基于分子印迹识别的 Fab 小分子抗体分离新方法研究

Small molecule antibody is one promising candidate for diversification of antibody industry. With smaller molecular weight, simple structure, strong penetration and low toxicity, it could overcome effectively the disadvantages of monoclonal antibody and gains increasing attention. However, the further development of research and industry is deeply restricted because of the high preparation cost of small molecule antibody. The absence of proper separation and purification methods is one important reason. Applicant will focus on the chromatography, the core technology of separation and purification, to carry on this project. Fab antibody will be chosen as the target. In view of its peculiar structure and size, the molecular simulation technique will be carried out first to screen ligands, and the mechanism between ligands and Fab will be analyzed. The rules of adsorption/desorption will be summarized as well. Then imprinting system will be screened further and surface molecule imprinting will be adopted to prepare imprinted adsorbent. The molecular imprinted recognition will be introduced to improve the selectivity of chromatography, and the synergistic mechanism of adsorption ligands and imprinting layer during the adsorption process will be discussed. After design and optimization of the separation process, a new chromatographic method for Fab with high efficiency will be founded. This project research will be a support for the further development of antibodies industry, and can be referenced in separation process for other small molecule antibodies and proteins.

小分子抗体是抗体产业实现多样化的重要对象，拥有较小的分子量，简单的结构，极强的组织穿透能力，较低的毒性，可以有效弥补单克隆抗体的不足，获得了越来越多的关注。但小分子抗体缺乏针对性的分离纯化手段，导致其制备成本高昂，严重制约了产研的进一步发展。申请人围绕分离纯化阶段的核心技术——层析进行研究：以Fab小分子抗体为对象，针对其特有的结构大小，通过分子模拟技术筛选适配Fab的吸附配基，探讨配基-Fab间分子间作用机制，总结吸附/解吸规律；进一步筛选合适印迹体系，以表面印迹法制备印迹层析介质，引入分子印迹识别能力，提升层析介质选择性，并探讨吸附配基及印迹层在吸附过程中的协同作用机制；最终设计并优化分离流程，构建Fab小分子抗体的高效分离新方法。本项目研究成果可借鉴于其他小分子抗体与蛋白质的分离过程设计，为抗体产业的进一步发展提供助力。

以抗体为代表的生物药受困于高昂的下游制备成本，制约其产研的进一步发展。本项目以分离纯化的核心技术，层析技术为突破口，在执行期间成功设计制备了混合模式/空间识别、离子交换/空间识别、疏水作用/空间识别等系列双模式识别介质，涵盖了主要的层析模式，进行了有效的革新优化。试验了新型介质的吸附容量、选择性等性能在静态和动态吸附条件下的变化规律，建立了以双模式识别层析为核心的分离过程；探讨了识别模式、环境因素与性能之间的关系。多篇研究论文发表在国内外权威刊物上，如：Biochemical Engineering Journal，Journal of Chemical & Engineering Data，Journal of Chemical Technology and Biotechnology等，发表SCI收录论文7篇，其中一区二区论文5篇。申请国家发明专利4项，授权1项。获省部级奖项2项。项目执行期间共培养2位硕士研究生，建立了较为广泛的国际学术交流与合作关系。