酶联适体分析法检测葡萄酒中的赭曲霉素A  

Enzyme-Linked Aptamer Assay for Detection of Ochratoxin A in Wine

作　　者：赵秋伶[1] 史素青[2] 

机构地区：[1]辽宁工程技术大学矿业技术学院,辽宁葫芦岛125105 [2]西北大学化学与材料学院,陕西西安710069

出　　处：《食品与生物技术学报》2015年第4期396-401,共6页Journal of Food Science and Biotechnology

基　　金：国家自然科学基金项目(21004047);陕西省青年科技新星人才项目(2014KJXX-62)

摘　　要：建立了竞争取代酶联适体分析方法,检测葡萄酒中的赭曲霉素A （OTA）.核酸适体和OTA特异性结合导致与核酸适体杂交的短链DNA 解链,解离的DNA 作为捕获元素,进一步特异性结合辣根过氧化物酶（HRP）,HRP催化四甲基二苯胺（TMB）底物显色,测定A450 nm与浓度的线性关系,确定OTA的检出限.考察了DNA包被浓度、杂交温度和封闭液等因素对检测的影响.结果表明,在优化的条件下,所建立的竞争取代酶联适体分析法对OTA检测有高灵敏度,检测限0.88 μg/L,线性范围1～100 μg/L在葡萄酒中添加时,加标回收率为92.03 %～106.6 %,相对标准偏差RSD（n=5）小于2.1 %,此方法可用于葡萄酒中OTA的快速测定.ompetitive enzyme-linked aptamer assay for Ochratoxin A(OTA) detection in wine wasinvestigated in this study. The binding of aptamer to the target OTA resulted in the dissociation of theDNA short chain from the aptamer, which was further captured by horseradish peroxidase(HRP). HRPcatalyzed 3,3',5,5'.- Tetramethylbenzidine(TMB) substrate and finally leaded to a characteristicchange that was detectable by the absorption of ultraviolet at 450 nm. The detection limit of OTA wasdetermined by the linear relationship between OTA concentration and the absorbance. The factorsinfluenced the effect of detection were investigated, including DNA concentration, temperature ofhybridization, and blocking buffer solution. This work described the high performance of thecompetitive enzyme-linked aptamer assay(ELAA) for the determination of OTA. Under the optimalcondition, the limit of detection attained was 0.88 μg/L and the linearity was in the range 1 to 100μg/L. For the OTA detection in wine, the recovery rate was between 92.03% and 106.6%, and therelative standard deviation(RSD, n=5) was within 2.1%. This study validates the competitive ELAA isa useful tool for a rapid detection of OTA in wine.

关 键 词：赭曲霉素A 竞争取代 酶联适体分析 葡萄酒 

分 类 号：TS262.6[轻工技术与工程—发酵工程]