| 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. |
- Alvarez Fernandez, Marcia, et al.
(författare)
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Enhancement of sulphide production in anaerobic packed bed bench-scale biofilm reactors by sulphate reducing bacteria
- 2006
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Ingår i: Biotechnology Letters. - : Springer Science and Business Media LLC. - 1573-6776 .- 0141-5492. ; 28:3, s. 175-181
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Tidskriftsartikel (refereegranskat)abstract
- Two biofilm reactors, using pumice stone and Poraver as biofilm supports, were run, based on the optimization of sulphide production using a factorial design. The maximum H2S concentrations reached were 10 and 15 mm, respectively, both being appropriate for metal precipitation in effluents. The set-up of the pumice stone biofilm reactor is suitable for application in the mining area in the Bolivian Andean region, where this material is widely available. The use of specific primers for sulphate-reducing bacteria groups permits the identification of the sulphide-producing bacteria present in biofilms.
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| 3. |
- 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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| 4. |
- Andersson, Jonatan, et al.
(författare)
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Isolation of potato proteins using simulated moving bed technology.
- 2008
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Ingår i: Biotechnology and Bioengineering. - : Wiley. - 1097-0290 .- 0006-3592. ; 101, s. 1256-1263
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Tidskriftsartikel (refereegranskat)abstract
- The simulated moving bed (SMB) concept of chromatography was applied to treat potato juice from production of starch. The aim was to harvest proteins. SMB offers possibilities to operate with different process strategies and in this study it was shown possible to harvest up to 80% of the protein in a process utilizing very little extra water besides that already present in the juice. After depleting protein from the juice in the adsorption step, the flow through was used to recondition the column after elution. The present study illustrates a new concept of applying chromatography as a capturing step of bulk products. Biotechnol. Bioeng. (c) 2008 Wiley Periodicals, Inc.
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| 5. |
- Andersson, Jonatan, et al.
(författare)
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Simulated moving bed technology with a simplified approach for protein purification - Separation of lactoperoxidase and lactoferrin from whey protein concentrate
- 2006
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Ingår i: Journal of Chromatography A. - : Elsevier BV. - 0021-9673. ; 1107:1-2, s. 88-95
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Tidskriftsartikel (refereegranskat)abstract
- Simulated moving bed (SMB) technology is a continuous chromatographic technique proven to have many advantages compared to conventional batch chromatography, such as: raised productivity and product concentration, reduced buffer consumption as well as more efficient use of raw material. In this study a 20 column SMB process for the separation of lactoperoxidase and lactoferrin from whey protein concentrate (WPC) was developed. A simplified approach with data from a single column experiment was used when designing the process. The SMB process data were compared to a theoretical scale-up of the breakthrough experiment reflecting the same 20 column set-up run in non-moving bed mode. The outcome of the comparison is a 48% raise in productivity, a 4.3 times decrease in buffer consumption, 6.5 times raise in target protein concentration with a raw material utilization which is slightly better for the SMB process.
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| 6. |
- Bansal, Vibha, et al.
(författare)
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Recovery of urokinase from integrated mammalian cell culture cryogel bioreactor and purification of the enzyme using p-aminobenzamidine affinity chromatography
- 2006
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Ingår i: Journal of Molecular Recognition. - : Wiley. - 1099-1352 .- 0952-3499. ; 19:4, s. 332-339
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Tidskriftsartikel (refereegranskat)abstract
- An integrated product recovery system was developed to separate urokinase from the cell culture broth of human kidney cells HT1080. Supermacroporous monolithic cryogels provided ideal matrices with respect to surface and flow properties for use as cell culture scaffold as well as for affinity chromatographic capture step of the enzyme in the integrated system. The urokinase was produced continuously in the reactor running for 4 weeks with continuous circulation of 500 ml of culture medium. The enzyme activity in the culture medium reached to 280 Plough units (PU)/mg protein. Cu(II)-iminodiacetic acid (IDA)-polyacrylamide (pAAm) cryogel column was used to capture urokinase by integrating with the gelatin-coupled pAAm-cryogel bioreactor for HT1080 cell culture. After removing the urokinase capture column from the integrated system the bound protein was eluted. The metal affinity capture step gave 4.5-fold purification of the enzyme thus achieving a specific activity of 1300 PU/mg protein. The enzyme eluate from Cu(11)-IDA-pAAm cryogel capture column was further purified on benzamidine-Sepharose affinity column. This step finally led to a homogeneous preparation of different forms of urokinase in two different ellution peaks with a best urokinase activity of 13 550 PU/mg of protein. As compared to initial activity in the cell culture broth, about 26.2- and 48.4-fold increase in specific activity was achieved with enzyme yields corresponding to 32% and 35% in two different peak fractions, respectively. Native electrophoresis and SDS-PAGE showed multiple protein bands corresponding to different forms of the urokinase, which were confirmed by Western blotting and zymography. Copyright (c) 2006 John Wiley & Sons, Ltd.
