| 1. |
- Ahlqvist, Josefin, et al.
(författare)
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Monitoring the production of inclusion bodies during fermentation and enzyme-linked immunosorbent assay analysis of intact inclusion bodies using cryogel minicolumn plates
- 2006
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Ingår i: Analytical Biochemistry. - : Elsevier BV. - 1096-0309 .- 0003-2697. ; 354:2, s. 229-237
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Tidskriftsartikel (refereegranskat)abstract
- A novel minicolumn chromatgraphic method to monitor the production of inclusion bodies during fermentation and anenzyme-linked immunosorbent assay (ELISA) system allowing direct analysis of the particles with surface-displayed antigens are described. A 33-kDa protein containing 306 amino acids with three sulfur bridges produced its inclusion bodies wits labeled with polyclonal antibodies against 15 amino acid (anti-A15) and 17 amino acid (anti-B17) residues at the N- and C-terminal ends of the protein, respectively. Labeled particles were bound to macroporous Monolithic protein A-cryogel adsorbents inserted into the open-ended wells of a 96-well plate (referred to as protein A-cryogel minicolumn plate). The concept behind this application is that the binding degree of inclusion bodies from lysed fermentation broth to the cryogel minicolumns increases with an increase in their concentration during fermentation. The technique allowed LIS to monitor the increase in the production levels of the inclusion bodies as the fermentation process progressed. The system also has a built-in quality parameter to ensure that the target protein has been fully expressed. Alternatively, inclusion bodies immobilized on phenyl-cryogel minicolumn plate were used in indirect ELISA based on anti-A15 and anti-B17 antibodies against terminal amino acid residues displayed oil the surface of inclusion bodies. Drainage-protected properties of the cryogel minicolumns allow performance of successive reactions with tested immunoglobulin G (IgG) samples and enzyme-conjugated secondary I-G and of enzymatic reaction within the adsorbent. (c) 2006 Elsevier Inc. All rights reserved.
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| 2. |
- Andac, M, et al.
(författare)
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Poly(hydroxyethyl methacrylate)-based macroporous hydrogels with disulfide cross-linker
- 2008
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Ingår i: Macromolecular Chemistry and Physics. - : Wiley. - 1521-3935 .- 1022-1352. ; 209:6, s. 577-584
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Tidskriftsartikel (refereegranskat)abstract
- Biodegradable supermacroporous PHEMA cryogels were produced by combining two cross-linkers, poly(ethylene glycol) diacrylate and a newly developed disulfide water soluble crosslinker, N,N'-bis(methacryloyl)-L-cystine. The biodegradable PHEMA cryogels were prepared with gel fraction yields up to 70% and were characterized by highly interconnected pores of micrometer size and good mechanical stability. When subjected to reductive agents like DTT, the biodegradable PHEMA cryogels disintegrated into small pieces. The rate of disintegration was controlled by the crosslinking density in the cryogels and the DTT concentration.
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| 3. |
- Babac, Ceyhun, et al.
(författare)
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Binding of antibodies to concanavalin A-modified monolithic cryogel
- 2006
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Ingår i: Reactive & Functional Polymers. - : Elsevier BV. - 1873-166X .- 1381-5148. ; 66:11, s. 1263-1271
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Tidskriftsartikel (refereegranskat)abstract
- Binding of human immunoglobulin-G (IgG) from aqueous solutions and human plasma to concanavalin A (Con A) immobilized poly(acrylamide-allyl glycidyl ether) [poly(AAm-AGE)] monolithic cryogel has been studied. Poly(AAm-AGE) cryogel was prepared by bulk polymerization which proceeds in aqueous solution of monomers frozen inside a column (cryo-polymerization). After thawing, the monolithic cryogel contains a continuous polymeric matrix having interconnected pores of 10-100 pm size. Con A was immobilized by covalent binding onto poly(AAm-AGE) cryogel via epoxy groups. The maximum IgG adsorption on the Con A-poly(AAm-AGE) cryogels was observed at pH 7.4 from aqueous solutions. The non-specific IgG adsorption onto the plain poly(AAm-AGE) adsorbents was about 0.25 mg/g. Up to 6.7 mg/g IgG were bound to Con A-poly(AAm-AGE) cryogels from aqueous solutions. The large pore size of the cryogel makes it possible to process blood cells without blocking the column. Higher adsorption capacity was observed from human plasma (up to 25.6 mg/g). Bound IgG was eluted using 2.0 M NaCl with a purity of 85%. Adsorption capacities of other blood proteins were obtained as 1.0 mg/g for fibrinogen and 1.7 mg/g for albumin. The total protein adsorption was determined as 28.6 mg/g. Con A-poly(AAm-AGE) cryogels was used for repetitive adsorption/desorption of IgG molecules without noticeable loss in IgG adsorption capacity after 10 cycles. (c) 2006 Elsevier B.V. All rights reserved.
