A biodegradable and cofactor self-sufficient aptazyme nanoprobe for amplified imaging of low-abundance protein in living cells

纳米探针 脱氧核酶 化学 辅因子 生物物理学 核酸酶 荧光团 DNA 荧光 生物化学 纳米技术 纳米颗粒 生物 物理 材料科学 量子力学
作者
Jiayao Xu,Lifang Yao,Xiaohong Zhong,Kun Hu,Shulin Zhao,Yong Huang
出处
期刊:Talanta [Elsevier]
卷期号:253: 123983-123983 被引量:4
标识
DOI:10.1016/j.talanta.2022.123983
摘要

Despite the progress on the analysis of proteins either in vitro or in vivo, detection and imaging of low-abundance proteins in living cells still remains challenging. Herein, a novel biodegradable and cofactor self-sufficient DNAzyme nanoprobe has been deve-loped for catalytic imaging of protein in living cells with signal amplification capacity. This DNAzyme nanoprobe is constructed by assembling a DNAzyme subunit-containing aptamer hairpin (HP), another DNAzyme subunit strand (DS), and the molecular beacon (MB) substrate strand onto pH-sensitive ZnO@polydopamine nanorods (ZnO@PDA NRs) that work as DNAzyme cofactor suppliers. Such a nanoprobe can facilitate cellular uptake of DNA molecules and protection of them from nuclease degradation as well as release of them in cells by lysosomal acid-triggered dissolution of ZnO@PDA NRs into Zn2+ as DNAzyme cofactor. Upon recognition and binding with the intracellular protein target, the stem of HP is opened, after which the opened HP hybridizes with DS and generates activated DNAzymes. Each activated DNAzyme can catalyze the cleavage of many MB substrates through true enzymatic multiple turnovers, resulting in the separation of the quenched fluorophore/quencher pair labeled in MB and the generation of significantly amplified fluorescence. Using nucleolin (NCL) as a model protein, this nanoprobe enables the analysis of NCL with a detection limit of 1.8 pM, which are at least two orders of magnitude lower than that of non-catalytic imaging probe. Moreover, it could accurately distinguish tumor cells and normal cells by live cell NCL imaging. And the experimental results are also further verified by flow cytometry assays. The developed nanoprobe can be easily extended to detect other biomolecules by the change of their corresponding aptamer sequences, thus providing a promising tool for highly sensitive imaging of low-abundance biomolecules in living cells.
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