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Sample records for maxam-gilbert workup denaturing

  1. Denatured fuel cycles

    International Nuclear Information System (INIS)

    Till, C.E.

    1979-01-01

    This paper traces the history of the denatured fuel concept and discusses the characteristics of fuel cycles based on the concept. The proliferation resistance of denatured fuel cycles, the reactor types they involve, and the limitations they place on energy generation potential are discussed. The paper concludes with some remarks on the outlook for such cycles

  2. [Diagnostic workup of fragrance allergy].

    Science.gov (United States)

    Geier, J; Uter, W

    2015-09-01

    The diagnostic workup of contact allergy to fragrances must not be limited to patch testing with the two well-established fragrance mixes. False-positive reactions to these mixes occur in up to 50 % of the patch tested patients. For the diagnostic work-up of positive reactions, and in cases of suspected fragrance allergy, patch testing with the single mix components and additional fragrances is mandatory. Frequently sensitizing fragrance materials are the 14 components of the two fragrance mixes and tree moss (Evernia furfuracea), ylang ylang oil (I + II; Cananga odorata), lemongrass oil (Cymbopogon schoenanthus), sandalwood oil (Santalum album), jasmine absolute (Jasminum spp.), and, less frequently, clove oil (Eugenia caryophyllus), cedarwood oil (Cedrus atlantica/deodara, Juniperus virginiana), Neroli oil (Citrus aurantium amara flower oil), salicylaldehyde, narcissus absolute (Narcissus spp.), and patchouli oil (Pogostemon cablin).

  3. Optimizing radiologic workup: An artificial intelligence approach

    International Nuclear Information System (INIS)

    Swett, H.A.; Rothschild, M.; Weltin, G.G.; Fisher, P.R.; Miller, P.L.

    1987-01-01

    The increasing complexity of diagnostic imaging is presenting an ever-expanding variety of radiologic test options to referring clinicians, making it more difficult for them to plan efficient workup. Diagnosis-oriented reimbursement systems are providing new incentives for hospitals and radiologists to use imaging modalities judiciously. This paper describes DxCON, a developmental artificial intelligence-based computer system, which gives advice to physicians about the optimum sequencing of radiologic tests. DxCON analyzes a physician's proposed workup plan and discusses its strengths and weaknesses. The domain chosen for this research is the imaging workup of obstructive jaundice

  4. Patterns of anaphylaxis after diagnostic workup

    DEFF Research Database (Denmark)

    Oropeza, Athamaica Ruiz; Bindslev-Jensen, Carsten; Broesby-Olsen, Sigurd

    2017-01-01

    BACKGROUND: Most published studies on anaphylaxis are retrospective or register based. Data on subsequent diagnostic work-up are sparse. We aimed to characterize patients seen with suspected anaphylaxis at the emergency care setting (ECS), after subsequent diagnostic work-up at our Allergy Center...... (AC). METHODS: Prospective study including patients from the ECS, Odense University Hospital, during May 2013-April 2014. Possible anaphylaxis cases were daily identified based on a broad search profile including history and symptoms in patient records, diagnostic codes and pharmacological treatments....... At the AC, all patients were evaluated according to international guidelines. RESULTS: Among 226 patients with suspected anaphylaxis, the diagnosis was confirmed in 124 (54.9%) after diagnostic work-up; 118 of the 124 fulfilled WAO/EAACI criteria of anaphylaxis at the ECS, while 6 were found among 46...

  5. Denaturing gradient gel electrophoresis

    International Nuclear Information System (INIS)

    Kocherginskaya, S.A.; Cann, I.K.O.; Mackie, R.I.

    2005-01-01

    It is worthwhile considering that only some 30 species make up the bulk of the bacterial population in human faeces at any one time based on the classical cultivation-based approach. The situation in the rumen is similar. Thus, it is practical to focus on specific groups of interest within the complex community. These may be the predominant or the most active species, specific physiological groups or readily identifiable (genetic) clusters of phylogenetically related organisms. Several 16S rDNA fingerprinting techniques can be invaluable for selecting and monitoring sequences or phylogenetic groups of interest and are described below. Over the past few decades, considerable attention was focussed on the identification of pure cultures of microbes on the basis of genetic polymorphisms of DNA encoding rRNA such as ribotyping, amplified fragment length polymorphism and randomly amplified polymorphic DNA. However, many of these methods require prior cultivation and are less suitable for use in analysis of complex mixed populations although important in describing cultivated microbial diversity in molecular terms. Much less attention was given to molecular characterization of complex communities. In particular, research into diversity and community structure over time has been revolutionized by the advent of molecular fingerprinting techniques for complex communities. Denaturing or temperature gradient gel electrophoresis (DGGE/TGGE) methods have been successfully applied to the analysis of human, pig, cattle, dog and rodent intestinal populations

  6. Patient-specific workup of adrenal incidentalomas

    Directory of Open Access Journals (Sweden)

    Romy R. de Haan

    Full Text Available Purpose: : To develop a clinical prediction model to predict a clinically relevant adrenal disorder for patients with adrenal incidentaloma. Materials and methods: : This retrospective study is approved by the institutional review board, with waiver of informed consent. Natural language processing is used for filtering of adrenal incidentaloma cases in all thoracic and abdominal CT reports from 2010 till 2012. A total of 635 patients are identified. Stepwise logistic regression is used to construct the prediction model. The model predicts if a patient is at risk for malignancy or hormonal hyperfunction of the adrenal gland at the moment of initial presentation, thus generates a predicted probability for every individual patient. The prediction model is evaluated on its usefulness in clinical practice using decision curve analysis (DCA based on different threshold probabilities. For patients whose predicted probability is lower than the predetermined threshold probability, further workup could be omitted. Results: : A prediction model is successfully developed, with an area under the curve (AUC of 0.78. Results of the DCA indicate that up to 11% of patients with an adrenal incidentaloma can be avoided from unnecessary workup, with a sensitivity of 100% and specificity of 11%. Conclusion: : A prediction model can accurately predict if an adrenal incidentaloma patient is at risk for malignancy or hormonal hyperfunction of the adrenal gland based on initial imaging features and patient demographics. However, with most adrenal incidentalomas labeled as nonfunctional adrenocortical adenomas requiring no further treatment, it is likely that more patients could be omitting from unnecessary diagnostics. Keywords: Adrenal incidentaloma, Patient-specific workup, Prediction model

  7. [Hypercoagulable workup in ophthalmology. When and what].

    Science.gov (United States)

    Muñoz-Negrete, F J; Casas-Lleras, P; Pérez-López, M; Rebolleda, G

    2009-07-01

    Most ophthalmologic disorders secondary to hypercoagulabe state are due to the confluence of congenital and adquired factors. A systematic workup is mandatory. Most of congenital coagulation disorders cause venous trombosis and are inherited autosomal dominantly. In order of frequency these are factor V Leiden mutation (activated protein C resistance), G20210A mutation of the prothrombin gen and protein C, protein S, and antithrombin III deficiencies. Sickle cell anemia can determine arerial and venous thrombosis. In relation with arterial occlusion, the markers most frequently involved are homcysteine fasting levels and the markers of antiphospholipid antibody syndrome. Both of them can also determine venous thrombosis. Several acquired factors can lead to hypoercoagulable state, especially hyperhomocysteinemia, antiphospholipid antibody syndrome, hepatic disease, alcohol and tobacco intake, oral contraceptives, immobilization, surgeries and malignancies. In central venous occlusion is only necessary to rule out hyperhomocysteinemia and antiphospholipid antibody syndrome in young patients without known risk factors. In central artery occlusion, hypercoagulable workup is only recommended for patients less than 50 years-old with unknown emboli source. In this cases protein C, protein S, and antithrombin III deficiencies, homocystein, sickle cell disease and antiphospholipid antibody syndrome will ruled out. In non arteritic ischemic optic neuropathy hypercoagulable work up is not necessary. In amaurosis fugax without known emboli source, it is recommended to rule out etiologies of arterial occlusion, especially antithrombin III deficiencies, homocystein, sickle cell disease and antiphospholipid antibody syndrome.

  8. 27 CFR 19.456 - Adding denaturants.

    Science.gov (United States)

    2010-04-01

    ... proprietor shall submit a flow diagram of the intended process or method of adding denaturants. (Sec. 201... OF THE TREASURY LIQUORS DISTILLED SPIRITS PLANTS Denaturing Operations and Manufacture of Articles...

  9. Neuroradiologic work-up of brain tumors

    International Nuclear Information System (INIS)

    Fishbein, D.S.

    1988-01-01

    The presence of an intracranial tumor may be suspected or deduced from the clinical history and examination, or it may be discovered incidentally during investigation of another disorder. Once the suggestion is raised, a variety of neuroradiologic techniques are available to define the extent and nature of the lesion. The studies performed may allow a tissue diagnosis to be presumed, may serve as a guide to proposed surgical therapy, or may allow the course of a previously diagnosed lesion to be followed. This chapter discusses the utility of common neuroradiologic techniques and their specific indications in the work-up of intracranial tumors. Emphasis is placed upon tests that are most frequently utilized and have the greatest value

  10. Single DNA denaturation and bubble dynamics

    DEFF Research Database (Denmark)

    Metzler, Ralf; Ambjörnsson, Tobias; Hanke, Andreas

    2009-01-01

    While the Watson-Crick double-strand is the thermodynamically stable state of DNA in a wide range of temperature and salt conditions, even at physiological conditions local denaturation bubbles may open up spontaneously due to thermal activation. By raising the ambient temperature, titration......, or by external forces in single molecule setups bubbles proliferate until full denaturation of the DNA occurs. Based on the Poland-Scheraga model we investigate both the equilibrium transition of DNA denaturation and the dynamics of the denaturation bubbles with respect to recent single DNA chain experiments...... for situations below, at, and above the denaturation transition. We also propose a new single molecule setup based on DNA constructs with two bubble zones to measure the bubble coalescence and extract the physical parameters relevant to DNA breathing. Finally we consider the interplay between denaturation...

  11. Single DNA denaturation and bubble dynamics

    International Nuclear Information System (INIS)

    Metzler, Ralf; Ambjoernsson, Tobias; Hanke, Andreas; Fogedby, Hans C

    2009-01-01

    While the Watson-Crick double-strand is the thermodynamically stable state of DNA in a wide range of temperature and salt conditions, even at physiological conditions local denaturation bubbles may open up spontaneously due to thermal activation. By raising the ambient temperature, titration, or by external forces in single molecule setups bubbles proliferate until full denaturation of the DNA occurs. Based on the Poland-Scheraga model we investigate both the equilibrium transition of DNA denaturation and the dynamics of the denaturation bubbles with respect to recent single DNA chain experiments for situations below, at, and above the denaturation transition. We also propose a new single molecule setup based on DNA constructs with two bubble zones to measure the bubble coalescence and extract the physical parameters relevant to DNA breathing. Finally we consider the interplay between denaturation bubbles and selectively single-stranded DNA binding proteins.

  12. Etiologic stroke subtypes: updated definition and efficient workup strategies.

    Science.gov (United States)

    Mehndiratta, Prachi; Chapman Smith, Sherita; Worrall, Bradford B

    2015-01-01

    Stroke affects approximately 16.9 million individuals per year worldwide and is the second leading cause of death. Stroke represents a family of related, but distinct subtypes. Classifying stroke subtypes must take into account various aspects of a standardized stroke workup to allow optimization of treatment and prevention strategies. Secondary prevention and pharmacologic treatment is tailored based on stroke mechanism. Additionally prognostication and recurrent risk also depends on stroke etiology. Efficient workup of stroke relies on a thorough history, clinical examination, imaging studies, and putative mechanism of stroke that lead the treating physician to a particular etiological path. Here , we provide the reader with updated definitions of etiologic ischemic stroke types as well as efficient workup strategies.

  13. Tertiary work-up of apparent treatment-resistant hypertension.

    Science.gov (United States)

    Heimark, Sondre; Eskås, Per Anders; Mariampillai, Julian Eek; Larstorp, Anne Cecilie K; Høieggen, Aud; Fadl Elmula, Fadl Elmula M

    2016-10-01

    Treatment-resistant hypertension (TRH) has regained attention with development of new methods for treatment. However, the prevalence of TRH varies considerably from primary to secondary and tertiary care. We aimed to assess the prevalence of true TRH in a population of patients with apparent TRH in a university hospital setting of tertiary work-up and also investigate reasons for poor BP control and evaluate how work-up can be performed in general practice and secondary care. In this cohort study, we characterize a study population from Oslo Renal Denervation (RDN) Study. Patients (n = 83) were referred for RDN from secondary care. All patients underwent thorough medical investigation and 24-h ambulatory blood pressure measurements (24ABPM) after directly observed therapy (DOT). We then assessed reasons for lack of BP control. Fifty-three of 83 patients did not have true TRH. Main reasons for non-TRH were poor drug adherence (32%), secondary hypertension (30%) and white coat hypertension (15%). Forty-seven percent achieved blood pressure control after DOT with subsequent 24ABPM. There were otherwise no statistically significant differences in patient characteristics between the true TRH and the non-TRH group. Despite being a highly selected cohort referred for tertiary work-up of apparent TRH, BP control was achieved or secondary causes were identified in almost two thirds of the patients. Thorough investigation according to guidelines and DOT with subsequent 24ABPM is needed in work-up of apparent TRH.

  14. Reversibility of partial denaturation of DNA

    International Nuclear Information System (INIS)

    Acuna, M.I.; Mingot, F.; Davila, C.A.

    1976-01-01

    The recovery of hypochromicity in a partially denatured DNA sample when salt concentration is suddenly increased at a intermediate stage of the thermal transition is studied. The results of CsCl gradient analysis, PEG/DEX partition analysis, behaviour in a new thermal transition hydrodynamic properties and transforming ability, support the view that the process is an intramolecular double chain denaturation. The degree of denaturation irreversibility is dependent on single chain molecular weight of DNA (discontinuities denisty) and upon the helicity value at which salt concentration jump is performed. Both dependences are formally interpreted according to Elton's model for base distribution in DNA. Kinetically the process behaves as being an hydrodynamically limited rewinding. (author)

  15. 9 CFR 325.13 - Denaturing procedures.

    Science.gov (United States)

    2010-01-01

    ... the appropriate agent shall be used to give the material a distinctive color, odor, or taste so that... thoroughly mixing therein denaturing oil, No. 2 fuel oil, brucine dissolved in a mixture of alcohol and pine... distinctive a color, odor, or taste that it cannot be confused with an article of human food. [35 FR 15605...

  16. Analyzing Protein Denaturation using Fast Differential Scanning Calorimetry

    NARCIS (Netherlands)

    Splinter, R.; Van Herwaarden, A.W.; Iervolino, E.; Vanden Poel, G.; Istrate, D.; Sarro, P.M.

    2012-01-01

    This paper investigates the possibility to measure protein denaturation with Fast Differential Scanning Calorimetry (FDSC). Cancer can be diagnosed by measuring protein denaturation in blood plasma using Differential Scanning Calorimetry (DSC). FDSC can reduce diagnosis time from hours to minutes,

  17. Guanidinium-induced denaturation by breaking of salt bridges

    NARCIS (Netherlands)

    Meuzelaar, H.; Panman, M.R.; Woutersen, S.

    2015-01-01

    Despite its wide use as a denaturant, the mechanism by which guanidinium (Gdm+) induces protein unfolding remains largely unclear. Herein, we show evidence that Gdm+ can induce denaturation by disrupting salt bridges that stabilize the folded conformation. We study the Gdm+-​induced denaturation of

  18. Hysteresis in pressure-driven DNA denaturation.

    Directory of Open Access Journals (Sweden)

    Enrique Hernández-Lemus

    Full Text Available In the past, a great deal of attention has been drawn to thermal driven denaturation processes. In recent years, however, the discovery of stress-induced denaturation, observed at the one-molecule level, has revealed new insights into the complex phenomena involved in the thermo-mechanics of DNA function. Understanding the effect of local pressure variations in DNA stability is thus an appealing topic. Such processes as cellular stress, dehydration, and changes in the ionic strength of the medium could explain local pressure changes that will affect the molecular mechanics of DNA and hence its stability. In this work, a theory that accounts for hysteresis in pressure-driven DNA denaturation is proposed. We here combine an irreversible thermodynamic approach with an equation of state based on the Poisson-Boltzmann cell model. The latter one provides a good description of the osmotic pressure over a wide range of DNA concentrations. The resulting theoretical framework predicts, in general, the process of denaturation and, in particular, hysteresis curves for a DNA sequence in terms of system parameters such as salt concentration, density of DNA molecules and temperature in addition to structural and configurational states of DNA. Furthermore, this formalism can be naturally extended to more complex situations, for example, in cases where the host medium is made up of asymmetric salts or in the description of the (helical-like charge distribution along the DNA molecule. Moreover, since this study incorporates the effect of pressure through a thermodynamic analysis, much of what is known from temperature-driven experiments will shed light on the pressure-induced melting issue.

  19. A correlated Walks' theory for DNA denaturation

    International Nuclear Information System (INIS)

    Mejdani, R.

    1994-08-01

    We have shown that by using a correlated Walks' theory for the lattice gas model on a one-dimensional lattice, we can study, beside the saturation curves obtained before for the enzyme kinetics, also the DNA denaturation process. In the limit of no interactions between sites the equation for melting curves of DNA reduces to the random model equation. Thus our leads naturally to this classical equation in the limiting case. (author). 22 refs, 3 figs

  20. Multidetector Computed Tomography in the Preoperative Workup of Hilar Cholangiocarcinoma

    International Nuclear Information System (INIS)

    Kim, Hyoung Jung; Lee, Dong Ho; Lim, Joo Won; Ko, Young Tae

    2009-01-01

    Hilar cholangiocarcinoma is associated with a dismal prognosis; however, curative resection may offer a chance of cure. Various factors should be considered in the surgical planning for curative resection. These factors include extent of bile duct involvement, relationship between portal vein and tumor involvement, diffuse hepato duodenal ligament infiltration, vascular invasion, lymph node metastasis, peritoneal seeding, and hepatic volume. Using high-quality volume data from multidetector-row computed tomography (MDCT) and adequate postprocessing images, radiologists can provide various types of information, imperative for curative resection of a hilar cholangiocarcinoma. This review illustrates the role of MDCT in the preoperative workup of hilar cholangiocarcinoma

  1. Denatured plutonium: a study of deterrent action. Final report

    International Nuclear Information System (INIS)

    Hutchins, B.A.

    1975-07-01

    The safeguarding of nuclear reactor fuel includes physical security methods as well as technological process options. The purpose of this study was to provide a preliminary evaluation of a technological option; the introduction of denaturing as a deterrent to illicit plutonium diversion. Denaturing is accomplished by coextracting some highly-radioactive fission products with the plutonium during reprocessing of spent fuel. The radioactive denaturant is always in companion with the plutonium through all subsequent fuel cycle steps - and serves as a deterrent to diversion or illicit usage of this fissile source. In concept the denaturing approach is simple and straightforward. This report provides a preliminary analysis of denaturing which can be achieved within the framework of present reprocessing technology. The impact of denaturing is indicated by comparison to a conventional (i.e., non-denatured) light water reacter cycle approach

  2. Hemichrome formation during hemoglobin Zurich denaturation

    International Nuclear Information System (INIS)

    Zago, M.A.; Costa, F.F.; Botura, C.; Baffa, O.

    1988-01-01

    Electron paramagnetic resonance (EPR)spectrum of hemoglobin Zurich, after oxidation, storage and heating, showed several absorption derives in the high field region (g ≅ 2) which are indicative of hemichrome formation. Characteristic visible spectra of hemichromes were observed for oxidized Hb Zurich and for its spontaneous precipitate. The proportional increase of EPR signals at g ≅ 2 and decrease at g = 6.37, the constant ratio of absorbance at 540 nm to 280 nm during heating, and the similarity of this ratio for spontaneously precipitated HbA and for Hb Zurich indicate that heme is not lost during the first steps of Hb Zurich denaturation. (author) [pt

  3. Splenomegaly - Diagnostic validity, work-up, and underlying causes

    DEFF Research Database (Denmark)

    Curovic Rotbain, Emelie; Lund Hansen, Dennis; Schaffalitzky de Muckadell, Ove

    2017-01-01

    to an internal medical department and can be caused by a large spectrum of diseases, including haematological diseases and liver cirrhosis. However, some patients remain without a causal diagnosis, despite extensive medical work-up. PATIENTS AND METHODS: We identified 129 patients through the DNRP, that had been...... given the ICD-10 splenomegaly diagnosis code in 1994-2013 at Odense University Hospital, Denmark, excluding patients with prior splenomegaly, malignant haematological neoplasia or liver cirrhosis. Medical records were reviewed for validity of the splenomegaly diagnosis, diagnostic work...... significant under-coding, it should be considered if supplementary data sources should be used in addition, in order to attain a more representative population. Haematological diseases were the most common cause, however in a large fraction of patients no causal diagnosis was found....

  4. Partially folded intermediates during trypsinogen denaturation

    Directory of Open Access Journals (Sweden)

    Martins N.F.

    1999-01-01

    Full Text Available The equilibrium unfolding of bovine trypsinogen was studied by circular dichroism, differential spectra and size exclusion HPLC. The change in free energy of denaturation was = 6.99 ± 1.40 kcal/mol for guanidine hydrochloride and = 6.37 ± 0.57 kcal/mol for urea. Satisfactory fits of equilibrium unfolding transitions required a three-state model involving an intermediate in addition to the native and unfolded forms. Size exclusion HPLC allowed the detection of an intermediate population of trypsinogen whose Stokes radii varied from 24.1 ± 0.4 Å to 26.0 ± 0.3 Å for 1.5 M and 2.5 M guanidine hydrochloride, respectively. During urea denaturation, the range of Stokes radii varied from 23.9 ± 0.3 Å to 25.7 ± 0.6 Å for 4.0 M and 6.0 M urea, respectively. Maximal intrinsic fluorescence was observed at about 3.8 M urea with 8-aniline-1-naphthalene sulfonate (ANS binding. These experimental data indicate that the unfolding of bovine trypsinogen is not a simple transition and suggest that the equilibrium intermediate population comprises one intermediate that may be characterized as a molten globule. To obtain further insight by studying intermediates representing different stages of unfolding, we hope to gain a better understanding of the complex interrelations between protein conformation and energetics.

  5. Simulated pressure denaturation thermodynamics of ubiquitin.

    Science.gov (United States)

    Ploetz, Elizabeth A; Smith, Paul E

    2017-12-01

    Simulations of protein thermodynamics are generally difficult to perform and provide limited information. It is desirable to increase the degree of detail provided by simulation and thereby the potential insight into the thermodynamic properties of proteins. In this study, we outline how to analyze simulation trajectories to decompose conformation-specific, parameter free, thermodynamically defined protein volumes into residue-based contributions. The total volumes are obtained using established methods from Fluctuation Solution Theory, while the volume decomposition is new and is performed using a simple proximity method. Native and fully extended ubiquitin are used as the test conformations. Changes in the protein volumes are then followed as a function of pressure, allowing for conformation-specific protein compressibility values to also be obtained. Residue volume and compressibility values indicate significant contributions to protein denaturation thermodynamics from nonpolar and coil residues, together with a general negative compressibility exhibited by acidic residues. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Reversible thermal denaturation of immobilized rhodanese

    International Nuclear Information System (INIS)

    Horowitz, P.; Bowman, S.

    1987-01-01

    For the first time, the enzyme rhodanese had been refolded after thermal denaturation. This was previously not possible because of the strong tendency for the soluble enzyme to aggregate at temperatures above 37 degrees C. The present work used rhodanese that was covalently coupled to a solid support under conditions that were found to preserve enzyme activity. Rhodanese was immobilized using an N-hydroxymalonimidyl derivative of Sepharose containing a 6-carbon spacer. The number of immobilized competent active sites was measured by using [ 35 S]SO 3 (2-) to form an active site persulfide that is the obligatory catalytic intermediate. Soluble enzyme was irreversibly inactivated in 10 min at 52 degrees C. The immobilized enzyme regained at least 30% of its original activity even after boiling for 20 min. The immobilized enzyme had a Km and Vmax that were each approximately 3 times higher than the corresponding values for the native enzyme. After preincubation at high temperatures, progress curves for the immobilized enzyme showed induction periods of up to 5 min before attaining apparently linear steady states. The pH dependence of the activity was the same for both the soluble and the immobilized enzyme. These results indicate significant stabilization of rhodanese after immobilization, and instabilities caused by adventitious solution components are not the sole reasons for irreversibility of thermal denaturation seen with the soluble enzyme. The results are consistent with models for rhodanese that invoke protein association as a major cause of inactivation of the enzyme. Furthermore, the induction period in the progress curves is consistent with studies which show that rhodanese refolding proceeds through intermediate states

  7. 27 CFR 19.464 - Denatured spirits inventories.

    Science.gov (United States)

    2010-04-01

    ... of Articles Inventories § 19.464 Denatured spirits inventories. Each proprietor shall take a physical inventory of all denatured spirits in the processing account at the close of each calendar quarter and at... inventories. 19.464 Section 19.464 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE...

  8. 27 CFR 20.144 - Packages of completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Packages of completely denatured alcohol. 20.144 Section 20.144 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS DISTRIBUTION AND USE OF DENATURED ALCOHOL AND RUM Sale...

  9. 27 CFR 20.261 - Records of completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Records of completely denatured alcohol. 20.261 Section 20.261 Alcohol, Tobacco Products and Firearms ALCOHOL AND TOBACCO TAX AND TRADE BUREAU, DEPARTMENT OF THE TREASURY LIQUORS DISTRIBUTION AND USE OF DENATURED ALCOHOL AND RUM...

  10. Workup and management of patients with frequent premature ventricular contractions.

    Science.gov (United States)

    Giles, Katie; Green, Martin S

    2013-11-01

    Premature ventricular contractions (PVCs) are a frequently encountered entity in clinical cardiology. They rarely affect prognosis or management. However, they might produce bothersome symptoms and, in select individuals with a high PVC burden, they might contribute to left ventricular (LV) dysfunction. Workup of patients with very frequent PVCs consists of a thorough history and physical examination to screen for underlying cardiac disease and potential triggers. Routine investigations include a standard 12-lead electrocardiogram, as well as an echocardiogram. A Holter monitor should be performed in those with severe symptoms, a history of syncope, or a malignant family history. Exercise stress testing has a role in evaluating for ischemia and in the assessment of patients with exertional symptoms. More advanced testing is not warranted if these initial investigations are reassuring. Referral to an arrhythmia specialist should be considered in patients with LV dysfunction whose PVC burden exceeds 15%. Frequent ventricular ectopy represents a rare, but reversible cause of LV dysfunction and these patients should be further evaluated for possible catheter ablation. Copyright © 2013 Canadian Cardiovascular Society. Published by Elsevier Inc. All rights reserved.