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| 7. |
- Birgisson, Hakon, et al.
(författare)
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Immobilization of a recombinant Escherichia coli producing a thermostable alpha-L-rhamnosidase: Creation of a bioreactor for hydrolyses of naringin
- 2007
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Ingår i: Enzyme and Microbial Technology. - : Elsevier BV. - 0141-0229. ; 40:5, s. 1181-1187
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Tidskriftsartikel (refereegranskat)abstract
- An U-L-rhamnosidase (E.C. 3.2.1.40) from a newly discovered thermophilic bacterium was expressed in Escherichia coli BL21 DE3 pRIL cells. The cells were immobilized in Ca2+-alginate beads. The temperature of 50 degrees C used in reactions, appeared to be sufficient for making the mesophilic strain porous enough for the substrate to access the cloned thermostable enzyme. Pretreatment of cells with heat or lysozyme prior to bead formation did not improve the results. The best cell concentration (w/w) for bead preparation was found to be 0.0 192 g ml(-1) and stability of beads increased if CaCl2 concentration in buffers and substrate was kept at 50 mM. In a 60 min assay, the optimal pH of the entrapped cells was found to be 7.8 and the optimal temperature 60 degrees C. By packing the beads in a column, a bioreactor for production Of L-rhamnose from naringin was created. Full degradation of 7.9 mM naringin could be reached by running the reactor at 1 ml min(-1) at 50 degrees C. The optimal running temperature of the reactor was found to be 50 degrees C and the reactor was fully stable over 3 days at that temperature. On the fourth day, substrate degradation capacity had decreased by 10-15%. (c) 2006 Elsevier Inc. All rights reserved.
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| 8. |
- 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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| 9. |
- 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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| 10. |
- Bohn, Irene, et al.
(författare)
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Effect of temperature decrease on the microbial population and process performance of a mesophilic anaerobic bioreactor
- 2007
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Ingår i: Environmental Technology. - 1479-487X. ; 28:8, s. 943-952
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Tidskriftsartikel (refereegranskat)abstract
- The effect of a temperature decrease from 33 degrees C to 12 degrees C was investigated for anaerobic digestion of crop residues. A laboratory-scale reactor (R,) was inoculated with mesophilic sludge and operated as continuously stirred fed-batch system at temperatures of 12 degrees C, 18 degrees C and 33 degrees C. Changes in the microbial populations of the sludge were followed by means of fluorescence in situ hybridization analysis. Methane was produced in R, at all temperatures. Stable long-term operation at 18 degrees C was achieved yielding 151 mlCH(4) gVS(added)(-1) at a rate of 108 mlCH(4) l(R)(-1)d(-1) once the microbial populations of the sludge had adapted to this temperature. After operation at 18 degrees C, the contents of R-0, was mixed and distributed into three smaller reactors, which were operated at 18 degrees C (R-18), 25 degrees C (R-25) and 37 degrees C (R-37) respectively. Methane production rates for R-37 and R-25 were 366 and 310 mlCH(4) l(R)(-1)d(-1), respectively, which were higher than the 215 mlCH(4) l(R)(-1)d(-1) obtained in R-0 when this was operated at 33 degrees C. Hydrolysis was found to decrease when temperature was decreased and especially below 25 degrees C. At temperatures below 16 degrees C, acidogenesis and methanogenesis were the rate-limiting steps. Adaptation of the mesophilic sludge to 18 degrees C was indicated by an increase in the ratio of Bacteria to total prokaryotes (sum of Archaea and Bacteria). This was thought to be caused by enrichment of Bacteria in the sludge, which appeared to be an important adaptation mechanism. During the adaptation, the Methanomicrobiales and Methanosarcinaceae populations increased relative to the total Archaea population whereas the Methanosaeta population decreased. The population changes were reflected by reactor performance.
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