bluuebunny/crossref_metadata_2025_split huggingface dataset:
Zur Querkraftgefährdung bestehender Spannbetonbrücken
Authors: Hegger Josef , Karakas Alexander , Pelke Eberhard , Schölch Udo 
Published: June 2010

AbstractZur Formulierung einer Handlungsanweisung wurden in Teil I die geänderten Lastannahmen für Brücken nach DIN Fachbericht 101 mit den Belastungsklassen nach alten Normen verglichen und gezeigt, wie anhand aktueller Verkehrszahlen die Lastansätze modifiziert werden können. Der aktuelle Brückenbestand in Hessen wurde mit der Straßeninformationsbank (Teilprojekt Bauwerksdaten) ausgewertet, um Klassifizierungs‐ und Bewertungskriterien zu erarbeiten. Zur Beurteilung des Querkraftwiderstandes wurden die Grundlagen der Querkraftbemessung nach DIN 1045, DIN 4227, DIN Fachbericht 102 und aus der Literatur gegenübergestellt.In Teil II werden Verfahren zur nachträglichen Verstärkung von Querschnitten und Tragsystemen mit zu geringer Querkrafttragfähigkeit zusammengestellt. Des Weiteren werden verschiedene Varianten der Modellierung verglichen, die bei der linearelastischen Schnittgrößenermittlung nach der Finite‐Elemente‐ Methode verwendet werden. Auf Grundlage der gewonnenen Erkenntnisse werden schließlich Empfehlungen für eine Handlungsanweisung zur Beurteilung querkraftgefährdeter Brückenbauwerke formuliert.Shear‐Vulnerability of Existing Pre‐Stressed Concrete BridgesIn Part I, to formulate an operation directive the revised load assumptions for bridges of DIN Technical Report 101 are compared with the loading classes according to old standards and it is demonstrated, how the loading models can be modified based on actual traffic data. The current bridge asset in the Federal State of Hesse is evaluated using the “Road Information Database (Sub‐Project: Structural Data)” in order to work out classification and evaluation criteria. Regarding the assessment of the shear load‐bearing capacity the fundamentals of the shear load design models of DIN 1045, DIN 4227, DIN TR 102 and from the literature are compared. In Part II, the procedures for the subsequent strengthening of cross‐sections and structural systems with a low shear load‐bearing capacity are compiled. In addition, different types of finite element modelling are compared used for the calculation of the linear‐elastic internal forces. Finally, based on the findings recommendations are formulated for an operation directive in regard to the assessment of shear‐vulnerable bridges.
Mouse brain full‐length cDNA library construction by negative selection of intact mRNAs
Authors: Wu Ning , Wu Huijuan , Li Yandong , Matand Kanyand 
Published: June 2010

AbstractSynthesis of full‐length cDNA libraries is an essential step for the study of gene function. The method for selecting the intact mRNA directly affects the number of full‐length transcripts. We have developed a novel method for intact mRNA selection based on the elimination of uncapped mRNAs. A negative‐selection strategy that removes both uncapped mRNA and other non‐mRNA molecules that present a phosphate at the 5'‐end has been applied in the mRNA purification procedures. Briefly, after performing a standard mRNA purification, a biotinylated oligoribonucleotide is ligated to the 5‐end phosphate of uncapped mRNAs. Streptavidin extraction is then performed to remove truncated and non‐mRNAs from the intact mRNAs. By comparing random sequencing results of mouse brain full‐length and standard cDNA libraries, there was a significant increase of full‐length clones with the modified procedure. The results showed that the full‐length library contained more than 68% full‐length clones with the 5'‐end positions ranging between –485 to +100 compared to the standard library with 33% of full‐length clones and 5'‐end positions ranging between –233 to +100. The data were analyzed using the t‐test with the significance level set at p<0.05. The statistical results showed that there were significant differences between two libraries in both 5'‐end position and mRNA size (p<0.05).
A small bispecific protein selected for orthogonal affinity purification
Authors: Alm Tove , Yderland Louise , Nilvebrant Johan , Halldin Anneli , Hober Sophia 
Published: June 2010

