中文摘要：抗独特型抗体（AIds）具有稳定的蛋白质构象，为免疫分析提供了一种替代传统抗原的方法。本研究从羊驼免疫噬菌体展示文库中筛选出抗黄曲霉毒素M1（AFM1）的抗独特型重链抗体（VHH）或与单克隆抗体(mAb) 2C9结合的纳米体VHH C4的可变域。VHH-C4在特异性和敏感性方面表现突出，IC50为0.25ng/mL，与其它黄曲霉毒素类似物几乎没有交叉反应。据悉，这是首个为AFM1研制抗独特型纳米体的报告。以VHH C4为包被抗原，建立了环境友好的竞争ELISA试剂盒。在0.10 - 0.60 ng/mL范围内呈良好的线性关系，检测限(LOD)为0.05 ng/mL。加标样品(纯牛奶、酸奶和奶粉)的回收率为84.1% - 119.1%。自然污染样品中，AFM1含量的ELISA测定结果与高效液相色谱法（HPLC）高度一致。可见，纳米体VHH-C4是一种很有前途的替代抗原，可用于乳制品中AFM1的免疫学检测。
外文摘要：Anti-idiotypic antibodies (AIds) display stable protein conformation, offering an alternative to conventional antigens in immunoassays. In this work, an anti-idiotypic variable domain of heavy-chain antibody (VHH) or nanobody named VHH C4 binding to monoclonal antibody (mAb) 2C9 against aflatoxin M-1 (AFM(1)) was screened out from an alpaca immunized phage-displayed library. VHH C4 shows outstanding features in specificity and sensitivity, the IC50 is 0.25 ng/mL and exhibit almost no cross-reactivity with other aflatoxin analogs. As our knowledge, it is the first report for developing an anti-idiotypic nanobody for AFM(1). An environment-friendly competitive ELISA was developed using VHH C4 as a coating antigen. The results revealed a linear range from 0.10 ng/mL to 0.60 ng/mL, with a limit of detection (LOD) of 0.05 ng/mL. The recovery of spiked samples (pure milk, yogurt, and milk powder) ranged from 84.1% to 119.1%. The ELISA results of AFM(1) content in naturally contaminated samples were in good agreement with those determined by high performance liquid chromatography (HPLC). The results demonstrated that the nanobody VHH C4 is a promising alternative antigen in developing immunological methods for the detection of AFM1 in dairy products.
外文关键词：Nanobody；Aflatoxin M-1；Phage-displayed；ELISA；Milk products
作者：Cai, C；Zhang, Q；Nidiaye, S；Yan, HL；Zhang, W；Tang, XQ；Li, PW
作者单位：Chinese Acad Agr Sci；Minist Agr & Rural Affairs
期刊名称：MICROCHEMICAL JOURNAL
期刊影响因子：3.594
出版年份：2021
出版刊次：167
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