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  • 1
    Book
    Book
    Berlin [u.a.] : Springer
    Part of " Advances in biochemical engineering, biotechnology"
    Keywords: Biotechnologie ; Chromatographie ; Chromatographie ; Biochemical engineering ; Chromatographic analysis ; Chromatography Collected Works methods ; Polymers Collected Works chemical synthesis ; Chromatographie ; Génie biochimique ; Aufsatzsammlung ; Biotechnologie ; Chromatographie ; Chromatographie
    Type of Medium: Book
    Pages: VI, 271 S. , Ill., graph. Darst.
    ISBN: 3540430423
    Series Statement: Advances in biochemical engineering, biotechnology 76
    RVK:
    RVK:
    Language: English
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Bioseparation 9 (2000), S. 359-368 
    ISSN: 1573-8272
    Keywords: Bioseparation ; clear lysate ; CHO cell culture ; E. coli ; Macro-Prep
    Source: Springer Online Journal Archives 1860-2000
    Topics: Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The chromatographic properties of three types of ceramic hydroxyapatite (HAP) are compared. All three materials were prepared by sintering the original precipitate, albeit at different temperatures (400, 700 and 1000°C for type I, II and III HAP, respectively). The three materials differed in pore size and pressure limits (both lowest for type I and highest for type III). Type I and II HAP had an average particle diameter of 20 μm. The particle size of the type III material was 40 μm. HAP-beads were slurry-packed into 4 × 25-mm stainless steel columns and investigated for the chromatographic isolation of plasmid DNA from clarified E. coli lysates and of a recombinant human antibody from CHO cell culture supernatants respectively. The chromatographic performance of the three types of HAP showed significant differences, which were correlated to the binding capacities of the materials for (linearized) plasmids of different size (4.7, 10.3 and 11.4 kb) and proteins of different isoelectric point (lysozyme, pI = 10.5; anti RhD antibody, pI = 8.3; β-lactoglobulin, pI = 4.9). The accessibility of the adsorptive surface (pore size) but also the types of binding sites on the HAP-surface (P/C-site ratio) are proposed as determining factors for the chromatographic behavior.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    ISSN: 0935-6304
    Keywords: Monolith ; continuous bed column ; convective interaction media ; membrane chromatography ; membrane adsorber ; HPMDC ; Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Monolithic stationary phases have revolutionized protein chromatography because they combine speed, capacity, and resolution in a unique manner. Since such stationary phases contain no particles but only flow-through pores, the usual mass transfer restrictions to the chromatography of large molecules are not observed and extremely fast separations become possible. Recently the area of application of monolith chromatography has been extended to the separation and analysis of small molecules and plasmid DNA. This review summarizes the state of art in high performance monolith and especially high performance monolithic disk chromatography (HPMDC). The current understanding of the theory of protein HPMDC is summarized, while an introduction to the evolving field of small molecule HPMDC is attempted. The basic differences between the monolithic disks and columns packed with conventional stationary phases (including perfusion and micropellicular particles) but also monolithic columns (porous rods) are outlined. Finally, the potential of HPMDC to analytical and preparative biochromatography is demonstrated by a discussion of recent applications of chromatographic disks for protein isolation and bioprocess analysis.
    Additional Material: 14 Ill.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Cytotechnology 21 (1996), S. 205-215 
    ISSN: 1573-0778
    Keywords: antithrombin III ; mammalian cell culture ; continuous culture ; capillary electrophoresis ; product monitoring ; recombinant protein ; temperature influence
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The importance of mammalian cell cultures for biotechnological production processes is steadily increasing, despite the high demands of these organisms on their culture conditions. Efforts towards a more efficient bioprocess generally concentrate on maximizing the culture's life time, the cell number, and the product concentration. Here recombinant BHK 21 c13 cells are used to produce rh-AT III, an anticoagulant of high therapeutic value. The influence of the process mode (batch, repeated batch, continuous perfusion) and the process temperature (30°C vs. 37°C) on the above mentioned parameters is investigated. It is possible to increase the length of the culture from 140 h (batch) to more than 500 h (continuous perfusion culture), while concomitantly increasing the cell density from 0.72 106/ml (batch) to 2.27 106/ml (repeated batch) and 2.87 106/ml (continuous perfusion culture). The accumulation of toxic metabolites, such as lactate, can be curtailed by reducing the bioreactor temperature from 37°C to 30°C during the later part of the exponential growth phase. Fast and reliable product monitoring became essential during process optimization. Capillary zone electrophoresis (CZE) in uncoated fused silica capillaries was studied for that purpose and compared to the standard ELISA. Under optimized conditions an AT III quantification could be done within 2 min with CZE. The detection limit was 5 μg/ml. A relative standard deviation of less than 0.9% was calculated. The detection limit could be lowered by one order of magnitude by using a two dimensional system, where an liquid chromatographic (LC) system is coupled to the CZE. Concomitantly the resolution is improved. The two-dimensional analysis required 5 min. Membrane adsorbers (MA) were used as stationary phase in the LC-system, to allow the application of high flow rates (5–10 ml/min). The correlation between the LC-CZE analysis and the standard AT III-ELISA was excellent, with r2: 0.965. Using the assay for at line product monitoring, it is shown, that the process temperature is of no consequence for the productivity whereas the process mode strongly influences this parameter.
