College of Chemistry and Chemical Engineering, Xinyang Key Laboratory of Functional Nanomaterials for Bioanalysis, Xinyang Normal University, NO. 237 in Nanhu Road, Xinyang, 464000, Henan, China.
Shandong Key Laboratory of Biochemical Analysis, Qingdao University of Science and Technology, Qingdao, 266042, China.
Anal Bioanal Chem. 2024 Sep;416(21):4759-4767. doi: 10.1007/s00216-024-05277-x. Epub 2024 Apr 22.
A highly efficient ratiometric electrochemiluminescence (ECL) immunoassay was explored by bidirectionally regulating the ECL intensity of two luminophors. The immunoassay was conducted in a split-type mode consisting of an ECL detection procedure and a sandwich immunoreaction. The ECL detection was executed using a dual-disk glassy carbon electrode modified with two potential-resolved luminophors (g-CN-Ag and Ru-MOF-Ag nanocomposites), and the sandwich immunoreaction using glucose oxidase (GOx)-modified SiO nanospheres as labels was carried out in a 96-well plate. The Ag nanoparticles (NPs) acted as bifunctional units both for triggering the resonance energy transfer (RET) with g-CN and for accelerating the electron transfer rate of the Ru-MOF-Ag ECL reaction. When the HO catalyzed by GOx in the 96-well plate was transferred to the dual-disk glass carbon electrode, the doped Ag NPs in the two luminophors could be etched, thus destroying the RET between CN and the accelerated reaction to Ru-MOF, resulting in an opposite trend in the ECL signal outputted from the dual disks. Using the ratio of the two signals for quantification, the constructed immunosensor for a model target, i.e. myoglobin, exhibited a low detection limit of 4.7 × 10 g/mL. The ingenious combination of ECL ratiometry, bifunctional Ag NPs, and a split-type strategy effectively reduces environmental and human errors, offering a more precise and sensitive analysis for complex samples.
一种高效的比率型电化学发光（ECL）免疫分析方法通过双向调节两种发光体的 ECL 强度来探索。免疫分析采用分体型模式进行，包括 ECL 检测程序和三明治免疫反应。ECL 检测使用双盘玻碳电极进行，该电极修饰有两种电位分辨发光体（g-CN-Ag 和 Ru-MOF-Ag 纳米复合材料），而使用葡萄糖氧化酶（GOx）修饰的 SiO2 纳米球作为标记的三明治免疫反应在 96 孔板中进行。Ag 纳米颗粒（NPs）既作为触发 g-CN 与 CN 的共振能量转移（RET）的双功能单元，又作为加速 Ru-MOF-Ag ECL 反应电子转移速率的双功能单元。当 GOx 在 96 孔板中催化的 HO 转移到双盘玻碳电极时，两种发光体中的掺杂 Ag NPs 可以被蚀刻，从而破坏 CN 与加速反应之间的 RET，导致双盘输出的 ECL 信号呈现相反的趋势。使用两种信号的比值进行定量，构建的模型靶标（即肌红蛋白）的免疫传感器表现出低至 4.7×10-10 g/mL 的检测限。ECL 比率法、双功能 Ag NPs 和分体型策略的巧妙结合有效减少了环境和人为误差，为复杂样品提供了更精确和敏感的分析。
