Oil Crops Research Institute of the Chinese Academy of Agricultural Sciences, Wuhan, 430062, People's Republic of China.
Key Laboratory of Biology and Genetic Improvement of Oil Crops, Ministry of Agriculture, Wuhan, 430062, People's Republic of China.
Sci Rep. 2017 Jun 28;7(1):4348. doi: 10.1038/s41598-017-04195-6.
Aspergillus and its poisonous mycotoxins are distributed worldwide throughout the environment and are of particular interest in agriculture and food safety. In order to develop a specific method for rapid detection of Aspergillus flavus to forecast diseases and control aflatoxins, a nanobody, PO8-VHH, highly reactive to A. flavus was isolated from an immunized alpaca nanobody library by phage display. The nanobody was verified to bind to the components of extracellular and intracellular antigen from both A. flavus and A. parasiticus. To construct a sandwich format immunoassay, polyclonal antibodies against Aspergillus were raised with rabbits. Finally, a highly selective nanobody-polyclonal antibody sandwich enzyme-linked immunosorbent assay was optimized and developed. The results revealed that the detection limits of the two fungi were as low as 1 μg mL, and that it is able to detect fungal concentrations below to 2 μg mg of peanut and maize grains in both artificially and naturally contaminated samples. Therefore, we here provided a rapid and simple method for monitoring Aspergillus spp. contamination in agricultural products.
曲霉菌及其有毒的真菌毒素在全球范围内分布于环境中，在农业和食品安全方面特别受到关注。为了开发一种快速检测黄曲霉菌的特异性方法，以预测疾病并控制黄曲霉毒素，本研究通过噬菌体展示技术从免疫羊驼纳米抗体文库中分离出一种对黄曲霉菌高度反应的纳米抗体 PO8-VHH。该纳米抗体被证实可与黄曲霉菌和寄生曲霉菌的细胞外和细胞内抗原成分结合。为了构建夹心型免疫测定法，用兔制备了针对曲霉菌的多克隆抗体。最后，对高度选择性的纳米抗体-多克隆抗体夹心酶联免疫吸附测定法进行了优化和开发。结果表明，两种真菌的检测限低至 1μg/mL，并且能够检测到人工和自然污染样品中花生和玉米颗粒中真菌浓度低至 2μg/mg。因此，本研究提供了一种快速简便的方法来监测农产品中曲霉菌属的污染。