AbstractA novel protein domain with dual affinity has been created by randomization and selection. The small alkali‐stabilized albumin‐binding domain (ABD*), used as scaffold to construct the library, has affinity to human serum albumin (HSA) and is constituted of 46 amino acids of which 11 were randomized. To achieve a dual binder, the binding site of the inherent HSA affinity was untouched and the randomization was made on the opposite side of the molecule. Despite its small size and randomization of almost a quarter of its amino acids, a bifunctional molecule, ABDz1, with ability to bind to both HSA and the Z2 domain/protein A was successfully selected using phage display. Moreover, the newly selected variant showed improved affinity for HSA compared to the parental molecule. This novel protein domain has been characterized regarding secondary structure and affinity to the two different ligands. The possibility for affinity purification on two different matrices has been investigated using the two ligands, the HSA matrix and the protein A‐based, MabSelect SuRe matrix, and the new protein domain was purified to homogeneity. Furthermore, gene fusions between the new domain and three different target proteins with different characteristics were made. To take advantage of both affinities, a purification strategy referred to as orthogonal affinity purification using two different matrices was created. Successful purification of all three versions was efficiently carried out using this strategy.
Cellulose hydrolysis in evolving substrate morphologies III: Time‐scale analysis
Authors: Zhou Wen , Xu Ying , Schüttler Heinz‐Bernd 
Published: June 2010

AbstractWe present a time‐scale analysis for the enzymatic hydrolysis of solid cellulosic substrates, based on our recently developed kinetic model (Zhou et al., 2009a, Biotechnol Bioeng 104:261–274; Zhou et al., 2009b, Biotechnol Bioeng 104:275–289) which incorporates both enzymatic chain fragmentation and hydrolytic time evolution of the solid substrate morphology. Analytical order‐of‐magnitude estimates of the relevant single‐layer chain depolymerization times are first discussed. These time‐scale estimates for pure and mixed enzyme systems can be employed to calculate the degree of synergy between endo‐ and exo‐acting enzymes in a mixed enzyme system. By the way of a quasi‐steady‐state approximation which allows for a greatly simplified analytical solution of the model, we also explain the origin and give order‐of‐magnitude estimates of the two characteristic hydrolysis time scales which arise in this model when the solid substrate morphology is taken into account. These analytically derived time‐scale relations explain how the embedding of cellulose chains in a solid substrate acts as a crucial rate‐limiting factor and results in a substantial slowing down of the hydrolytic conversion process, compared to a hypothetical substrate of immediately enzyme‐accessible, isolated chains. The analytical time‐scale results are verified by numerical simulations and compared to experimental observations. Biotechnol. Bioeng. 2010;107: 224–234. © 2010 Wiley Periodicals, Inc.
Measurement of key metabolic enzyme activities in mammalian cells using rapid and sensitive microplate‐based assays
Authors: Janke R. , Genzel Y. , Wahl A. , Reichl U. 
Published: June 2010

AbstractSensitive microplate‐based assays to determine low levels of key enzyme activities in mammalian cells are presented. The enzyme platform consists of four cycling assays to measure the activity of 28 enzymes involved in central carbon and glutamine metabolism. The sensitivity limit of all cycling assays was between 0.025 and 0.4 nmol product. For the detection of glutaminase activity, a new glutamate cycle system involving the enzymes glutamate dehydrogenase and aspartate transaminase was established. The relative standard deviation of the method was found to be 1.7% with a limit of detection of 8.2 pmol and a limit of quantitation of 24.8 pmol. Hence, cell extracts could be highly diluted to reduce interferences caused by other components in the extract, which in addition minimized underestimates or overestimates of actual enzyme activities. Since substrate concentrations could be maintained at a nearly constant level throughout the assay product accumulation during the reaction was low, which minimized product inhibition. As an example, the enzyme platform was used to investigate maximum enzyme activities of stationary‐phase MDCK cells grown in serum‐containing GMEM medium as typically used in influenza vaccine production. Biotechnol. Bioeng. 2010;107: 566–581. © 2010 Wiley Periodicals, Inc.
Construction of cellobiose phosphorylase variants with broadened acceptor specificity towards anomerically substituted glucosides
Authors: De Groeve Manu R.M. , Remmery Laurens , Van Hoorebeke Annelies , Stout Jan , Desmet Tom , Savvides Savvas N. , Soetaert Wim 
Published: June 2010

