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101.
Schriebl K Trummer E Weik R Lattenmayer C Müller D Kunert R Katinger H Vorauer-Uhl K 《Electrophoresis》2007,28(12):2100-2107
For biotechnological research, development, and production various analytical methods are required to determine the quality of the target product. In this context, the determination of isoforms is state-of-the-art; however, the majority of applied techniques are more qualitative than quantitative. To address this fact, we evaluated different post- and pre-electrophoretic staining dyes for their applicability on linear IPG gels using recombinant human erythropoietin as a model protein. Each evaluated dyes was able to detect all isoforms reproducibly, but CyDyes were found to be the most promising. Using CyDyes, up to three samples can be focused on the same lane under identical electrophoretic conditions, thus, a fast, reproducible, sensitive and quantitative isoform determination can be performed. To illustrate the practical relevance, quantitative CyDye technique was used for the characterization of our model protein, recombinant human Epo-Fc. This method makes it possible to determine the isoform pattern of nonpurified supernatants as well as purified proteins. We conclude that quantitative pre-electrophoretic staining IEF using CyDyes is a fast, simple, accurate method to determine isoforms, which can be used in research, development, and manufacturing for product quality analysis, e.g., clone screening, process optimization, and purification monitoring. 相似文献
102.
Musyanovych A Rossmanith R Tontsch C Landfester K 《Langmuir : the ACS journal of surfaces and colloids》2007,23(10):5367-5376
Carboxyl and amino-functionalized polystyrene latex particles were synthesized by the miniemulsion copolymerization of styrene and acrylic acid or 2-aminoethyl methacrylate hydrochloride (AEMH). The reaction was started by using an oil-soluble initiator, such as 2,2'-azobis(2-methylbutyronitrile) (V-59). The effect of the functional monomer content and type of surfactant (non-ionic versus ionic) on the particle size and particle size distribution was investigated by dynamic light scattering (DLS) and transmission electron microscopy (TEM). A bimodal particle size distribution was observed for functionalized latex particles prepared in the presence of the non-ionic surfactant (i.e., Lutensol AT-50) when 1 wt % of acrylic acid or 3 wt % of AEMH as a comonomer was employed. The copolymer particle nucleation was studied by using a highly hydrophobic fluorescent dye. From the obtained results, the formation of bimodal particle size distribution may be attributed to a budding-like effect, which takes place during the earlier stage of polymerization and is caused by the additional stabilizing energy originated from the ionic groups of a functional polymer. The reaction mechanism of particle formation in the presence of non-ionic and ionic surfactants has been proposed. The amount of the surface functional groups was determined from polyelectrolyte titration data. 相似文献
103.
Schmelzer CE Schöps R Reynell L Ulbrich-Hofmann R Neubert RH Raith K 《Journal of chromatography. A》2007,1166(1-2):108-115
Numerous peptides obtained by enzymatic digestion of food proteins have been reported to exhibit biological activities. In this study, the focus was placed on peptides of beta-casein from bovine milk after a gastro-analogous in vitro digestion with pepsin, a protease with broad specificity. In order to study the time course of the digestion, the process was stopped after specific times and the samples were subjected to HPLC separation followed by matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) and nanoelectrospray (nanoESI) quadrupole time-of-flight (qTOF) mass spectrometry. A combined sequencing approach using de novo interpretation and databases was employed. Overall, 100% of the beta-casein sequence was covered by identifying 125 peptides of 4-84 residues in length, including 3 phosphorylated species. The results show that the peptic hydrolysis starts at the C-terminus of the protein. The release of known bioactive peptides from beta-casein following the peptic digestion under simulated gastric conditions is unlikely with a few exceptions. Furthermore, an amino acid variation was found, providing evidence for the existence of an additional genetic variant of beta-casein. 相似文献
104.
In this paper, novel hollow polyelectrolyte multilayer tubes from poly(diallyldimethylammonium chloride) (PDADMAC), poly(styrene sulfonate) (PSS), and poly(allylamine hydrochloride) (PAH) were prepared: Readily available glass fiber templates are coated with polyelectrolytes using the layer-by-layer technique, followed by subsequent fiber dissolution. Depending on the composition of the polymeric multilayer, stable hollow tubes or tubes showing a pearling instability are observed. This instability corresponds to the Rayleigh instability and is a consequence of an increased mobility of the polyelectrolyte chains within the multilayer. The well-defined stable tubes were characterized with fluorescence microscopy, confocal laser scanning microscopy, and atomic force microscopy (AFM). The tubes were found to be remarkably free of defects, which results in an impermeable tube wall for even low molecular weight molecules. The mechanical properties of the tubes were determined with AFM force spectroscopy in water, and because continuum mechanical models apply, the Young's modulus of the wall material was determined. Additionally, scaling relations for the dependency of tube stiffness on diameter and wall thickness were validated. Because both parameters can be experimentally controlled by our approach, the deformability of the tubes can be varied over a broad range and adjusted for the particular needs. 相似文献
105.