  11. Diagnostic workup in carotid stenosis - a neurologist's perspective

    International Nuclear Information System (INIS)

    Rosenkranz, Michael; Gerloff, Christian

    2010-01-01

    Carotid artery stenosis is associated with the risk of stroke, myocardial infarction, and vascular death. In selected patients, revascularization of carotid narrowing by endarterectomy may reduce the risk of stroke distal to the stenosis. Carotid artery stenting has evolved as a potential alternative to endarterectomy. Four randomized clinical trials comparing safety and efficacy of endarterectomy versus stenting of symptomatic carotid stenosis have been published in recent years, but there remains some uncertainty about the implications of these trials for clinical routine. Both carotid stenting and endarterectomy are based on different treatment strategies which may result in different specific risk factors associated with each procedure. Hence, the procedural risk of either modality varies not only with the skills of the surgeon or the interventionalist but may depend on patient characteristics. It appears that the most important question is not whether one revascularization modality is superior but for which patient one modality is better than the other. A comprehensive diagnostic workup of patients with carotid stenosis based on a broad panel of covariates that affect the risk of vascular events may improve selection of patients for carotid revascularization and may help to decide for whom one revascularization modality is likely to be better than the other. (orig.)

  12. Guanidinium-Induced Denaturation by Breaking of Salt Bridges.

    Science.gov (United States)

    Meuzelaar, Heleen; Panman, Matthijs R; Woutersen, Sander

    2015-12-07

    Despite its wide use as a denaturant, the mechanism by which guanidinium (Gdm(+) ) induces protein unfolding remains largely unclear. Herein, we show evidence that Gdm(+) can induce denaturation by disrupting salt bridges that stabilize the folded conformation. We study the Gdm(+) -induced denaturation of a series of peptides containing Arg/Glu and Lys/Glu salt bridges that either stabilize or destabilize the folded conformation. The peptides containing stabilizing salt bridges are found to be denatured much more efficiently by Gdm(+) than the peptides containing destabilizing salt bridges. Complementary 2D-infrared measurements suggest a denaturation mechanism in which Gdm(+) binds to side-chain carboxylate groups involved in salt bridges. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Effect of radiologists' diagnostic work-up volume on interpretive performance.

    Science.gov (United States)

    Buist, Diana S M; Anderson, Melissa L; Smith, Robert A; Carney, Patricia A; Miglioretti, Diana L; Monsees, Barbara S; Sickles, Edward A; Taplin, Stephen H; Geller, Berta M; Yankaskas, Bonnie C; Onega, Tracy L

    2014-11-01

    To examine radiologists' screening performance in relation to the number of diagnostic work-ups performed after abnormal findings are discovered at screening mammography by the same radiologist or by different radiologists. In an institutional review board-approved HIPAA-compliant study, the authors linked 651 671 screening mammograms interpreted from 2002 to 2006 by 96 radiologists in the Breast Cancer Surveillance Consortium to cancer registries (standard of reference) to evaluate the performance of screening mammography (sensitivity, false-positive rate [ FPR false-positive rate ], and cancer detection rate [ CDR cancer detection rate ]). Logistic regression was used to assess the association between the volume of recalled screening mammograms ("own" mammograms, where the radiologist who interpreted the diagnostic image was the same radiologist who had interpreted the screening image, and "any" mammograms, where the radiologist who interpreted the diagnostic image may or may not have been the radiologist who interpreted the screening image) and screening performance and whether the association between total annual volume and performance differed according to the volume of diagnostic work-up. Annually, 38% of radiologists performed the diagnostic work-up for 25 or fewer of their own recalled screening mammograms, 24% performed the work-up for 0-50, and 39% performed the work-up for more than 50. For the work-up of recalled screening mammograms from any radiologist, 24% of radiologists performed the work-up for 0-50 mammograms, 32% performed the work-up for 51-125, and 44% performed the work-up for more than 125. With increasing numbers of radiologist work-ups for their own recalled mammograms, the sensitivity (P = .039), FPR false-positive rate (P = .004), and CDR cancer detection rate (P women recalled per cancer detected from 17.4 for 25 or fewer mammograms to 24.6 for more than 50 mammograms. Increases in work-ups for any radiologist yielded significant

  14. Effect of Radiologists’ Diagnostic Work-up Volume on Interpretive Performance

    Science.gov (United States)

    Anderson, Melissa L.; Smith, Robert A.; Carney, Patricia A.; Miglioretti, Diana L.; Monsees, Barbara S.; Sickles, Edward A.; Taplin, Stephen H.; Geller, Berta M.; Yankaskas, Bonnie C.; Onega, Tracy L.

    2014-01-01

    Purpose To examine radiologists’ screening performance in relation to the number of diagnostic work-ups performed after abnormal findings are discovered at screening mammography by the same radiologist or by different radiologists. Materials and Methods In an institutional review board–approved HIPAA-compliant study, the authors linked 651 671 screening mammograms interpreted from 2002 to 2006 by 96 radiologists in the Breast Cancer Surveillance Consortium to cancer registries (standard of reference) to evaluate the performance of screening mammography (sensitivity, false-positive rate [FPRfalse-positive rate], and cancer detection rate [CDRcancer detection rate]). Logistic regression was used to assess the association between the volume of recalled screening mammograms (“own” mammograms, where the radiologist who interpreted the diagnostic image was the same radiologist who had interpreted the screening image, and “any” mammograms, where the radiologist who interpreted the diagnostic image may or may not have been the radiologist who interpreted the screening image) and screening performance and whether the association between total annual volume and performance differed according to the volume of diagnostic work-up. Results Annually, 38% of radiologists performed the diagnostic work-up for 25 or fewer of their own recalled screening mammograms, 24% performed the work-up for 0–50, and 39% performed the work-up for more than 50. For the work-up of recalled screening mammograms from any radiologist, 24% of radiologists performed the work-up for 0–50 mammograms, 32% performed the work-up for 51–125, and 44% performed the work-up for more than 125. With increasing numbers of radiologist work-ups for their own recalled mammograms, the sensitivity (P = .039), FPRfalse-positive rate (P = .004), and CDRcancer detection rate (P work-ups for any radiologist yielded significant increases in FPRfalse-positive rate (P = .011) and CDRcancer detection rate (P

  15. Thermal denaturation of A-DNA

    International Nuclear Information System (INIS)

    Valle-Orero, J; Wildes, A R; Theodorakopoulos, N; Cuesta-López, S; Peyrard, M; Garden, J-L; Danilkin, S

    2014-01-01

    The DNA molecule can take various conformational forms. Investigations focus mainly on the so-called ‘B-form’, schematically drawn in the famous paper by Watson and Crick [1]. This is the usual form of DNA in a biological environment and is the only form that is stable in an aqueous environment. Other forms, however, can teach us much about DNA. They have the same nucleotide base pairs for ‘building blocks’ as B-DNA, but with different relative positions, and studying these forms gives insight into the interactions between elements under conditions far from equilibrium in the B-form. Studying the thermal denaturation is particularly interesting because it provides a direct probe of those interactions which control the growth of the fluctuations when the ‘melting’ temperature is approached. Here we report such a study on the ‘A-form’ using calorimetry and neutron scattering. We show that it can be carried further than a similar study on B-DNA, requiring the improvement of thermodynamic models for DNA. (paper)

  16. Single-molecule denaturation mapping of DNA in nanofluidic channels

    DEFF Research Database (Denmark)

    Reisner, Walter; Larsen, Niels Bent; Silahtaroglu, Asli

    2010-01-01

    Here we explore the potential power of denaturation mapping as a single-molecule technique. By partially denaturing YOYO (R)-1-labeled DNA in nanofluidic channels with a combination of formamide and local heating, we obtain a sequence-dependent "barcode" corresponding to a series of local dips...... and peaks in the intensity trace along the extended molecule. We demonstrate that this structure arises from the physics of local denaturation: statistical mechanical calculations of sequence-dependent melting probability can predict the barcode to be observed experimentally for a given sequence...

  17. Microwave-enhanced folding and denaturation of globular proteins

    DEFF Research Database (Denmark)

    Bohr, Henrik; Bohr, Jakob

    2000-01-01

    It is shown that microwave irradiation can affect the kinetics of the folding process of some globular proteins, especially beta-lactoglobulin. At low temperature the folding from the cold denatured phase of the protein is enhanced, while at a higher temperature the denaturation of the protein from...... its folded state is enhanced. In the latter case, a negative temperature gradient is needed for the denaturation process, suggesting that the effects of the microwaves are nonthermal. This supports the notion that coherent topological excitations can exist in proteins. The application of microwaves...

  18. Light water reactors with a denatured thorium fuel cycle

    International Nuclear Information System (INIS)

    1978-05-01

    Discussed in this paper is the performance of denatured thorium fuel cycles in PWR plants of conventional design, such as those currently in operation or under construction. Although some improvement in U 3 O 8 utilization is anticipated in PWRs optimized explicitly for the denatured thorium fuel cycle, this paper is limited to a discussion of the performance of denatured thorium fuels in conventional PWRs and consequently the data presented is representative of the use of thorium fuel in existing PWRs or those presently under construction. In subsequent sections of this paper, the design of the PWR, its performance on the denatured thorium fuel cycle, safety, accident and environmental considerations, and technological status and R and D requirements are discussed

  19. Accuracy and consequences of same-day, invasive lung cancer workup

    DEFF Research Database (Denmark)

    Madsen, Kirsten Riis; Høegholm, Asbjørn; Bodtger, Uffe

    2016-01-01

    BACKGROUND: Though widely used, little is known about accuracy and efficacy of same-day, invasive workup of suspected lung cancer. OBJECTIVE: To evaluate the accuracy and efficacy of same-day, invasive lung cancer workup (diagnosis and mediastinal staging), and to identify differences between...... patients without (Group A) or with (Group B) need for resampling. METHODS: A retrospective study was performed on all consecutive patients referred for surgical treatment for localised lung cancer after invasive diagnostic and staging workup at our unit. Data were extracted from electronic medical files...... pulmonary disease. Tumour located in right upper lobe was associated with need for resampling. DISCUSSION: Our retrospective study suggests that same-day, invasive workup for lung cancer is safe, accurate, and efficacious in reducing time to therapy, even in patients with small lesions and low tumour burden....

  20. Thermal denaturation of type I collagen vitrified gels

    International Nuclear Information System (INIS)

    Xia, Zhiyong; Calderon-Colon, Xiomara; Trexler, Morgana; Elisseeff, Jennifer; Guo, Qiongyu

    2012-01-01

    Highlights: ► We analyzed the denaturation of vitrigels synthesized under different conditions. ► Overall denaturation kinetics consisted of both reversible and irreversible steps. ► More stable vitrigels were formed under high level of vitrification. - Abstract: The denaturation kinetics of type I collagen vitrigels synthesized under different vitrification time and temperature were analyzed by the classical Kissinger approach and the advanced model free kinetics (AMFK) using the Vyazovkin algorithm. The AMFK successfully elucidated the overall denaturation into reversible and irreversible processes. Depending on vitrification conditions, the activation energy for the irreversible process ranged from 100 to 200 kJ/mol, and the reversible enthalpy ranged from 250 to 300 kJ/mol. All of these values increased with the vitrification time and temperature, indicating that a more stable and complex structure formed with increased vitrification. The classical Kissinger method predicted the presence of a critical temperate of approximately 60 °C for the transition between reversible and irreversible processes. Scanning electron microscopy revealed the presence of fibril structures in vitrigels both before and after full denaturation; however the fibrils had became thicker and rougher after denaturation.

  1. "Cooperative collapse" of the denatured state revealed through Clausius-Clapeyron analysis of protein denaturation phase diagrams.

    Science.gov (United States)

    Tischer, Alexander; Machha, Venkata R; Rösgen, Jörg; Auton, Matthew

    2018-02-19

    Protein phase diagrams have a unique potential to identify the presence of additional thermodynamic states even when non-2-state character is not readily apparent from the experimental observables used to follow protein unfolding transitions. Two-state analysis of the von Willebrand factor A3 domain has previously revealed a discrepancy in the calorimetric enthalpy obtained from thermal unfolding transitions as compared with Gibbs-Helmholtz analysis of free energies obtained from the Linear Extrapolation Method (Tischer and Auton, Prot Sci 2013; 22(9):1147-60). We resolve this thermodynamic conundrum using a Clausius-Clapeyron analysis of the urea-temperature phase diagram that defines how ΔH and the urea m-value interconvert through the slope of c m versus T, (∂cm/∂T)=ΔH/(mT). This relationship permits the calculation of ΔH at low temperature from m-values obtained through iso-thermal urea denaturation and high temperature m-values from ΔH obtained through iso-urea thermal denaturation. Application of this equation uncovers sigmoid transitions in both cooperativity parameters as temperature is increased. Such residual thermal cooperativity of ΔH and the m-value confirms the presence of an additional state which is verified to result from a cooperative phase transition between urea-expanded and thermally-compact denatured states. Comparison of the equilibria between expanded and compact denatured ensembles of disulfide-intact and carboxyamidated A3 domains reveals that introducing a single disulfide crosslink does not affect the presence of the additional denatured state. It does, however, make a small thermodynamically favorable free energy (∼-13 ± 1 kJ/mol) contribution to the cooperative denatured state collapse transition as temperature is raised and urea concentration is lowered. The thermodynamics of this "cooperative collapse" of the denatured state retain significant compensations between the enthalpy and entropy contributions to the overall

  2. Heavy water reactors on the denatured thorium cycles

    International Nuclear Information System (INIS)

    1978-05-01

    This paper presents preliminary technical and economic data to INFCE on the denatured U-233/Thorium fuel cycle for use in early comparisons of alternate nuclear systems. The once-through uranium fuel cycle is discussed in a companion paper. In presenting this preliminary information at this time, it is recognized that there are several other denatured thorium fuel cycles of potential interest, such as the U-235/thorium cycle which could be implemented at an earlier date. Information on these alternate cycles is currently being developed, and will be provided to INFCE when available

  3. On the radioimmunological determination of native and heat denaturated protein

    International Nuclear Information System (INIS)

    Menzel, E.J.; Glatz, F.; Technische Univ., Vienna

    1981-01-01

    Precipitation radioimmunoassay, solid phase radioimmunoassay and passive hemagglutination were examined for their efficiency in the determination of native or denaturated soy proteins. Native as well as autoclaved soy protein could be determined quantitatively in the precipitation radioimmunoassay, using antisera directed against the native product. In the solid phase technique only the autoclaved soy protein could be detected with high sensitivity. In the passive hemagglutination reaction, no agglutination could be observed with erythrocytes coated with autoclaved soy protein. Only antisera against the denaturated (autoclaved) soy protein agglutinated these erythrocytes. (orig.) [de

  4. GPs' perspectives on the diagnostic work-up in patients with shoulder pain: a qualitative study.

    Science.gov (United States)

    Ottenheijm, Ramon P G; Hesselmans, Nicolle J J M; Kemper, Anouk; Moser, Albine; de Bie, Rob A; Dinant, Geert-Jan; Cals, Jochen W L

    2014-06-01

    The diagnostic work-up of patients with shoulder pain in general practice is complex. General practitioners' (GPs) guidelines advise a pragmatic diagnostic work-up in which additional imaging has a limited role. However, diagnostic ultrasounds are increasingly ordered by GPs, which seems to reflect complexity in management of shoulder pain. This study aimed to explore GPs' perspectives on the diagnostic work-up of patients with shoulder pain. This study has a qualitative exploratory design with an inductive approach and was carried out in Dutch general practice. The study population consisted of 18 Dutch GPs who were sampled purposefully with a spread in clinical experience and ordering diagnostic ultrasound. Data were gathered by means of semi-structured interviews and analysed following principles of the constant comparative method. Three main categories with subcategories emerged that captured the diagnostic work-up of shoulder pain: variety in diagnostic classifications [(non-)specific diagnosis and interdisciplinary differences], establishing strategies for diagnostic work-up (use of existing tools and motives to deviate from existing tools), and strategies dealing with diagnostic uncertainties (accepting diagnostic uncertainties, diagnostic imaging tests, and interdisciplinary consultation and referral). Despite the availability of evidence-based shoulder guidelines, GPs experience uncertainties during diagnostic work-up and apply different strategies when dealing with these uncertainties. At some point, GPs as well as patients seem to have a need for a specific diagnosis. Currently, there appears to be little agreement if, or in which phase of shoulder pain, diagnostic ultrasound is useful or indicated. © 2014 John Wiley & Sons, Ltd.

  5. Denaturation of membrane proteins and hyperthermic cell killing

    NARCIS (Netherlands)

    Burgman, Paulus Wilhelmus Johannes Jozef

    1993-01-01

    Summarizing: heat induced denaturation of membrane proteins is probably related to hyperthermic cell killing. Induced resistance of heat sensitive proteins seems to be involved in the development of thermotolerance. Although many questions remain still to be answered, it appears that HSP72, when

  6. Role of cyclobutane dimers in UV-denaturation of DNA

    International Nuclear Information System (INIS)

    Zavil'gel'skij, G.B.; Zuev, A.V.

    1978-01-01

    UV irradiation of double-stranded DNA produces local denatured regions. The evidence presented indicates that these single-stranded regions arise from photoproducts other than pyrimidine dimers. The irradiation of T2 DNA at 8x10 4 erg/mm 2 (254 nm) produces 6-8% thymine dimers, amd Tsub(mel) drops by 12-14 deg C, accompanied by a significant broadening of the transition profile. The kinetics of denatured region formation and lowering Tsub(mel) corresponds to that of formation of crosslinkages and differs markedly from the kinetics of formation of cyclobutane pyrimidine dimers. Treatment of UV-irradiated DNA with light in the presence of yeast photoreactivating enzyme monomerizes almost all thymine dimers but does not change the Tsub(mel). Local denatured regions are detected in UV-irradiated DNA and are absent from AcPhM-sensibilized DNA, which contains 20-25% thymine dimers, as determined by the accridine orange fluorescence technique. S1 nuclease from Aspergillis oryzae produces single-strand breaks in UV-irradiated DNA of phage PM2 but is not active on AcPhM-treated PM2 DNA, which contains about 50 thymine dimers. It is supposed that the formation of a cyclobutane dimer only weakens the hydrogen bonds in the AT base pair rather than breaks them. Local denatured regions are thought to arise from the accumulation in UV-irradiated DNA (254 nm) of the sufficient number of photoproducts with impaired ability to base pairing

  7. Cold denaturation of the HIV-1 protease monomer

    DEFF Research Database (Denmark)

    Rösner, Heike Ilona; Caldarini, Martina; Prestel, Andreas

    2017-01-01

    The HIV-1-protease is a complex protein which in its active form adopts a homodimer dominated by -sheet structures. We have discovered a cold-denatured state of the monomeric subunit of HIV-1-protease which is populated above 0ºC and therefore directly accessible to various spectroscopic approac...

  8. Evaluation of denaturing gradient gel electrophoresis (DGGE) used ...

    African Journals Online (AJOL)

    Denaturing gradient gel electrophoresis (DGGE) is a powerful method used to study structure of bacterial communities, without cultivation, based on the diversity of the genes coding for ribosomal RNA. However, the results are strongly dependent on the respective target region of the used primer systems. Therefore, three ...

  9. Workup of Human Blood Samples for Deep Sequencing of HIV-1 Genomes

    NARCIS (Netherlands)

    Cornelissen, Marion; Gall, Astrid; van der Kuyl, Antoinette; Wymant, Chris; Blanquart, François; Fraser, Christophe; Berkhout, Ben

    2018-01-01

    We describe a detailed protocol for the manual workup of blood (plasma/serum) samples from individuals infected with the human immunodeficiency virus type 1 (HIV-1) for deep sequence analysis of the viral genome. The study optimizing the assay was performed in the context of the BEEHIVE (Bridging

  10. Diagnostic work-up and loss of tuberculosis suspects in Jogjakarta, Indonesia.

    Science.gov (United States)

    Ahmad, Riris Andono; Matthys, Francine; Dwihardiani, Bintari; Rintiswati, Ning; de Vlas, Sake J; Mahendradhata, Yodi; van der Stuyft, Patrick

    2012-02-15

    Early and accurate diagnosis of pulmonary tuberculosis (TB) is critical for successful TB control. To assist in the diagnosis of smear-negative pulmonary TB, the World Health Organisation (WHO) recommends the use of a diagnostic algorithm. Our study evaluated the implementation of the national tuberculosis programme's diagnostic algorithm in routine health care settings in Jogjakarta, Indonesia. The diagnostic algorithm is based on the WHO TB diagnostic algorithm, which had already been implemented in the health facilities. We prospectively documented the diagnostic work-up of all new tuberculosis suspects until a diagnosis was reached. We used clinical audit forms to record each step chronologically. Data on the patient's gender, age, symptoms, examinations (types, dates, and results), and final diagnosis were collected. Information was recorded for 754 TB suspects; 43.5% of whom were lost during the diagnostic work-up in health centres, 0% in lung clinics. Among the TB suspects who completed diagnostic work-ups, 51.1% and 100.0% were diagnosed without following the national TB diagnostic algorithm in health centres and lung clinics, respectively. However, the work-up in the health centres and lung clinics generally conformed to international standards for tuberculosis care (ISTC). Diagnostic delays were significantly longer in health centres compared to lung clinics. The high rate of patients lost in health centres needs to be addressed through the implementation of TB suspect tracing and better programme supervision. The national TB algorithm needs to be revised and differentiated according to the level of care.

  11. Incidence of malignancy in patients with pleural effusion referred for workup by pulmonologists

    DEFF Research Database (Denmark)

    Lindegaard, Dennis V.; Reuter, Simon; Laursen, Christian B.

    2016-01-01

    Introduction Pleural effusion (PE) is a common condition. Malignancy accounts for lt;25% in general populations. The proportion is unknown in patients referred to pulmonologists for workup.Finding malignant cells in PE by thoracentesis suggests metastatic and incurable disease making further tests...

  12. Efficiency of Staging Work-Ups in the Evaluation of Carcinoma of the Uterine Cervix

    International Nuclear Information System (INIS)

    Kim, Jae Sung; Ha, Sung Whan

    1991-01-01

    A series of 510 patients with carcinoma of the uterine cervix given the curative radiation therapy from March 1979 through December 1986 was evaluated to determine the value of intravenous pyelography(IVP), cystoscopy, sigmoidoscopy, and abdomino-pelvic CT as staging work-up prior to treatment. On IVP and cystoscopy, 10.7%(49/456) and 5/3%(24/452) showed abnormality, respectively, but only 0.7% (3/413) did on signoidoscopy. As a result of these work-ups prerequisite to FIGO staging, twenty six (5.1%) out of 510 patients were upstaged from the stage determined by the findings of physical examination alone. The proportions of upstaging in each stage were as follows none in stage IB(35), IIA (89) and IIIA(8), 7.9%(20/252) in stage IIB(14 patients to FIGO Stage IIIB, 6 patients to FIGO stage IVA), and 4.8% (6/126) in stage IIIB (all to FIGO stage IVA). Positive findings of staging work-ups were found only in patients with advanced stages of stage IIB or over determined by physical examination alone but not in those with earlier stages. CT was performed in 337 patients. CT detected pelvic lymph node (LN) enlargement in 25.2% (85/337) and paraaortic LN enlargement in 7.4% (25/337). Pelvic LN positivity was well correlated with increasing stage but paraaortic LN positivity was not. In the evaluation of parametrial involvement, CT findings were in accordance with those of physical examination only in 65.6%(442/674). When compared with endoscopic studies, CT had much lower positive predictive value than negative predictive value in the evaluation of adjacent organ invasion. The staging work-ups should be individualized by the disease extent of each patient, and then the efficiency of work-ups may be increased without compromising the appropriate FIGO staging and treatment

  13. Spectral shift controlled reactors, denatured U-233/thorium cycle

    International Nuclear Information System (INIS)

    1978-05-01

    This paper presents technical and economic data on the SSCR which may be of use in the International Fuel Cycle Evaluation Program to intercompare alternative nuclear systems. Included in this paper are data on the denatured U-233/thorium cycle. This cycle shows a proliferation advantage over more classical thorium fuel cycle (e.g., highly-enriched U-235/thorium or plutonium/thorium) due to the elimination of chemically-separable, concentrated fissile material from unirradiated nuclear fuel. The U-233 is denatured by mixing with depleted uranium to a concentration no greater than 12 w/o. An exogenous source of U-233 is assumed in this paper, since U-233 does not occur in nature and only a limited supply has been produced to date for research and development work

  14. Vibrational energy relaxation: proposed pathway of fast local chromatin denaturation

    International Nuclear Information System (INIS)

    Harder, D.; Greinert, R.

    2002-01-01

    The molecular mechanism responsible for the a component of exchange-type chromosome aberrations, of chromosome fragmentation and of reproductive cell death is one of the unsolved issues of radiation biology. Under review is whether vibrational energy relaxation in the constitutive biopolymers of chromatin, induced by inelastic energy deposition events and mediated via highly excited vibrational states, may provide a pathway of fast local chromatin denaturation, thereby producing the severe DNA lesion able to interact chemically with other, non-damaged chromatin. (author)

  15. Preparation of denatured protein bone sterilized with gamma radiation

    International Nuclear Information System (INIS)

    Luna Z, D.

    2005-01-01

    The bone is one of the tissues more transplanted in the entire world by that the bone necessity for transplant every day becomes bigger. In the Bank of tissues Radio sterilized of the ININ the amnion and the pig skin are routinely processed. The tissue with which will be continued is with bone. Due to that in our country it doesn't have enough bone of human origin for the necessities required in the bone transplant, an option is the bone of bovine. Of this bone one can obtain denatured protein bone, with the same characteristics of the denatured protein human bone, the one which has been proven that it has good acceptance and incorporation in the human body when is transplanted. The method for the obtaining of the denatured protein bone of bovine, with the confirmation of the final product by means of X-ray diffraction is described. The radiosterilization of this bone with gamma rays and the determination of the lead content. (Author)

  16. Diagnostic work-up and loss of tuberculosis suspects in Jogjakarta, Indonesia

    Directory of Open Access Journals (Sweden)

    Ahmad Riris

    2012-02-01

    Full Text Available Abstract Background Early and accurate diagnosis of pulmonary tuberculosis (TB is critical for successful TB control. To assist in the diagnosis of smear-negative pulmonary TB, the World Health Organisation (WHO recommends the use of a diagnostic algorithm. Our study evaluated the implementation of the national tuberculosis programme's diagnostic algorithm in routine health care settings in Jogjakarta, Indonesia. The diagnostic algorithm is based on the WHO TB diagnostic algorithm, which had already been implemented in the health facilities. Methods We prospectively documented the diagnostic work-up of all new tuberculosis suspects until a diagnosis was reached. We used clinical audit forms to record each step chronologically. Data on the patient's gender, age, symptoms, examinations (types, dates, and results, and final diagnosis were collected. Results Information was recorded for 754 TB suspects; 43.5% of whom were lost during the diagnostic work-up in health centres, 0% in lung clinics. Among the TB suspects who completed diagnostic work-ups, 51.1% and 100.0% were diagnosed without following the national TB diagnostic algorithm in health centres and lung clinics, respectively. However, the work-up in the health centres and lung clinics generally conformed to international standards for tuberculosis care (ISTC. Diagnostic delays were significantly longer in health centres compared to lung clinics. Conclusions The high rate of patients lost in health centres needs to be addressed through the implementation of TB suspect tracing and better programme supervision. The national TB algorithm needs to be revised and differentiated according to the level of care.

