基于酶辅助靶标循环的适体传感器灵敏检测中药中黄曲霉毒素B1  

Sensitive Detection of Aflatoxin B1 in Traditional Chinese Medicine Using an Aptasensor Based on Enzyme-assisted Target Recycling

作　　者：吴昊[1] 吴军[1] 王洪勇[1] 刘娅灵[1] 韩国庆 李文超 施青云 邹霈[1,2] WU Hao;WU Jun;WANG Hong-yong;LIU Ya-ling;HAN Guo-qing;LI Wen-chao;SHI Qing-yun;ZOU Pei(NHC Key Laboratory of Nuclear Medicine,Jiangsu Key Laboratory of Molecular Nuclear Medicine,Jiangsu Institute of Nuclear Medicine,Wuxi 214063,China;School of Pharmacy,Nanjing Medical University,Nanjing 211166,China)

机构地区：[1]国家卫健委核医学重点实验室江苏省分子核医学重点实验室江苏省原子医学研究所,江苏无锡214063 [2]南京医科大学药学院,江苏南京211166

出　　处：《分析测试学报》2021年第1期92-98,共7页Journal of Instrumental Analysis

基　　金：国家自然科学基金项目(81771905);无锡市中医药管理局中医药科技项目(ZYKJ201903)。

摘　　要：该文基于酶辅助靶标循环信号放大策略构建了用于黄曲霉毒素B1(AFB1)高灵敏检测的化学发光适体传感器。以G-四链体/氯化血红素DNA酶为信号分子设计了免标记的适体探针H1-S1和发夹探针H2。适体探针结合目标AFB1,在核酸外切酶I辅助下,触发靶标循环反应产生发夹H1。发夹H1与H2杂交,释放出完整的G-四链体序列,并进一步与氯化血红素结合形成G-四链体/氯化血红素DNA酶。DNA酶通过催化氧化鲁米诺-H2O2化学发光体系产生化学发光信号,实现AFB1的放大检测。在最优实验条件下,化学发光强度与AFB1质量浓度的对数在0.001~100 ng/mL范围内呈良好的线性关系,相关系数(r2)为0.9955,检出限为0.93 pg/mL,回收率为93.7%~107%。该适体传感器操作简单、灵敏度高、特异性好,在黄曲霉毒素污染检测方面具有良好的应用前景。In this paper,a chemiluminescence aptasensor for the highly sensitive detection of aflatoxin B1(AFB1)was constructed based on the strategy of enzyme-assisted target recycling amplification.The label-free aptamer probe H1-S1 and hairpin probe H2 were well designed using G-quadruplex/hemin DNAzyme as the signal molecule.The aptamer probe bound to the target AFB1,which triggered the target recycling amplification reaction with the help of exonuclease I to generate hairpin H1.The hairpin H1 was hybridized with H2,releasing the complete G-quadruplex sequence.The G-quadruplex sequence combined with hemin to form G-quadruplex/hemin DNAzyme,which catalyzed the oxidation of luminol-H2O2 chemiluminescence system to generate chemiluminescence signals to achieve the amplified detection of AFB1.Under the optimal experimental conditions,the chemiluminescence intensity exhibited a good linear relationship(r2=0.9955)with the logarithm of AFB1 concentration in the range of 0.001-100 ng/mL.The detection limit was 0.93 pg/mL,and the recoveries were in the range of 93.7%-107%.With the advantages of simple operation,high sensitivity and good specificity,this aptasensor has a good application prospect in the detection of aflatoxin contamination.

关 键 词：黄曲霉毒素B1(AFB1) 化学发光 靶标循环 G-四链体 DNA酶 中药 

分 类 号：O657.1[理学—分析化学] O625.3[理学—化学]