Depletion of algal toxin-contaminated water using selective biofilters based on plant-produced antibodies (plantibodies)

虽然利用转基因植物进行生物修复或污染部位的原位清理已经有一段时间了，但迄今为止，很少有人注意到植物中抗体的产生及其在选择性消耗中的离体应用。因此，本研究将以微囊藻毒素为例，在植物体内低成本生产出高仿射和特异性的藻类毒素抗体。其基础是在以前的研究项目中产生的单克隆抗体（Mab 10E7，物种：小鼠）。可变结构域的序列将被确定，为植物优化并亚克隆到合适的植物转化载体中，这些转化载体已经包含恒定的抗体序列。此外，将构建包含不同标签序列和融合蛋白的scFv片段。将使用叶基（烟草）和种子基（大麦）系统。亲和纯化的植物产生的抗体（植物抗体）将使用微滴板- elisa和表面等离子体共振（SPR）详细表征其结合特性。该小鼠单克隆抗体将作为参考。为了确保未来应用的成本效益，将对大致纯化的馏分（依次进行pH和温度处理，然后过滤）进行升级测试。将植物体部分固定在合适的基质上，例如膜、多孔聚合物单体或多孔玻璃中，将使用添加了微囊藻毒素的模型水样来确定它们的消耗应用。

Although the use of genetically modified plants for bioremediation, or the in situ cleaning of contaminated sites, has been known for quite some time, little attention has so far been paid to the production of antibodies in plants and their ex vivo application in selective depletion. Therefore, highly affine and specific antibodies against algal toxins using microcystin as an example will be produced in plants at low cost within this research project. The basis is a monoclonal antibody (Mab 10E7, species: mouse) generated in a former research project. The sequence of the variable domains will be determined, optimised for plants and subcloned into suitable plant transformation vectors, which already contain constant antibody sequences. In addition, a scFv fragment containing different tag sequences and fusion proteins will be constructed. Leaf-based (tobacco) as well as seed-based (barley) systems will be used.Affinity-purified plant-produced antibodies (plantibodies) will be characterised in detail for their binding properties using microtitre plate-ELISA and surface plasmon resonance (SPR). The monoclonal mouse antibody will be used as reference. To assure cost-efficiency for future applications, roughly purified fractions (sequential pH and temperature treatment followed by filtration) will be tested for the upscaling. Following immobilisation of the plantibody fractions on suitable substrates, for instance membranes, porous polymer monoliths or in porous glasses, their application for depletion will be defined using model water samples spiked fortified with microcystins.