  17. Balloon-assisted enteroscopy for suspected Meckel’s diverticulum and indefinite diagnostic imaging workup

    Science.gov (United States)

    Gomes, Guilherme Francisco; Bonin, Eduardo Aimore; Noda, Rafael William; Cavazzola, Leandro Totti; Bartholomei, Thiago Ferreira

    2016-01-01

    Meckel’s diverticulum (MD) is estimated to affect 1%-2% of the general population, and it represents a clinically silent finding of a congenital anomaly in up to 85% of the cases. In adults, MD may cause symptoms, such as overt occult lower gastrointestinal bleeding. The diagnostic imaging workup includes computed tomography scan, magnetic resonance imaging enterography, technetium 99m scintigraphy (99mTc) using either labeled red blood cells or pertechnetate (known as the Meckel’s scan) and angiography. The preoperative detection rate of MD in adults is low, and many patients ultimately undergo exploratory laparoscopy. More recently, however, endoscopic identification of MD has been possible with the use of balloon-assisted enteroscopy via direct luminal access, which also provides visualization of the diverticular ostium. The aim of this study was to review the diagnosis by double-balloon enteroscopy of 4 adults with symptomatic MD but who had negative diagnostic imaging workups. These cases indicate that balloon-assisted enteroscopy is a valuable diagnostic method and should be considered in adult patients who have suspected MD and indefinite findings on diagnostic imaging workup, including negative Meckel’s scan. PMID:27803776

  18. Membrane bridging and hemifusion by denaturated Munc18.

    Directory of Open Access Journals (Sweden)

    Yi Xu

    Full Text Available Neuronal Munc18-1 and members of the Sec1/Munc18 (SM protein family play a critical function(s in intracellular membrane fusion together with SNARE proteins, but the mechanism of action of SM proteins remains highly enigmatic. During experiments designed to address this question employing a 7-nitrobenz-2-oxa-1,3-diazole (NBD fluorescence de-quenching assay that is widely used to study lipid mixing between reconstituted proteoliposomes, we observed that Munc18-1 from squid (sMunc18-1 was able to increase the apparent NBD fluorescence emission intensity even in the absence of SNARE proteins. Fluorescence emission scans and dynamic light scattering experiments show that this phenomenon arises at least in part from increased light scattering due to sMunc18-1-induced liposome clustering. Nuclear magnetic resonance and circular dichroism data suggest that, although native sMunc18-1 does not bind significantly to lipids, sMunc18-1 denaturation at 37 °C leads to insertion into membranes. The liposome clustering activity of sMunc18-1 can thus be attributed to its ability to bridge two membranes upon (perhaps partial denaturation; correspondingly, this activity is hindered by addition of glycerol. Cryo-electron microscopy shows that liposome clusters induced by sMunc18-1 include extended interfaces where the bilayers of two liposomes come into very close proximity, and clear hemifusion diaphragms. Although the physiological relevance of our results is uncertain, they emphasize the necessity of complementing fluorescence de-quenching assays with alternative experiments in studies of membrane fusion, as well as the importance of considering the potential effects of protein denaturation. In addition, our data suggest a novel mechanism of membrane hemifusion induced by amphipathic macromolecules that does not involve formation of a stalk intermediate.

  19. Strategies for denaturing the weapons-grade plutonium stockpile

    International Nuclear Information System (INIS)

    Buckner, M.R.; Parks, P.B.

    1992-10-01

    In the next few years, approximately 50 metric tons of weapons-grade plutonium and 150 metric tons of highly-enriched uranium (HEU) may be removed from nuclear weapons in the US and declared excess. These materials represent a significant energy resource that could substantially contribute to our national energy requirements. HEU can be used as fuel in naval reactors, or diluted with depleted uranium for use as fuel in commercial reactors. This paper proposes to use the weapons-grade plutonium as fuel in light water reactors. The first such reactor would demonstrate the dual objectives of producing electrical power and denaturing the plutonium to prevent use in nuclear weapons

  20. U.S. leans toward denatured thorium cycle

    International Nuclear Information System (INIS)

    Smock, R.

    1977-01-01

    Denatured thorium appears to be the most promising among the nonproliferating alternatives to the plutonium cycle, which the Carter Administration is trying to cancel. Criteria for a better system include uranium utilization comparable to current light water reactors and minimal separation of fissile material into the waste stream. Comparisons with other systems conclude that thorium is preferable because it can lead to an acceptable fast breeder. The thorium cycle can be placed in energy centers for sensitive facilities and can also be introduced into ongoing light water systems. Reprocessing can be handled in the centers, where thorium can be mixed with plutonium for use in reactors within the center, while light water reactors operate on the outside. Any fuel leaving the center would be unsuitable for weapons. Later adaptation to in-center fast breeders will extend energy supplies, although a thorium breeder will be less efficient than a plutonium fast breeder. Denatured thorium is a technical answer to a complex political problem, but those in the nuclear industry see the U.S. goal of a nonproliferating fuel as futile in the light of world politics and breeder efforts in other countries

  1. Cooperative unfolding of apolipoprotein A-1 induced by chemical denaturation.

    Science.gov (United States)

    Eckhardt, D; Li-Blatter, X; Schönfeld, H-J; Heerklotz, H; Seelig, J

    2018-05-25

    Apolipoprotein A-1 (Apo A-1) plays an important role in lipid transfer and obesity. Chemical unfolding of α-helical Apo A-1 is induced with guanidineHCl and monitored with differential scanning calorimetry (DSC) and CD spectroscopy. The unfolding enthalpy and the midpoint temperature of unfolding decrease linearly with increasing guanidineHCl concentration, caused by the weak binding of denaturant. At room temperature, binding of 50-60 molecules guanidineHCl leads to a complete Apo A-1 unfolding. The entropy of unfolding decreases to a lesser extent than the unfolding enthalpy. Apo A-1 chemical unfolding is a dynamic multi-state equilibrium that is analysed with the Zimm-Bragg theory modified for chemical unfolding. The chemical Zimm-Bragg theory predicts the denaturant binding constant K D and the protein cooperativity σ. Chemical unfolding of Apo A-1 is two orders of magnitude less cooperative than thermal unfolding. The free energy of thermal unfolding is ~0.2 kcal/mol per amino acid residue and ~1.0 kcal/mol for chemical unfolding at room temperature. The Zimm-Bragg theory calculates conformational probabilities and the chemical Zimm-Bragg theory predicts stretches of α-helical segments in dynamic equilibrium, unfolding and refolding independently and fast. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  2. Thermal denaturation of sunflower globulins in low moisture conditions

    International Nuclear Information System (INIS)

    Rouilly, A.; Orliac, O.; Silvestre, F.; Rigal, L.

    2003-01-01

    DSC analysis in pressure resisting pans of sunflower oil cake makes appear the endothermic peak of sunflower globulins denaturation. Its temperature decreases from 189.5 to 119.9 deg. C while the corresponding enthalpy increases from 2.6 to 3.3 J/g of sample, or from 6.7 to 12.2 J/g of dry protein, when the samples moisture content varies from 0 to 30.0% of the total weight. The plot of the denaturation temperature versus the moisture content is not linear but has a rounded global shape and seems to follow the hydration behavior of the proteins, modeled with the sorption isotherm. As it can be seen on scanning electron microscopy (SEM) micrographs, protein corpuscles 'melt' after such a thermal treatment and large aggregates form by coagulation. Moisture dependence of the 'fusion' temperature of native proteic organization, in low moisture conditions, offers so a new characterization method for the use of vegetable proteins in agro-materials

  3. Thermal denaturation of sunflower globulins in low moisture conditions

    Energy Technology Data Exchange (ETDEWEB)

    Rouilly, A.; Orliac, O.; Silvestre, F.; Rigal, L

    2003-03-05

    DSC analysis in pressure resisting pans of sunflower oil cake makes appear the endothermic peak of sunflower globulins denaturation. Its temperature decreases from 189.5 to 119.9 deg. C while the corresponding enthalpy increases from 2.6 to 3.3 J/g of sample, or from 6.7 to 12.2 J/g of dry protein, when the samples moisture content varies from 0 to 30.0% of the total weight. The plot of the denaturation temperature versus the moisture content is not linear but has a rounded global shape and seems to follow the hydration behavior of the proteins, modeled with the sorption isotherm. As it can be seen on scanning electron microscopy (SEM) micrographs, protein corpuscles 'melt' after such a thermal treatment and large aggregates form by coagulation. Moisture dependence of the 'fusion' temperature of native proteic organization, in low moisture conditions, offers so a new characterization method for the use of vegetable proteins in agro-materials.

  4. Acetic acid denaturing pulsed field capillary electrophoresis for RNA separation.

    Science.gov (United States)

    Li, Zhenqing; Dou, Xiaoming; Ni, Yi; Sumitomo, Keiko; Yamaguchi, Yoshinori

    2010-10-01

    Based on our previous work of in-capillary denaturing polymer electrophoresis, we present a study of RNA molecular separation up to 6.0 kilo nucleotide by pulsed field CE. This is the first systematic investigation of electrophoresis of a larger molecular mass RNA in linear hydroxyethylcellulose (HEC) under pulsed field conditions. The parameters that may influence the separation performance, e.g. gel polymer concentration, modulation depth and pulse frequency, are analyzed in terms of resolution and mobility. For denaturing and separating RNA in the capillary simultaneously, 2 M acetic acid was added into the HEC polymer to serve as separation buffer. Result shows that (i) in pulsed field conditions, RNA separation can be achieved in a wide range of concentration of HEC polymer, and RNA fragments between 0.3 and 0.6 kilo nucleotide are sensitive to the polymer concentration; (ii) under certain pulsed field conditions, RNA fragments move linearly as the modulation depth increases; (iii) 12.5 Hz is the resonance frequency for RNA reorientation time and applied frequency.

  5. Chronic unexplained diarrhea: what to do when the initial workup is negative?

    Science.gov (United States)

    Talley, Nicholas J

    2008-01-01

    Diagnosing the patient with chronic diarrhea can still be a major challenge despite modern testing. It is useful initially to consider what may be the likely underlying pathophysiologic explanation for diarrhea from the history (osmotic, secretory, inflammatory, or motility related) and let this information drive the management algorithm. However, gastroenterologists frequently ask how far the workup should go and when a confident diagnosis of functional diarrhea can be made. In contrast to diarrhea-predominant irritable bowel syndrome, the evidence that functional diarrhea exists as a real diagnostic entity is actually remarkably unconvincing.

  6. Denaturation/Renaturation of Organophosphorus Acid Anhydrolase (OPAA) Using Guanidinium Hydrochloride and Urea

    National Research Council Canada - National Science Library

    Ong, K. K; Sun, Z; Cheng, T. C; Wei, Y; Yuan, J. M; Yin, R

    2004-01-01

    .... Using organophosphorus acid anhydrolase (OPAA) as the model protein, a guanidinium hydrochloride and urea denaturation/renaturation study was conducted and measured both optically and enzymatically...

  7. Denaturation/Renaturation of Organophosphorus Acid Anhydrolase (OPAA) Using Guanidinium Hydrochloride and Urea

    National Research Council Canada - National Science Library

    Ong, K. K; Sun, Z; Cheng, T. C; Wei, Y; Yuan, J. M; Yin, R

    2004-01-01

    ...; thereby indicating conformational changes. Similar results were obtained with circular dichroism as the peak representing the alpha-helix conformation decreased as denaturant concentration was increased...

  8. Visualization of early events in acetic acid denaturation of HIV-1 protease: a molecular dynamics study.

    Directory of Open Access Journals (Sweden)

    Aditi Narendra Borkar

    Full Text Available Protein denaturation plays a crucial role in cellular processes. In this study, denaturation of HIV-1 Protease (PR was investigated by all-atom MD simulations in explicit solvent. The PR dimer and monomer were simulated separately in 9 M acetic acid (9 M AcOH solution and water to study the denaturation process of PR in acetic acid environment. Direct visualization of the denaturation dynamics that is readily available from such simulations has been presented. Our simulations in 9 M AcOH reveal that the PR denaturation begins by separation of dimer into intact monomers and it is only after this separation that the monomer units start denaturing. The denaturation of the monomers is flagged off by the loss of crucial interactions between the α-helix at C-terminal and surrounding β-strands. This causes the structure to transit from the equilibrium dynamics to random non-equilibrating dynamics. Residence time calculations indicate that denaturation occurs via direct interaction of the acetic acid molecules with certain regions of the protein in 9 M AcOH. All these observations have helped to decipher a picture of the early events in acetic acid denaturation of PR and have illustrated that the α-helix and the β-sheet at the C-terminus of a native and functional PR dimer should maintain both the stability and the function of the enzyme and thus present newer targets for blocking PR function.

  9. CT virtual reality in the preoperative workup of malunited distal radius fractures: preliminary results

    International Nuclear Information System (INIS)

    Rieger, Michael; Gruber, Hannes; Jaschke, Werner R.; Gabl, Markus; Mallouhi, Ammar

    2005-01-01

    Our objective was to evaluate the usefulness of CT virtual preoperative planning in the surgical repositioning of malunited distal radius fracture. Eleven patients with malunited distal radius fracture underwent multislice CT of both wrists. A preoperative workup was performed in a virtual reality environment created from the CT data sets. Virtual planning comprised three main procedures, carrying out the virtual osteotomy of the radius, prediction of the final position of the distal radius after osteotomy and computer-assisted manufacturing of a repositioning device, which was later placed at the surgical osteotomy site to reposition objectively the distal radius fragment before fixation with the osteosynthesis. All patients tolerated the surgical procedure well. During surgery, the orthopedic surgeons were not required in any of the cases to alter the position of the distal radius that was determined by the repositioning device. At postoperative follow-up, the anatomic relationship of the distal radius was restored (radial inclination, 21.4 ; volar tilt, 10.3 ; ulnar variance, 0.5 mm). Clinically, a significant improvement of pronation (P=0.012), supination (P=0.01), flexion (P=0.001) and extension (P=0.006) was achieved. Pain decreased from 54 to 7 points. CT virtual reality is a valuable adjunct for the preoperative workup and surgical reposition of malunited distal radius fractures. (orig.)

  10. Non-traumatic cortical subarachnoid haemorrhage: diagnostic work-up and aetiological background

    Energy Technology Data Exchange (ETDEWEB)

    Spitzer, C.; Kosinski, C.M. [University Hospital of RWTH Aachen, Department of Neurology, Aachen (Germany); Mull, M. [University Hospital of RWTH Aachen, Department of Neuroradiology, Aachen (Germany); Rohde, V. [University Hospital of RWTH Aachen, Department of Neurosurgery, Aachen (Germany)

    2005-07-01

    Only 15% of all subarachnoid haemorrhages (SAHs) are not of aneurysmal origin. Among those, circumscribed SAHs along the cortical convexity are rare and have only been described in singular case reports so far. Here, we present a collection of 12 cases of SAH along the convexity, of non-traumatic origin. Over a period of 10 years, 12 cases of circumscribed SAH along the convexity were identified at our clinic. The clinical presentations, neuroradiological SAH characteristics, further diagnostic work-up to identify the underlying aetiologies, the therapy and clinical outcome were analysed. The patients' chief complaints were unspecific cephalgia, focal or generalised seizures and focal neurological deficits. Typical signs of basal SAH, such as nuchal rigidity, thunderclap-headache or alteration of consciousness, were rare. Magnetic resonance imaging (MRI) and digital subtraction angiography (DSA) revealed different aetiologies, namely postpartal posterior encephalopathy (three), cerebral vasculitis (two), dural sinus thrombosis (two), cortical venous thrombosis (one), intracerebral abscesses (one) and cerebral cavernoma (one). Two cases remained unresolved. Treatment of the underlying disease and symptomatic medication led to good clinical outcome in almost all cases. On the basis of these findings, we demonstrate that the clinical presentation, localisation and aetiology of cortical SAH differ clearly from other SAHs. A diagnostic work-up with MRI and eventually DSA is essential. Mostly, the causative disease can be identified, and specific treatment allows a favourable outcome. (orig.)

  11. Value of flexible bronchoscopy in the pre-operative work-up of solitary pulmonary nodules.

    Science.gov (United States)

    Schwarz, Carsten; Schönfeld, Nicolas; Bittner, Roland C; Mairinger, Thomas; Rüssmann, Holger; Bauer, Torsten T; Kaiser, Dirk; Loddenkemper, Robert

    2013-01-01

    The diagnostic value of flexible bronchoscopy in the pre-operative work-up of solitary pulmonary nodules (SPN) is still under debate among pneumologists, radiologists and thoracic surgeons. In a prospective observational manner, flexible bronchoscopy was routinely performed in 225 patients with SPN of unknown origin. Of the 225 patients, 80.5% had lung cancer, 7.6% had metastasis of an extrapulmonary primary tumour and 12% had benign aetiology. Unsuspected endobronchial involvement was found in 4.4% of all 225 patients (or in 5.5% of patients with lung cancer). In addition, flexible bronchoscopy clarified the underlying aetiology in 41% of the cases. The bronchoscopic biopsy results from the SPN were positive in 84 (46.5%) patients with lung cancer. Surgery was cancelled due to the results of flexible bronchoscopy in four cases (involvement of the right main bronchus (impaired pulmonary function did not allow pneumonectomy) n=1, small cell lung cancer n=1, bacterial pneumonia n=2), and the surgical strategy had to be modified to bilobectomy in one patient. Flexible bronchoscopy changed the planned surgical approach in five cases substantially. These results suggest that routine flexible bronchoscopy should be included in the regular pre-operative work-up of patients with SPN.

  12. Geriatric work-up in the Nordic countries. The Nordic approach to comprehensive geriatric assessment

    DEFF Research Database (Denmark)

    Sletvold, O; Tilvis, R; Jonsson, A

    1996-01-01

    A group established by the Nordic professors of geriatrics has developed a position document presenting a shared and updated review of geriatric work-up as a way of comprehensive geriatric assessment in the Nordic countries. The main intention is that the document will serve as support and help f...... languages, and the translated versions should ideally have been subjected to validity and reliability testing. However, so far no scale meets these demands regarding all the five Nordic languages.......A group established by the Nordic professors of geriatrics has developed a position document presenting a shared and updated review of geriatric work-up as a way of comprehensive geriatric assessment in the Nordic countries. The main intention is that the document will serve as support and help...... for the clinician concerned with hospital based geriatric medicine. It may also be useful for quality control and teaching. Not least, it may be useful for health professionals other than geriatricians. To some extent, the position of geriatric medicine in the Nordic countries varies between the countries. However...

  13. Irreversible denaturation of maltodextrin glucosidase studied by differential scanning calorimetry, circular dichroism, and turbidity measurements.

    Science.gov (United States)

    Goyal, Megha; Chaudhuri, Tapan K; Kuwajima, Kunihiro

    2014-01-01

    Thermal denaturation of Escherichia coli maltodextrin glucosidase was studied by differential scanning calorimetry, circular dichroism (230 nm), and UV-absorption measurements (340 nm), which were respectively used to monitor heat absorption, conformational unfolding, and the production of solution turbidity. The denaturation was irreversible, and the thermal transition recorded at scan rates of 0.5-1.5 K/min was significantly scan-rate dependent, indicating that the thermal denaturation was kinetically controlled. The absence of a protein-concentration effect on the thermal transition indicated that the denaturation was rate-limited by a mono-molecular process. From the analysis of the calorimetric thermograms, a one-step irreversible model well represented the thermal denaturation of the protein. The calorimetrically observed thermal transitions showed excellent coincidence with the turbidity transitions monitored by UV-absorption as well as with the unfolding transitions monitored by circular dichroism. The thermal denaturation of the protein was thus rate-limited by conformational unfolding, which was followed by a rapid irreversible formation of aggregates that produced the solution turbidity. It is thus important to note that the absence of the protein-concentration effect on the irreversible thermal denaturation does not necessarily means the absence of protein aggregation itself. The turbidity measurements together with differential scanning calorimetry in the irreversible thermal denaturation of the protein provided a very effective approach for understanding the mechanisms of the irreversible denaturation. The Arrhenius-equation parameters obtained from analysis of the thermal denaturation were compared with those of other proteins that have been reported to show the one-step irreversible thermal denaturation. Maltodextrin glucosidase had sufficiently high kinetic stability with a half-life of 68 days at a physiological temperature (37°C).

  14. Uranium production in thorium/denatured uranium fueled PWRs

    International Nuclear Information System (INIS)

    Arthur, W.B.

    1977-01-01

    Uranium-232 buildup in a thorium/denatured uranium fueled pressurized water reactor, PWR(Th), was studied using a modified version of the spectrum-dependent zero dimensional depletion code, LEOPARD. The generic Combustion Engineering System 80 reactor design was selected as the reactor model for the calculations. Reactors fueled with either enriched natural uranium and self-generated recycled uranium or uranium from a thorium breeder and self-generated recycled uranium were considered. For enriched natural uranium, concentrations of 232 U varied from about 135 ppM ( 232 U/U weight basis) in the zeroth generation to about 260 ppM ( 232 U/U weight basis) at the end of the fifth generation. For the case in which thorium breeder fuel (with its relatively high 232 U concentration) was used as reactor makeup fuel, concentrations of 232 U varied from 441 ppM ( 232 U/U weight basis) at discharge from the first generation to about 512 ppM ( 232 U/U weight basis) at the end of the fifth generation. Concentrations in freshly fabricated fuel for this later case were 20 to 35% higher than the discharge concentration. These concentrations are low when compared to those of other thorium fueled reactor types (HTGR and MSBR) because of the relatively high 238 U concentration added to the fuel as a denaturant. Excellent agreement was found between calculated and existing experimental values. Nevertheless, caution is urged in the use of these values because experimental results are very limited, and the relevant nuclear data, especially for 231 Pa and 232 U, are not of high quality

  15. The generation of denatured reactor plutonium by different options of the fuel cycle

    Energy Technology Data Exchange (ETDEWEB)

    Broeders, C.H.M.; Kessler, G. [Inst. for Neutron Physics and Reactor Technology, Research Center Karlsruhe (Germany)

    2006-11-15

    Denatured (proliferation resistant) reactor plutonium can be generated in a number of different fuel cycle options. First denatured reactor plutonium can be obtained if, instead of low enriched U-235 PWR fuel, re-enriched U-235/U-236 from reprocessed uranium is used (fuel type A). Also the envisaged existing 2,500 t of reactor plutonium (being generated world wide up to the year 2010), mostly stored in intermediate fuel storage facilities at present, could be converted during a transition phase into denatured reactor plutonium by the options fuel type B and D. Denatured reactor plutonium could have the same safeguards standard as present low enriched (<20% U-235) LWR fuel. It could be incinerated by recycling once or twice in PWRs and subsequently by multi-recycling in FRs (CAPRA type or IFRs). Once denatured, such reactor plutonium could remain denatured during multiple recycling. In a PWR, e.g., denatured reactor plutonium could be destroyed at a rate of about 250 kg/GWey. While denatured reactor plutonium could be recycled and incinerated under relieved IAEA safeguards, neptunium would still have to be monitored by the IAEA in future for all cases in which considerable amounts of neptunium are produced. (orig.)

  16. Heat denaturation of soy glycinin : Influence of pH and ionic strength on molecular structure

    NARCIS (Netherlands)

    Lakemond, C.M.M.; Jongh, de H.H.J.; Hessing, M.; Gruppen, H.; Voragen, A.G.J.

    2000-01-01

    The 7S/11S glycinin equilibrium as found in Lakemond et al. (J. Agric. Food Chem. 2000, 48, xxxx-xxxx) at ambient temperatures influences heat denaturation. It is found that the 7S form of glycinin denatures at a lower temperature than the 11S form, as demonstrated by a combination of calorimetric

  17. 27 CFR 20.148 - Manufacture of articles with completely denatured alcohol.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Manufacture of articles with completely denatured alcohol. 20.148 Section 20.148 Alcohol, Tobacco Products and Firearms ALCOHOL... ALCOHOL AND RUM Sale and Use of Completely Denatured Alcohol § 20.148 Manufacture of articles with...

  18. Cation-Induced Stabilization and Denaturation of DNA Origami Nanostructures in Urea and Guanidinium Chloride.

    Science.gov (United States)

    Ramakrishnan, Saminathan; Krainer, Georg; Grundmeier, Guido; Schlierf, Michael; Keller, Adrian

    2017-11-01

    The stability of DNA origami nanostructures under various environmental conditions constitutes an important issue in numerous applications, including drug delivery, molecular sensing, and single-molecule biophysics. Here, the effect of Na + and Mg 2+ concentrations on DNA origami stability is investigated in the presence of urea and guanidinium chloride (GdmCl), two strong denaturants commonly employed in protein folding studies. While increasing concentrations of both cations stabilize the DNA origami nanostructures against urea denaturation, they are found to promote DNA origami denaturation by GdmCl. These inverse behaviors are rationalized by a salting-out of Gdm + to the hydrophobic DNA base stack. The effect of cation-induced DNA origami denaturation by GdmCl deserves consideration in the design of single-molecule studies and may potentially be exploited in future applications such as selective denaturation for purification purposes. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  19. Interim assessment of the denatured 233U fuel cycle: feasibility and nonproliferation characteristics

    International Nuclear Information System (INIS)

    Abbott, L.S.; Bartine, D.E.; Burns, T.J.

    1979-12-01

    A fuel cycle that employs 233 U denatured with 238 U and mixed with thorium fertile material is examined with respect to its proliferation-resistance characteristics and its technical and economic feasibility. The rationale for considering the denatured 233 U fuel cycle is presented, and the impact of the denatured fuel on the performance of Light-Water Reactors, Spectral-Shift-Controlled Reactors, Gas-Cooled Reactors, Heavy-Water Reactors, and Fast Breeder Reactors is discussed. The scope of the R, D and D programs to commercialize these reactors and their associated fuel cycles is also summarized and the resource requirements and economics of denatured 233 U cycles are compared to those of the conventional Pu/U cycle. In addition, several nuclear power systems that employ denatured 233 U fuel and are based on the energy center concept are evaluated

  20. Use of anionic denaturing detergents to purify insoluble proteins after overexpression

    Directory of Open Access Journals (Sweden)

    Schlager Benjamin

    2012-12-01

    Full Text Available Background Many proteins form insoluble protein aggregates, called “inclusion bodies”, when overexpressed in E. coli. This is the biggest obstacle in biotechnology. Ever since the reversible denaturation of proteins by chaotropic agents such as urea or guanidinium hydrochloride had been shown, these compounds were predominantly used to dissolve inclusion bodies. Other denaturants exist but have received much less attention in protein purification. While the anionic, denaturing detergent sodiumdodecylsulphate (SDS is used extensively in analytical SDS-PAGE, it has rarely been used in preparative purification. Results Here we present a simple and versatile method to purify insoluble, hexahistidine-tagged proteins under denaturing conditions. It is based on dissolution of overexpressing bacterial cells in a buffer containing sodiumdodecylsulfate (SDS and whole-lysate denaturation of proteins. The excess of detergent is removed by cooling and centrifugation prior to affinity purification. Host- and overexpressed proteins do not co-precipitate with SDS and the residual concentration of detergent is compatible with affinity purification on Ni/NTA resin. We show that SDS can be replaced with another ionic detergent, Sarkosyl, during purification. Key advantages over denaturing purification in urea or guanidinium are speed, ease of use, low cost of denaturant and the compatibility of buffers with automated FPLC. Conclusion Ionic, denaturing detergents are useful in breaking the solubility barrier, a major obstacle in biotechnology. The method we present yields detergent-denatured protein. Methods to refold proteins from a detergent denatured state are known and therefore we propose that the procedure presented herein will be of general application in biotechnology.