    Type of Medium: Electronic Resource
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Cytotechnology 30 (1999), S. 159-168 
    ISSN: 1573-0778
    Keywords: continuous bed column ; displacement chromatography ; lipopolysaccharide ; plasmid DNA ; SMA model
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract The preparation of plasmid DNA at large scale constitutes a pressing problem in bioseparation. This paper describes a first investigation of displacement chromatography as a means to separate plasmid DNA (4.7 kb) from E. coli lipopolysaccharides and protein (holo transferrin), respectively. Displacement chromatography has advantages in this regard, since the substance mixture is resolved into rectangular zones of the individual components rather than into peaks. Thus a higher total concentration can be maintained in the pooled product fractions. Hydroxyapatite (type I and II) and anion exchange stationary phases were included in the experiments. In addition to a conventional anion exchange column packed with porous particles, the recently introduced continuous bed UNOTM anion exchange column was investigated. No DNA purification was possible with either hydroxyapatite material. Conventional particle based columns in general were not suited to the separation of any two substances varying considerably in molecular mass, e.g. plasmid DNA and standard protein. Presumably, the direct competition for the binding sites, which is essential in displacement chromatography, was restricted by the size dependency of the accessible stationary phase surface area in this case. Better results were obtained with the continuous bed column, in which the adsorptive surface coincides with the walls of the flow through pores. As a result the accessible surface does not vary as much with the size of the interacting molecules as for the conventional stationary phase materials. Sharper transitions were also observed between substance zones recovered from the UNOTM column. The steric mass action model was used to aid method development in case of the anion exchange approach. While further research in obviously necessary, displacement chromatography on continuous bed columns has been shown to be capable of separating plasmid DNA from typical impurities.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Cytotechnology 6 (1991), S. 55-63 
    ISSN: 1573-0778
    Keywords: flow injection analysis ; high cell density ; hybridoma cells ; on-line monitoring of monoclonal antibody formation ; on-line glucose and lactate measurements ; perfusion cultivation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract An automated flow injection system for on-line analysis of proteins in real fermentation fluids was developed by combining the principles of stopped-flow, merging zones flow injection analysis (FIA) with antigen-antibody reactions. IgG in the sample reacted with its corresponding antibody (a-IgG) in the reagent solution. Formation of insoluble immunocomplexes resulted in an increase of the turbidity which was determined photometrically. This system was used to monitor monoclonal antibody production in high cell density perfusion culture of hybridoma cells. Perfusion was performed with a newly developed static filtration unit equipped with hydrophilic microporous tubular membranes. Different sampling devices were tested to obtain a cell-free sample stream for on-line product anlysis of high molecular weight (e.g., monoclonal antibodies) and low molecular weight (e.g., glucose, lactate) medium components. In fermentation fluids a good correlation (coefficient: 0.996) between the FIA method and an ELISA test was demonstrated. In a high density perfusion cultivation process mAb formation was succesfully monitored on-line over a period of 400 h using a reliable sampling system. Glucose and lactate were measured over the same period of time using a commercially available automatic analyser based on immobilized enzyme technology.
