UNS:Thermal control of avidity for separation of biologicals

1511227 RaoNorth卡罗莱纳State UAffinity色谱法利用固定在表面上的分子或配体与目标物质之间的高度特异性相互作用。在所提出的工作中，亲和吸附剂将使用相同的合成方案用所提出的分子或配体产生，用于小规模和大规模分离。预期这些亲和吸附剂具有耐用性，并且在灭菌后可多次重复使用。然而，预计生产具有高价值生物制品的一次性应用吸附剂仍将具有成本效益。将与北卡罗来纳州的一家州立公司合作，探索开发用于复杂生物制品的商业膜分离系统。为了鉴定温度敏感的亲和配体，将评价响应于温度变化而改变构象的基于肽的支架。将产生两种模型靶标的亲和配体：在造血干细胞上表达的CD117和线粒体蛋白TOM 22。随后，将在细胞和线粒体分离的背景下评估这些配体的有效性。因此，将筛选肽的组合文库以鉴定模型靶标的肽结合剂。还将使用产生的肽构建热响应配体。随后，将评估温度对这些配体的结合亲和力的影响。最后，将在分离CD 117+细胞和线粒体的背景下评价用所选配体官能化的磁珠的使用。

1511227RaoNorth Carolina State UAffinity chromatography exploits the highly specific interaction between molecules or ligands immobilized on a surface and the target species of interest. In the proposed work affinity adsorbents will be generated with the proposed molecules or ligands for both small and large scale separations using the same synthesis scheme. These affinity adsorbents are expected to be robust and amenable to multiple re-uses after sterilization. Yet, it is expected that production of adsorbents for single-use applications with high value biologicals will still be cost-effective. Collaboration with a North Carolina State-based company will be undertaken to explore the development of commercial membrane-based separation systems for complex biologicals. To identify temperature-sensitive affinity ligands, a peptide-based scaffold that changes conformation in response to changes in temperature will be evaluated. Affinity ligands for two model targets will be generated: CD117, which is expressed on hematopoietic stem cells, and TOM22, a mitochondrial protein. Subsequently, the effectiveness of these ligands will be evaluated in the context of cell and mitochondrial separation. Accordingly, combinatorial libraries of peptides will be screened to identify peptides binders for the model targets. Thermo-responsive ligands will also be constructed using the peptides generated. Subsequently, the effect of temperature on the binding affinity of these ligands will be evaluated. Finally, the use of magnetic beads functionalized with the selected ligands will be evaluated in the context of separation of CD117+ cells and mitochondria.