  1. [Functional childhood gastrointestinal disorders. III. Constipation and solitary encopresis; diagnostic work-up and therapy].

    Science.gov (United States)

    van Ginkel, R; Büller, H A; Heymans, H S; Taminiau, J A; Boeckxstaens, G E; Benninga, M A

    2003-06-28

    A detailed medical history in combination with a thorough physical examination, including rectal examination, form the cornerstone in the diagnostic work-up for children with functional defecation disorders. Additional investigations are often not informative and have only minor diagnostic or therapeutic implications. Medical therapy in children with functional constipation and solitary encopresis is primarily based on clinical experience. In both patient groups, the role of education, the use of diary cards and toilet training is important. In some patients behaviour interventions are important. Oral laxatives are the basis of treatment of children with functional constipation, whereas they are contra-indicated in children with solitary encopresis. In both groups, biofeedback training appears to be of little additional benefit. Long-term follow-up of children with functional defecation disorders shows that complaints continue far beyond puberty in many children.

  2. Effects of Diagnostic Work-Up on Medical Decision-Making for Canine Urinary Tract Infection

    DEFF Research Database (Denmark)

    Sørensen, T M; Bjørnvad, C R; Cordoba, G

    2018-01-01

    dipstick (99%), microscopic examination of urine (80%) and bacterial culture (56%). Fifty-one percent of dogs had urinary tract infection (UTI) based on reference QBC. Appropriate DTT was made for 62% of the dogs, while 36% were over-prescribed and 2% under-prescribed. Inappropriate use of second......-line agents was found in 57% of the UTI cases. Performing microscopy-but not culture-significantly impacted DTT (P = 0.039) while no difference was seen in COT (P = 0.67). The accuracy of in-house microscopy and culture were 64.5 and 77%, respectively. CONCLUSIONS AND CLINICAL IMPORTANCE: Over......-prescription of antibiotics was common among dogs with suspected UTI, regardless of the diagnostic work-up performed. Test inaccuracy under practice conditions and incoherence between diagnostic test results and decision-making both explained inappropriate and unnecessary use of antibiotics....

  3. Staging of untreated nasopharyngeal carcinoma with PET/CT: comparison with conventional imaging work-up

    International Nuclear Information System (INIS)

    Ng, Shu-Hang; Chan, Sheng-Chieh; Yen, Tzu-Chen; Liu, Feng-Yuan; Chang, Joseph Tung-Chieh; Fan, Kang-Hsing; Liao, Chun-Ta; Ko, Sheung-Fat; Chin, Shu-Chyn; Hsu, Cheng-Lung

    2009-01-01

    We prospectively compared PET/CT and conventional imaging for initial staging of nasopharyngeal carcinoma (NPC). A total of 111 patients with histologically proven NPC were investigated with PET/CT and conventional imaging (head-and-neck MRI, chest X-ray, abdominal ultrasound, and bone scan) before treatment. The respective findings were reviewed independently and then compared with each other. With regard to T staging, PET/CT showed a discrepancy with head-and-neck MRI in 36 (32.4%) of the study subjects. With regard to N staging, PET/CT showed a discrepancy with head-and-neck MRI in 15 (13.5%) patients. Among the discordant cases, MRI was superior in demonstrating tumor involvement in the parapharyngeal space, skull base, intracranial area, sphenoid sinus, and retropharyngeal nodes while PET/CT was superior in demonstrating neck nodal metastasis. PET/CT disclosed 13 of 16 patients with distant malignancy compared with four patients disclosed by conventional imaging work-up. The false-positive rate of PET/CT was 18.8%. PET/CT correctly modified M staging in eight patients (7.2%) and disclosed a second primary lung malignancy in one patient (0.9%). In NPC patients, MRI appears to be superior to PET/CT for the assessment of locoregional invasion and retropharyngeal nodal metastasis. PET/CT is more accurate than MRI for determining cervical nodal metastasis and should be the better reference for the neck status. PET/CT has an acceptable diagnostic yield and a low false-positive rate for the detection of distant malignancy and can replace conventional work-up to this aim. PET/CT and head-and-neck MRI are suggested for the initial staging of NPC patients. (orig.)

  4. How to tackle tremor – systematic review of the literature and diagnostic work-up

    Directory of Open Access Journals (Sweden)

    Arthur W.G. Buijink

    2012-10-01

    Full Text Available BackgroundTremor is the most prevalent movement disorder in clinical practice. It is defined as involuntary, rhythmic, oscillatory movements. The diagnostic process of patients with tremor can be laborious and challenging, and a clear, systematic overview of available diagnostic techniques is lacking. Tremor can be a symptom of many diseases, but can also represent a distinct disease entity.ObjectiveThe objective of this review is to give a clear, systematic and step-wise overview of the diagnostic work-up of a patient with tremor. The clinical relevance and value of available laboratory tests in patients with tremor will be explored.MethodsWe systematically searched through EMBASE. The retrieved articles were supplemented by articles containing relevant data or provided important background information. Studies that were included investigated the value and/or usability of diagnostic tests for tremor.ResultsIn most patients, history and clinical examination by an experienced movement disorders neurologist are sufficient to establish a correct diagnosis, and further ancillary examinations will not be needed. Ancillary investigation should always be guided by tremor type(s present and other associated signs and symptoms. The main ancillary examination techniques currently are electromyography and SPECT imaging. Unfortunately, many techniques have not been studied in large prospective, diagnostic studies to be able to determine important variables like sensitivity and specificity.ConclusionWhen encountering a patient with tremor, history and careful clinical examination should guide the diagnostic process. Adherence to the diagnostic work-up provided in this review will help the diagnostic process of these patients.

  5. Membranous Nephropathy and Anti-Podocytes Antibodies: Implications for the Diagnostic Workup and Disease Management.

    Science.gov (United States)

    Pozdzik, Agnieszka; Brochériou, Isabelle; David, Cristina; Touzani, Fahd; Goujon, Jean Michel; Wissing, Karl Martin

    2018-01-01

    The discovery of circulating antibodies specific for native podocyte antigens has transformed the diagnostic workup and greatly improved management of idiopathic membranous nephropathy (iMN). In addition, their identification has clearly characterized iMN as a largely autoimmune disorder. Anti-PLA2R1 antibodies are detected in approximately 70% to 80% and anti-THSD7A antibodies in only 2% of adult patients with iMN. The presence of anti-THSD7A antibodies is associated with increased risk of malignancy. The assessment of PLA2R1 and THSD7A antigen expression in glomerular immune deposits has a better sensitivity than measurement of the corresponding autoantibodies. Therefore, in the presence of circulating anti-podocytes autoantibodies and/or enhanced expression of PLA2R1 and THSD7A antigens MN should be considered as primary MN (pMN). Anti-PLA2R1 or anti-THSD7A autoantibodies have been proposed as biomarkers of autoimmune disease activity and their blood levels should be regularly monitored in pMN to evaluate disease activity and predict outcomes. We propose a revised clinical workup flow for patients with MN that recommends assessment of kidney biopsy for PLA2R1 and THSD7A antigen expression, screening for circulating anti-podocytes antibodies, and assessment for secondary causes, especially cancer, in patients with THSD7A antibodies. Persistence of anti-podocyte antibodies for 6 months or their increase in association with nephrotic proteinuria should lead to the introduction of immunosuppressive therapies. Recent data have reported the efficacy and safety of new specific therapies targeting B cells (anti-CD20 antibodies, inhibitors of proteasome) in pMN which should lead to an update of currently outdated treatment guidelines.

  6. In the workup of patients with obscure gastrointestinal bleed, does 64-slice MDCT have a role?

    International Nuclear Information System (INIS)

    Kulkarni, Chinmay; Moorthy, Srikanth; Sreekumar, KP; Rajeshkannan, R; Nazar, PK; Sandya, CJ; Sivasubramanian, S; Ramchandran, PV

    2012-01-01

    The purpose was to prospectively determine the sensitivity of 64-slice MDCT in detecting and diagnosing the cause of obscure gastrointestinal bleed (OGIB). Our study included 50 patients (male 30, female 20) in the age range of 3–82 years (average age: 58.52 years) who were referred to our radiology department as part of their workup for clinically evident gastrointestinal (GI) bleed or as part of workup for anemia (with and without positive fecal occult blood test). All patients underwent conventional upper endoscopy and colonoscopy before undergoing CT scan. Following a noncontrast scan, all patients underwent triple-phase contrast CT scan using a 64-slice CT scan system. The diagnostic performance of 64-slice MDCT was compared to the results of capsule endoscopy, 99m-technetium-labeled red blood cell scintigraphy (99mTc-RBC scintigraphy), digital subtraction angiography, and surgery whenever available. CT scan showed positive findings in 32 of 50 patients. The sensitivity, specificity, positive predictive value, and negative predictive values of MDCT for detection of bleed were 72.2%, 42.8%, 81.2%, and 44.4%, respectively. Capsule endoscopy was done in 15 patients and was positive in 10 patients; it had a sensitivity of 71.4%. Eleven patients had undergone 99mTc-RBC scintigraphy prior to CT scan, and the result was positive in seven patients (sensitivity 70%). Digital subtraction angiography was performed in only eight patients and among them all except one patient showed findings consistent with the lesions detected on MDCT. MDCT is a sensitive and noninvasive tool that allows rapid detection and localization of OGIB. It can be used as the first-line investigation in patients with negative endoscopy and colonoscopy studies. MDCT and capsule endoscopy have complementary roles in the evaluation of OGIB

  7. Calorimetric studies of the thermal denaturation of cytochrome c peroxidase

    International Nuclear Information System (INIS)

    Kresheck, G.C.; Erman, J.E.

    1988-01-01

    Two endotherms are observed by differential scanning calorimetry during the thermal denaturation of cytochrome c peroxidase at pH 7.0. The transition midpoint temperatures (t/sub m/) were 43.9 +- 1.4 and 63.3 +- 1.6 0 C, independent of concentration. The two endotherms were observed at all pH values between 4 and 8, with the transition temperatures varying with pH. Precipitation was observed between pH 4 and 6, and only qualitative data are presented for this region. The thermal unfolding of cytochrome c peroxidase was sensitive to the presence and ligation state of the heme. Only a single endotherm was observed for the unfolding of the apoprotein, and this transition was similar to the high-temperature transition in the holoenzyme. Addition of KCN to the holoenzyme increases the midpoint of the high-temperature transition whereas the low-temperature transition was increased upon addition of KF. Binding of the natural substrate ferricytochrome c to the enzyme increases the low-temperature transition by 4.8 +- 1.3 0 C but has no effect on the high-temperature transition at pH 7. The presence of cytochrome c peroxidase decreases the stability of cytochrome c, and both proteins appear to unfold simultaneously. The results are discussed in terms of the two domains evident in the X-ray crystallographic structure of cytochrome c peroxidase

  8. Renaturation of the androgen receptor after denaturation in SDS

    International Nuclear Information System (INIS)

    Johnson, M.P.; Young, C.Y.F.; Rowley, D.R.; Tindall, D.J.

    1986-01-01

    Renaturation of the steroid binding activity of receptor proteins is a potentially useful tool for their purification and analysis. Cytosol was prepared from rat Dunning prostate tumor in buffer containing molybdate and then denatured by addition of SDS buffer and heating. Aliquots were precipitated in cold acetone and the resulting pellets were washed and solubilized with a small volume of solution containing a chaotropic agent such as 6M guanidine, 8M urea, or 5M sodium iodide. After a 20-minute incubation, samples were diluted 20-fold with buffer containing 4nM [ 3 H]dihydrotestosterone with or without excess unlabeled dihydrotestosterone. Diluted samples were incubated at 0 0 C for varying periods of time prior to assay of bound radioactivity using hydroxylapatite. A time-course of renaturation after exposure to guanidine showed a steady increase of specific binding activity during the first 7 hrs post-dilution that remained stable up to 22 hrs. Experiments with guanidine consistently demonstrated that 25-50% of binding activity was recoverable. Preliminary results using urea or sodium iodide were similar. Efforts to optimize recovery and to characterize the renatured androgen receptor are in progress

  9. Thermally responsive silicon nanowire arrays for native/denatured-protein separation

    International Nuclear Information System (INIS)

    Wang Hongwei; Wang Yanwei; Yuan Lin; Wang Lei; Yang Weikang; Wu Zhaoqiang; Li Dan; Chen Hong

    2013-01-01

    We present our findings of the selective adsorption of native and denatured proteins onto thermally responsive, native-protein resistant poly(N-isopropylacrylamide) (PNIPAAm) decorated silicon nanowire arrays (SiNWAs). The PNIPAAm–SiNWAs surface, which shows very low levels of native-protein adsorption, favors the adsorption of denatured proteins. The amount of denatured-protein adsorption is higher at temperatures above the lower critical solution temperature (LCST) of PNIPAAm. Temperature cycling surrounding the LCST, which ensures against thermal denaturation of native proteins, further increases the amount of denatured-protein adsorption. Moreover, the PNIPAAm–SiNWAs surface is able to selectively adsorb denatured protein even from mixtures of different protein species; meanwhile, the amount of native proteins in solution is kept nearly at its original level. It is believed that these results will not only enrich current understanding of protein interactions with PNIPAAm-modified SiNWAs surfaces, but may also stimulate applications of PNIPAAm–SiNWAs surfaces for native/denatured protein separation. (paper)

  10. 27 CFR 19.41 - Claims on spirits, denatured spirits, articles, or wines lost or destroyed in bond.

    Science.gov (United States)

    2010-04-01

    ..., denatured spirits, articles, or wines lost or destroyed in bond. 19.41 Section 19.41 Alcohol, Tobacco... DISTILLED SPIRITS PLANTS Taxes Claims § 19.41 Claims on spirits, denatured spirits, articles, or wines lost..., relating to the destruction or loss of spirits, denatured spirits, articles, or wines in bond, shall be...

  11. Urea-temperature phase diagrams capture the thermodynamics of denatured state expansion that accompany protein unfolding

    Science.gov (United States)

    Tischer, Alexander; Auton, Matthew

    2013-01-01

    We have analyzed the thermodynamic properties of the von Willebrand factor (VWF) A3 domain using urea-induced unfolding at variable temperature and thermal unfolding at variable urea concentrations to generate a phase diagram that quantitatively describes the equilibrium between native and denatured states. From this analysis, we were able to determine consistent thermodynamic parameters with various spectroscopic and calorimetric methods that define the urea–temperature parameter plane from cold denaturation to heat denaturation. Urea and thermal denaturation are experimentally reversible and independent of the thermal scan rate indicating that all transitions are at equilibrium and the van't Hoff and calorimetric enthalpies obtained from analysis of individual thermal transitions are equivalent demonstrating two-state character. Global analysis of the urea–temperature phase diagram results in a significantly higher enthalpy of unfolding than obtained from analysis of individual thermal transitions and significant cross correlations describing the urea dependence of and that define a complex temperature dependence of the m-value. Circular dichroism (CD) spectroscopy illustrates a large increase in secondary structure content of the urea-denatured state as temperature increases and a loss of secondary structure in the thermally denatured state upon addition of urea. These structural changes in the denatured ensemble make up ∼40% of the total ellipticity change indicating a highly compact thermally denatured state. The difference between the thermodynamic parameters obtained from phase diagram analysis and those obtained from analysis of individual thermal transitions illustrates that phase diagrams capture both contributions to unfolding and denatured state expansion and by comparison are able to decipher these contributions. PMID:23813497

  12. Protein denaturation and functional properties of Lenient Steam Injection heat treated whey protein concentrate

    DEFF Research Database (Denmark)

    Dickow, Jonatan Ahrens; Kaufmann, Niels; Wiking, Lars

    2012-01-01

    Whey protein concentrate (WPC) was heat treated by use of the novel heat treatment method of Lenient Steam Injection (LSI) to elucidate new functional properties in relation to heat-induced gelation of heat treated WPC. Denaturation was measured by both DSC and FPLC, and the results of the two...... methods were highly correlated. Temperatures of up to 90 °C were applicable using LSI, whereas only 68 °C could be reached by plate heat exchange before coagulation/fouling. Denaturation of whey proteins increased with increasing heat treatment temperature up to a degree of 30–35% denaturation at 90 °C...

  13. Optimizing diagnostic workup in the DRG environment: Dynamic algorithms and minimizing radiologic costs may cost your hospital money

    International Nuclear Information System (INIS)

    Saint-Louis, L.A.; Henschke, C.I.; Balter, S.; Whalen, J.P.; Balter, P.

    1987-01-01

    In certain diagnosis-related group (DRG) categories, the availability of sufficient CT scanners or of new equipment, such as MR equipment, can expedite the definitive workup. This will reduce the average length of stay and hospital cost. We analyzed the total hospital and radiologic charges by DRG category for all patients admitted to our hospital in 1985 and 1986. Although the cost per procedure is relatively high, the radiologic component is a small percentage of total hospital costs (median, 3%; maximum, <10%). The authors developed alternative diagnostic algorithms for radiologic-intensive DRG categories. Different diagnostic algorithms proposed for the same clinical problems were compared analytically in terms of impact on the hospital (cost, equipment availability, and length of stay). An example is the workup for FUO. Traditional approach uses plain x-rays and gallium scans and only uses CT when localizing symptoms are present. An alternative approach is to perform CT only. Although more CT examinations would be required, there is considerable reduction in the length of hospital stay and in overall charges. Neurologic and thoracic workups will be given as examples of classes or problems that can be addressed analytically: sequencing of the workup; prevalence; patient population; resource of allocation; and introduction of new imaging modality

  14. Liver CT for vascular mapping during radioembolisation workup : comparison of an early and late arterial phase protocol

    NARCIS (Netherlands)

    van den Hoven, Andor F; Braat, Manon N G J A; Prince, Jip F; van Doormaal, Pieter J; van Leeuwen, Maarten S; Lam, Marnix G E H; van den Bosch, Maurice A A J

    OBJECTIVES: To compare right gastric (RGA) and segment 4 artery (A4) origin detection rates during radioembolisation workup between early and late arterial phase liver CT protocols. METHODS: 100 consecutive patients who underwent liver CT between May 2012-January 2015 with early or late arterial

  15. Contribution of ictal- and interictal brain SPECT to the diagnostic work-up of epileptic patients

    International Nuclear Information System (INIS)

    Dondi, M.; Salgarello, M.; Zoboli, S.; Cidda, C.; Nanni, C.; Rubboli, G.; Meletti, S.; Volpi, L.; Tassinari, C.A.

    2002-01-01

    Aim of the study: We aimed at assessing the contribution of brain SPECT to the diagnostic work-up of patients admitted to the Intensive Epilepsy Monitoring Unit (IEMU) by evaluating concordance of SPECT results with clinical diagnosis (DX) at IEMU admittance (Adm-DX) and at hospital discharge (Disch-DX). Materials and methods: 48 consecutive patients were enrolled in this study and submitted to both ictal and inter-ictal brain SPECT, carried out by means of a three-head system. Before ictal studies, patients were video-EEG monitored in the IEMU. 740 MBq of Tc99m ECD were injected during seizures and imaging performed within 45-60 minutes. For interictal studies, injection was given after at least a 24-hours seizure-free interval. Slices were reconstructed along the orbito-meatal line as well as along the temporal cut. Possible epileptic foci were identified by visual comparison of ictal and interictal studies. Clinical data: Out of the 48 cases, 27 were diagnosed as temporal lobe epilepsies (TLE). Of these, 15 had an Adm-DX of lateralization, whereas 12 were not lateralized according to standardized clinical and EEG criteria. Frontal lobe epilepsy (FLE) was diagnosed in 11 patients, another group of 3 was classified as cryptogenetic (CRYP), while the remaining 7 cases did not fall into any of the previous groups (OTH). SPECT results: in TLE, ictal/interictal SPECT results were congruent with Adm-Dx in 13/27 cases (48%) whereas congruence was found with 25 out of 27 (92%) of Disch-DX. In FLE, concordance between SPECT and clinical diagnosis remained unchanged (9/11 in comparison to either Adm-DX or Disch-DX). CRYPT patients had no lateralization at Adm-DX, while Dis-DX was concordant with SPECT in 2/3 cases. The group labeled as OTH showed concordance SPECT with Adm-DX in 0/7 cases, as opposed to 5/7 of Disch-DX. On the whole, SPECT results were concordant with Adm-DX in 34/48 cases (71%) but concordance was much higher with Dis-DX (41/48; 85%). Conclusions: Brain

  16. Studying pressure denaturation of a protein by molecular dynamics simulations.

    Science.gov (United States)

    Sarupria, Sapna; Ghosh, Tuhin; García, Angel E; Garde, Shekhar

    2010-05-15

    Many globular proteins unfold when subjected to several kilobars of hydrostatic pressure. This "unfolding-up-on-squeezing" is counter-intuitive in that one expects mechanical compression of proteins with increasing pressure. Molecular simulations have the potential to provide fundamental understanding of pressure effects on proteins. However, the slow kinetics of unfolding, especially at high pressures, eliminates the possibility of its direct observation by molecular dynamics (MD) simulations. Motivated by experimental results-that pressure denatured states are water-swollen, and theoretical results-that water transfer into hydrophobic contacts becomes favorable with increasing pressure, we employ a water insertion method to generate unfolded states of the protein Staphylococcal Nuclease (Snase). Structural characteristics of these unfolded states-their water-swollen nature, retention of secondary structure, and overall compactness-mimic those observed in experiments. Using conformations of folded and unfolded states, we calculate their partial molar volumes in MD simulations and estimate the pressure-dependent free energy of unfolding. The volume of unfolding of Snase is negative (approximately -60 mL/mol at 1 bar) and is relatively insensitive to pressure, leading to its unfolding in the pressure range of 1500-2000 bars. Interestingly, once the protein is sufficiently water swollen, the partial molar volume of the protein appears to be insensitive to further conformational expansion or unfolding. Specifically, water-swollen structures with relatively low radii of gyration have partial molar volume that are similar to that of significantly more unfolded states. We find that the compressibility change on unfolding is negligible, consistent with experiments. We also analyze hydration shell fluctuations to comment on the hydration contributions to protein compressibility. Our study demonstrates the utility of molecular simulations in estimating volumetric properties

  17. How Chain Length and Charge Affect Surfactant Denaturation of Acyl Coenzyme A Binding Protein (ACBP)

    DEFF Research Database (Denmark)

    Andersen, Kell Kleiner; Otzen, Daniel

    2009-01-01

    maltoside (DDM). The aim has been to determine how surfactant chain length and micellar charge affect the denaturation mechanism. ACBP denatures in two steps irrespective of surfactant chain length, but with increasing chain length, the potency of the denaturant rises more rapidly than the critical micelle......Using intrinsic tryptophan fluorescence, equilibria and kinetics of unfolding of acyl coenzyme A binding protein (ACBP) have been investigated in sodium alkyl sulfate surfactants of different chain length (8-16 carbon atoms) and with different proportions of the nonionic surfactant dodecyl...... constants increases linearly with denaturant concentration below the cmc but declines at higher concentrations. Both shortening chain length and decreasing micellar charge reduce the overall kinetics of unfolding and makes the dependence of unfolding rate constants on surfactant concentration more complex...

  18. On the Use of Potential Denaturing Agents for Ethanol in Direct Ethanol Fuel Cells

    Directory of Open Access Journals (Sweden)

    Domnik Bayer

    2011-01-01

    Full Text Available Acidic or alkaline direct ethanol fuel cells (DEFCs can be a sustainable alternative for power generation if they are fuelled with bio-ethanol. However, in order to keep the fuel cheap, ethanol has to be exempted from tax on spirits by denaturing. In this investigation the potential denaturing agents fusel oil, tert-butyl ethyl ether, and Bitrex were tested with regard to their compatibility with fuel cells. Experiments were carried out both in sulphuric acid and potassium hydroxide solution. Beside, basic electrochemical tests, differential electrochemical mass spectrometry (DEMS and fuel cell tests were conducted. It was found that fusel oil is not suitable as denaturing agent for DEFC. However, tert-butyl ethyl ether does not seem to hinder the ethanol conversion as much. Finally, a mixture of tert-butyl ethyl ether and Bitrex can be proposed as promising candidate as denaturing agent for use in acidic and alkaline DEFC.

  19. The expression and proangiogenic effect of nucleolin during the recovery of heat-denatured HUVECs.

    Science.gov (United States)

    Liang, Pengfei; Jiang, Bimei; Lv, Chunliu; Huang, Xu; Sun, Li; Zhang, Pihong; Huang, Xiaoyuan

    2013-10-01

    The present study aims to examine the expression patterns and roles of nucleolin during the recovery of heat-denatured human umbilical vein endothelial cells (HUVECs). Deep partial thickness burn model in Sprague-Dawley rats and the heat denatured cell model (52°C, 35s) were used. The expression of nucleolin was measured using Western blot analysis and real-time PCR. Angiogenesis was assessed using in vitro parameters including endothelial cell proliferation, transwell migration assay, and scratched wound healing. Gene transfection and RNA interference approaches were employed to investigate the roles of nucleolin. Nucleolin mRNA and protein expression showed a time-dependent increase during the recovery of heat-denatured dermis and HUVECs. Heat-denaturation time-dependently promoted cell growth, adhesion, migration, scratched wound healing and formation of tube-like structures in HUVECs. These effects of heat denaturation on endothelial wound healing and formation of tube-like structures were prevented by knockdown of nucleolin, whereas over-expression of nucleolin increased cell growth, migration, and formation of tube-like structures in cultured HUVEC endothelial cells. In addition, we found that the expression of vascular endothelial growth factor (VEGF) increased during the recovery of heat-denatured dermis and HUVECs, and nucleolin up-regulated VEGF in HUVECs. The present study reveals that the expression of nucleolin is up-regulated, and plays a pro-angiogenic role during the recovery of heat-denatured dermis and its mechanism is probably dependent on production of VEGF. We find a novel and important pro-angiogenic role of nucleolin during the recovery of heat-denatured dermis. Copyright © 2013 Elsevier B.V. All rights reserved.

