Functionalized Membranes for Membrane Chromatography

用于大规模纯化单克隆抗体的Ritchie技术对于新疗法变得越来越重要。 亲和层析是有效的，但费用高和低通量使大规模操作复杂化。 膜色谱法提供更高的通量，并且如果可以改善容量、选择性和孔径分布，则膜色谱法将是极好的选择。 该提案的重点是使用功能化膜来满足这些挑战中的每一个。 官能化膜提供优异的通量和高容量，因为吸附位点位于孔中的聚合物接枝物上，而不是孔表面上。 在所提出的研究中，吸附位点是基于苯丙氨酸-132/酪氨酸-133基序的类似物，在固定在微滤膜孔中的聚合物刷中沿沿着每个官能化链多次掺入。 聚合物刷是通过一系列取代苯乙烯的顺序阳离子聚合形成的。 氯甲基苯乙烯重复单元的甘氨酸官能化将提供间隔子以限制空间位阻并抑制吸附抗体的变性。 牛免疫球蛋白和牛血清白蛋白的单独和竞争性吸附将用作容量和选择性的试验系统。 功能化的聚合物刷将被提取，并通过1H NMR和X射线晶体学进行充分的非原位表征。 此外，将测试功能化膜的孔径分布，以表征孔径变窄。 这项研究将导致改进的膜吸附分离蛋白质和抗体。 此外，它将导致在序列聚合中的发现，以产生定制的结构，非肽聚合物的测序，以及合成亲和配体的设计。 这项研究的结果将在经过评审的期刊出版物上以及在区域和国家会议上的发言中广泛传播。

CBET-0756460RitchieTechnologies for large scale purification of monoclonal antibodies are becoming increasingly important for new therapies. Affinity chromatography is effective, but expense and low throughput complicate large scale operation. Membrane chromatography provides higher throughput, and would be an excellent option if the capacity, selectivity, and pore size distribution can be improved. The focus of this proposal is to meet each of these challenges using functionalized membranes. Functionalized membranes provide excellent throughput and high capacity because adsorption sites are located on polymer grafts in the pores, rather than on the pore surface. In the proposed research, the adsorption sites are based on an analog for the phenylalanine-132/tyrosine-133 motif, incorporated multiple times along each functionalized chain in a polymer brush immobilized in the pores of a microfiltration membrane. The polymer brush is formed by sequential cationic polymerization of a series of substituted styrenes. Glycine functionalization of chloromethyl styrene repeat units will provide spacers to limit steric hindrance and inhibit denaturing of the adsorbed antibody. Individual and competitive sorption of bovine immunoglobulin and bovine serum albumin will be used as a test system for capacity and selectivity. The functionalized polymer brushes will be extracted and fully characterized ex-situ by 1H NMR and X-ray crystallography. In addition, functionalized membranes will be tested for pore size distribution to characterize narrowing of the pore size. This research will result in improved membrane adsorbers for protein and antibody separation. In addition, it will lead to discoveries in sequential polymerization to generate customized structures, sequencing of non-peptide polymers, and design of synthetic affinity ligands. The results of this research will be broadly disseminated in refereed journal publications and in presentations at regional and national meetings.