    Type of Medium: Electronic Resource
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  • 7
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    Electrophoresis 14 (1993), S. 753-758 
    ISSN: 0173-0835
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Plasmid DNA prepared from cultivation samples of recombinant Escherichia coli was analyzed by capillary gel electrophoresis. We used this method to control the genetic stability during a fed-batch culture. The plasmid DNA and a standard DNA mixture with molecular weights in the size range of 3000-22 000 base pairs (bp) were analyzed in capillaries that were filled with solutions of non-cross-linked polyacrylamide. With this method the plasmid DNA from recombinant E. coli cultivation samples was analyzed within 30 min. Separation parameters such as gel concentration, capillary length and current were optimized for that purpose. The method was modified to allow the analysis of proteins. Crude cell lysate samples could be screened for the protein pattern within 15 min in sodium dodecyl sulfate-polyacrylamide filled capillaries. The method was also used to verify product purity after the down-stream process.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    Electrophoresis 18 (1997), S. 1899-1905 
    ISSN: 0173-0835
    Keywords: Endotoxin ; Lipopolysaccharide ; Pyrogen ; Gram-negative bacteria ; Indirect UV detection ; Capillary electrophoresis ; Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Chemistry and Pharmacology
    Notes: Endotoxins are part of the outer membrane of gram-negative bacteria such as E. coli. Upon entering the blood stream, they cause a violent, sometimes life-threatening, response of the immune system. Endotoxins are lipopolysaccharides (LPS), lacking optically active groups, and their detection in the underivatized state can be difficult. In this paper the potential of capillary electrophoresis (CE) for LPS analysis is investigated. By using a standard phosphate buffer method, concentrations down to 100 μg/mL can be detected within 6 min. The detection limit can be lowered by one order of magnitude by using a sodium dodecyl sulfate (SDS)/borate buffer, pH 9.2. In this buffer, the SDS serves to homogenize the size of the LPS aggregates, while the borate forms complexes with the diol groups of the molecule, thereby enhancing its optical activity. The formation of LPS-affinity complexes with the UV-active polymyxin B or labeling of the LPS with a fluorophore (fluorescein isothiocyanate) was unsuccessful. Best results, in terms of detection limit and speed, were obtained with an indirect UV-detection CE method. By using a strongly UV-active electrophoresis buffer, endotoxins could be detected as “negative” peaks. In this case, a detection limit of 3 μg/mL (35 pM) was determined. Proteins and other UV-active substances did not disturb the assay, since they generated no detectable signals. The indirect UV detection was used to quantify the residual LPS content of a DNA preparation from E. coli.
    Additional Material: 9 Ill.
    Type of Medium: Electronic Resource
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  • 9
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    Chemie Ingenieur Technik - CIT 66 (1994), S. 1585-1592 
    ISSN: 0009-286X
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Integrated Biotechnological Production Processes. Manufacturing processes for ethanol and butanol are investigated with respect to reduction of product inhibition by integration with reversed osmosis, pervaporation, extraction, adsorption, flash evaporation, vacuum evaporation, and gas stripping. The integration of lactic acid formation with precipitation, extraction, electrodialysis, ion excange, and crystallization are considered. Adsorption, extraction and crystallization are examined as possible integrated recovery processes for antibiotics production. The possible future integrated use of adsorption, chromatography, membrane chromatography, extraction and flotation for protein production processes is discussed.
    Additional Material: 1 Tab.
    Type of Medium: Electronic Resource
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  • 10
    Electronic Resource
    Electronic Resource
    Bognor Regis [u.a.] : Wiley-Blackwell
    ISSN: 0887-624X
    Keywords: group transfer polymerization ; poly-N,N-diethylacrylamide ; thermosensitive water-solubility ; MALDI-MS ; affinity precipitation ; Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The conditions for preparing poly-N,N-diethylacrylamide by group transfer polymerization (GTP) were investigated. While electrophiles did not catalyze the reaction, various nucleophilic substances could be used for that purpose. By using an appropriate initiator, either an ester or a carboxylic acid end group could be formed. The highest yields in the first case were obtained using tetrabutylammonium acetate and dimethylketene methyl trimethylsilyl acetal as catalyst and initiator, respectively, while the use of the corresponding bistrimethylsilyl compound as initiator gave polymers, albeit at lower yields, which carried the acidic end group. The 1H-NMR, 13C-NMR, and IR spectra of the polymers were taken and used together with information obtained with soft-ionization mass spectrometric methods (MALDI-MS, ESI-MS, and FD-MS) to elucidate molecule structure, apparent molecular weight distribution, polydispersity, and possible mechanisms of the termination reaction. The poly-N,N-diethylacrylamide prepared, showed an inverse temperature dependency in its water-solubility, with a lower critical solution temperature between 29.8°C and 30°C. © 1994 John Wiley & Sons, Inc.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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