  20. On the Use of Potential Denaturing Agents for Ethanol in Direct Ethanol Fuel Cells

    OpenAIRE

    Domnik Bayer; Florina Jung; Birgit Kintzel; Martin Joos; Carsten Cremers; Dierk Martin; Jörg Bernard; Jens Tübke

    2011-01-01

    Acidic or alkaline direct ethanol fuel cells (DEFCs) can be a sustainable alternative for power generation if they are fuelled with bio-ethanol. However, in order to keep the fuel cheap, ethanol has to be exempted from tax on spirits by denaturing. In this investigation the potential denaturing agents fusel oil, tert-butyl ethyl ether, and Bitrex were tested with regard to their compatibility with fuel cells. Experiments were carried out both in sulphuric acid and potassium hydroxide solution...

  1. Detection of human DNA polymorphisms with a simplified denaturing gradient gel electrophoresis technique.

    OpenAIRE

    Noll, W W; Collins, M

    1987-01-01

    Single base pair differences between otherwise identical DNA molecules can result in altered melting behavior detectable by denaturing gradient gel electrophoresis. We have developed a simplified procedure for using denaturing gradient gel electrophoresis to detect base pair changes in genomic DNA. Genomic DNA is digested with restriction enzymes and hybridized in solution to labeled single-stranded probe DNA. The excess probe is then hybridized to complementary phage M13 template DNA, and th...

  2. The Use of Cardiac Magnetic Resonance Imaging in the Diagnostic Workup and Treatment of Atrial Fibrillation

    Directory of Open Access Journals (Sweden)

    Peter Haemers

    2012-01-01

    Full Text Available Atrial fibrillation (AF is the most common cardiac arrhythmia and imposes a huge clinical and economic burden. AF is correlated with an increased morbidity and mortality, mainly due to stroke and heart failure. Cardiovascular imaging modalities, including echocardiography, computed tomography (CT, and cardiovascular magnetic resonance (CMR, play a central role in the workup and treatment of AF. One of the major advantages of CMR is the high contrast to noise ratio combined with good spatial and temporal resolution, without any radiation burden. This allows a detailed assessment of the structure and function of the left atrium (LA. Of particular interest is the ability to visualize the extent of LA wall injury. We provide a focused review of the value of CMR in identifying the underlying pathophysiological mechanisms of AF, its role in stroke prevention and in the guidance of radiofrequency catheter ablation. CMR is a promising technique that could add valuable information for therapeutic decision making in specific subpopulations with AF.

  3. Spinal cysts. Diagnostic workup and therapy; Spinale Zysten. Diagnostik und Therapie

    Energy Technology Data Exchange (ETDEWEB)

    Simgen, A. [Universitaetsklinikum des Saarlandes, Klinik fuer Diagnostische und Interventionelle Neuroradiologie, Homburg/Saar (Germany)

    2018-02-15

    Spinal cysts can be classified as meningeal, not meningeal, and tumor-associated cysts. Due to the widespread availability of high-resolution computed tomography and magnet resonance imaging, spinal cysts can be detected with high sensitivity these days. Concerning the variety of potential cystic differential diagnoses, a precise classification is difficult and can often only be realized after surgical inspection or histological examination. Spinal cysts are generally incidental findings during a routine diagnostic workup and need no further therapy. Surgical treatment can be necessary if the spinal cyst reaches a certain size and causes neurological symptoms due to the compression of the spinal cord or the nerve root. (orig.) [German] Spinale Zysten koennen in meningeale, nichtmeningeale und tumorassoziierte Zysten eingeteilt werden. Durch die weite Verbreitung von hochaufloesenden Computer- und Magnetresonanztomographen koennen spinale Zysten heutzutage mit einer hohen Sensitivitaet erkannt werden. Eine genaue Klassifikation kann sich unter der Vielzahl der moeglichen zystischen Differenzialdiagnosen schwierig gestalten und ist haeufig nur durch eine chirurgische Inspektion oder die histologische Untersuchung moeglich. Meistens werden spinale Zysten bei der Routinediagnostik als Zufallsbefunde entdeckt und benoetigen keine weitere Therapie. Erreichen sie allerdings eine gewisse Groesse, koennen sie raumfordernd auf das Myelon oder einzelne Nervenwurzeln wirken und somit ausgepraegte neurologische Symptome verursachen. In solchen Faellen ist ein chirurgisches Vorgehen zur Resektion einer spinalen Zyste notwendig. (orig.)

  4. Imaging techniques used in the diagnostic workup of acute venous thromboembolic disease.

    Science.gov (United States)

    Tilve-Gómez, A; Rodríguez-Fernández, P; Trillo-Fandiño, L; Plasencia-Martínez, J M

    Early diagnosis is one of the most important factors affecting the prognosis of pulmonary embolism (PE); however, the clinical presentation of PE is often very unspecific and it can simulate other diseases. For these reasons, imaging tests, especially computed tomography angiography (CTA) of the pulmonary arteries, have become the keystone in the diagnostic workup of PE. The wide availability and high diagnostic performance of pulmonary CTA has led to an increase in the number of examinations done and a consequent increase in the population's exposure to radiation and iodinated contrast material. Thus, other techniques such as scintigraphy and venous ultrasonography of the lower limbs, although less accurate, continue to be used in certain circumstances, and optimized protocols have been developed for CTA to reduce the dose of radiation (by decreasing the kilovoltage) and the dose of contrast agents. We describe the technical characteristics and interpretation of the findings for each imaging technique used to diagnose PE and discuss their advantages and limitations; this knowledge will help the best technique to be chosen for each case. Finally, we comment on some data about the increased use of CTA, its clinical repercussions, its "overuse", and doubts about its cost-effectiveness. Copyright © 2016 SERAM. Publicado por Elsevier España, S.L.U. All rights reserved.

  5. Trends in standard workup performed by pediatric subspecialists for the diagnosis of adolescent polycystic ovary syndrome.

    Science.gov (United States)

    Powers, Sarah E; Uliassi, Nicole W; Sullivan, Shannon D; Tuchman, Lisa K; Mehra, Rinku; Gomez-Lobo, Veronica

    2015-02-01

    The purpose of this study is to identify trends in the clinical workup, diagnosis, and treatment of polycystic ovary syndrome by pediatric endocrinologists, pediatric gynecologists, and adolescent medicine specialists. Retrospective chart review. Tertiary care medical center. Females aged 11-18 y who were evaluated for PCOS from June 2009 to October 2011 were included. Any patients with coexisting diagnoses of other primary etiology for amenorrhea were excluded. Patients were identified by ICD-9 codes for PCOS, hypersecretion of ovarian androgens, irregular menses, hirsutism, oligomenorrhea, or amenorrhea. 261 patients were included: 144 from endocrinology, 9 from gynecology, and 108 from adolescent pediatric practices. There were no significant differences in the androgen labs ordered by the subspecialties. Gynecologists ordered pelvic ultrasonography for 89% (n = 8) of patients, compared to 9% (n = 10) by adolescent medicine specialists and 24% (n = 34) by endocrinologists (P PCOS with metformin (58%, n = 66), compared to gynecologists (14%, n = 1) and adolescent medicine specialists (5%, n = 3) (P PCOS are evident among pediatric subspecialties, reflecting lack of standardized care for adolescents. Quantifying outcomes based on diagnostic and therapeutic approaches are important next steps. Copyright © 2015 North American Society for Pediatric and Adolescent Gynecology. Published by Elsevier Inc. All rights reserved.

  6. Interim assessment of the denatured 233U fuel cycle: feasibility and nonproliferation characteristics

    International Nuclear Information System (INIS)

    Abbott, L.S.; Bartine, D.E.; Burns, T.J.

    1978-12-01

    A fuel cycle that employs 233 U denatured with 238 U and mixed with thorium fertile material is examined with respect to its proliferation-resistance characteristics and its technical and economic feasibility. The rationale for considering the denatured 233 U fuel cycle is presented, and the impact of the denatured fuel on the performance of Light-Water Reactors, Spectral-Shift-Controlled Reactors, Gas-Cooled Reactors, Heavy-Water Reactors, and Fast Breeder Reactors is discussed. The scope of the R, D and D programs to commercialize these reactors and their associated fuel cycles is also summarized and the resource requirements and economics of denatured 233 U cycles are compared to those of the conventional Pu/U cycle. In addition, several nuclear power systems that employ denatured 233 U fuel and are based on the energy center concept are evaluated. Under this concept, dispersed power reactors fueled with denatured or low-enriched uranium fuel are supported by secure energy centers in which sensitive activities of the nuclear cycle are performed. These activities include 233 U production by Pu-fueled transmuters (thermal or fast reactors) and reprocessing. A summary chapter presents the most significant conclusions from the study and recommends areas for future work

  7. Interim assessment of the denatured /sup 233/U fuel cycle: feasibility and nonproliferation characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Abbott, L.S.; Bartine, D.E.; Burns, T.J. (eds.)

    1978-12-01

    A fuel cycle that employs /sup 233/U denatured with /sup 238/U and mixed with thorium fertile material is examined with respect to its proliferation-resistance characteristics and its technical and economic feasibility. The rationale for considering the denatured /sup 233/U fuel cycle is presented, and the impact of the denatured fuel on the performance of Light-Water Reactors, Spectral-Shift-Controlled Reactors, Gas-Cooled Reactors, Heavy-Water Reactors, and Fast Breeder Reactors is discussed. The scope of the R, D and D programs to commercialize these reactors and their associated fuel cycles is also summarized and the resource requirements and economics of denatured /sup 233/U cycles are compared to those of the conventional Pu/U cycle. In addition, several nuclear power systems that employ denatured /sup 233/U fuel and are based on the energy center concept are evaluated. Under this concept, dispersed power reactors fueled with denatured or low-enriched uranium fuel are supported by secure energy centers in which sensitive activities of the nuclear cycle are performed. These activities include /sup 233/U production by Pu-fueled transmuters (thermal or fast reactors) and reprocessing. A summary chapter presents the most significant conclusions from the study and recommends areas for future work.

  8. Mutation of charged residues to neutral ones accelerates urea denaturation of HP-35.

    Science.gov (United States)

    Wei, Haiyan; Yang, Lijiang; Gao, Yi Qin

    2010-09-16

    Following the studies of urea denaturation of β-hairpins using molecular dynamics, in this paper, molecular dynamics simulations of two peptides, a 35 residue three helix bundle villin headpiece protein HP-35 and its doubly norleucine-substituent mutant (Lys24Nle/Lys29Nle) HP-35 NleNle, were undertaken in urea solutions to understand the molecular mechanism of urea denaturation of α-helices. The mutant HP-35 NleNle was found to denature more easily than the wild type. During the expansion of the small hydrophobic core, water penetration occurs first, followed by that of urea molecules. It was also found that the initial hydration of the peptide backbone is achieved through water hydrogen bonding with the backbone CO groups during the denaturation of both polypeptides. The mutation of the two charged lysine residues to apolar norleucine enhances the accumulation of urea near the hydrophobic core and facilitates the denaturation process. Urea also interacts directly with the peptide backbone as well as side chains, thereby stabilizing nonnative conformations. The mechanism revealed here is consistent with the previous study on secondary structure of β-hairpin polypeptide, GB1, PEPTIDE 1, and TRPZIP4, suggesting that there is a general mechanism in the denaturation of protein backbone hydrogen bonds by urea.

  9. 27 CFR 19.32 - Assessment of tax on spirits, denatured spirits, or wines in bond which are lost, destroyed or...

    Science.gov (United States)

    2010-04-01

    ... spirits, denatured spirits, or wines in bond which are lost, destroyed or removed without authorization... spirits, denatured spirits, or wines in bond which are lost, destroyed or removed without authorization. When spirits, denatured spirits, or wines in bond are lost or destroyed (except spirits, denatured...

  10. Myelodysplastic syndromes: advantages of a combined cytogenetic and molecular diagnostic workup.

    Science.gov (United States)

    Ciabatti, Elena; Valetto, Angelo; Bertini, Veronica; Ferreri, Maria Immacolata; Guazzelli, Alice; Grassi, Susanna; Guerrini, Francesca; Petrini, Iacopo; Metelli, Maria Rita; Caligo, Maria Adelaide; Rossi, Simona; Galimberti, Sara

    2017-10-03

    In this study we present a new diagnostic workup for the myelodysplastic syndromes (MDS) including FISH, aCGH, and somatic mutation assays in addition to the conventional cytogenetics (CC). We analyzed 61 patients by CC, FISH for chromosome 5, 7, 8 and PDGFR rearrangements, aCGH, and PCR for ASXL1, EZH2, TP53, TET2, RUNX1, DNMT3A, SF3B1 somatic mutations. Moreover, we quantified WT1 and RPS14 gene expression levels, in order to find their possible adjunctive value and their possible clinical impact. CC analysis showed 32% of patients with at least one aberration. FISH analysis detected chromosomal aberrations in 24% of patients and recovered 5 cases (13.5%) at normal karyotype (two 5q- syndromes, one del(7) case, two cases with PDGFR rearrangement). The aGCH detected 10 "new" unbalanced cases in respect of the CC, including one with alteration of the ETV6 gene. After mutational analysis, 33 patients (54%) presented at least one mutation and represented the only marker of clonality in 36% of all patients. The statistical analysis confirmed the prognostic role of CC either on overall or on progression-free-survival. In addition, deletions detected by aCGH and WT1 over-expression negatively conditioned survival. In conclusion, our work showed that 1) the addition of FISH (at least for chr. 5 and 7) can improve the definition of the risk score; 2) mutational analysis, especially for the TP53 and SF3B1, could better define the type of MDS and represent a "clinical warning"; 3) the aCGH use could be probably applied to selected cases (with suboptimal response or failure).

  11. Acute testicular torsion in children: the role of sonography in the diagnostic workup.

    Science.gov (United States)

    Gunther, P; Schenk, J P; Wunsch, R; Holland-Cunz, S; Kessler, U; Troger, J; Waag, K L

    2006-11-01

    Acute testicular torsion in children is an emergency and has to be diagnosed urgently. Doppler sonography is increasingly used in imaging the acute scrotum. Nevertheless, in uncertain cases, surgical exploration is required. In this study, we attempted to define the role of Doppler sonography in the diagnostic workup of the acutely painful scrotum. All patients admitted between 1999 and 2005 with acute scrotal pain were included. After clinical assessment, patients were imaged by Doppler sonography with a ''high-end'' instrument. In cases of absent arterial perfusion of the testis in Doppler sonography, surgical exploration was carried out. Patients with unaffected perfusion were followed clinically by ultrasound for up to 2 years. Sixty-one infants and children aged 1 day to 17 years (median: 7.9 years) were included. In 14 cases, sonography demonstrated absent central perfusion, with abnormal parenchymal echogenicity in six. Absence of venous blood flow together with reduction of central arterial perfusion was found in one infant. In these 15 patients, surgical exploration confirmed testicular torsion. Among the other 46 patients, we found four cases with increased testicular perfusion and 27 with increased perfusion of the epididymis. In one infant, a testicular tumour was found sonographically, and orchiectomy confirmed diagnosis of a teratoma. Follow-up examinations of the conservatively treated patients showed good clinical outcome with physiologic central perfusion as well as normal echogenic pattern of both testes. No case of testicular torsion was missed. By means of Doppler sonography, an unequivocal statement regarding testicular perfusion was possible in all cases. The initial Doppler diagnosis was confirmed by operative evaluation and follow-up ultrasound. Testicular torsion can therefore be excluded by correctly performed ultrasound with modern equipment.

  12. Y Choromosomal Microdeletion Screening in The Workup of Male Infertility and Its Current Status in India

    Directory of Open Access Journals (Sweden)

    Ramaswamy Suganthi

    2014-01-01

    Full Text Available Spermatogenesis is an essential stage in human male gamete development, which is regulated by many Y chromosome specific genes. Most of these genes are centred in a specific region located on the long arm of the human Y chromosome known as the azoospermia factor region (AZF. Deletion events are common in Y chromosome because of its peculiar structural organization. Astonishingly, among the several known genetic causes of male infertility, Y chromosomal microdeletions emerged as the most frequent structural chromosome anomaly associated with the quantitative reduction of sperm. The development of assisted reproductive techniques (ART like intra-cytoplasmic sperm injection (ICSI and testicular sperm extraction (TESE helps to bypass the natural barriers of fertilization, but it increases the concern about the transmission of genetic defects. Experimental evidence suggested that the men with Y chromosomal microdeletions vertically transmitted their deletion as well as related fertility disorders to their offspring via these ART techniques. In India, infertility is on alarming rise. ART centres have opened up in virtually every state but still most of the infertility centres in India do not choose to perform Y chromosomal microdeletion diagnosis because of some advanced theoretical reasons. Moreover, there is no consensus among the clinicians about the diagnosis and management of Y chromosomal microdeletion defects. The current review discusses thoroughly the role of Y chromosome microdeletion screening in the workup of male infertility, its significance as a diagnostic test, novel approaches for screening Y deletions and finally a systematic review on the current status of Y chromosome microdeletion deletion screening in India.

  13. Is a Double Contrast Barium Enema Necessary in Patients who Undergo a CT Colonography as a Preoperative Work-up?

    International Nuclear Information System (INIS)

    Lee, Ji Won; Lee, Soon Jin; Choi, Dong Il; Kim, Hee Jung; Chun, Ho Kyung; Yun, Seong Hyeon; Jeon, Yong Hwan; Park, Seung Bae; Kim, Min Ju

    2009-01-01

    To determine the efficiency of CT colonography (CTC), as compared with the double contrast barium enema (DCBE) for the preoperative work-up of colorectal neoplasms. A total of 39 patients underwent both a CTC and DCBE before surgery. Three abdominal radiologists and three colorectal surgeons retrospectively reviewed virtual double contrast (VDC) and DCBE images, regarding each examination for localization, conspicuity, extent and morphology of neoplasms. We reviewed the radiological reports of the CTC and DCBE for the polyps. In the case of both VDC and DCBE, 40 cancers were detected in 39 patients. According to abdominal radiologists, VDC showed the same ability to identify DCBE for localization of neoplasms (p 0.05). The performance of the CTC is comparable to the DCBE for the localization of main mass and polyps in patients with colorectal neoplasm. Barium enema don't seem to be necessary for patients who undergo a CTC as preoperative work-up

  14. Diagnostic Workup of Paediatric Inflammatory Bowel Disease Patients In Europe: Results of A 5-Year Audit of The EUROKIDS Registry

    DEFF Research Database (Denmark)

    de Bie, Charlotte I; Buderus, Stephan; Sandhu, Bhupinder K

    2011-01-01

    ), ileocolonoscopy, and (except in cases of definitive ulcerative colitis (UC)) adequate imaging of the small bowel. To audit and evaluate the diagnostic workup of paediatric IBD patients in Europe, the Working Group created EUROKIDS, a prospective, web-based registry of newly diagnosed paediatric IBD patients...... patients were correctly registered. Both OGD and ileocolonoscopy had been performed in 64% of all patients, and increased significantly from year 1 (52 %) to year 5 (71%, p¿...

  15. Improving work-up of the abnormal mammogram through organized assessment: results from the ontario breast screening program.

    Science.gov (United States)

    Quan, May Lynn; Shumak, Rene S; Majpruz, Vicky; Holloway, Claire M D; O'Malley, Frances P; Chiarelli, Anna M

    2012-03-01

    Women with an abnormal screening mammogram should ideally undergo an organized assessment to attain a timely diagnosis. This study evaluated outcomes of women undergoing work-up after abnormal mammogram through a formal breast assessment affiliate (BAA) program with explicit care pathways compared with usual care (UC) using developed quality indicators for screening mammography programs. Between January 1 and December 31, 2007, a total of 320,635 women underwent a screening mammogram through the Ontario Breast Screening Program (OBSP), of whom 25,543 had an abnormal result requiring further assessment. Established indicators assessing timeliness, appropriateness of follow-up, and biopsy rates were compared between women who were assessed through either a BAA or UC using χ(2) analysis. Work-up of the abnormal mammogram for patients screened through a BAA resulted in a greater proportion of women attaining a definitive diagnosis within the recommended time interval when a histologic diagnosis was required. In addition, use of other quality measures including specimen radiography for both core biopsies and surgical specimens and preoperative core needle biopsy was greater in BAA facilities. These findings support future efforts to increase the number of BAAs within the OBSP, because the pathways and reporting methods associated with them result in improvements in our ability to provide timely and appropriate care for women requiring work-up of an abnormal mammogram.

  16. Pseudomonas aeruginosa cytochrome c551 denaturation by five systematic urea derivatives that differ in the alkyl chain length.

    Science.gov (United States)

    Kobayashi, Shinya; Fujii, Sotaro; Koga, Aya; Wakai, Satoshi; Matubayasi, Nobuyuki; Sambongi, Yoshihiro

    2017-07-01

    Reversible denaturation of Pseudomonas aeruginosa cytochrome c 551 (PAc 551 ) could be followed using five systematic urea derivatives that differ in the alkyl chain length, i.e. urea, N-methylurea (MU), N-ethylurea (EU), N-propylurea (PU), and N-butylurea (BU). The BU concentration was the lowest required for the PAc 551 denaturation, those of PU, EU, MU, and urea being gradually higher. Furthermore, the accessible surface area difference upon PAc 551 denaturation caused by BU was found to be the highest, those by PU, EU, MU, and urea being gradually lower. These findings indicate that urea derivatives with longer alkyl chains are stronger denaturants. In this study, as many as five systematic urea derivatives could be applied for the reversible denaturation of a single protein, PAc 551 , for the first time, and the effects of the alkyl chain length on protein denaturation were systematically verified by means of thermodynamic parameters.

  17. Phagocytosis by macrophages mediated by receptors for denatured proteins - dependence on tyrosine protein kinases

    Directory of Open Access Journals (Sweden)

    M.R. Hespanhol

    2002-03-01

    Full Text Available Previous studies have demonstrated that some components of the leukocyte cell membrane, CR3 (Mac-1, CD11b/CD18 and p150/95, are able to bind to denatured proteins. Thus, it is of interest to know which effector functions of these cells can be triggered by these receptors when they interact with particles or surfaces covered with denatured proteins. In the present study we analyzed their possible role as mediators of phagocytosis of red cells covered with denatured bovine serum albumin (BSA by mouse peritoneal macrophages. We observed that a macrophages are able to recognize (bind to these red cells, b this interaction can be inhibited by denatured BSA in the fluid phase, c there is no phagocytosis of these particles by normal macrophages, d phagocytosis mediated by denatured BSA can be, however, effectively triggered in inflammatory macrophages induced by glycogen or in macrophages activated in vivo with LPS, and e this phagocytic capacity is strongly dependent on the activity of tyrosine protein kinases in its signal transduction pathway, as demonstrated by using three kinds of enzyme inhibitors (genistein, quercetin and herbimycin A.

  18. Alcohol-induced structural transitions in the acid-denatured Bacillus licheniformis α-amylase

    Directory of Open Access Journals (Sweden)

    Adyani Azizah Abd Halim

    2017-01-01

    Full Text Available Alcohol-induced structural changes in the acid-denatured Bacillus licheniformis α-amylase (BLA at pH 2.0 were studied by far-ultra violet circular dichroism, intrinsic, three-dimensional and 8-anilino-1-naphthalene sulfonic acid (ANS fluorescence, acrylamide quenching and thermal denaturation. All the alcohols used in this study produced partial refolding in the acid-denatured BLA as evident from the increased mean residue ellipticity at 222 nm, increased intrinsic fluorescence and decreased ANS fluorescence. The order of effectiveness of these alcohols to induce a partially folded state of BLA was found to be: 2,2,2-trifluoroethanol/tert-butanol > 1-propanol/2-propanol > 2-chloroethanol > ethanol > methanol. Three-dimensional fluorescence and acrylamide quenching results obtained in the presence of 5.5 M tert-butanol also suggested formation of a partially folded state in the acid-denatured BLA. However, 5.5 M tert-butanol-induced state of BLA showed a non-cooperative thermal transition. All these results suggested formation of a partially folded state of the acid-denatured BLA in the presence of these alcohols. Furthermore, their effectiveness was found to be guided by their chain length, position of methyl groups and presence of the substituents.

  19. Drying and denaturation characteristics of whey protein isolate in the presence of lactose and trehalose.

    Science.gov (United States)

    Haque, M Amdadul; Chen, Jie; Aldred, Peter; Adhikari, Benu

    2015-06-15

    The denaturation kinetics of whey protein isolate (WPI), in the presence and absence of lactose and trehalose, was quantified in a convective air-drying environment. Single droplets of WPI, WPI-lactose and WPI-trehalose were dried in conditioned air (2.5% RH, 0.5m/s air velocity) at two temperatures (65°C and 80°C) for 500s. The initial solid concentration of these solutions was 10% (w/v) in all the samples. Approximately 68% of WPI was denatured when it was dried in the absence of sugars. Addition of 20% trehalose prevented the irreversible denaturation of WPI at both temperatures. Thirty percent lactose was required to prevent denaturation of WPI at 65°C and the same amount of lactose protected only 70% of WPI from denaturation at 80°C. The secondary structures of WPI were found to be altered by the drying-induced stresses, even in the presence of 20% trehalose and 30% lactose. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Neutronics calculations for denatured molten salt reactors: Assessing resource requirements and proliferation-risk attributes

    International Nuclear Information System (INIS)

    Ahmad, Ali; McClamrock, Edward B.; Glaser, Alexander

    2015-01-01

    Highlights: • We study the proliferation-risk and resource attributes of denatured MSRs. • MSRs offer significantly better resource efficiency compared to light-water reactors. • Denatured single-fluid MSRs reactors offer promising non-proliferation attributes. - Abstract: Molten salt reactors (MSRs) are often advocated as a radical but worthwhile alternative to traditional reactor concepts based on solid fuels. This article builds upon the existing research into MSRs to model and simulate the operation of thorium-fueled single-fluid and two-fluid reactors. The analysis is based on neutronics calculations and focuses on denatured MSR systems. Resource utilization and basic proliferation-risk attributes are compared to those of standard light-water reactors. Depending on specific design choices, even fully denatured reactors could reduce uranium and enrichment requirements by a factor of 3–4. Overall, denatured single-fluid designs appear as the most promising candidate technology minimizing both design complexity and overall proliferation risks despite being somewhat less attractive from the perspective of resource utilization

  1. Feasibility of dynamic MR-hysterosalpingography for the diagnostic work-up of infertile women

    International Nuclear Information System (INIS)

    Winter, Leopold; Gluecker, Thomas; Steinbrich, Wolfgang; Pegios, Wassilios; Steimann, Sabine; De Geyter, Christian; Froehlich, Johannes M.