AbstractThe general application of glycoside phosphorylases such as cellobiose phosphorylase (CP) for glycoside synthesis is hindered by their relatively narrow substrate specificity. We have previously reported on the creation of Cellulomonas uda CP enzyme variants with either modified donor or acceptor specificity. Remarkably, in this study it was found that the donor mutant also displays broadened acceptor specificity towards several β‐glucosides. Triple mutants containing donor (T508I/N667A) as well as acceptor mutations (E649C or E649G) also display a broader acceptor specificity than any of the parent enzymes. Moreover, further broadening of the acceptor specificity has been achieved by site‐saturation mutagenesis of residues near the active site entrance. The best enzyme variant contains the additional N156D and N163D mutations and is active towards various alkyl β‐glucosides, methyl α‐glucoside and cellobiose. In comparison with the wild‐type C. uda CP enzyme, which cannot accept anomerically substituted glucosides at all, the obtained increase in substrate specificity is significant. The described CP enzyme variants should be useful for the synthesis of cellobiosides and other glycosides with prebiotic and pharmaceutical properties. Biotechnol. Bioeng. 2010;107: 413–420. © 2010 Wiley Periodicals, Inc.
A metal‐repressed promoter from gram‐positive Bacillus subtilis is highly active and metal‐induced in gram‐negative Cupriavidus metallidurans
Authors: Ribeiro‐dos‐Santos Gabriela , Biondo Ronaldo , Quadros Oeber de Freitas , Vicente Elisabete José , Schenberg Ana Clara Guerrini 
Published: June 2010

AbstractA synthetic version of the metal‐regulated gene A (mrgA) promoter from Bacillus subtilis, which in this Gram‐positive bacterium is negatively regulated by manganese, iron, cobalt, or copper turned out to promote high level of basal gene expression that is further enhanced by Co(II), Cd(II), Mn(II), Zn(II), Cu(II), or Ni(II), when cloned in the Gram‐negative bacterium Cupriavidus metallidurans. Promoter activity was monitored by expression of the reporter gene coding for the enhanced green fluorescent protein (EGFP), and cellular intensity fluorescence was quantified by flow cytometry. Expression levels in C. metallidurans driven by the heterologous promoter, here called pan, ranged from 20‐ to 53‐fold the expression level driven by the Escherichia coli lac promoter (which is constitutively expressed in C. metallidurans), whether in the absence or presence of metal ions, respectively. The pan promoter did also function in E. coli in a constitutive pattern, regardless of the presence of Mn(II) or Fe(II). In conclusion, the pan promoter proved to be a powerful tool to express heterologous proteins in Gram‐negative bacteria, especially in C. metallidurans grown upon high levels of toxic metals, with potential applications in bioremediation. Biotechnol. Bioeng. 2010;107: 469–477. © 2010 Wiley Periodicals, Inc.
Modeling the competition between aggregation and self‐assembly during virus‐like particle processing
Authors: Ding Yong , Chuan Yap Pang , He Lizhong , Middelberg Anton P.J. 
Published: June 2010

AbstractUnderstanding and controlling aggregation is an essential aspect in the development of pharmaceutical proteins to improve product yield, potency and quality consistency. Even a minute quantity of aggregates may be reactogenic and can render the final product unusable. Self‐assembly processing of virus‐like particles (VLPs) is an efficient method to quicken the delivery of safe and efficacious vaccines to the market at low cost. VLP production, as with the manufacture of many biotherapeutics, is susceptible to aggregation, which may be minimized through the use of accurate and practical mathematical models. However, existing models for virus assembly are idealized, and do not predict the non‐native aggregation behavior of self‐assembling viral subunits in a tractable nor useful way. Here we present a mechanistic mathematical model describing VLP self‐assembly that accounts for partitioning of reactive subunits between the correct and aggregation pathways. Our results show that unproductive aggregation causes up to 38% product loss by competing favorably with the productive nucleation of self‐assembling subunits, therefore limiting the availability of nuclei for subsequent capsid growth. The protein subunit aggregation reaction exhibits an apparent second‐order concentration dependence, suggesting a dimerization‐controlled agglomeration pathway. Despite the plethora of possible assembly intermediates and aggregation pathways, protein aggregation behavior may be predicted by a relatively simple yet realistic model. More importantly, we have shown that our bioengineering model is amenable to different reactor formats, thus opening the way to rational scale‐up strategies for products that comprise biomolecular assemblies. Biotechnol. Bioeng. 2010;107: 550–560. © 2010 Wiley Periodicals, Inc.
Sialylation enhancement of CTLA4‐Ig fusion protein in Chinese hamster ovary cells by dexamethasone
Authors: Jing Ying , Qian Yueming , Li Zheng Jian 
Published: June 2010

