Teng Xu, Gui Lingyan, Liang Weiyuan, Liu Tao, Chen Jiuyang, Liu Yue, Li Qiguang, Zeng Jun, Liang Yaru, Li Linhai
Department of Laboratory Medicine, The Affiliated Qingyuan Hospital (Qingyuan People's Hospital), Guangzhou Medical University, Qingyuan 511518, China.
Department of Pharmacy, The Third People's Hospital of Chengdu, Chengdu 610031, China.
Anal Chem. 2025 Apr 1;97(12):6661-6669. doi: 10.1021/acs.analchem.4c06593. Epub 2025 Mar 19.
The wash-free method represents a promising strategy for enhancing the detection efficiency of automated chemiluminescent (CL) immunoassay analyzers. Herein, a novel separation-free and wash-free immunoassay was developed for the first time based on the distance-driven CL technique. In the CL immunoassay, the well-designed Au-Co metal nanoclusters (NCs) exhibited excellent peroxidase-like activity and good stability, allowing for efficient catalysis of luminol or its analogue (ABEI)-HO system even at low concentrations. Furthermore, the large specific surface area of Au-Co NCs facilitated the accommodation of a greater number of antibodies, thereby enhancing the capture of antigens and achieving dual amplification of the CL signal. The distance-driven CL technique relied on the formation of sandwich-type immunocomplexes. Upon the generation of hydroxyl radicals and superoxide anion radicals through the catalytic decomposition of HO by Au-Co NCs, the ABEI within sandwich-type immunocomplexes could efficiently react with these radicals, leading to a significant enhancement in CL signals. Furthermore, C-reactive protein (CRP) was chosen as the model analyte to evaluate the practicability of the proposed immunoassay. Notably, the proposed immunoassay presented high sensitivity, selectivity, reproducibility, and stability, successfully determining CRP in serum samples with recoveries of 96.55-106.29%. Accordingly, the proposed strategy with the advantage of separation-free, wash-free, and reliable characteristics could drastically simplify the detection operation steps and enhance the detection efficiency, which would make significant advances in the revolution of traditional CL immunoassay.
免冲洗方法是提高自动化化学发光（CL）免疫分析仪器检测效率的一种很有前景的策略。在此，首次基于距离驱动化学发光技术开发了一种新型的免分离和免冲洗免疫分析方法。在化学发光免疫分析中，精心设计的金-钴金属纳米簇（NCs）表现出优异的类过氧化物酶活性和良好的稳定性，即使在低浓度下也能有效催化鲁米诺或其类似物（ABEI）-HO体系。此外，金-钴纳米簇的大比表面积有利于容纳更多的抗体，从而增强对抗原的捕获并实现化学发光信号的双重放大。距离驱动化学发光技术依赖于夹心型免疫复合物的形成。通过金-钴纳米簇催化分解HO产生羟基自由基和超氧阴离子自由基后，夹心型免疫复合物中的ABEI能与这些自由基有效反应，导致化学发光信号显著增强。此外，选择C反应蛋白（CRP）作为模型分析物来评估所提出的免疫分析方法的实用性。值得注意的是，所提出的免疫分析方法具有高灵敏度、选择性、重现性和稳定性，成功测定了血清样品中的CRP，回收率为96.55-106.29%。因此，所提出的具有免分离、免冲洗和可靠特点的策略可以极大地简化检测操作步骤并提高检测效率，这将在传统化学发光免疫分析的变革中取得重大进展。