    2010-01-01

    Background: Tubal disturbances often contribute to infertility. Conventional hysterosalpingography (HSG) is considered as standard in the assessment of the patency of the fallopian tubes, but requires ionizing radiation and is restricted to the imaging of endoluminal structures. Purpose: To evaluate dynamic magnetic resonance-HSG (dMR-HSG) in the diagnostic work-up in patients with infertility. Material and Methods: Thirty-seven consecutive infertile women underwent dMR-HSG: 20 ml of gadolinium-polyvidone solution (18.4 mM Dotarem 1:20 with polyvidone) were injected intracervically through a 5-Charriere balloon catheter while acquiring five consecutive flash-3D T1-weighted MR sequences with fat saturation. Two experienced readers assessed image quality and anatomic-pathologic correlations prospectively. The relevance of results was evaluated in the clinical context of each patient. Patient comfort was evaluated with a standardized questionnaire. Results: dMR-HSG was successfully completed in 33/37 patient with an average study time of 45 min. In 4 of 37 patients the catheter became dislodged during the examination, resulting in two complete diagnostic failures. Failure in another two patients was due to preliminary termination because of excessive pain and discomfort during the application of the contrast solution. The uterine cavity was completely visualized and bilateral fallopian tube patency was confirmed by dMR-HSG in 27 of 33 patients. Bilateral tubal occlusion was diagnosed in one of the remaining six patients and was confirmed by laparoscopy. Successful selective tubal catheterization was performed in one additional patient with unilateral and one patient with bilateral fallopian tube occlusion. In three cases, the catheter became dislocated at the end of the examination without demonstration of tubal patency. Since all three patients refused diagnostic laparoscopy and conventional HSG, possible bilateral occlusions of the fallopian tubes could not be

  2. Feasibility of dynamic MR-hysterosalpingography for the diagnostic work-up of infertile women

    Energy Technology Data Exchange (ETDEWEB)

    Winter, Leopold; Gluecker, Thomas; Steinbrich, Wolfgang; Pegios, Wassilios (Dept. of Radiology, Univ. Hospital Basel (Switzerland)), e-mail: winterl@uhbs.ch; Steimann, Sabine; De Geyter, Christian (Women' s Hospital, Univ. Hospital Basel (Switzerland)); Froehlich, Johannes M. (Guerbet AG, Zuerich (Switzerland))

    2010-07-15

    Background: Tubal disturbances often contribute to infertility. Conventional hysterosalpingography (HSG) is considered as standard in the assessment of the patency of the fallopian tubes, but requires ionizing radiation and is restricted to the imaging of endoluminal structures. Purpose: To evaluate dynamic magnetic resonance-HSG (dMR-HSG) in the diagnostic work-up in patients with infertility. Material and Methods: Thirty-seven consecutive infertile women underwent dMR-HSG: 20 ml of gadolinium-polyvidone solution (18.4 mM Dotarem 1:20 with polyvidone) were injected intracervically through a 5-Charriere balloon catheter while acquiring five consecutive flash-3D T1-weighted MR sequences with fat saturation. Two experienced readers assessed image quality and anatomic-pathologic correlations prospectively. The relevance of results was evaluated in the clinical context of each patient. Patient comfort was evaluated with a standardized questionnaire. Results: dMR-HSG was successfully completed in 33/37 patient with an average study time of 45 min. In 4 of 37 patients the catheter became dislodged during the examination, resulting in two complete diagnostic failures. Failure in another two patients was due to preliminary termination because of excessive pain and discomfort during the application of the contrast solution. The uterine cavity was completely visualized and bilateral fallopian tube patency was confirmed by dMR-HSG in 27 of 33 patients. Bilateral tubal occlusion was diagnosed in one of the remaining six patients and was confirmed by laparoscopy. Successful selective tubal catheterization was performed in one additional patient with unilateral and one patient with bilateral fallopian tube occlusion. In three cases, the catheter became dislocated at the end of the examination without demonstration of tubal patency. Since all three patients refused diagnostic laparoscopy and conventional HSG, possible bilateral occlusions of the fallopian tubes could not be

  3. Nonsurgical Transurethral Radiofrequency Collagen Denaturation: Results at Three Years after Treatment

    Directory of Open Access Journals (Sweden)

    Denise M. Elser

    2011-01-01

    Full Text Available Objective. To assess treatment efficacy and quality of life in women with stress urinary incontinence 3 years after treatment with nonsurgical transurethral radiofrequency collagen denaturation. Methods. This prospective study included 139 women with stress urinary incontinence due to bladder outlet hypermobility. Radiofrequency collagen denaturation was performed using local anesthesia in an office setting. Assessments included incontinence quality of life (I-QOL and urogenital distress inventory (UDI-6 instruments. Results. In total, 139 women were enrolled and 136 women were treated (mean age, 47 years. At 36 months, intent-to-treat analysis (n=139 revealed significant improvements in quality of life. Mean I-QOL score improved 17 points from baseline (P=.0004, while mean UDI-6 score improved (decreased 19 points (P=.0005. Conclusions. Transurethral collagen denaturation is a low-risk, office-based procedure that results in durable quality-of-life improvements in a significant proportion of women for as long as 3 years.

  4. Intrinsic alterations in the partial molar volume on the protein denaturation: surficial Kirkwood-Buff approach.

    Science.gov (United States)

    Yu, Isseki; Takayanagi, Masayoshi; Nagaoka, Masataka

    2009-03-19

    The partial molar volume (PMV) of the protein chymotrypsin inhibitor 2 (CI2) was calculated by all-atom MD simulation. Denatured CI2 showed almost the same average PMV value as that of native CI2. This is consistent with the phenomenological question of the protein volume paradox. Furthermore, using the surficial Kirkwood-Buff approach, spatial distributions of PMV were analyzed as a function of the distance from the CI2 surface. The profiles of the new R-dependent PMV indicate that, in denatured CI2, the reduction in the solvent electrostatic interaction volume is canceled out mainly by an increment in thermal volume in the vicinity of its surface. In addition, the PMV of the denatured CI2 was found to increase in the region in which the number density of water atoms is minimum. These results provide a direct and detailed picture of the mechanism of the protein volume paradox suggested by Chalikian et al.

  5. Denaturing of plutonium by transmutation of minor-actinides for enhancement of proliferation resistance

    International Nuclear Information System (INIS)

    Sagara, Hiroshi; Saito, Masaki; Peryoga, Yoga; Ezoubtchenko, Alexey; Takivayev, Alan

    2005-01-01

    Feasibility study for the plutonium denaturing by utilizing minor-actinide transmutation in light water reactors has been performed. And the intrinsic feature of proliferation resistance of plutonium has been discussed based on IAEA's publication and Kessler's proposal. The analytical results show that not only 238 Pu but also other plutonium isotopes with even-mass-number have very important role for denaturing of plutonium due to their relatively large critical mass and noticeably high spontaneous fission neutron generation. With the change of the minor-actinide doping ratio in U-Pu mix oxide fuel and moderator to fuel ratio, it is found that the reactor-grade plutonium from conventional light water reactors can be denatured to satisfy the proliferation resistance criterion based on the Kessler's proposal but not to be sufficient for the criterion based on IAEA's publication. It has been also confirmed that all the safety coefficients take negative value throughout the irradiation. (author)

  6. Detection of human DNA polymorphisms with a simplified denaturing gradient gel electrophoresis technique

    International Nuclear Information System (INIS)

    Noll, W.W.; Collins, M.

    1987-01-01

    Single base pair differences between otherwise identical DNA molecules can result in altered melting behavior detectable by denaturing gradient gel electrophoresis. The authors have developed a simplified procedure for using denaturing gradient gel electrophoresis to detect base pair changes in genomic DNA. Genomic DNA is digested with restriction enzymes and hybridized in solution to labeled single-stranded probe DNA. The excess probe is then hybridized to complementary phage M13 template DNA, and the reaction mixture is electrophoresed on a denaturing gradient gel. Only the genomic DNA probe hybrids migrate into the gel. Differences in hybrid mobility on the gel indicate base pair changes in the genomic DNA. They have used this technique to identify two polymorphic sites within a 1.2-kilobase region of human chromosome 20. This approach should greatly facilitate the identification of DNA polymorphisms useful for gene linkage studies and the diagnosis of genetic diseases

  7. Distribution, transition and thermodynamic stability of protein conformations in the denaturant-induced unfolding of proteins.

    Science.gov (United States)

    Bian, Liujiao; Ji, Xu

    2014-01-01

    Extensive and intensive studies on the unfolding of proteins require appropriate theoretical model and parameter to clearly illustrate the feature and characteristic of the unfolding system. Over the past several decades, four approaches have been proposed to describe the interaction between proteins and denaturants, but some ambiguity and deviations usually occur in the explanation of the experimental data. In this work, a theoretical model was presented to show the dependency of the residual activity ratio of the proteins on the molar denaturant concentration. Through the characteristic unfolding parameters ki and Δmi in this model, the distribution, transition and thermodynamic stability of protein conformations during the unfolding process can be quantitatively described. This model was tested with the two-state unfolding of bovine heart cytochrome c and the three-state unfolding of hen egg white lysozyme induced by both guanidine hydrochloride and urea, the four-state unfolding of bovine carbonic anhydrase b induced by guanidine hydrochloride and the unfolding of some other proteins induced by denaturants. The results illustrated that this model could be used accurately to reveal the distribution and transition of protein conformations in the presence of different concentrations of denaturants and to evaluate the unfolding tendency and thermodynamic stability of different conformations. In most denaturant-induced unfolding of proteins, the unfolding became increasingly hard in next transition step and the proteins became more unstable as they attained next successive stable conformation. This work presents a useful method for people to study the unfolding of proteins and may be used to describe the unfolding and refolding of other biopolymers induced by denaturants, inducers, etc.

  8. The efficacy of denaturing actinide elements as a means of decreasing materials attractiveness

    Energy Technology Data Exchange (ETDEWEB)

    Hase, K.R.; Bathke, C.G. [Los Alamos National Laboratory: P.O. Box 1663, Los Alamos, NM 87545 (United States); Ebbinghaus, B.B.; Sleaford, B.W.; Robel, M. [Lawrence Livermore National Laboratory: P.O. Box 808, Livermore, CA 94551 (United States); Collins, B.A.; Prichard, A.W. [Pacific Northwest National Laboratory: P.O. Box 999, Richland, WA 99352 (United States)

    2013-07-01

    This study considers the concept of denaturing as applied to the actinide elements present in spent fuel as a means to reduce materials attractiveness. Highly attractive materials generally have low values of bare critical mass, heat content, and dose. To denature an attractive element, its spent-fuel isotopic composition (isotopic vector) is intentionally modified by introducing sufficient quantities of a significantly less attractive isotope to dilute the concentration of a highly attractive isotope so that the overall attractiveness of the element is reduced. The authors used FOM (Figure of Merit) formula as the material attractiveness metric for their parametric determination of the attractiveness of the Pu and U. Materials attractiveness needs to be considered in three distinct phases in the process to construct a nuclear explosive device (NED): the acquisition phase, processing phase, and utilization phase. The results show that denaturing uranium with {sup 238}U is actually an effective means of reducing the attractiveness. For uranium with a large minority of {sup 235}U, a mixture of 80% {sup 238}U to 20% {sup 235}U is required to reduce the attractiveness to low. For uranium with a large concentration of {sup 233}U, a mixture of 88% {sup 238}U to 12% {sup 233}U is required to reduce the attractiveness to low. The results also show that denaturing plutonium with {sup 238}Pu is less effective than denaturing uranium with {sup 238}U. Using {sup 238}Pu as the denaturing agent would require 80% or more by mass in order to reduce the attractiveness to low. No amount of {sup 240}Pu is enough to reduce the plutonium attractiveness below medium. The combination of {sup 238}Pu and {sup 240}Pu would require approximately 70% {sup 238}Pu and 25% {sup 240}Pu by mass to reduce the plutonium attractiveness to low.

  9. The efficacy of denaturing actinide elements as a means of decreasing materials attractiveness

    International Nuclear Information System (INIS)

    Hase, K.R.; Bathke, C.G.; Ebbinghaus, B.B.; Sleaford, B.W.; Robel, M.; Collins, B.A.; Prichard, A.W.

    2013-01-01

    This study considers the concept of denaturing as applied to the actinide elements present in spent fuel as a means to reduce materials attractiveness. Highly attractive materials generally have low values of bare critical mass, heat content, and dose. To denature an attractive element, its spent-fuel isotopic composition (isotopic vector) is intentionally modified by introducing sufficient quantities of a significantly less attractive isotope to dilute the concentration of a highly attractive isotope so that the overall attractiveness of the element is reduced. The authors used FOM (Figure of Merit) formula as the material attractiveness metric for their parametric determination of the attractiveness of the Pu and U. Materials attractiveness needs to be considered in three distinct phases in the process to construct a nuclear explosive device (NED): the acquisition phase, processing phase, and utilization phase. The results show that denaturing uranium with 238 U is actually an effective means of reducing the attractiveness. For uranium with a large minority of 235 U, a mixture of 80% 238 U to 20% 235 U is required to reduce the attractiveness to low. For uranium with a large concentration of 233 U, a mixture of 88% 238 U to 12% 233 U is required to reduce the attractiveness to low. The results also show that denaturing plutonium with 238 Pu is less effective than denaturing uranium with 238 U. Using 238 Pu as the denaturing agent would require 80% or more by mass in order to reduce the attractiveness to low. No amount of 240 Pu is enough to reduce the plutonium attractiveness below medium. The combination of 238 Pu and 240 Pu would require approximately 70% 238 Pu and 25% 240 Pu by mass to reduce the plutonium attractiveness to low

  10. Using fluorescence correlation spectroscopy to study conformational changes in denatured proteins.

    Science.gov (United States)

    Sherman, Eilon; Itkin, Anna; Kuttner, Yosef Yehuda; Rhoades, Elizabeth; Amir, Dan; Haas, Elisha; Haran, Gilad

    2008-06-01

    Fluorescence correlation spectroscopy (FCS) is a sensitive analytical tool that allows dynamics and hydrodynamics of biomolecules to be studied under a broad range of experimental conditions. One application of FCS of current interest is the determination of the size of protein molecules in the various states they sample along their folding reaction coordinate, which can be accessed through the measurement of diffusion coefficients. It has been pointed out that the analysis of FCS curves is prone to artifacts that may lead to erroneous size determination. To set the stage for FCS studies of unfolded proteins, we first show that the diffusion coefficients of small molecules as well as proteins can be determined accurately even in the presence of high concentrations of co-solutes that change the solution refractive index significantly. Indeed, it is found that the Stokes-Einstein relation between the measured diffusion coefficient and solution viscosity holds even in highly concentrated glycerol or guanidinium hydrochloride (GuHCl) solutions. These measurements form the basis for an investigation of the structure of the denatured state of two proteins, the small protein L and the larger, three-domain protein adenylate kinase (AK). FCS is found useful for probing expansion in the denatured state beyond the unfolding transition. It is shown that the denatured state of protein L expands as the denaturant concentration increases, in a process akin to the transition from a globule to a coil in polymers. This process continues at least up to 5 M GuHCl. On the other hand, the denatured state of AK does not seem to expand much beyond 2 M GuHCl, a result that is in qualitative accord with single-molecule fluorescence histograms. Because both the unfolding transition and the coil-globule transition of AK occur at a much lower denaturant concentration than those of protein L, a possible correlation between the two phenomena is suggested.

  11. Complement-fixing antibodies against denatured HLA and MICA antigens are associated with antibody mediated rejection.

    Science.gov (United States)

    Cai, Junchao; Terasaki, Paul I; Zhu, Dong; Lachmann, Nils; Schönemann, Constanze; Everly, Matthew J; Qing, Xin

    2016-02-01

    We have found antibodies against denatured HLA class I antigens in the serum of allograft recipients which were not significantly associated with graft failure. It is unknown whether transplant recipients also have denatured HLA class II and MICA antibodies. The effects of denatured HLA class I, class II, and MICA antibodies on long-term graft outcome were further investigated based on their ability to fix complement c1q. In this 4-year retrospective cohort study, post-transplant sera from 975 kidney transplant recipients were tested for antibodies against denatured HLA/MICA antigens and these antibodies were further classified based on their ability to fix c1q. Thirty percent of patients had antibodies against denatured HLA class I, II, or MICA antigens. Among them, 8.5% and 21.5% of all patients had c1q-fixing and non c1q-fixing antibodies respectively. There was no significant difference on graft survival between patients with or without antibodies against denatured HLA/MICA. However, when these antibodies were further classified according to their ability to fix c1q, patients with c1q-fixing antibodies had a significantly lower graft survival rate than patients without antibodies or patients with non c1q-fixing antibodies (p=0.008). In 169 patients who lost renal grafts, 44% of them had c1q-fixing antibodies against denatured HLA/MICA antigens, which was significantly higher than that in patients with functioning renal transplants (25%, pantibodies were more significantly associated with graft failure caused by AMR (72.73%) or mixed AMR/CMR (61.9%) as compared to failure due to CMR (35.3%) or other causes (39.2%) (p=0.026). Transplant recipients had antibodies against denatured HLA class I, II, and MICA antigens. However, only c1q-fixing antibodies were associated with graft failure which was related to antibody mediated rejection. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Advanced radiological work-up as an adjunct to decision in early reconstructive surgery in brachial plexus injuries

    Directory of Open Access Journals (Sweden)

    Björkman Anders

    2010-07-01

    Full Text Available Abstract Background As neurophysiologic tests may not reveal the extent of brachial plexus injury at the early stage, the role of early radiological work-up has become increasingly important. The aim of the study was to evaluate the concordance between the radiological and clinical findings with the intraoperative findings in adult patients with brachial plexus injuries. Methods Seven consecutive male patients (median age 33; range 15-61 with brachial plexus injuries, caused by motor cycle accidents in 5/7 patients, who underwent extensive radiological work-up with magnetic resonance imaging (MRI, computed tomography myelography (CT-M or both were included in this retrospective study. A total of 34 spinal nerve roots were evaluated by neuroradiologists at two different occasions. The degree of agreement between the radiological findings of every individual nerve root and the intraoperative findings was estimated by calculation of kappa coefficient (К-value. Using the operative findings as a gold standard, the accuracy, sensitivity, specificity, positive predictive value (PPV and negative predictive value (NPV of the clinical findings and the radiological findings were estimated. Results The diagnostic accuracy of radiological findings was 88% compared with 65% for the clinical findings. The concordance between the radiological findings and the intraoperative findings was substantial (К = 0.76 compared with only fair (К = 0.34 for the clinical findings. There were two false positive and two false negative radiological findings (sensitivity and PPV of 0.90; specificity and NPV of 0.87. Conclusions The advanced optimized radiological work-up used showed high reliability and substantial agreement with the intraoperative findings in adult patients with brachial plexus injury.

  13. Cost-effectiveness of functional cardiac imaging in the diagnostic work-up of coronary heart disease.

    Science.gov (United States)

    Pletscher, Mark; Walker, Simon; Moschetti, Karine; Pinget, Christophe; Wasserfallen, Jean-Blaise; Greenwood, John P; Schwitter, Juerg; Girardin, François R

    2016-07-01

    The aim of this study was to assess the cost-effectiveness of eight common diagnostic work-up strategies for coronary heart disease (CHD) in patients with stable angina symptoms in Switzerland. A decision analytical model was used to perform a cost-effectiveness comparison of eight common multitest strategies to diagnose CHD using combinations of four diagnostic techniques: exercise treadmill test (ETT), single-photon emission computed tomography (SPECT), cardiac magnetic resonance imaging (CMR), and coronary angiography (CA). We used a Markov state transition model to extrapolate the results over a life-time horizon, from a third-party payer perspective. We used a CHD prevalence rate of 39% in patients and a base-case scenario with 60-year-old male patients with intermediate symptom severity Canadian Cardiovascular Society grading of angina pectoris 2 and at least one cardiovascular (CV) risk factor but without a history of myocardial infarction and without need for revascularization. Among the eight work-up strategies, one strategy was dominant, i.e. least costly and most effective: ETT followed by CMR if the ETT result was inconclusive and then CA if the CMR result was positive or inconclusive. The CMR features a favourable balance between false-negative diagnoses, associated with an elevated risk of CV events, and false-positive diagnoses, leading to unnecessary CA and related mortality. Key parameters guiding the diagnostic strategy are the prevalence of CHD in patients with angina symptoms and the diagnostic costs of CA and CMR. Cardiac magnetic resonance imaging appears to be a cost-effective work-up strategy compared with other regimens using SPECT or direct CA. Cardiac magnetic resonance imaging should be more widely recommended as a diagnostic procedure for patients with suspected angina symptoms.

  14. Role of 3.0 Tesla magnetic resonance hysterosalpingography in the diagnostic work-up of female infertility.

    Science.gov (United States)

    Cipolla, Valentina; Guerrieri, Daniele; Pietrangeli, Daniela; Santucci, Domiziana; Argirò, Renato; de Felice, Carlo

    2016-09-01

    Imaging evaluation plays a crucial role in the diagnostic work-up of female infertility. In recent years, the possibility to evaluate tubal patency using 1.5 Tesla magnetic resonance (1.5T MR) has been studied. To assess the feasibility of 3.0 Tesla magnetic resonance (3.0T MR) hysterosalpingography and its role in the diagnostic work-up of female infertility and to evaluate if this fast "one-stop-shop" imaging approach should be proposed as a first-line examination. A total of 116 infertile women were enrolled in this prospective study; all underwent 3.0T MR hysterosalpingography. After standard imaging of the pelvis, tubal patency was assessed by acquiring 3D dynamic time-resolved T1-weighted (T1W) sequences during manual injection of 4-5 mL of contrast solution consisting of gadolinium and normal sterile saline. Images were evaluated by two radiologists with different experience in MR imaging (MRI). The examination was successfully completed in 96.5% of cases, failure rate was 3.5%. Dynamic sequences showed bilateral tubal patency in 64.3%, unilateral tubal patency in 25.9%, and bilateral tubal occlusion in 9.8%. Extratubal abnormalities were found in 69.9% of patients. Comprehensive analysis of morphological and dynamic sequences showed extratubal abnormalities in 43.1% of patients with bilateral tubal patency. 3.0T MR hysterosalpingography is a feasible, simple, fast, safe, and well-tolerated examination, which allows evaluation of tubal patency and other pelvic causes of female infertility in a single session, and it may thus represent a "one-stop-shop" solution in female infertility diagnostic work-up. © The Foundation Acta Radiologica 2015.

  15. Unravelling the hydrophobicity of urea in water using thermodiffusion: implications for protein denaturation.

    Science.gov (United States)

    Niether, Doreen; Di Lecce, Silvia; Bresme, Fernando; Wiegand, Simone

    2018-01-03

    Urea is widely used as a protein denaturant in aqueous solutions. Experimental and computer simulation studies have shown that it dissolves in water almost ideally at high concentrations, introducing little disruption in the water hydrogen bonded structure. However, at concentrations of the order of 5 M or higher, urea induces denaturation in a wide range of proteins. The origin of this behaviour is not completely understood, but it is believed to stem from a balance between urea-protein and urea-water interactions, with urea becoming possibly hydrophobic at a specific concentration range. The small changes observed in the water structure make it difficult to connect the denaturation effects to the solvation properties. Here we show that the exquisite sensitivity of thermodiffusion to solute-water interactions allows the identification of the onset of hydrophobicity of urea-water mixtures. The hydrophobic behaviour is reflected in a sign reversal of the temperature dependent slope of the Soret coefficient, which is observed, both in experiments and non-equilibrium computer simulations at ∼5 M concentration of urea in water. This concentration regime corresponds to the one where abrupt changes in the denaturation of proteins are commonly observed. We show that the onset of hydrophobicity is intrinsically connected to the urea-water interactions. Our results allow us to identify correlations between the Soret coefficient and the partition coefficient, log P, hence establishing the thermodiffusion technique as a powerful approach to study hydrophobicity.

  16. Thermal denaturation of protein studied by terahertz time-domain spectroscopy

    Science.gov (United States)

    Fu, Xiuhua; Li, Xiangjun; Liu, Jianjun; Du, Yong; Hong, Zhi

    2012-12-01

    In this study, the absorption spectra of native or thermal protein were measured in 0.2-1.4THz using terahertz time-domain spectroscopy (THz-TDS) system at room temperature, their absorption spectra and the refractive spectra were obtained. Experimental results indicate that protein both has strong absorption but their characteristics were not distinct in the THz region, and the absorption decreased during thermal denatured state. In order to prove protein had been denatured, we used Differential scanning calorimeter (DSC) measured their denatured temperature, from their DSC heating traces, collagen Td=101℃, Bovine serum albumin Td=97℃. While we also combined the Fourier transform infrared spectrometer (FTIR) to investigate their secondary and tertiary structure before and after denatuation, but the results did not have the distinct changes. We turned the absorption spectra and the refractive spectra to the dielectric spectra, and used the one-stage Debye model simulated the terahertz dielectric spectra of protein before and after denaturation. This research proved that the terahertz spectrum technology is feasible in testing protein that were affected by temperature or other factors which can provide theoretical foundation in the further study about the THz spectrum of protein and peptide temperature stability.

  17. Mesoscopic modeling of DNA denaturation rates: Sequence dependence and experimental comparison

    Energy Technology Data Exchange (ETDEWEB)

    Dahlen, Oda, E-mail: oda.dahlen@ntnu.no; Erp, Titus S. van, E-mail: titus.van.erp@ntnu.no [Department of Chemistry, Norwegian University of Science and Technology (NTNU), Høgskoleringen 5, Realfagbygget D3-117 7491 Trondheim (Norway)

    2015-06-21

    Using rare event simulation techniques, we calculated DNA denaturation rate constants for a range of sequences and temperatures for the Peyrard-Bishop-Dauxois (PBD) model with two different parameter sets. We studied a larger variety of sequences compared to previous studies that only consider DNA homopolymers and DNA sequences containing an equal amount of weak AT- and strong GC-base pairs. Our results show that, contrary to previous findings, an even distribution of the strong GC-base pairs does not always result in the fastest possible denaturation. In addition, we applied an adaptation of the PBD model to study hairpin denaturation for which experimental data are available. This is the first quantitative study in which dynamical results from the mesoscopic PBD model have been compared with experiments. Our results show that present parameterized models, although giving good results regarding thermodynamic properties, overestimate denaturation rates by orders of magnitude. We believe that our dynamical approach is, therefore, an important tool for verifying DNA models and for developing next generation models that have higher predictive power than present ones.

  18. Resistance to DNA denaturation in irradiated Chinese hamster V79 fibroblasts is linked to cell shape

    International Nuclear Information System (INIS)

    Olive, P.L.; Vanderbyl, S.; MacPhail, S.H.

