Ge Shengjie, Wang Yujie, Li Zhiyue, Lu Bin, Zhu Jinhua, Lu Hongmei, Cao Xiaowei, Qian Yayun
Department of Clinical Medicine, Medical College, Yangzhou University, Yangzhou, 225001, P. R. China.
Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou University, Yangzhou, P. R. China.
J Mater Chem B. 2022 Nov 9;10(43):8931-8944. doi: 10.1039/d2tb01520j.
In this study, we apply catalytic hairpin assembly (CHA) as the signal amplification strategy for the quantification of carcinoembryonic antigen (CEA) and cytokeratin fragment antigen 21-1 (CYFRA21-1) with a surface-enhanced Raman scattering (SERS) microfluidic chip (LoC-SERS) as the carrier. Herein, antibody-DNA conjugates are designed to assist the application of CHA amplification in protein detection. In the presence of protein biomarkers, antibody-DNA conjugates can specifically bind to the target proteins, forming the antigen@antibody-DNA conjugates. The terminal free part of the DNA on the conjugates can trigger the CHA events to connect SERS nanotags to capture nanoprobes. Then, micro-magnet can gather the CHA products in a rectangular chamber, resulting in the aggregation of SERS nanotags, which can ultimately generate abundant "hot spots" for SERS signal enhancement. Using this strategy, CEA and CYFRA21-1 can be successfully determined with a limit of detection (LOD) as low as pg mL, much lower than recently reported methods. Meanwhile, a non-small cell lung cancer (NSCLC)-xenografted mouse model was established, and SERS was applied to analyze the expression level of CEA and CYFRA21-1 in tumorigenesis and development. The comparison between SERS results and those of the ELISA method demonstrated a high degree of consistency, suggesting that the proposed CHA-assisted LoC-SERS device has satisfying accuracy. Thus, introducing the CHA strategy the design of antibody-DNA conjugates opens new gates to ultra-sensitive and specific SERS detection of protein biomarkers.
在本研究中，我们应用催化发夹组装（CHA）作为信号放大策略，以表面增强拉曼散射（SERS）微流控芯片（芯片型SERS）为载体，对癌胚抗原（CEA）和细胞角蛋白片段抗原21-1（CYFRA21-1）进行定量分析。在此，设计了抗体-DNA偶联物以辅助CHA扩增在蛋白质检测中的应用。在存在蛋白质生物标志物的情况下，抗体-DNA偶联物可特异性结合靶蛋白，形成抗原@抗体-DNA偶联物。偶联物上DNA的末端游离部分可触发CHA事件，将SERS纳米标签连接到捕获纳米探针上。然后，微磁体可将CHA产物聚集在一个矩形腔室中，导致SERS纳米标签聚集，最终产生大量用于SERS信号增强的“热点”。使用该策略，CEA和CYFRA21-1能够成功测定，检测限（LOD）低至pg/mL，远低于最近报道的方法。同时，建立了非小细胞肺癌（NSCLC）异种移植小鼠模型，并应用SERS分析CEA和CYFRA21-1在肿瘤发生和发展过程中的表达水平。SERS结果与ELISA方法结果的比较显示出高度一致性，表明所提出的CHA辅助芯片型SERS装置具有令人满意的准确性。因此，将CHA策略引入抗体-DNA偶联物的设计为蛋白质生物标志物的超灵敏和特异性SERS检测打开了新的大门。
