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Öğe Antibody purification with protein A attached supermacroporous poly(hydroxyethyl methacrylate) cryogel(Elsevier, 2009) Alkan, Hueseyin; Bereli, Nilay; Baysal, Zuebeyde; Denizli, AdilImmunoglobulin G (IgG) Purification from human plasma with protein A attached supermacroporous poly(hydroxyethyl methacrylate) [PHEMA] cryogel has been studied. PHEMA cryogel was prepared by bulk polymerization which proceeds in aqueous solution of monomer frozen inside a plastic syringe (cryo-polymerization). After thawing, the PHEMA cryogel contains a Continuous matrix having interconnected pores of 10-200 mu m size. Protein was covalently attached onto the PHEMA cryogel via cyanogen bromide (CNBr) activation. The maximum IgG adsorption oil the PHEMA/protein A cryogel Was found to be 83.2 mg/g at pH 7.4 from aqueous Solutions. The non-specific IgG adsorption onto the PHEMA cryogel was about 0.38 mg/g. The macropore size of the cryogel makes it possible to process blood cells without blocking the Column. Higher adsorption capacity was observed from human plasma (Lip to 88.1 mg/g). Adsorbed IgG was eluted using 0.1 M glycine-HCl buffer (pH 3.5) with a purity of 85%. PHEMA-protein A cryogel was used for repetitive adsorption/desorption of IgG without noticeable loss in IgG adsorption capacity after 10 cycles. PHEMA-protein A cryogel showed several advantages Such as simpler preparation procedure, good selectivity for IgG Purification from human plasma and good stability throughout repeated adsorption-desorption cycles. (C) 2009 Elsevier B.V. All rights reserved.Öğe Molecularly imprinted cryogel as a pH-responsive delivery system for doxorubicin(Taylor & Francis Inc, 2017) Cetin, Kemal; Alkan, Huseyin; Bereli, Nilay; Denizli, AdilIn this study, implantable and degradable molecularly imprinted cryogel was prepared for pH-responsive delivery of doxorubicin. Cryogel discs were synthesized using amino acid-based functional monomer with HEMA and gelatin. The molecularly imprinted discs were characterized by scanning electron microscopy, Fourier transform infrared spectroscopy, degradation and swelling tests. In vitro delivery experiments were carried out in order to examine the effects of medium pH and drug content. The degree of degradation of composite cryogels was found to be 83.45 +/- 1.86% after 56days. The release profiles of DOX from molecularly imprinted cryogel discs exhibit a biphasic delivery. It was observed that an initial burst release step from 0 to 12h was followed by a slower and sustained release. Release rate of DOX from cryogel discs increased in more acidic conditions. Kinetic studies showed that a combination of diffusion and erosion control is mainly responsible from the general release behaviors of molecularly imprinted cryogel discs.Öğe Poly(Hydroxyethyl methacrylate) immunoaffinity cryogel column for the purification of human immunoglobulin M(MDPI Multidisciplinary Digital Publishing Institute, 2020) Bakhshpour, Monireh; Topçu, Aykut Arif; Bereli, Nilay; Alkan, Huseyin; Denizli, AdilHuman immunoglobulin M (hIgM) antibodies are considered as hopeful tools for diseases therapy. Therefore, chromatography approaches are used to purify hIgM with a single step. In this study, we prepared a poly(hydroxyethyl methacrylate) based immunoaffinity p(HEMA-I) cryogel column by using cyanamide to immobilize antihuman immunoglobulin on the p(HEMA) cryogel for purification of hIgM in aqueous solution and artificial human plasma. The characterization of the p(HEMA) cryogel column was performed by using a scanning electron microscope (SEM), micro-computerized tomography (µ-CT), Fourier transform infrared spectroscopy (FTIR), swelling degree and macro-porosity. Further, the optimizations of various parameters were performed such as, pH, ionic strength, temperature and concentration of hIgM in aqueous solutions. In addition, the Langmuir adsorption model was supported by experimental results. Maximum adsorbed amount of hIgM corresponded to 11.1 mg/g at pH 5.75 [morpholino ethanesulfonic acid (MES buffer)]. Our results indicated that the p(HEMA-I) cryogel column can be reused at least 10 times without significant loss in adsorption capacity. As a natural source, artificial human plasma was selected for hIgM adsorption and the purity of hIgM was evaluated using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE).Öğe Selective removal of the autoantibodies from rheumatoid arthritis patient plasma using protein A carrying affinity cryogels(Elsevier, 2010) Alkan, Huseyin; Bereli, Nilay; Baysal, Zubeyde; Denizli, AdilRheumatoid arthritis is a chronic, progressive, deabilitating autoimmune disease that occurs in approximately 1% of adults. Rheumatoid arthritis is characterized by chronic polyarthritis and destruction of multiple joints. In this study, IgM-antibody removal from human plasma with supermacroporous poly(hydroxyethyl methacrylate) [PHEMA] cryogel carrying protein A has been evaluated. The PHEMA cryogel was prepared by bulk polymerization which proceeds in an aqueous solution of monomer frozen inside a plastic syringe (cryo-polymerization). After thawing, the PHEMA cryogel contains a continuous matrix having interconnected macropores of 10-200 mu m size. Pore volume in the PHEMA cryogel was 71.6%. Protein A molecules were covalently immobilized onto the PHEMA cryogel via cyanogen bromide (CNBr) activation. The PHEMA cryogel was contacted with blood in in vitro system for the determination of blood-compatibility. The supermacroporous structure of the PHEMA cryogel makes it possible to process blood cells without blocking the cryogel column. IgM-antibody adsorption capacity decreased significantly with the increase of the plasma flow-rate. The maximum IgM-antibody adsorption amount was 42.7 mg/g. IgM-antibody molecules could be repeatedly adsorbed and eluted without noticeable loss in the IgM-antibody adsorption amount. (C) 2010 Elsevier B.V. All rights reserved.
