基于量子点标记的赭曲霉毒素A快速、高灵敏荧光免疫层析检测方法的建立及应用

赭曲霉毒素A（ochratoxin A,OTA）具有肾毒性、致畸性、致癌性和免疫毒性,广泛存在于各种粮食作物及其副产品中,是食品和饲料原料的重要污染物,可在人类及动物体内蓄积,在已知发现的真菌毒素中,重要性和危害性仅次于黄曲霉毒素。本研究通过采用量子点荧光微球（quantum dots,QDs）标记OTA单克隆抗体,并基于免疫层析原理,优化、建立了OTA高灵敏荧光免疫层析检测方法（FICGA）,15min即可实现对农产品中OTA污染的快速定量检测。该方法检测下限（IC₁₀）达到0.04ng/mL,检测区间（IC₂₀-IC₈₀）为0.05-0.59ng/mL,半数抑制率（IC₅₀）为0.18ng/mL。与OTA类似物OTB、OTC交叉反应性为7.3%和11.9%,对其他常见真菌毒素AFB₁、ZEN、FB₁和DON均无交叉反应。在玉米、面粉和大豆样本中的加标回收率可达83.2%-117.8%,与LC-MS/MS同时对天然样本中OTA含量的检测结果表明,两种方法相关性良好。本研究建立的FICGA快速、灵敏,可满足基层单位和现场的快速检测需求,具有很好的应用前景。

Rapid and sensitive detection of ochratoxin A by using a quantum dots-based immunochromatographic assay

Ochratoxin A (OTA) produced by Aspergillus and Penicillium species, is one of the most widespread toxic members of ochratoxins and has been shown to be nephrotoxic, teratogenic, carcinogenic and immunotoxic to several animal species. Rapid detection of OTA is important to prevent contaminated food from entering into the food chain. A rapid and sensitive fluorescent immunochromatographic assay (FICGA) based on the use of quantum dots (QDs) was prototyped and applied for detecting OTA in cereal samples. The detection signal was enhanced which in turn improved the sensitivity of the assay. The limit of detection of FICGA was 0.04ng/mL while the IC₅₀ was 0.18ng/mL. The linear working range was 0.05-0.59ng/mL. The cross-reactivities to the OTA analogues, ochratoxin B and ochratoxin C, were 7.3% and 11.9% respectively. No cross-reactivity (<0.01%) was observed to other co-occurring mycotoxins (AFB₁, ZEN, FB₁ and DON). The recovery rates in spiked corn, wheat and soybean samples were 83.2%-117.8%, and the intra-day and inter-day relative standard deviations were <10%. Simultaneous analysis of commercial samples (corn, wheat, and feedstuff) showed a good correlation between FICGA and liquid chromatography tandem mass spectrometry (LC/MS-MS). Hence, this developed quantum dots-based FICGA shows great potential for rapid, sensitive, simple and cost-effective quantitative detection of OTA in food safety control.