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| 4. |
- Baydemir, Goezde, et al.
(författare)
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Bilirubin recognition via molecularly imprinted supermacroporous cryogels
- 2009
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Ingår i: Colloids and Surfaces B: Biointerfaces. - : Elsevier BV. - 1873-4367 .- 0927-7765. ; 68:1, s. 33-38
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Tidskriftsartikel (refereegranskat)abstract
- Recent years molecular imprinting has received considerable attention as an excellent and simple approach to recognize small molecules and bioactive substances. The aim of this study is to prepare the bilirubin-imprinted supermacroporous cryogels which can be used for the adsorption of bilirubin from human plasma. N-methacryloyl(L)-tyrosinemethylester (MAT) was chosen as the preorganization monomer. In the first step, bilirubin was complexed with MAT and the bilirubin-imprinted poly(hydroxyethyl methacrylate-N-methacryloyl-(L)-tyrosine methylester) [BR-MIP] cryogel was produced by free radical polymerization initiated by N,N,N',N'-tetramethylene diamine (TEMED) and ammonium persulfate (APS) pair in an ice bath. After that, the template molecules (i.e., bilirubin) were removed from the polymeric structure using sodium carbonate and sodium hydroxide. The maximum bilirubin adsorption amount was 3.6 mg/g polymer. The relative selectivity coefficients of the BR-MIP cryogel for bilirubin/cholesterol and bilirubin/testosterone mixtures were 7.3 and 3.2 times greater than non-imprinted poly(HEMA-MAT) [NIP] cryogel, respectively. The BR-MIP cryogel could be used many times without decreasing bilirubin adsorption amount significantly. Therefore, as a reusable carrier possessing high selectivity, BR-MIP cryogel has a potential candidate as a clinical hemoperfusion material. (C) 2008 Elsevier B.V. All rights reserved.
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| 5. |
- Baydemir, Goezde, et al.
(författare)
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Supermacroporous poly(hydroxyethyl methacrylate) based cryogel with embedded bilirubin imprinted particles
- 2009
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Ingår i: Reactive & Functional Polymers. - : Elsevier BV. - 1873-166X .- 1381-5148. ; 69:1, s. 36-42
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Tidskriftsartikel (refereegranskat)abstract
- Molecular imprinted polymers are artificial, template-made materials with the ability to recognize and to specifically bind the target molecule. The aim of this study is to prepare supermacroporous cryogel with embedded bilirubin-imprinted particles which can be used for the selective removal of bilirubin from human plasma. N-methacryloyl-(L)-tyrosinemethylester (MAT) was chosen as the pre-organization monomer. In the first step, bilirubin was complexed with MAT and the bilirubin-imprinted poly(hydroxyethyl methacrylate-N-methacryloly-(L)-tyrosine methyl-ester) [MIP] monolith was produced by bulk polymerization. MIP monolith was smashed and the particles ground and sieved through 100 pm sieves. In the second step. the supermacroporous poly(hydroxyethyl methacrylate) (PHEMA) cryogel with embedded MIP particles [PHEMA/MIP composite cryogel] was produced by free radical polymerization initiated by N.N,N',N'-tetramethylene diamine (TEMED) and ammonium persulfate (APS) pair in an ice bath. After that, the template (i.e., bilirubin) molecules were removed using sodium carbonate and sodium hydroxide. Compared with the PHEMA cryogel (0.2 mg/g polymer), the bilirubin adsorption capacity of the PHEMA/MIP composite cryogel (10.3 mg/g polymer) was improved significantly due to the embedded MIP particles into the polymeric matrix. The relative selectivity coefficients of PHEMA/MIP composite cryogel for bilirubin/cholesterol and bilirubin/testosterone were 8.6 and 4.1 times greater than the PHEMA cryogel, respectively. The PHEMA/MIP composite cryogel could be used many times without decreasing the bilirubin adsorption capacity significantly. (c) 2008 Elsevier Ltd. All rights reserved.