    1991-01-01

    Exponentially growing Chinese hamster V79-171b lung fibroblasts seeded at high density on plastic (approximately 7 x 10(3) cells/cm2) flatten, elongate, and produce significant amounts of extracellular fibronectin. When lysed in weak alkali/high salt, the rate of DNA denaturation following exposure to ionizing radiation is exponential. Conversely, cells plated at low density (approximately 7 x 10(2) cells/cm2) on plastic are more rounded 24 h later, produce little extracellular fibronectin, and display unusual DNA denaturation kinetics after X-irradiation. DNA in these cells resists denaturation, as though constraints to DNA unwinding have developed. Cell doubling time and distribution of cells in the growth cycle are identical for both high and low density cultures as is cell survival in response to radiation damage. The connection between DNA conformation and cell shape was examined further in low density cultures grown in conditioned medium. Under these conditions, cells at low density were able to elongate, and DNA denaturation of low density cultures was identical to that of high density cultures. Conversely, cytochalasin D, which interferes with actin polymerization causing cells to round up and release fibronectin, allowed development of constraints in high density cultures. These results suggest that DNA conformation is sensitive to changes in cell shape which result when cells are grown in different environments. However, these changes in DNA conformation detected by the DNA unwinding assay do not appear to play a direct role in radiation-induced cell killing

  19. 27 CFR 20.178 - Marks and brands on containers of specially denatured spirits.

    Science.gov (United States)

    2010-04-01

    ... 27 Alcohol, Tobacco Products and Firearms 1 2010-04-01 2010-04-01 false Marks and brands on... Dealers § 20.178 Marks and brands on containers of specially denatured spirits. (a) Required marks. Each... officer, or (2) Consist of a brand name, or consist of caution notices, or consist of other material...

  20. Isoenergic modification of whey protein structure by denaturation and crosslinking using transglutaminase

    DEFF Research Database (Denmark)

    Stender, Emil G. P.; Koutina, Glykeria; Almdal, Kristoffer

    2018-01-01

    Transglutaminase (TG) catalyzes formation of covalent bonds between lysine and glutamine side chains and has applications in manipulation of food structure. Physical properties of a whey protein mixture (SPC) denatured either at elevated pH or by heat-treatment and followed by TG catalyzed...

  1. [Inactivating Effect of Heat-Denatured Lysozyme on Murine Norovirus in Bread Fillings].

    Science.gov (United States)

    Takahashi, Michiko; Yasuda, Yuka; Takahashi, Hajime; Takeuchi, Akira; Kuda, Takashi; Kimura, Bon

    2018-01-01

    In this study, we investigated the viability of murine norovirus strain 1 (MNV-1), a surrogate for human norovirus, in bread fillings used for making stuffed buns and pastries. The inactivating effect of heat-denatured lysozyme, which was recently reported to have an antiviral effect, on MNV-1 contaminating the bread fillings was also examined. MNV-1 was inoculated into two types of fillings (chocolate cream, marmalade jam) at 4.5 log PFU/g, and the bread fillings were stored at 4℃ for 5 days. MNV-1 remained viable in the bread fillings during storage. However, addition of 1% heat-denatured lysozyme to the fillings resulted in a decrease of MNV-1 infectivity immediately after inoculation, in both fillings. On the fifth day of storage, MNV-1 infectivity was decreased by 1.2 log PFU/g in chocolate cream and by 0.9 log PFU/g in marmalade jam. Although the mechanism underlying the anti-norovirus effect of heat-denatured lysozyme has not been clarified, our results suggest that heat-denatured lysozyme can be used as an inactivating agent against norovirus in bread fillings.

  2. Improvement of fragment and primer selection for mutation detection by denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Wu, Y; Hayes, VM; Osinga, J; Mulder, IM; Looman, MWG; Buys, CHCM; Hofstra, RMW

    1998-01-01

    Denaturing gradient gel electrophoresis (DGGE) is one of the most powerful methods for mutation detection currently available. For successful application the appropriate selection of PCR fragments and PCR primers is crucial. The sequence of interest should always be within the domain with the lowest

  3. Comparative analyses of amplicon migration behavior in differing denaturing gradient gel electrophoresis (DGGE) systems

    Science.gov (United States)

    Thornhill, D. J.; Kemp, D. W.; Sampayo, E. M.; Schmidt, G. W.

    2010-03-01

    Denaturing gradient gel electrophoresis (DGGE) is commonly utilized to identify and quantify microbial diversity, but the conditions required for different electrophoretic systems to yield equivalent results and optimal resolution have not been assessed. Herein, the influence of different DGGE system configuration parameters on microbial diversity estimates was tested using Symbiodinium, a group of marine eukaryotic microbes that are important constituents of coral reef ecosystems. To accomplish this, bacterial clone libraries were constructed and sequenced from cultured isolates of Symbiodinium for the ribosomal DNA internal transcribed spacer 2 (ITS2) region. From these, 15 clones were subjected to PCR with a GC clamped primer set for DGGE analyses. Migration behaviors of the resulting amplicons were analyzed using a range of conditions, including variation in the composition of the denaturing gradient, electrophoresis time, and applied voltage. All tests were conducted in parallel on two commercial DGGE systems, a C.B.S. Scientific DGGE-2001, and the Bio-Rad DCode system. In this context, identical nucleotide fragments exhibited differing migration behaviors depending on the model of apparatus utilized, with fragments denaturing at a lower gradient concentration and applied voltage on the Bio-Rad DCode system than on the C.B.S. Scientific DGGE-2001 system. Although equivalent PCR-DGGE profiles could be achieved with both brands of DGGE system, the composition of the denaturing gradient and application of electrophoresis time × voltage must be appropriately optimized to achieve congruent results across platforms.

  4. Urea and Guanidinium Induced Denaturation of a Trp-Cage Miniprotein

    Czech Academy of Sciences Publication Activity Database

    Heyda, Jan; Kožíšek, Milan; Bednárová, Lucie; Thompson, G.; Konvalinka, Jan; Vondrášek, Jiří; Jungwirth, Pavel

    2011-01-01

    Roč. 115, č. 28 (2011), s. 8910-8924 ISSN 1520-6106 R&D Projects: GA MŠk LC512; GA ČR GA203/08/0114 Institutional research plan: CEZ:AV0Z40550506 Keywords : trp-cage denaturation * urea * guanidinium * molecular dynamics Subject RIV: CC - Organic Chemistry Impact factor: 3.696, year: 2011

  5. A Role For Ca 2+ in the Thermal and Urea Denaturation of ...

    African Journals Online (AJOL)

    Giant African snail (Achatina achatina) becomes dormant (aestivate) under harsh environmental conditions like dry seasons. During this period the animal accumulates urea and is faced with thermal death. The stability towards thermal and urea denaturation of haemocyanin from aestivating and nonaestivating A. achatina ...

  6. Kinetic evidence for a two-stage mechanism of protein denaturation by guanidinium chloride.

    Science.gov (United States)

    Jha, Santosh Kumar; Marqusee, Susan

    2014-04-01

    Dry molten globular (DMG) intermediates, an expanded form of the native protein with a dry core, have been observed during denaturant-induced unfolding of many proteins. These observations are counterintuitive because traditional models of chemical denaturation rely on changes in solvent-accessible surface area, and there is no notable change in solvent-accessible surface area during the formation of the DMG. Here we show, using multisite fluorescence resonance energy transfer, far-UV CD, and kinetic thiol-labeling experiments, that the guanidinium chloride (GdmCl)-induced unfolding of RNase H also begins with the formation of the DMG. Population of the DMG occurs within the 5-ms dead time of our measurements. We observe that the size and/or population of the DMG is linearly dependent on [GdmCl], although not as strongly as the second and major step of unfolding, which is accompanied by core solvation and global unfolding. This rapid GdmCl-dependent population of the DMG indicates that GdmCl can interact with the protein before disrupting the hydrophobic core. These results imply that the effect of chemical denaturants cannot be interpreted solely as a disruption of the hydrophobic effect and strongly support recent computational studies, which hypothesize that chemical denaturants first interact directly with the protein surface before completely unfolding the protein in the second step (direct interaction mechanism).

  7. Interaction of ATP with acid-denatured cytochrome c via coupled folding-binding mechanism

    International Nuclear Information System (INIS)

    Ahluwalia, Unnati; Deep, Shashank

    2012-01-01

    Highlights: ► Interaction between ATP and cyt c takes place via coupled binding–folding mechanism. ► Binding of ATP to cyt c is endothermic. ► GTP and CTP induce similar level of helicity in acid-denatured cyt c as with ATP. ► Compactness induced by ATP is far greater than ADP or AMP. - Abstract: The non-native conformations of the cytochrome c (cyt c) are believed to play key roles in a number of physiological processes. Nucleotides are supposed to act as allosteric effectors in these processes by regulating structural transitions among different conformations of cyt c. To understand the interaction between acid denatured cytochrome c and nucleotides, spectroscopic and calorimetric techniques were utilized to observe the structural features of the induced conformation and the energetics of interaction of acid denatured cyt c with different nucleotides. Structure induction in the acid denatured cyt c was observed on the addition of the ∼1 mM nucleotide tri-phosphates (ATP/GTP/CTP) at 25 °C, however, not in the presence of 1 mM nucleotide mono and diphosphates. ATP-bound cyt c at pH 2.0 is likely to have a conformation that has intact α-helical domain. However, Met80-Fe(III) axial bond is still ruptured. Observed thermodynamics reflect interaction between nucleotide and cyt c via coupled binding–folding mechanism. DSC data suggest the preferential binding of the ATP to the folded conformation with respect to the acid denatured cyt c. ITC data indicate that the exothermic folding of cyt c was accompanied by endothermic binding of ATP to cyt c.

  8. Quantitative assessments of the distinct contributions of polypeptide backbone amides versus sidechain groups to chain expansion via chemical denaturation

    Science.gov (United States)

    Holehouse, Alex S.; Garai, Kanchan; Lyle, Nicholas; Vitalis, Andreas; Pappu, Rohit V.

    2015-01-01

    In aqueous solutions with high concentrations of chemical denaturants such as urea and guanidinium chloride (GdmCl) proteins expand to populate heterogeneous conformational ensembles. These denaturing environments are thought to be good solvents for generic protein sequences because properties of conformational distributions align with those of canonical random coils. Previous studies showed that water is a poor solvent for polypeptide backbones and therefore backbones form collapsed globular structures in aqueous solvents. Here, we ask if polypeptide backbones can intrinsically undergo the requisite chain expansion in aqueous solutions with high concentrations of urea and GdmCl. We answer this question using a combination of molecular dynamics simulations and fluorescence correlation spectroscopy. We find that the degree of backbone expansion is minimal in aqueous solutions with high concentrations denaturants. Instead, polypeptide backbones sample conformations that are denaturant-specific mixtures of coils and globules, with a persistent preference for globules. Therefore, typical denaturing environments cannot be classified as good solvents for polypeptide backbones. How then do generic protein sequences expand in denaturing environments? To answer this question, we investigated the effects of sidechains using simulations of two archetypal sequences with amino acid compositions that are mixtures of charged, hydrophobic, and polar groups. We find that sidechains lower the effective concentration of backbone amides in water leading to an intrinsic expansion of polypeptide backbones in the absence of denaturants. Additional dilution of the effective concentration of backbone amides is achieved through preferential interactions with denaturants. These effects lead to conformational statistics in denaturing environments that are congruent with those of canonical random coils. Our results highlight the role of sidechain-mediated interactions as determinants of the

  9. Transurethral radiofrequency collagen denaturation for the treatment of women with urinary incontinence.

    Science.gov (United States)

    Kang, Diana; Han, Julia; Neuberger, Molly M; Moy, M Louis; Wallace, Sheila A; Alonso-Coello, Pablo; Dahm, Philipp

    2015-03-18

    Transurethral radiofrequency collagen denaturation is a relatively novel, minimally invasive device-based intervention used to treat individuals with urinary incontinence (UI). No systematic review of the evidence supporting its use has been published to date. To evaluate the efficacy of transurethral radiofrequency collagen denaturation, compared with other interventions, in the treatment of women with UI.Review authors sought to compare the following.• Transurethral radiofrequency collagen denaturation versus no treatment/sham treatment.• Transurethral radiofrequency collagen denaturation versus conservative physical treatment.• Transurethral radiofrequency collagen denaturation versus mechanical devices (pessaries for UI).• Transurethral radiofrequency collagen denaturation versus drug treatment.• Transurethral radiofrequency collagen denaturation versus injectable treatment for UI.• Transurethral radiofrequency collagen denaturation versus other surgery for UI. We conducted a systematic search of the Cochrane Incontinence Group Specialised Register (searched 19 December 2014), EMBASE and EMBASE Classic (January 1947 to 2014 Week 50), Google Scholar and three trials registries in December 2014, along with reference checking. We sought to identify unpublished studies by handsearching abstracts of major gynaecology and urology meetings, and by contacting experts in the field and the device manufacturer. Randomised and quasi-randomised trials of transurethral radiofrequency collagen denaturation versus no treatment/sham treatment, conservative physical treatment, mechanical devices, drug treatment, injectable treatment for UI or other surgery for UI in women were eligible. We screened search results and selected eligible studies for inclusion. We assessed risk of bias and analysed dichotomous variables as risk ratios (RRs) with 95% confidence intervals (CIs) and continuous variables as mean differences (MDs) with 95% CIs. We rated the quality of

  10. The role of PET in initial work-up and evaluation after therapy in patients with carcinoma of unknown primary

    Energy Technology Data Exchange (ETDEWEB)

    Ryoo, Baek Yeol; Kang, Yoon Koo

    1998-12-01

    The carcinoma of unknown primary occupied 5 - 10 % of all malignancies. It is heterogenous in origin and has poor prognosis. The indentification of primary site and definition of involved area are more helpful in the management. The efficacy of positron emission tomography (PET) with fluorine-18- fluorodeoxyglucose (F18-FDG) positron emission tomography (PET) with fluorine-18-fluorodeoxyglucose (F18-FDG) was evaluated in several tumors such as breast, pancreas and head and neck cancers. In carcinoma of unknown primary, it was reported that the concentration of FDG was increased in tumor tissues, and that PET with F18-FDG may be much helpful in identifying primary site and defining involved area. The authors evaluated the usefulness of PET with F18-FDG in initial work-up and in evaluation after radical therapy for the patients with carcinoma of unknown primary. The visual analysis of FDG-PET would be helpful in identifying primary site and defining involved area. In detecting recurrent of residual lesions, FDG-PET seemed to be less helpful than conventional diagnostic work-up. But more studies with larger number of cases and longer follow-up were required. The results of this study can be bases for the direction of future studies for the usefulness of PET in carcinoma of unknown primary.

  11. The role of PET in initial work-up and evaluation after therapy in patients with carcinoma of unknown primary

    International Nuclear Information System (INIS)

    Ryoo, Baek Yeol; Kang, Yoon Koo

    1998-12-01

    The carcinoma of unknown primary occupied 5 - 10 % of all malignancies. It is heterogenous in origin and has poor prognosis. The indentification of primary site and definition of involved area are more helpful in the management. The efficacy of positron emission tomography (PET) with fluorine-18- fluorodeoxyglucose (F18-FDG) positron emission tomography (PET) with fluorine-18-fluorodeoxyglucose (F18-FDG) was evaluated in several tumors such as breast, pancreas and head and neck cancers. In carcinoma of unknown primary, it was reported that the concentration of FDG was increased in tumor tissues, and that PET with F18-FDG may be much helpful in identifying primary site and defining involved area. The authors evaluated the usefulness of PET with F18-FDG in initial work-up and in evaluation after radical therapy for the patients with carcinoma of unknown primary. The visual analysis of FDG-PET would be helpful in identifying primary site and defining involved area. In detecting recurrent of residual lesions, FDG-PET seemed to be less helpful than conventional diagnostic work-up. But more studies with larger number of cases and longer follow-up were required. The results of this study can be bases for the direction of future studies for the usefulness of PET in carcinoma of unknown primary

  12. The Appropriate Use of Neuroimaging in the Diagnostic Work-Up of Dementia

    Science.gov (United States)

    2014-01-01

    Background Diagnosis of dementia is challenging and requires both ruling out potentially treatable underlying causes and ruling in a diagnosis of dementia subtype to manage patients and suitably plan for the future. Objectives This analysis sought to determine the appropriate use of neuroimaging during the diagnostic work-up of dementia, including indications for neuroimaging and comparative accuracy of alternative technologies. Data Sources A literature search was performed using Ovid MEDLINE, Ovid MEDLINE In-Process and Other Non-Indexed Citations, Ovid Embase, the Wiley Cochrane Library, and the Centre for Reviews and Dissemination database, for studies published between 2000 and 2013. Review Methods Data on diagnostic accuracy and impact on clinical decision making were abstracted from included studies. Quality of evidence was assessed using GRADE. Results The search yielded 5,374 citations and 15 studies were included. Approximately 10% of dementia cases are potentially treatable, though less than 1% reverse partially or fully. Neither prediction rules nor clinical indications reliably select the subset of patients who will likely benefit from neuroimaging. Clinical utility is highest in ambiguous cases or where dementia may be mixed, and lowest for clinically diagnosed Alzheimer disease or clinically excluded vascular dementia. There is a lack of evidence that MRI is superior to CT in detecting a vascular component to dementia. Accuracy of structural imaging is moderate to high for discriminating different types of dementia. Limitations There was significant heterogeneity in estimates of diagnostic accuracy, which often prohibited a statistical summary of findings. The quality of data reported by studies prohibited calculation of likelihood ratios in the present analysis. No studies from primary care were found; thus, generalizability beyond tertiary care settings may be limited. Conclusions A diagnosis of reversible dementia is rare. Imaging has the most

  13. Denaturing of single electrospun fibrinogen fibers studied by deep ultraviolet fluorescence microscopy.

    Science.gov (United States)

    Kim, Jeongyong; Song, Hugeun; Park, Inho; Carlisle, Christine R; Bonin, Keith; Guthold, Martin

    2011-03-01

    Deep ultraviolet (DUV) microscopy is a fluorescence microscopy technique to image unlabeled proteins via the native fluorescence of some of their amino acids. We constructed a DUV fluorescence microscope, capable of 280 nm wavelength excitation by modifying an inverted optical microscope. Moreover, we integrated a nanomanipulator-controlled micropipette into this instrument for precise delivery of picoliter amounts of fluid to selected regions of the sample. In proof-of-principle experiments, we used this instrument to study, in situ, the effect of a denaturing agent on the autofluorescence intensity of single, unlabeled, electrospun fibrinogen nanofibers. Autofluorescence emission from the nanofibers was excited at 280 nm and detected at ∼350 nm. A denaturant solution was discretely applied to small, select sections of the nanofibers and a clear local reduction in autofluorescence intensity was observed. This reduction is attributed to the dissolution of the fibers and the unfolding of proteins in the fibers. Copyright © 2010 Wiley-Liss, Inc.

  14. Theoretical aspects of pressure and solute denaturation of proteins: A Kirkwood-buff-theory approach

    Science.gov (United States)

    Ben-Naim, Arieh

    2012-12-01

    A new approach to the problem of pressure-denaturation (PD) and solute-denaturation (SD) of proteins is presented. The problem is formulated in terms of Le Chatelier principle, and a solution is sought in terms of the Kirkwood-Buff theory of solutions. It is found that both problems have one factor in common; the excluded volumes of the folded and the unfolded forms with respect to the solvent molecules. It is shown that solvent-induced effects operating on hydrophilic groups along the protein are probably the main reason for PD. On the other hand, the SD depends on the preferential solvation of the folded and the unfolded forms with respect to solvent and co-solvent molecules.

  15. Teaching what one does not know: strangeness and denaturation in (autobiographical narrations

    Directory of Open Access Journals (Sweden)

    Jorge Luiz da Cunha

    2014-01-01

    Full Text Available The thematic focus in this text are the estrangement/denaturation processes in (autobiographical narrations. The aim of this study was to reflect on the possibility to promote estrangement/denatura - tion in (autobiographical writings made by teenagers in the space/ time of the classroom environment. The methodological proposal consisted on developing (autobiographical writings by students from sociology classes in High School. A total of 138 teenagers from a public school, attending the first school trimester in the year 2013, have participated in the study. The concepts of estrangement/de - naturation are located in the anthropology field and, the work with (autobiographical narrations is located in the socio-clinic perspec - tives and of biographization processes. The results indicate that (autobiographical narrations provide estrangements/denaturation and go towards teaching what one does not know. We can, then, conclude that this possibility, as an educational act, may generate knowledge suspension to self-inventiveness.

  16. Two-dimensional salt and temperature DNA denaturation analysis using a magnetoresistive sensor

    DEFF Research Database (Denmark)

    Rizzi, Giovanni; Dufva, Martin; Hansen, Mikkel Fougt

    2017-01-01

    We present a microfluidic system and its use to measure DNA denaturation curves by varying the temperature or salt (Na+) concentration. The readout is based on real-time measurements of DNA hybridization using magnetoresistive sensors and magnetic nanoparticles (MNPs) as labels. We report the first...... melting curves of DNA hybrids measured as a function of continuously decreasing salt concentration at fixed temperature and compare them to the corresponding curves obtained vs. temperature at fixed salt concentration. The magnetoresistive sensor platform provided reliable results under varying....... The results demonstrate that concentration melting provides an attractive alternative to temperature melting in on-chip DNA denaturation experiments and further show that the magnetoresistive platform is attractive due to its low cross-sensitivity to temperature and liquid composition....

  17. Native and denatured bovine serum albumin. D.c. polarography, stripping voltammetry and constant current chronopotentiometry

    Czech Academy of Sciences Publication Activity Database

    Ostatná, Veronika; Uslu, B.; Dogan, B.; Ozkan, S.; Paleček, Emil

    2006-01-01

    Roč. 593, č. 1-2 (2006), s. 172-178 ISSN 0022-0728 R&D Projects: GA AV ČR(CZ) IAA500040513; GA MŠk(CZ) LC06035 Institutional research plan: CEZ:AV0Z50040507 Keywords : protein electrochemistry * bovine serum albumin * native and denatured proteins Subject RIV: BO - Biophysics Impact factor: 2.339, year: 2006

  18. Micro-CT Imaging of Denatured Chitin by Silver to Explore Honey Bee and Insect Pathologies

    Science.gov (United States)

    Butzloff, Peter R.

    2011-01-01

    Background Chitin and cuticle coatings are important to the environmental and immune defense of honey bees and insect pollinators. Pesticides or environmental effects may target the biochemistry of insect chitin and cuticle coating. Denaturing of chitin involves a combination of deacetylation, intercalation, oxidation, Schweiger-peeling, and the formation of amine hydrochloride salt. The term “denatured chitin” calls attention to structural and property changes to the internal membranes and external carapace of organisms so that some properties affecting biological activities are diminished. Methodology/Principal Findings A case study was performed on honey bees using silver staining and microscopic computer-tomographic x-ray radiography (micro-CT). Silver nitrate formed counter-ion complexes with labile ammonium cations and reacted with amine hydrochloride. Silver was concentrated in the peritrophic membrane, on the abdomen, in the glossa, at intersegmental joints (tarsi), at wing attachments, and in tracheal air sacs. Imaged mono-esters and fatty acids from cuticle coating on external surfaces were apparently reduced by an alcohol pretreatment. Conclusions/Significance The technique provides 3-dimensional and sectional images of individual honey bees consistent with the chemistries of silver reaction and complex formation with denatured chitin. Environmental exposures and influences such as gaseous nitric oxide intercalant, trace oxidants such as ozone gas, oligosachharide salt conversion, exposure to acid rain, and chemical or biochemical denaturing by pesticides may be studied using this technique. Peritrophic membranes, which protect against food abrasion, microorganisms, and permit efficient digestion, were imaged. Apparent surface damage to the corneal lenses of compound eyes by dilute acid exposure consistent with chitin amine hydrochloride formation was imaged. The technique can contribute to existing insect pathology research, and may provide an

  19. Micro-CT imaging of denatured chitin by silver to explore honey bee and insect pathologies.

    Directory of Open Access Journals (Sweden)

    Peter R Butzloff

    Full Text Available BACKGROUND: Chitin and cuticle coatings are important to the environmental and immune defense of honey bees and insect pollinators. Pesticides or environmental effects may target the biochemistry of insect chitin and cuticle coating. Denaturing of chitin involves a combination of deacetylation, intercalation, oxidation, Schweiger-peeling, and the formation of amine hydrochloride salt. The term "denatured chitin" calls attention to structural and property changes to the internal membranes and external carapace of organisms so that some properties affecting biological activities are diminished. METHODOLOGY/PRINCIPAL FINDINGS: A case study was performed on honey bees using silver staining and microscopic computer-tomographic x-ray radiography (micro-CT. Silver nitrate formed counter-ion complexes with labile ammonium cations and reacted with amine hydrochloride. Silver was concentrated in the peritrophic membrane, on the abdomen, in the glossa, at intersegmental joints (tarsi, at wing attachments, and in tracheal air sacs. Imaged mono-esters and fatty acids from cuticle coating on external surfaces were apparently reduced by an alcohol pretreatment. CONCLUSIONS/SIGNIFICANCE: The technique provides 3-dimensional and sectional images of individual honey bees consistent with the chemistries of silver reaction and complex formation with denatured chitin. Environmental exposures and influences such as gaseous nitric oxide intercalant, trace oxidants such as ozone gas, oligosachharide salt conversion, exposure to acid rain, and chemical or biochemical denaturing by pesticides may be studied using this technique. Peritrophic membranes, which protect against food abrasion, microorganisms, and permit efficient digestion, were imaged. Apparent surface damage to the corneal lenses of compound eyes by dilute acid exposure consistent with chitin amine hydrochloride formation was imaged. The technique can contribute to existing insect pathology research, and may

  20. Denaturing gradient gel electrophoresis profiling of bacterial communities composition in Arabian Sea

    Digital Repository Service at National Institute of Oceanography (India)

    Singh, S.K.; Ramaiah, N.

    of Environmental Biology circleshadowdwnMay 2011circleshadowdwn Introduction The bacteria play a major role in carbon dynamics of marine ecosystems and, the importance of heterotrophic bacteria in marine ecosystem functioning is very well recognized (Azam et al..., 2008). Denaturing gradient gel-electrophoressis (DGGE) based fingerprinting helps estimate the numbers of dominant phylotype in a given sample (Muyzer et al., 1993). Very diverse bacterial assemblages such as those in the soils present many bands...

  1. Native and denatured forms of proteins can be discriminated at edge plane carbon electrodes

    Czech Academy of Sciences Publication Activity Database

    Ostatná, Veronika; Černocká, Hana; Kurzatkowska, K.; Paleček, Emil

    2012-01-01

    Roč. 735, JUL (2012), s. 31-36 ISSN 0003-2670 R&D Projects: GA AV ČR(CZ) KJB100040901; GA ČR(CZ) GAP301/11/2055; GA MŠk(CZ) ME09038 Institutional research plan: CEZ:AV0Z50040702 Keywords : protein denaturation * carbon electrodes * edge plane pyrolytic graphite Subject RIV: BO - Biophysics Impact factor: 4.387, year: 2012

  2. Multifocal peritoneal splenosis in Tc-99m-labeled heat-denatured red blood cell scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Min Ki; Hwang, Kyung Hoon; Choe, Won Sick [Gachon University Gil Medical Center, Incheon (Korea, Republic of)

    2006-06-15

    A 44-year-old man with a past medical history of splenectomy came to hospital because of epigastric pain abdominopelvic computed tomography(CT) showed a soft tissue mass and multifocal variable-sized nodules as well as finding suggestive of cholecystitis. Subsequently, he underwent Tc-99m-labeled heat- denatured red blood cell(RBC) scintigraphy to evaluate the mass and nodules. The scintigraphy confirmed multifocal peritoneal splenosis in the abdominopelvic cavity.

