Ratiometric ECL sensor based onApt‑AuNS@Lu nanoprobe foranalyzing cell swelling‑induced ATP release
Ratiometric ECL sensor based onApt‑AuNS@Lu nanoprobe foranalyzing cell swelling‑induced ATP release
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基于 Apt™AuNS@Lu 纳米探针的比率 ECL 传感器用于分析细胞肿胀诱导的 ATP 释放
DOI：
10.1007/s00604-022-05491-3
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发表时间：
2022
影响因子：
5.7
通讯作者：
Peihui Yang
中科院分区：
文献类型：
--
作者：
Fan Zhou;Mingxing Xiao;Defen Feng;Peihui Yang
构建了一种新的基于金纳米星(AuNSS)载体的比率计量型电化学发光(ECL)体系，用于检测低渗诱导的人肝癌细胞株HepG2细胞中ATP的释放。AUNS@Lu纳米探针作为阳极发光体，K2S2O8作为阴极发光体和阳极共反应物。采用比表面积大的AUNS吸附大量的鲁米诺形成固体探针，并作为亲和载体固定三磷酸腺苷适配子(APT)。由于AUNS具有良好的导电性和催化活性，得到的纳米复合材料(APT-AUNS@Lu)与共反应物K2S2O8在 + 0.4nV(vs.Ag/AgCl2)处产生了很强的电化学发光信号。此外，氧化石墨烯还被还原到氧化铟锡(ITO)电极上，以促进电子转移。随后，通过自聚合形成聚多巴胺(PDA)膜，提高了电极表面的稳定性和附着力。通过在RGO/PDA表面吸附大量的AuNSs来固定化ATP捕获适配子(APTC)。当该传感器在apt-AUNS@Lu和靶ATP的混合液中孵育时，apt-AUNS@Lu的ECL信号随着ATP浓度的增加而增加，而K2S2O8的信号则随着ATP浓度的增加而减弱。用这两个发光团的比值来定量测定ATP。线性范围为5~250nM，检出限为1.4nM(3σ/s)。该方法已成功应用于0.45%氯化钠低渗溶液刺激的人肝癌细胞株的三磷酸腺苷释放分析。结果表明，三磷酸腺苷的释药动力学曲线呈S形，在10min内快速释放，然后缓慢释放。与等渗组相比，低渗组细胞内的三磷酸腺苷浓度为3.7 ± 0.3nM(n= 3)，下降了40.3%，细胞外的三磷酸腺苷浓度为2 3.4 ± 1.2nM(n= 3)，低渗10min时，细胞内三磷酸腺苷的释放增加了9.2倍，这与商业的酶联免疫吸附试验结果一致。所提出的策略将有利于扩大ECL技术在细胞生物学功能研究中的应用。
A novel ratiometric electrochemiluminescence (ECL) system based on gold nanostars (AuNSs) support was constructed for the determination of hypotonicity-induced ATP release from HepG2 cells. AuNS@Lu nanoprobe was used as anodic luminophore and K2S2O8as cathodic luminophore as well as anodic co-reactant. AuNS with the large specific surface was adopted to adsorb plentiful luminol to form solid-state probe and as affinity support to immobilize ATP aptamer (Apt). The obtained nanocomposite (Apt-AuNS@Lu) generated a strong ECL signal at + 0.4 V (vs. Ag/AgCl) with co-reactant K2S2O8, because of excellent conductivity and catalytic activity of AuNS. Furthermore, graphene oxide was reduced onto indium tin oxide (ITO) electrodes to facilitate the electron transfer. Following, polydopamine (PDA) film was formed via self-polymerization, improving stability and adhesion of the electrode surface. To immobilize ATP capture aptamer (AptC), abounding AuNSs were attached to RGO/PDA surface. When the sensor was incubated in the mixture solution of Apt-AuNS@Lu and target ATP, the ECL signal of Apt-AuNS@Lu increased with the increase of ATP concentration, meanwhile, the signal of K2S2O8declined. The ratio of the two luminophores was used for the quantitative determination of ATP. The linear range was 5 to 250 nM, and the limit of detection was 1.4 nM at (3σ)/S. The method was successfully applied to analyze ATP release from HepG2 cells stimulated by 0.45% NaCl hypotonic solution. The results showed that the release kinetics profile of ATP had a sigmoidal shape with rapid release within 10 min and then slowed. Compared to the isotonic groups, the intracellular ATP concentration was 3.7 ± 0.3 µM (n= 3) decreasing by 40.3% and the extracellular was 23.4 ± 1.2 nM (n= 3) increasing by 9.2 times in the hypotonicity for 10 min, which showed ATP release from cells and good agreement with commercial ELISA test. The proposed strategy would be beneficial to broadening application of ECL technology in studying cell biological functions.Graphical abstract