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| 6. |
- Bereli, Nilay, et al.
(författare)
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Protein recognition via ion-coordinated molecularly imprinted supermacroporous cryogels
- 2008
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1190:1-2, s. 18-26
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Tidskriftsartikel (refereegranskat)abstract
- Molecular imprinting is a method for making selective binding sites in synthetic polymers using a molecular template. The aim of this study is to prepare lysozyme-imprinted supermacroporous cryogels which can be used for the purification of lysozyme (Lyz) from egg white. N-Methacryloyl-(L)-histiclinemethylester (MAH) was chosen as the metal-coordinating monomer. In the first step, Cu2+ was complexed with MAH and the lysozyme-imprinted poly(HEMA-MAH) [Lyz-MIP] cryogel were produced by free radical polymerization initiated by N,N,N',N'-tetramethylene diamine (TEMED) in an ice bath. After that, the template (i.e., lysozyme) was removed using 0.05 M phosphate buffer containing 1 M NaCl (pH 8.0). The maximum lysozyme adsorption capacity was 22.9 mg/g polymer. The relative selectivity coefficients of Lyz-MIP cryogel for lysozyme/bovine serum albumin and lysozyme/cytochrome c were 4.6 and 3.2 times greater than non-imprinted poly(HEMA-MAH) (NIP) cryogel, respectively. Purification of lysozyme from egg white was also monitored by determining the lysozyme activity using Micrococcus lysodeikticus as substrate. The purity of the desorbed lysozyme was about 94% with recovery about 86%. The Lyz-MIP cryogel could be used many times without decreasing the adsorption capacity significantly.
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| 7. |
- Bloch, K, et al.
(författare)
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Functional activity of insulinoma cells (INS-1E) and pancreatic islets cultured in agarose cryogel sponges
- 2005
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Ingår i: Journal of Biomedical Materials Research. Part A. - : Wiley. - 1552-4965 .- 1549-3296. ; 75A:4, s. 802-809
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Tidskriftsartikel (refereegranskat)abstract
- Here, we describe the preparation, structure, and properties of cryogel sponges, which represent a new type of macroporous biomaterial for tissue engineering. Cryogels were produced through freeze-thawing techniques, either from agarose alone or from agarose with grafted gelatin. The aim of this study was to evaluate agarose cryogel sponges as scaffolds for Culturing both isolated pancreatic islets and insulinoma cells (INS-IE). In order to evaluate the effect of cell entrapment in artificial scaffolds, cell function reflected by insulin secretion and content was studied in cells cultivated for a 2-week period either in Culture plastic plates or in cryogel sponge disks. Our results show that tumor-derived INS-1E cells grown either on plastic or on cryogels do not differ in their proliferation, morphology, insulin release, and intracellular insulin content. However, isolated pancreatic islets cultivated on cryogels sponge show 15-fold higher basal insulin secretion at 3.0 mM glucose than islets cultivated on plastic plates and fail to respond to stimulation with 16.7 mM glucose. In addition, these islets have about 2-fold lower insulin content compared to those grown in plastic plates. It is possible that the cell dysfunction noted in these in vitro experiments is due to the effect of the limited oxygen supply to the islets cultivated in cryogel sponge. Further in vivo Studies are needed to clarify the nature of such an observation since according to previous reports, agarose and gelatin induce new vessel formation supporting enhanced oxygen supply. (c) 2005 Wiley Periodicals, Inc.
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| 8. |
- Boelgen, Nimet, et al.
