Nie Wei, Zhang Ruoxian, Hu Chao, Jin Tengchuan, Wei Xi, Cui Hua
Key Laboratory of Precision and Intelligent Chemistry, iChEM (Collaborative Innovation Center of Chemistry for Energy Materials), Department of Chemistry, University of Science and Technology of China, Hefei, Anhui, 230026, PR China.
Laboratory of Structural Immunology, CAS Key Laboratory of Innate Immunity and Chronic Diseases, CAS Center for Excellence in Molecular Cell Science, Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei, Anhui, 230027, PR China.
Biosens Bioelectron. 2023 Nov 15;240:115662. doi: 10.1016/j.bios.2023.115662. Epub 2023 Sep 1.
SARS-CoV-2 antigen detection plays a key role in the rapid diagnosis of COVID-19. However, current clinically used immunoassays are often limited by assay throughput, sensitivity, accuracy, and field operating conditions. To address these challenges, we constructed a self-enhanced electrochemiluminescence (ECL) array chip (SEAC) for highly sensitive and label-free detection of SARS-CoV-2 nucleocapsid protein (N protein) with a facile and portable assay setup. Firstly, the self-enhanced ECL nanomaterials with inherent film-forming properties were synthesized by co-doping Ru(bpy) and polyethyleneimine (PEI) in silica nanoparticles (Ru/PEI@SiO). Secondly, a resistance-induced potential difference-based single-electrode electrochemical system (SEES) was adapted to serve as the electrode array to facilitate one-step assembly without the need for chip alignment. Thirdly, the chip electrode array was functionalized with the synthesized self-enhanced ECL emitters and captured antibodies. In addition, a portable detection box equipped with a smartphone was 3D-printed to serve as the chip holder and "dark room" for imaging acquisition. The SEAC performance was validated with N protein with a limit of detection (LOD) of 0.47 pg/mL in the range of 1-10,000 pg/mL. Furthermore, the chip successfully detected the viral antigen residue as low as 1.92 pg/mL from diluted rehabilitation patients' serum samples. This is the first study reporting label-free detection of SARS-Cov-2 N protein based on a self-enhanced ECL immunosensor, which provides a novel facile method for highly sensitive diagnosis of COVID-19 with high throughput, portability, and low cost.
严重急性呼吸综合征冠状病毒2（SARS-CoV-2）抗原检测在2019冠状病毒病（COVID-19）的快速诊断中起着关键作用。然而，目前临床使用的免疫测定法常常受到检测通量、灵敏度、准确性和现场操作条件的限制。为应对这些挑战，我们构建了一种自增强电化学发光（ECL）阵列芯片（SEAC），用于以简便且便携的检测装置对SARS-CoV-2核衣壳蛋白（N蛋白）进行高灵敏度和无标记检测。首先，通过在二氧化硅纳米颗粒（Ru/PEI@SiO）中共掺杂钌（联吡啶）（Ru(bpy)）和聚乙烯亚胺（PEI）合成具有固有成膜特性的自增强ECL纳米材料。其次，采用基于电阻诱导电位差的单电极电化学系统（SEES）作为电极阵列，以利于一步组装，无需芯片对齐。第三，用合成的自增强ECL发光体和捕获抗体对芯片电极阵列进行功能化。此外，还3D打印了一个配备智能手机的便携式检测盒，用作芯片支架和成像采集的“暗室”。在1-10000 pg/mL范围内，用N蛋白验证了SEAC的性能，检测限（LOD）为0.47 pg/mL。此外，该芯片成功检测到来自康复患者稀释血清样本中低至1.92 pg/mL的病毒抗原残留。这是第一项基于自增强ECL免疫传感器无标记检测SARS-CoV-2 N蛋白的研究，为COVID-19的高通量、便携且低成本的高灵敏度诊断提供了一种新颖简便的方法。
