基于DNA杂交指示剂和银纳米棒阵列芯片构建氯霉素SERS适配体传感器的研究

doi:10.3964/j.issn.1000-0593(2023)11-3445-07

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摘要：氯霉素（CAP）是一种人工合成的抗生素，通过与细菌的核糖体进行结合而抑制蛋白质合成达到抑菌目的。长期摄入残留CAP动源性食品会导致人体发生贫血及白血病，也会使机体产生耐药性严重危害人类健康，许多国家规定不得在畜产品中检出CAP，因此，设计一种更快速简便且高特异性的CAP检测方法具有重要意义。以CAP的巯基化适配体（SH-Apt）修饰银纳米棒阵列芯片（芯片）作SERS基底、以DNA杂交指示剂亚甲基蓝（MB）作为拉曼信号分子，利用CAP、CAP适配体互补链（cDNA）与CAP适配体（Apt）竞争结合的关系，构建了一种新型的高特异性CAP-SERS适配体传感器。利用扫描电子显微镜（SEM）、X射线衍射（XRD）及EDS能谱仪对芯片及CAP-SERS适配体传感器进行表征，结果表明芯片表面均匀分布大量银元素且证明CAP-SERS适配体传感器成功制备。在室温条件下对CAP标准样进行检测，传感器相关性能的考察结果表明，在0.001～10 ng·mL^-1范围之间，CAP浓度的对数值（logc）与1 624 cm^-1处的SERS信号强度（I_SERS）存在良好的线性关系：I_SERS=-971logc+1 983(R²=0.991)。拉曼增强因子EF=1.01×10⁷，检测限为0.2 pg·mL^-1（S/N=3），进一步表明该基底具有良好的拉曼增强效果。应用该传感器分别检测CAP片剂、人血清及猪血清中的CAP，并进行加标试验，结果令人满意，回收率在91.2%~120.5%之间，RSD（n=3）在0.97%~8.1%之间，证明该传感器有良好的准确性。该传感器具有制作简单、灵敏度高、选择性强、重现性和稳定性好以及检测速度快等优点，可望应用于实际样品中CAP的快速定量检测，为检测CAP提供了一种新思路。

关键词：亚甲基蓝；适配体；表面增强拉曼光谱；适配体传感器；氯霉素

Abstract：Chloramphenicol (CAP) is a synthetic antibiotic that inhibits protein synthesis by binding to the ribosomes of bacteria to achieve the purpose of antibacterial. Long-term intake of residual CAP animal-derived food can lead to anemia and leukemia in the human body and can also cause the body to develop drug resistance, which will seriously endanger human health. Many countries have regulations prohibiting the detection of CAP in livestock products. Therefore, designing a more rapid, simple and highly specific CAP detection method is of great significance. In this paper, the thiolated aptamer of CAP (SH-Apt) was used to modify the silver nanorod array chip(chip)as the SERS substrate, and the DNA hybridization indicator methylene blue (MB) as the Raman reporter, A novel high-specificity CAP-SERS aptamer sensor was constructed by utilizing the competitive binding relationship between CAP, CAP aptamer complementary strand (cDNA) and CAP aptamer (Apt). The chip and CAP-SERS aptamer sensor were characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) and EDS spectroscopy. The results showed that a large amount of silver elements were uniformly distributed on the surface of the chip and the CAP-SERS aptamer sensor was successfully prepared. Detection of CAP standards at room temperature, the results of the sensor-related performance analysis suggest that, with an increase of CAP concentration (0.001～10 ng·mL^-1) caused the decreased SERS signal at 1 624 cm^-1(I_SERS) intensity (I_SERS=-971logc+1 983). A good negative correlation (R²=0.991) was achieved, and the limit of detection (LOD) was as low as 0.2 pg·mL^-1 (S/N=3), the Raman enhancement factor EF=1.01×10⁷. Further indicating that the substrate has good Raman enhancement. The sensor is used to detect individual CAP tablets and CAP in human and pig sera, spik experiments were performed, and the results were satisfactory. The recovery and relative standard deviations(RSD) were 91.2%～120.5% and 0.97%～8.1%, which proved that the sensor had good accuracy. The sensor is intended to be used for the rapid quantitative detection of CAP in real samples because of its simple manufacture, high sensitivity, strong selectivity, good reproducibility, good stability, and fast detection speed. It provides a new idea for detecting CAP.

Key words：Methylene blue; Aptamer; Surface-Enhanced Raman Scattering; Aptamer sensor;Chloramphenicol

收稿日期: 2022-06-23 修订日期: 2022-09-18

中图分类号:

O657.37

基金资助: 国家自然科学基金项目（81860633），广西自然科学基金项目（2019GXNSFDA245025，2020GXNSFAA297168）资助

通讯作者: 李利军，冯军 E-mail: lilijun0562@sina.com；hxpfengjun@163.com

作者简介: 郭贺媛熙，女，1997年生，广西科技大学生物与化学工程学院硕士研究生 e-mail: 1448567030@qq.com

引用本文:

郭贺媛熙，李利军，冯军，林鑫，李睿. 基于DNA杂交指示剂和银纳米棒阵列芯片构建氯霉素SERS适配体传感器的研究[J]. 光谱学与光谱分析, 2023, 43(11): 3445-3451.
GUO He-yuanxi, LI Li-jun, FENG Jun, LIN Xin, LI Rui. A SERS-Aptsensor for Detection of Chloramphenicol Based on DNA Hybridization Indicator and Silver Nanorod Array Chip. SPECTROSCOPY AND SPECTRAL ANALYSIS, 2023, 43(11): 3445-3451.

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