(författare)
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Cryogelation for preparation of novel biodegradable tissue-engineering scaffolds
- 2007
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Ingår i: Journal of Biomaterials Science. Polymer Edition. - : Informa UK Limited. - 0920-5063 .- 1568-5624. ; 18:9, s. 1165-1179
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Tidskriftsartikel (refereegranskat)abstract
- 2-Hydroxyethyl methacrylate-L-lactate (HEMA-LLA) and HEMA-L-lactate-dextran (HEMA-LLA-D) were synthesized. H-1-NMR confirmed the formation of these oligomers and macromers. Cryogels with different pore structures were prepared using different amounts of HEMA, HEMA-LLA and HEMA-LLA-D by a cryogelation technique. SEM micrographs exhibited pore morphologies. Cryogels were highly porous with interconnected pore structures, opaque, spongy and highly elastic. It was possible to compress them to remove the water in the pores and to return to their original form just by immersing them in water in few minutes, which was quite reproducible. Their swelling abilities, compressive strengths and degradation in buffer solutions were found to be related with their structural properties which was controlled by changing the cryogelation recipe.
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| 9. |
- Bolgen, Nimet, et al.
(författare)
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Tissue responses to novel tissue engineering biodegradable cryogel scaffolds: An animal model
- 2009
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Ingår i: Journal of Biomedical Materials Research. Part A. - : Wiley. - 1552-4965 .- 1549-3296. ; 91A:1, s. 60-68
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Tidskriftsartikel (refereegranskat)abstract
- Biodegradable macroporous cryogels with highly open and interconnected pore structures were produced from dextran modified with oligo L-lactide bearing hydroxyethylmethacrylate (HEMA) end groups in moderately frozen solutions. Tissue responses to these novel scaffolds were evaluated in rats after dorsal subcutaneous implantation, iliac submuscular implantation, auricular implantation, or in calvarial defect model. In no case, either necrosis or foreign body reaction was observed during histological studies. The cryogel scaffolds integrated with the surrounding tissue and the formation of a new tissue were accompanied with significant ingrowth of connective tissue cells and new blood vessels into the cryogel. The tissue responses were significantly lower in auricular and calvarial implantations when compared with the subcutanous and the submuscular implantations. The degradation of the scaffold was slower in bone comparing to soft tissues. The biodegradable cryogels are highly biocompatible and combine extraordinary properties including having soft and elastic nature, open porous structure, and very rapid and controllable swelling. Therefore, the cryogels could be promising candidates for further clinical applications in tissue regeneration. (C) 2008 Wiley Periodicals, Inc. J Biomed Mater Res 91 A: 60-68, 2009
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| 10. |
- Dainiak, Maria, et al.
(författare)
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Affinity cryogel monoliths for screening for optimal separation conditions and chromatographic separation of cells
- 2006
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1123:2, s. 145-150
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Tidskriftsartikel (refereegranskat)abstract
- Suitable conditions for separating cells using a chromatographic procedure were evaluated in parallel chromatography on minicolumns. A 96-hole minicolumn plate filled with cryogel monoliths (18.8 mm x 7.1 mm O) with immobilized concanavalin A was used. Chromatographic columns (113 mm x 7.1 mm O) were used for chromatographic resolution of a mixture of Saccharomyces cerevisiae and Escherichia coli cells. Separation of a cell mixture containing equal amounts of cells of both types performed in a column format under the determined optimal conditions, resulted in a quantitative capture of applied S. cerevisiae cells, while E. coli passed through the column. Bound S. cerevisiae cells were released by flow-induced detachment and by compression of the adsorbent in the presence of 0.3 M methyl U-D-manno-pyranoside. The flowthrough and the eluted fractions were analyzed by plate counting and by registering metabolic activity of S. cerevisiae cells in the eluted fractions after capturing on ConA-cryogel monoliths in a 96-minicolumn plate format. The flowthrough fraction contained E. coli cells with nearly 100% purity, whereas the fraction eluted by compression of the adsorbent contained viable S. cerevisiae cells with 95% purity. Thus, an efficient chromatographic separation of cells was achieved using affinity cryogel column.
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