Institute of Medicinal Plant Development, Chinese Academy of Medical Science & Peking Union Medical College, Beijing, 100193, PR China.
Analyst. 2012 Jan 7;137(1):229-36. doi: 10.1039/c1an15487g. Epub 2011 Nov 2.
Although anti-zearalenone (ZEN) antibodies have been widely prepared, these antibodies cross-react with α-zearalenol (α-ZOL), β-zearalenol (β-ZOL), zearalanone (ZAN), α-zearalanol (α-ZAL) and β-zearalanol (β-ZAL). To overcome this problem and improve the specificity of immunoassays, we produced anti-ZEN antibodies based on a ZEN-cationic protein conjugate. In this study, ZEN was coupled with cationic bovine serum albumin (cBSA) via a Mannich reaction. After BALB/c mice were immunized with ZEN-cBSA, an immunological response was rapidly induced. The titers of the polyclonal antisera and monoclonal antibody were 30,000 and 20,000, respectively. Cross-reactivity (CR) values of the anti-ZEN polyclonal antisera and monoclonal antibody with the 5 analogs were <7% and <2%, respectively. An indirect competitive enzyme-linked immunosorbent assay based on the monoclonal anti-ZEN antibody was established. The recovery rates of ZEN in spiked cereal and feed were in the range of 80%-120% with coefficients of variation <15%. The intra-assay variation and inter-assay variation in assay buffer were both <5%. Therefore, the results demonstrated a feasible approach for preparing highly specific, higher titer and more rapidly induced antibodies against ZEN by using a ZEN-cBSA conjugate as the immunogen instead of currently used immunogens.
尽管已经制备了许多抗玉米赤霉烯酮（ZEN）抗体，但这些抗体与α-玉米赤霉醇（α-ZOL）、β-玉米赤霉醇（β-ZOL）、玉米赤霉酮（ZAN）、α-玉米赤霉烯醇（α-ZAL）和β-玉米赤霉烯醇（β-ZAL）有交叉反应。为了解决这个问题并提高免疫分析的特异性，我们基于 ZEN-阳离子蛋白缀合物制备了抗 ZEN 抗体。在这项研究中，ZEN 通过曼尼希反应与阳离子牛血清白蛋白（cBSA）偶联。用 ZEN-cBSA 免疫 BALB/c 小鼠后，迅速引起免疫反应。多克隆抗血清和单克隆抗体的效价分别为 30,000 和 20,000。多克隆抗血清和单克隆抗体与 5 种类似物的交叉反应（CR）值均<7%和<2%。建立了基于单克隆抗 ZEN 抗体的间接竞争酶联免疫吸附测定法。ZEN 在添加的谷物和饲料中的回收率在 80%-120%范围内，变异系数<15%。在测定缓冲液中的日内和日间变异均<5%。因此，结果表明，使用 ZEN-cBSA 缀合物作为免疫原而不是目前使用的免疫原，是一种可行的方法，可以制备针对 ZEN 的高特异性、高效价和更快诱导的抗体。