AbstractThe importance of glycoprotein sialic acid levels is well known, as increased levels have been shown to increase in vivo serum half‐life profiles. Here we demonstrate for the first time that dexamethasone (DEX) was capable of improving the sialylation of a CTLA4‐Ig fusion protein produced by Chinese hamster ovary (CHO) cells. DEX was shown to enhance the intracellular addition of sialic acid by sialyltransferases as well as reduce extracellular removal of sialic acid by sialidase cleavage. We illustrated that DEX addition resulted in increased expression of the glycosyltransferases α2,3‐sialyltransferase (α2,3‐ST) and β1,4‐galactosyltransferase (β1,4‐GT) in CHO cells. Based upon our previous results showing DEX addition increased culture cell viability, we confirmed here that cultures treated with DEX also resulted in decreased sialidase activity. Addition of the glucocorticoid receptor (GR) antagonist mifepristone (RU‐486) was capable of blocking the increase in sialylation by DEX which further supports that DEX affected sialylation as well as provides evidence that the sialylation enhancement effects of DEX on recombinant CHO cells occurred through the GR. Finally, the effects of DEX on increasing sialylation were then confirmed in 5‐L controlled bioreactors. Addition of 1 µM DEX to the bioreactors on day 2 resulted in harvests with average increases of 16.2% for total sialic acid content and 15.8% in the protein fraction with N‐linked sialylation. DEX was found to be a simple and effective method for increasing sialylation of this CTLA4‐Ig fusion protein expressed in CHO cells. Biotechnol. Bioeng. 2010;107: 488–496. © 2010 Wiley Periodicals, Inc.
Profiling ofN‐glycosylation gene expression in CHO cell fed‐batch cultures
Authors: Wong Danny Chee Furng , Wong Niki Soo Ching , Goh John Soo Yang , May Lee May , Yap Miranda Gek Sim 
Published: June 2010

AbstractOne of the goals of recombinant glycoprotein production is to achieve consistent glycosylation. Although many studies have examined the changes in the glycosylation quality of recombinant protein with culture, very little has been done to examine the underlying changes in glycosylation gene expression as a culture progresses. In this study, the expression of 24 genes involved inN‐glycosylation were examined using quantitative RT PCR to gain a better understanding of recombinant glycoprotein glycosylation during production processes. Profiling of theN‐glycosylation genes as well as concurrent analysis of glycoprotein quality was performed across the exponential, stationary and death phases of a fed‐batch culture of a CHO cell line producing recombinant human interferon‐γ (IFN‐γ). Of the 24N‐glycosylation genes examined, 21 showed significant up‐ or down‐regulation of gene expression as the fed‐batch culture progressed from exponential, stationary and death phase. As the fed‐batch culture progressed, there was also an increase in less sialylated IFN‐γ glycoforms, leading to a 30% decrease in the molar ratio of sialic acid to recombinant IFN‐γ. This correlated with decreased expression of genes involved with CMP sialic acid synthesis coupled with increased expression of sialidases. Compared to batch culture, a low glutamine fed‐batch strategy appears to need a 0.5 mM glutamine threshold to maintain similarN‐glycosylation genes expression levels and to achieve comparable glycoprotein quality. This study demonstrates the use of quantitative real time PCR method to identify possible “bottlenecks” or “compromised” pathways inN‐glycosylation and subsequently allow for the development of strategies to improve glycosylation quality. Biotechnol. Bioeng. 2010;107: 516–528. © 2010 Wiley Periodicals, Inc.
Measuring knowledge of the insanity defense: Scale construction and validation
Authors: Daftary‐Kapur Tarika , Groscup Jennifer L. , O'Connor Maureen , Coffaro Frank , Galietta Michele 
Published: June 2010

AbstractGiven the influence of social conformity and prejudice, defendants pleading not guilty by reason of insanity face the significant challenges of securing fair and impartial juries. Attitudes and knowledge of the insanity defense are factors that may influence levels of impartiality. In the light of this, we set out to develop a scale to examine knowledge levels of the insanity defense and their influence on decision‐making. Two studies were conducted to construct a scale designed to assess laypersons' knowledge of the insanity defense. Items measuring knowledge of the insanity defense were based on Perlin's (1995) insanity defense myths. The first study identified particular items in need of revision and subscales that required the development of additional items in order to improve reliability and construct validity in the second study. The second study used the revised scale, demonstrating improved validity and reliability. The scale also had acceptable predictive validity with reference to insanity defense verdicts. Copyright © 2010 John Wiley & Sons, Ltd.
Radio frequency radiation causes no nonthermal damage in enzymes and living cells
Authors: Fortune Jennifer A. , Wu Bae‐Ian , Klibanov Alexander M. 
Published: June 2010