We present simulations of the mechanism of secondary nucleation of polymer crystallization, based on a new model accounting for the microscopic kinetics of attaching and detaching. As the key feature of the model, we introduced multibody-interaction parameters that establish correlations between the attaching and detaching rate constants and the resulting thickness and width of the crystalline lamella. Using MATLAB and Monte Carlomethod, we followed the evolution of the secondary nuclei as a function of various multibody-interaction parameters. We identified three different growth progressions of the crystal:(i) Widening, (ii) thickening and (iii) simultaneously thickening and widening of lamellar crystals, controlled by the corresponding kinetic parameters. 相似文献
106.
Arnold M Bittermann H Kalbfuss-Zimmermann B Neumann T Schmidt K Sekul R Hilbrig F Ludolph H Freitag R 《Journal of chromatography. A》2011,1218(29):4649-4659
Libraries of small molecules were searched for Fc-fragment selective binders to a recombinant human antibody ("MDJ8″, IgG(1)-subtype, κ-light chain) via SPR-based screening of chemical microarrays. Identified hit structures were immobilised on NHS-activated Sepharose for the determination of MDJ8 binding and selectivity versus typical proteineous impurities represented by the spend cell culture supernatant. Columns were packed and the most promising ligands further characterized in terms of binding constants, binding kinetics, as well as dynamic and equilibrium binding capacities. The performance of the best ligand, 2A10, was compared to standard Protein A chromatography. Using ligand 2A10 antibody capture from unprocessed cell culture supernatants was possible at similar recovery yield (>90%), purity (>80%), and eluting concentration (approximately 1 g/L) as with Protein A. Affinity constants (K(d)) of 2A10 were an order of magnitude higher than for the Protein A material, but still in the nM-range, while maximum binding capacities and binding kinetics were in the same order of magnitude. Ligand 2A10 was also able to capture a murine monoclonal antibody, again with similar efficiency as Protein A, as well as a number of humanised therapeutic antibodies. Antibody elution from the 2A10 column was possible using the Protein A standard protocol, i.e. 100mM glycine HCl pH 3.0, but also at near physiological pH, when some organic solvent or modifiers were present. Ligand 2A10 thus constitutes a cheaper, more robust alternative to Protein A as possible generic antibody binder. Moreover, the outlined approach to ligand selection could in principle by used to create suitable affinity ligands for other high value biotech products. 相似文献
107.
Alaa Fahmy Renate Mix Andreas Sch?nhals J?rg Friedrich 《Plasma Chemistry and Plasma Processing》2012,32(4):767-780
Polystyrene (PS) spin coated thin films were modified by O2 and Ar plasma as well as by UV irradiation treatments. The modified PS samples were compared with plasma polymerized and commercial polystyrene. The effects of plasma (O2 and Ar) and UV irradiation treatments on the surface and the bulk properties of the polymer layers were discussed. The surface properties were evaluated by X-ray Photoelectron Spectroscopy and Contact angle measurements and the bulk properties were investigated by FTIR and dielectric relaxation spectroscopy. As a result only one second treatment time was sufficient to modify the surface. However, this study was also dedicated to understand the effect of plasma and plasma irradiation on the deposited layers of plasma polymers. The dielectric measurements showed that the plasma deposited films were not thermally stable and underwent an undesired post-plasma chemical oxidation. 相似文献
108.
Abstract In this article the numerical approximation of solutions of Itô stochastic delay differential equations is considered. We construct stochastic linear multi-step Maruyama methods and develop the fundamental numerical analysis concerning their 𝕃 p -consistency, numerical 𝕃 p -stability and 𝕃 p -convergence. For the special case of two-step Maruyama schemes we derive conditions guaranteeing their mean-square consistency. 相似文献
109.
The paper consists of two parts. In the first part of the paper, we proposed a procedure to estimate local errors of low order
methods applied to solve initial value problems in ordinary differential equations (ODEs) and index-1 differential-algebraic
equations (DAEs). Based on the idea of Defect Correction we developed local error estimates for the case when the problem
data is only moderately smooth, which is typically the case in stochastic differential equations. In this second part, we
will consider the estimation of local errors in context of mean-square convergent methods for stochastic differential equations
(SDEs) with small noise and index-1 stochastic differential-algebraic equations (SDAEs). Numerical experiments illustrate
the performance of the mesh adaptation based on the local error estimation developed in this paper.
The first author acknowledges support by the BMBF-project 03RONAVN and the second author support by the Austrian Science Fund
Project P17253. 相似文献
110.
Hoffmann K Mix R Friedrich JF Buschmann HJ Resch-Genger U 《Journal of fluorescence》2009,19(2):229-237
Polypropylene supports were functionalized by plasma-deposition of polymeric allylamine layers. The surface amino groups generated
were wet-chemically reacted with xanthene dyes resulting in fluorescent polymer films. The effect of polymer-attachment of
the dyes on their emission features was studied fluorometrically and different methods were tested to improve the fluorescence
properties of the films. Modification with cucurbit[6]uril (CB6) yields a moderately enhanced fluorescence as well as an improved
photostability. The observed effect is most likely due to CB6-induced rigidization of the linker molecules which seems to
reduce fluorescence quenching dye–dye and fluorophore–surface interactions. 相似文献