhances the efficient uptake of the nanoprobe by cancer cells and enables the nanoprobe to selectively accumulate in lysosomes, achieving endogenous HClO imaging in living cells through fluorescence ratiometric imaging.

中文翻译：

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零串扰荧光聚合物纳米探针的工程学，用于活细胞中溶酶体次氯酸的自参考比例成像

零串扰自参考比例荧光探针在活细胞中进行精确分子成像的合理设计仍然是一个重大挑战。在本文中，我们报告了使用单波长激发对溶酶体次氯酸（HClO）成像具有自参考比例效应的零串扰荧光聚合物纳米探针（命名为ZC-FPN）。新设计的聚合物纳米探针（ZC-FPN）是通过简单的共沉淀方法通过将两种两性嵌段共聚物自组装而制备的，这两种共聚物与卟啉荧光团（HClO响应单元）和萘二甲酰亚胺荧光团共价连接（参考单位）。值得注意的是，ZC-FPN表现出前所未有的两个发射峰（〜191 nm）分离，可以避免发射位移型比例式探针的光谱串扰问题，对高分辨率生物成像非常有利。此外，所制备的ZC-FPN还显示出良好的水分散性，高选择性，出色的长期荧光稳定性（> 10周）和低细胞毒性。更重要的是，氨基部分的正电荷增强了癌细胞对纳米探针的有效吸收，并使纳米探针能够选择性地蓄积在溶酶体中，通过荧光比率成像在活细胞中实现内源性HClO成像。