AbstractThe ability of radio frequency radiation (RFR) to exert irreversible nonthermal (i.e., not caused by accompanying heat) effects on biologics has been widely debated due to a relative paucity of comprehensive critical details in published reports dealing with this issue. In this study, we used rigorous control over experimental conditions to determine whether continuous RFR nonthermally affects commercially important enzymes and live bacterial and human cells using three most commonly used frequencies in current RF identification technology, namely 2.45 GHz, 915 MHz, and 13.56 MHz. Diverse biological samples were exposed to RFR under deliberately harsh conditions to increase the likelihood of observing such effects should they exist. Enzymatic activities of horseradish peroxidase and β‐galactosidase in aqueous solution exhibited no statistically discernable consequences of even very intense RFR. Likewise, with putative thermal effects excluded, the viabilities of bacteria (both gram‐positive and gram‐negative) and of human cells were not detectably compromised by such an RFR exposure. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010
Cryopreservation of human bone marrow‐derived mesenchymal stem cells with reduced dimethylsulfoxide and well‐defined freezing solutions
Authors: Liu Yang , Xu Xia , Ma Xuehu , Martin‐Rendon Enca , Watt Suzanne , Cui Zhanfeng 
Published: June 2010

AbstractThe aim of this study is to investigate the feasibility of using well defined, serum‐free freezing solutions with a reduced level of dimethylsulfoxide (DMSO) of 7.5, 5, and 2.5% (v/v) in the combination with polyethylene glycol (PEG) or trehalose to cryopreserve human bone marrow‐derived mesenchymal stem cells (hBMSCs), a main source of stem cells for cell therapy and tissue engineering. The standard laboratory freezing protocol of around 1°C/min was used in the experiments. The efficiency of 1,2‐propandiol on cryopreservation of hBMSCs was explored. We measured the post‐thawing cell viability and early apoptotic behaviors, cell metabolic activities, and growth dynamics. Cell morphology and osteogenic, adipogenic and chondrogenic differentiation capability were also tested after cryopreservation. The results showed that post‐thawing viability of hBMSCs in 7.5% DMSO (v/v), 2.5% PEG (w/v), and 2% bovine serum albumin (BSA) (w/v) was comparable with that obtained in conventional 10% DMSO, that is, 82.9 ± 4.3% and 82.7 ± 3.7%, respectively. In addition, 5% DMSO (v/v) with 5% PEG (w/v) and 7.5% 1,2‐propandiol (v/v) with 2.5% PEG (w/v) can provide good protection to hBMSCs when 2% albumin (w/v) is present. Enhanced cell viability was observed with the addition of albumin to all tested freezing solutions. © 2010 American Institute of Chemical Engineers Biotechnol. Prog., 2010
Late aortic perforation with an atriasept device resulting in life‐threatening tamponade
Authors: Murphy John C. , Walsh Simon J. , Spence, Mark S. 
Published: June 2010

AbstractPatients with compelling evidence of cryptogenic stroke due to patent foreamen ovale (PFO) are increasingly likely to be offered percutaneous closure of the defect. With improvements in technique there is now a high procedural success rate with low rates of periprocedural complications. Late complications are also rare, but include late perforation of the aortic root, which is rapidly progressive and potentially fatal. This has lead to the development of lower profile devices, which aim to reduce the risk of both early and late complications. At this stage, it is not clear if lower profile devices have safety profiles which are superior to the more established devices. We report the first case of late perforation of the aortic sinus by the lower profile Atriasept (Cardia) device presenting as life threatening cardiac tamponade in an adult who previously underwent percutaneous PFO closure. © 2010 Wiley‐Liss, Inc.
Cardiac shunt calculations made easy: A case‐based approach
Authors: Gössl Mario , Rihal Charanjit S. 
Published: June 2010

AbstractOn the basis of an interesting case of a 40‐year‐old lady with an atrial septal defect, we describe a simple approach for complex shunt volume calculations based on the Fick principle using hemodynamic data obtained in the catheterization laboratory. Understanding Fick's principle allows determination of all relevant flows, flow ratio, and resistances without having to resort to memorization of complex formulae. Those findings significantly impact the managements of patients like ours with hemodynamically and clinically relevant shunting.© 2010 Wiley‐Liss, Inc.