比率型荧光纸芯片快速检测赭曲霉毒素A

注册 English

比率型荧光纸芯片快速检测赭曲霉毒素A

林浩 , 贺璇 , 陈燕 , 庞鲁羿 , 牛磊 , 付秀丽

引用本文: 林浩, 贺璇, 陈燕, 庞鲁羿, 牛磊, 付秀丽. 比率型荧光纸芯片快速检测赭曲霉毒素A[J]. 分析化学, 2022, 50(9): 1336-1344. doi: 10.19756/j.issn.0253-3820.221127 shu

shu

Citation: LIN Hao, HE Xuan, CHEN Yan, PANG Lu-Yi, NIU Lei, FU Xiu-Li. A Rapid Ratiometric Fluorescence Biosensor for Detection of Ochratoxin A Based on Paper Chip[J]. Chinese Journal of Analytical Chemistry, 2022, 50(9): 1336-1344. doi: 10.19756/j.issn.0253-3820.221127 shu

shu

比率型荧光纸芯片快速检测赭曲霉毒素A

林浩 ^1, ,
贺璇 ^1, ,
陈燕 ^2, ,
庞鲁羿 ^1, ,
牛磊 ^1, ,
付秀丽 ^{1,
,}

1.
烟台大学化学化工学院, 烟台 264005;
2.
山东农业工程学院资源与环境工程学院, 济南 250100

通讯作者: 付秀丽,E-mail:fuxiuli@ytu.edu.cn

基金项目:
山东省高等学校青年创新团队发展计划项目（No.2009CB724100）和国家自然科学基金项目（Nos.61801274，21705139）资助。

摘要: 采用双荧光染料构建了一种新型比率型荧光纸芯片,将荧光染料Cy3和Cy5分别作为荧光供体和荧光受体,以二者间荧光共振能量转移(FRET)引起的荧光变化实现对赭曲霉毒素A (OTA)的一步法快速灵敏检测。将标记Cy3基团的核酸适配体(Aptamer)和标记Cy5基团的辅助DNA (aDNA)同时与纸芯片表面的互补DNA (CDNA)形成特定双链结构而发生FRET,导致Cy3荧光减弱而Cy5荧光增强。当存在OTA时,Aptamer与OTA结合后脱离双链结构,Cy3和Cy5两个荧光团远离,Cy3荧光增强而Cy5荧光减弱。实验结果表明,此系统的比率信号F₅₆₇/F₆₆₉(F₅₆₇/F₆₆₉为Cy3在567 nm处的荧光强度值与Cy5在669 nm处荧光强度值的比值)与OTA浓度在10-300 nmol/L范围内呈良好的线性响应,检出限(S/N=3)为5.6 nmol/L,花生和红酒样品中OTA的加标回收率为92.7%~107.6%。此传感器为食品中OTA等霉菌毒素污染检测提供了一种高效便捷的新方法。

关键词:

English

A Rapid Ratiometric Fluorescence Biosensor for Detection of Ochratoxin A Based on Paper Chip

LIN Hao ^1, ,
HE Xuan ^1, ,
CHEN Yan ^2, ,
PANG Lu-Yi ^1, ,
NIU Lei ^1, ,
FU Xiu-Li ^{1,
,}

1.
School of Chemistry and Chemical Engineering, Yantai University, Yantai 264005, China;
2.
School of Resources and Environmental Engineering, Shandong Agriculture and Engineering University, Jinan 250100, China

Corresponding author: FU Xiu-Li, fuxiuli@ytu.edu.cn

Received Date: 15 March 2022
Revised Date: 20 May 2022
Fund Project:
Supported by the Qingchuang Science and Technology program of Shandong Province, China (No.2019KJF029) and the National Natural Science Foundation of China (Nos.61801274, 21705139).

Abstract: A novel ratiometric fluorescent paper chip was constructed using dual fluorescent dyes based on fluorescence resonance energy transfer (FRET) between donor Cy3 and acceptor Cy5. The Cy3-labeled aptamer and Cy5-labeled auxiliary DNA (aDNA) were simultaneously combined with the complementary DNA (cDNA) pre-modified on the surface of paper chip to form a specific double stranded structure. After forming double stranded structure, a ratiometric fluorescence signal change was realized by FRET between donor Cy3 and acceptor Cy5. In this case, the fluorescence intensity of Cy3 decreased accompanied by the increasing of the fluorescence intensity of Cy5. In the presence of ochratoxin A (OTA), the specific binding of aptamer toward OTA led to Cy3-labeled aptamer detaching from the double stranded structure, which resulted in an increasing distance between Cy3 and Cy5. Therefore, the fluorescence intensity of Cy3 increased while the fluorescence intensity of Cy5 decreased. The fluorescence response ratio values (F₅₆₇/F₆₆₉:the fluorescence intensity of Cy3 at 567 nm to the fluorescence intensity of Cy5 at 669 nm) exhibited a good linear response with OTA concentration in the range of 10-300 nmol/L and the detection limit was 5.6 nmol/L. The designed ratiometric fluorescent paper chip was applied to detect OTA in spiked peanut and red wine samples, and the recoveries were 92.7%-107.6%. This sensor provided an efficient and convenient method for detection of mycotoxin in food.

Key words:

Paper chip
/ Ratiometric fluorescence
/ Fluorescence resonance energy transfer
/ Ochratoxin A
/ Aptamer

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