Tez Türü: Doktora
Tezin Yürütüldüğü Kurum: Sivas Cumhuriyet Üniversitesi, Fen Bilimleri Enstitüsü, Fen Bilimleri Enstitüsü, Türkiye
Tez Danışmanı: Adil Denizli,Ferda Candan
Tezin Onay Tarihi: 2009
Tezin Dili: Türkçe
Açık Arşiv Koleksiyonu: AVESİS Açık Erişim Koleksiyonu
Desteklendiği Program: Diğer
Özet:
In this study, purification of human immunoglobulin-G (IgG) from aqueous solutions and human serum were studied by using Con A immobilized m-poly-glycidyl methacrylate (GMA) monosize microbeads. M-poly(GMA) monosizes microbeads were prepared by dispersion polymerization in the presence of Fe3O4 nanopowder. FT-IR, 1H NMR, SEM, ESR, VSM were used for characterization of the m-poly(GMA) microbeads.The epoxy ring of m-poly(GMA) microbeads were opened in alkali conditions. Con A was immobilized by covalent binding onto epoxy ring opened m-poly(GMA) beads via hidroxyl groups. Maximum Con A immobilization value was 12.5 mg g-1 at pH 7.4 (0,1 M phosphate buffer), at 4 oC. The maximum IgG adsorption capacity on m-poly(GMA)-Con A beads in batch system and MSFB system were 35.5 and 37.7 mg g-1, respectively. Maximum adsorption of IgG was observed at pH 6.0 (0.1 M phosphate buffer). In MSFB system, flow rate and magnetic field effect was also studied. The adsorption capacity decreased drastically from 37.7 to 10.9 mg g-1 with the increase of the flow rate from 1.0 to 4.0 mL min-1. When magnetic field was increased, adsorbed IgG was decreased from 37.7 to 18.0 mg g-1. In both system, desorption of adsorbed IgG on polymeric microbeads and reusability of microbeads were investigated by using ethylene glycole + glucose as desorption agent.The maximum IgG adsorption capacity on m-poly(GMA)-Con A beads from human serum was found 48 mg g-1. 2.0 M NaCl was used for desorption of adsorbed IgG on m-poly(GMA)-Con A microbeads from human serum. The purity of IgG was investigated by SDS-PAGE. All the measurements were performed by using ELISA techniques. Spectroflourimetric studies were done for investigating structural change of IgG at the course of adsorption and desorption studies.Last part of this study include of SPR (Surface Plasmon Resonance) kinetic studies between Con A and IgG molecules. For this aim, Con A was immobilized on SPR chip followed by detecting of plasmone angle and finally kinetic studies were done. In order to determine the kinetic and binding constant three different isotherms, Langmuir, Freundlich and Langmuir-Freundlich, were applied to kinetic data. As seen from the results Langmuir isotherm has high correlation constant.Key Words: m-poly(GMA), Con A immobilization, dispersion polymerization, lectin affinity chromatography, IgG purification.