  3. Equilibrium unfolding of A. niger RNase: pH dependence of chemical and thermal denaturation.

    Science.gov (United States)

    Kumar, Gundampati Ravi; Sharma, Anurag; Kumari, Moni; Jagannadham, Medicherla V; Debnath, Mira

    2011-08-01

    Equilibrium unfolding of A. niger RNase with chemical denaturants, for example GuHCl and urea, and thermal unfolding have been studied as a function of pH using fluorescence, far-UV, near-UV, and absorbance spectroscopy. Because of their ability to affect electrostatic interactions, pH and chemical denaturants have a marked effect on the stability, structure, and function of many globular proteins. ANS binding studies have been conducted to enable understanding of the folding mechanism of the protein in the presence of the denaturants. Spectroscopic studies by absorbance, fluorescence, and circular dichroism and use of K2D software revealed that the enzyme has α + β type secondary structure with approximately 29% α-helix, 24% β-sheet, and 47% random coil. Under neutral conditions the enzyme is stable in urea whereas GuHCl-induced equilibrium unfolding was cooperative. A. niger RNase has little ANS binding even under neutral conditions. Multiple intermediates were populated during the pH-induced unfolding of A. niger RNase. Urea and temperature-induced unfolding of A. niger RNase into the molten globule-like state is non-cooperative, in contrast to the cooperativity seen with the native protein, suggesting the presence of two parts/domains, in the molecular structure of A. niger RNase, with different stability that unfolds in steps. Interestingly, the GuHCl-induced unfolding of the A state (molten globule state) of A. niger RNase is unique, because a low concentration of denaturant not only induces structural change but also facilitates transition from one molten globule like state (A(MG1)) into another (I(MG2)).

  4. Thermal stability of chemically denatured green fluorescent protein (GFP) A preliminary study

    Energy Technology Data Exchange (ETDEWEB)

    Nagy, Attila; Malnasi-Csizmadia, Andras; Somogyi, Bela; Lorinczy, Denes

    2004-02-09

    Green fluorescent protein (GFP) is a light emitter in the bioluminescence reaction of the jellyfish Aequorea victoria. The protein consist of 238 amino acids and produces green fluorescent light ({lambda}{sub max}=508 nm), when irradiated with near ultraviolet light. The fluorescence is due to the presence of chromophore consisting of an imidazolone ring, formed by a post-translational modification of the tripeptide -Ser{sup 65}-Tyr{sup 66}-Gly{sup 67}-, which buried into {beta}-barrel. GFP is extremely compact and heat stable molecule. In this work, we present data for the effect of chemical denaturing agent on the thermal stability of GFP. When denaturing agent is applied, global thermal stability and the melting point of the molecule is decreases, that can be monitored with differential scanning calorimetry. The results indicate, that in 1-6 M range of GuHCl the melting temperature is decreasing continuously from 83 to 38 deg. C. Interesting finding, that the calculated calorimetric enthalpy decreases with GuHCl concentration up to 3 M (5.6-0.2 kJ mol{sup -1}), but at 4 M it jumps to 8.4 and at greater concentration it is falling down to 1.1 kJ mol{sup -1}. First phenomena, i.e. the decrease of melting point with increasing GuHCl concentration can be easily explained by the effect of the extended chemical denaturation, when less and less amount of heat required to diminish the remaining hydrogen bonds in {beta}-barrel. The surprising increase of calorimetric enthalpy at 4 M concentration of GuHCl could be the consequence of a dimerization or a formation of stable complex between GFP and denaturing agent as well as a precipitation at an extreme GuHCl concentration. We are planning further experiments to elucidate fluorescent consequence of these processes.

  5. Added diagnostic value of magnetoencephalography (MEG) in patients suspected for epilepsy, where previous, extensive EEG workup was unrevealing

    DEFF Research Database (Denmark)

    Duez, Lene; Beniczky, Sándor; Tankisi, Hatice

    2016-01-01

    was calculated from the cases where abnormalities were seen in MEG but not EEG. RESULTS: Twenty-two patients had the diagnosis epilepsy according to the reference standard. MEG-EEG detected abnormalities, and supported the diagnosis in nine of the 22 patients with the diagnosis epilepsy at one-year follow......OBJECTIVE: To elucidate the possible additional diagnostic yield of MEG in the workup of patients with suspected epilepsy, where repeated EEGs, including sleep-recordings failed to identify abnormalities. METHODS: Fifty-two consecutive patients with clinical suspicion of epilepsy and at least three...... normal EEGs, including sleep-EEG, were prospectively analyzed. The reference standard was inferred from the diagnosis obtained from the medical charts, after at least one-year follow-up. MEG (306-channel, whole-head) and simultaneous EEG (MEG-EEG) was recorded for one hour. The added sensitivity of MEG...

  6. Preanalytical blood sample workup for cell-free DNA analysis using Droplet Digital PCR for future molecular cancer diagnostics.

    Science.gov (United States)

    van Ginkel, Joost H; van den Broek, Daan A; van Kuik, Joyce; Linders, Dorothé; de Weger, Roel; Willems, Stefan M; Huibers, Manon M H

    2017-10-01

    In current molecular cancer diagnostics, using blood samples of cancer patients for the detection of genetic alterations in plasma (cell-free) circulating tumor DNA (ctDNA) is an emerging practice. Since ctDNA levels in blood are low, highly sensitive Droplet Digital PCR (ddPCR) can be used for detecting rare mutational targets. In order to perform ddPCR on blood samples, a standardized procedure for processing and analyzing blood samples is necessary to facilitate implementation into clinical practice. Therefore, we assessed the technical sample workup procedure for ddPCR on blood plasma samples. Blood samples from healthy individuals, as well as lung cancer patients were analyzed. We compared different methods and protocols for sample collection, storage, centrifugation, isolation, and quantification. Cell-free DNA (cfDNA) concentrations of several wild-type targets and BRAF and EGFR-mutant ctDNA concentrations quantified by ddPCR were primary outcome measurements. Highest cfDNA concentrations were measured in blood collected in serum tubes. No significant differences in cfDNA concentrations were detected between various time points of up to 24 h until centrifugation. Highest cfDNA concentrations were detected after DNA isolation with the Quick cfDNA Serum & Plasma Kit, while plasma isolation using the QIAamp Circulating Nucleic Acid Kit yielded the most consistent results. DdPCR results on cfDNA are highly dependent on multiple factors during preanalytical sample workup, which need to be addressed during the development of this diagnostic tool for cancer diagnostics in the future. © 2017 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  7. Optimizing the pre-referral workup for gastroenterology and hepatology specialty care: consensus using the Delphi method.

    Science.gov (United States)

    Ho, Chanda K; Boscardin, Christy K; Gleason, Nathaniel; Collado, Don; Terdiman, Jonathan; Terrault, Norah A; Gonzales, Ralph

    2016-02-01

    Specialty care referrals have doubled in the last decade. Optimization of the pre-referral workup by a primary care doctor can lead to a more efficient first specialty visit with the patient. Guidance regarding pre-referral laboratory testing is a first step towards improving the specialty referral process. Our aim was to establish consensus regarding appropriate pre-referral workup for common gastrointestinal and liver conditions. The Delphi method was used to establish local consensus for recommending certain laboratory tests prior to specialty referral for 13 clinical conditions. Seven conditions from The University of Michigan outpatient referral guidelines were used as a baseline. An expert panel of three PCPs and nine gastroenterologists from three academic hospitals participated in three iterative rounds of electronic surveys. Each panellist ranked each test using a 5-point Likert scale (strongly disagree to strongly agree). Local panellists could recommend additional tests for the initial diagnoses, and also recommended additional diagnoses needing guidelines: iron deficiency anaemia, abdominal pain, irritable bowel syndrome, fatty liver disease, liver mass and cirrhosis. Consensus was defined as ≥70% of experts scoring ≥4 (agree or strongly agree). Applying Delphi methodology to extrapolate externally developed referral guidelines for local implementation resulted in considerable modifications. For some conditions, many tests from the external group were eliminated by the local group (abdominal bloating; iron deficiency anaemia; irritable bowel syndrome). In contrast, for chronic diarrhoea, abnormal liver enzymes and viral hepatitis, all/most original tests were retained with additional tests added. For liver mass, fatty liver disease and cirrhosis, there was high concordance among the panel with few tests added or eliminated. Consideration of externally developed referral guidelines using a consensus-building process leads to significant local

  8. Denaturation of proteins by surfactants studied by the Taylor dispersion analysis.

    Directory of Open Access Journals (Sweden)

    Aldona Jelińska

    Full Text Available We showed that the Taylor Dispersion Analysis (TDA is a fast and easy to use method for the study of denaturation proteins. We applied TDA to study denaturation of β-lactoglobulin, transferrin, and human insulin by anionic surfactant sodium dodecyl sulfate (SDS. A series of measurements at constant protein concentration (for transferrin was 1.9 x 10-5 M, for β- lactoglobulin was 7.6 x 10-5 M, and for insulin was 1.2 x 10-4 M and varying SDS concentrations were carried out in the phosphate-buffered saline (PBS. The structural changes were analyzed based on the diffusion coefficients of the complexes formed at various surfactant concentrations. The concentration of surfactant was varied in the range from 1.2 x 10-4 M to 8.7 x 10-2 M. We determined the minimum concentration of the surfactant necessary to change the native conformation of the proteins. The minimal concentration of SDS for β-lactoglobulin and transferrin was 4.3 x 10-4 M and for insulin 2.3 x 10-4 M. To evaluate the TDA as a novel method for studying denaturation of proteins we also applied other methods i.e. electronic circular dichroism (ECD and dynamic light scattering (DLS to study the same phenomenon. The results obtained using these methods were in agreement with the results from TDA.

  9. Chemical Denaturants Smoothen Ruggedness on the Free Energy Landscape of Protein Folding.

    Science.gov (United States)

    Malhotra, Pooja; Jethva, Prashant N; Udgaonkar, Jayant B

    2017-08-08

    To characterize experimentally the ruggedness of the free energy landscape of protein folding is challenging, because the distributed small free energy barriers are usually dominated by one, or a few, large activation free energy barriers. This study delineates changes in the roughness of the free energy landscape by making use of the observation that a decrease in ruggedness is accompanied invariably by an increase in folding cooperativity. Hydrogen exchange (HX) coupled to mass spectrometry was used to detect transient sampling of local energy minima and the global unfolded state on the free energy landscape of the small protein single-chain monellin. Under native conditions, local noncooperative openings result in interconversions between Boltzmann-distributed intermediate states, populated on an extremely rugged "uphill" energy landscape. The cooperativity of these interconversions was increased by selectively destabilizing the native state via mutations, and further by the addition of a chemical denaturant. The perturbation of stability alone resulted in seven backbone amide sites exchanging cooperatively. The size of the cooperatively exchanging and/or unfolding unit did not depend on the extent of protein destabilization. Only upon the addition of a denaturant to a destabilized mutant variant did seven additional backbone amide sites exchange cooperatively. Segmentwise analysis of the HX kinetics of the mutant variants further confirmed that the observed increase in cooperativity was due to the smoothing of the ruggedness of the free energy landscape of folding of the protein by the chemical denaturant.

  10. Modified denatured lysozyme effectively solubilizes fullerene c60 nanoparticles in water

    Science.gov (United States)

    Siepi, Marialuisa; Politi, Jane; Dardano, Principia; Amoresano, Angela; De Stefano, Luca; Monti, Daria Maria; Notomista, Eugenio

    2017-08-01

    Fullerenes, allotropic forms of carbon, have very interesting pharmacological effects and engineering applications. However, a very low solubility both in organic solvents and water hinders their use. Fullerene C60, the most studied among fullerenes, can be dissolved in water only in the form of nanoparticles of variable dimensions and limited stability. Here the effect on the production of C60 nanoparticles by a native and denatured hen egg white lysozyme, a highly basic protein, has been systematically studied. In order to obtain a denatured, yet soluble, lysozyme derivative, the four disulfides of the native protein were reduced and exposed cysteines were alkylated by 3-bromopropylamine, thus introducing eight additional positive charges. The C60 solubilizing properties of the modified denatured lysozyme proved to be superior to those of the native protein, allowing the preparation of biocompatible highly homogeneous and stable C60 nanoparticles using lower amounts of protein, as demonstrated by dynamic light scattering, transmission electron microscopy and atomic force microscopy studies. This lysozyme derivative could represent an effective tool for the solubilization of other carbon allotropes.

  11. Evaluation of gasoline-denatured ethanol as a carbon source for denitrification.

    Science.gov (United States)

    Kazasi, Anna; Boardman, Gregory D; Bott, Charles B

    2013-06-01

    In this study concerning denitrification, the performance of three carbon sources, methanol (MeOH), ethanol (EtOH) and gasoline-denatured ethanol (dEtOH), was compared and evaluated on the basis of treatment efficiency, inhibition potential and cost. The gasoline denaturant considered here contained mostly aliphatic compounds and little of the components that typically boost the octane rating, such as benzene, toluene, ethylbenzene and xylenes. Results were obtained using three lab-scale SBRs operated at SRT of 12.0 +/- 0.9 days. After biomass was acclimated, denitrification rates with dEtOH were similar to those of EtOH (201 +/- 50 and 197 +/- 28 NO3-N/g MLVSS x d, respectively), and higher than those of MeOH (165 +/- 49 mg NO3-N/g MLVSS x d). The denaturant did not affect biomass production, nitrification or denitrification. Effluent soluble COD concentrations were always less than the analytical detection limit. Although the cost of dEtOH ($2.00/kg nitrate removed) was somewhat higher than that of methanol ($1.63/kg nitrate removed), the use of dEtOH is very promising and utilities will have to decide if it is worth paying a little extra to take advantage of its benefits.

  12. Comparison of membrane electroporation and protein denature in response to pulsed electric field with different durations.

    Science.gov (United States)

    Huang, Feiran; Fang, Zhihui; Mast, Jason; Chen, Wei

    2013-05-01

    In this paper, we compared the minimum potential differences in the electroporation of membrane lipid bilayers and the denaturation of membrane proteins in response to an intensive pulsed electric field with various pulse durations. Single skeletal muscle fibers were exposed to a pulsed external electric field. The field-induced changes in the membrane integrity (leakage current) and the Na channel currents were monitored to identify the minimum electric field needed to damage the membrane lipid bilayer and the membrane proteins, respectively. We found that in response to a relatively long pulsed electric shock (longer than the membrane intrinsic time constant), a lower membrane potential was needed to electroporate the cell membrane than for denaturing the membrane proteins, while for a short pulse a higher membrane potential was needed. In other words, phospholipid bilayers are more sensitive to the electric field than the membrane proteins for a long pulsed shock, while for a short pulse the proteins become more vulnerable. We can predict that for a short or ultrashort pulsed electric shock, the minimum membrane potential required to start to denature the protein functions in the cell plasma membrane is lower than that which starts to reduce the membrane integrity. Copyright © 2013 Wiley Periodicals, Inc.

  13. Radioimmunoassay and heat denaturation enzyme assay for the detection of Tay-Sachs heterozygotes during pregnancy

    International Nuclear Information System (INIS)

    Nguyen, C.; Gold, R.J.M.; Mahuran, D.; Lowden, J.A.

    1981-01-01

    Tay-Sachs disease results from a loss of activity of hexosaminidase A (HEX A) in body tissues and fluids. Heterozygotes for the disease are usually identified by their relatively low ratio of heat-labile HEX A to total hexosaminidase. During pregnancy an intermediate isoenzyme (HEX I) increases in activity in serum and obscures the heterozygote status. HEX I does not increase in leucocytes, tears and other body tissues but because of technical difficulties in these assays the authors examined the feasibility of using a radioimmunoassay for HEX A. By univariate analysis, the heat denaturation assay gave a lower cost of misclassification for non-pregnant normals while RIA did so for pregnant normals. A combination of both tests led to reduced cost of misclassification compared to either alone. Bayesian analysis of bivariate gaussian density functions for heat denaturation and for radioimmunoassays of HEX isoenzymes was employed to calculate misclassification frequencies. Among the parameters examined, HEX A measured by RIA and % HEX A by heat-denaturation assay were the two having the best discriminatory power. (Auth.)

  14. Isothermal chemical denaturation of large proteins: Path-dependence and irreversibility.

    Science.gov (United States)

    Wafer, Lucas; Kloczewiak, Marek; Polleck, Sharon M; Luo, Yin

    2017-12-15

    State functions (e.g., ΔG) are path independent and quantitatively describe the equilibrium states of a thermodynamic system. Isothermal chemical denaturation (ICD) is often used to extrapolate state function parameters for protein unfolding in native buffer conditions. The approach is prudent when the unfolding/refolding processes are path independent and reversible, but may lead to erroneous results if the processes are not reversible. The reversibility was demonstrated in several early studies for smaller proteins, but was assumed in some reports for large proteins with complex structures. In this work, the unfolding/refolding of several proteins were systematically studied using an automated ICD instrument. It is shown that: (i) the apparent unfolding mechanism and conformational stability of large proteins can be denaturant-dependent, (ii) equilibration times for large proteins are non-trivial and may introduce significant error into calculations of ΔG, (iii) fluorescence emission spectroscopy may not correspond to other methods, such as circular dichroism, when used to measure protein unfolding, and (iv) irreversible unfolding and hysteresis can occur in the absence of aggregation. These results suggest that thorough confirmation of the state functions by, for example, performing refolding experiments or using additional denaturants, is needed when quantitatively studying the thermodynamics of protein unfolding using ICD. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Effects of high pressure freezing (HPF) on denaturation of natural actomyosin extracted from prawn (Metapenaeus ensis).

    Science.gov (United States)

    Cheng, Lina; Sun, Da-Wen; Zhu, Zhiwei; Zhang, Zhihang

    2017-08-15

    Effects of protein denaturation caused by high pressure freezing, involving Pressure-Factors (pressure, time) and Freezing-Factors (temperature, phase transition, recrystallization, ice crystal types), are complicated. In the current study, the conformation and functional changes of natural actomyosin (NAM) under pressure assisted freezing (PAF, 100,150,300,400,500MPa P -20°C/25min ), pressure shift freezing (PSF, 200MPa P -20°C/25min ), and immersion freezing ( 0.1MPa P -20°C/5min ) after pressure was released to 0.1MPa, as compared to normal immersion freezing process (IF, 0.1MPa P -20°C/30min ). Results indicated that PSF ( 200MPa P -20°C/30min ) could reduce the denaturation of frozen NAM and a pressure of 300MPa was the critical point to induce such a denaturation. During the periods of B→D in PSF or B→C→D in PAF, the generation and growth of ice crystals played an important role on changing the secondary and tertiary structure of the treated NAM. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Clinical impact of 18F-FDG-PET/CT in the extra cardiac work-up of patients with infective endocarditis

    DEFF Research Database (Denmark)

    Asmar, Ali; Ozcan, Cengiz; Diederichsen, Axel C P

    2014-01-01

    OBJECTIVE: The purpose of this study was to assess the clinical importance of (18)F-FDG-PET/CT used in the extra cardiac work-up of patients with infective endocarditis (IE). BACKGROUND: IE is a serious condition with a significant mortality. Besides the degree of valvular involvement, the progno......OBJECTIVE: The purpose of this study was to assess the clinical importance of (18)F-FDG-PET/CT used in the extra cardiac work-up of patients with infective endocarditis (IE). BACKGROUND: IE is a serious condition with a significant mortality. Besides the degree of valvular involvement...

  17. Exact method for numerically analyzing a model of local denaturation in superhelically stressed DNA

    International Nuclear Information System (INIS)

    Fye, R.M.; Benham, C.J.

    1999-01-01

    Local denaturation, the separation at specific sites of the two strands comprising the DNA double helix, is one of the most fundamental processes in biology, required to allow the base sequence to be read both in DNA transcription and in replication. In living organisms this process can be mediated by enzymes which regulate the amount of superhelical stress imposed on the DNA. We present a numerically exact technique for analyzing a model of denaturation in superhelically stressed DNA. This approach is capable of predicting the locations and extents of transition in circular superhelical DNA molecules of kilobase lengths and specified base pair sequences. It can also be used for closed loops of DNA which are typically found in vivo to be kilobases long. The analytic method consists of an integration over the DNA twist degrees of freedom followed by the introduction of auxiliary variables to decouple the remaining degrees of freedom, which allows the use of the transfer matrix method. The algorithm implementing our technique requires O(N 2 ) operations and O(N) memory to analyze a DNA domain containing N base pairs. However, to analyze kilobase length DNA molecules it must be implemented in high precision floating point arithmetic. An accelerated algorithm is constructed by imposing an upper bound M on the number of base pairs that can simultaneously denature in a state. This accelerated algorithm requires O(MN) operations, and has an analytically bounded error. Sample calculations show that it achieves high accuracy (greater than 15 decimal digits) with relatively small values of M (M<0.05N) for kilobase length molecules under physiologically relevant conditions. Calculations are performed on the superhelical pBR322 DNA sequence to test the accuracy of the method. With no free parameters in the model, the locations and extents of local denaturation predicted by this analysis are in quantitatively precise agreement with in vitro experimental measurements

  18. Preparation of denatured sup(99m)Tc labeled HSA aerosols of different median diameters for various imaging studies

    Energy Technology Data Exchange (ETDEWEB)

    Raghunath, B.; Kotrappa, P.; Soni, P.S.; Ganatra, R.D. (Bhabha Atomic Research Centre, Bombay (India))

    1982-02-01

    The preparation of denatured sup(99m)Tc-labelled human serum albumin (HSA) aerosols of different median diameters is described using the BARC (Bhabha Atomic Research Centre) dry aerosol generation and delivery system. The applications of these radioactive aerosols are demonstrated in aerosol scintigraphy of lungs, mucociliary movement studies and lymphoscintigraphy in rabbits. It is concluded that the BARC system gives a simplified, rapid and versatile procedure for generation of denatured volume tagged HSA aerosols for a variety of clinical applications.

  19. Fluctuating partially native-like topologies in the acid denatured ensemble of autolysis resistant HIV-1 protease.

    Science.gov (United States)

    Rout, Manoj Kumar; Hosur, Ramakrishna V

    2009-02-01

    Folding, in-vivo, starts from a denatured state and thus the nature of the denatured state would play an important role in directing the folding of a protein. We report here NMR characterization of the acid-denatured state of a mutant of HIV-1 protease, designed to prevent autolysis (Q7K, L33I, L63I) and to prevent cysteine oxidation (C67A and C95A). Secondary chemical shifts, TALOS analysis of chemical shifts and (15)N relaxation data (R(1), R(2), NOE) coupled with AABUF and hydrophobicity calculations, suggest formation of hydrophobic clusters and possibility of some partially native-like topologies in the acid denatured state of the protease. The structural and dynamics characteristics of the acid denatured PR seem to be considerably different from those of the guanidine or urea denatured states of some variants of PR. These would have implications for the folding and auto-processing of the enzyme in-vivo.

  20. [Results from a general training hospital for the implementation of a diagnostic workup for pulmonary embolism according to the Dutch Institute for Health Care Improvement

    NARCIS (Netherlands)

    Kamphuisen, P.W.; Jacobs, E.M.G.; Mol, J.J.; Rijnders, A.J.; Ullmann, E.F.

    2002-01-01

    OBJECTIVE: To evaluate the active implementation of the Dutch Institute for Healthcare Improvement's guideline for the diagnostic work-up for pulmonary embolism in a general training hospital, and to analyse reasons for not following the guideline strategy. DESIGN: Partly retrospective and partly

  1. The use of a rapid D-dimer blood test in the diagnostic work-up for pulmonary embolism: a management study

    NARCIS (Netherlands)

    de Groot, M. R.; van Marwijk Kooy, M.; Pouwels, J. G.; Engelage, A. H.; Kuipers, B. F.; Büller, H. R.

    1999-01-01

    D-dimer assays have a potential to rule out pulmonary embolism in case of a normal test result. We studied the clinical utility of incorporating the SimpliRED D-dimer test result and clinical probability in the routine diagnostic work-up of patients with suspected acute pulmonary embolism. In a

  2. Contribution of CSF cytology in the diagnostic work-up of breast cancer patients with neurological symptoms: a retrospective analysis over two decades.

    NARCIS (Netherlands)

    Wauters, Carla; Poelen, J.; Mulder, I.; Venderink, D.; Strobbe, L.J.; Wesseling, P.

    2012-01-01

    The aim of this study was to evaluate the contribution of cytological analysis of cerebrospinal fluid (CSF) in the diagnostic work-up of breast cancer patients who present with neurological symptoms suspected for central nervous system (CNS) metastases. In the period 1989-2009, a total of 81

  3. Comparison of a gene expression profiling strategy to standard clinical work-up for determination of tumour origin in cancer of unknown primary (CUP)

    DEFF Research Database (Denmark)

    Ades, Felipe; de Azambuja, Evandro; Daugaard, Gedske

    2013-01-01

    CupPrint® is a genomic signature able to identify 47 different cancer types. The aim of our study was to compare the accuracy of this genomic signature to that of a full clinical work-up in diagnosing the primary tumour site. Patients with newly diagnosed, untreated metastatic tumours were eligible...

  4. Children presenting in delayed fashion after minor head trauma with scalp swelling: do they require further workup?

    Science.gov (United States)

    Sellin, Jonathan N; Moreno, Amee; Ryan, Sheila L; Lam, Sandi K; Donaruma-Kwoh, Marcella; Luerssen, Thomas G; Jea, Andrew

    2017-04-01

    It is common to evaluate children who have sustained minor head trauma with computed tomography (CT) of the head. Scalp swelling, in particular, has been associated with intracranial injury. A subset of patients, however, present in delayed fashion, often days after the head trauma, as soft tissue edema progresses and their caregiver notices scalp swelling. We explore the value of further workup in this setting. We conducted a retrospective review of a prospectively collected cohort of children ≤24 months of age presenting to the Texas Children's Hospital with scalp swelling more than 24 h following a head trauma. Cases were collected over a 2-year study period from June 1, 2014 to May 31, 2016. Seventy-six patients comprising 78 patient encounters were included in our study. The mean age at presentation was 8.8 months (range 3 days-24 months). All patients had noncontrast CT of the head as part of their evaluation by emergency medicine, as well as screening for nonaccidental trauma (NAT) by the Child Protection Team. The most common finding on CT head was a linear/nondisplaced skull fracture (SF) with associated extra-axial hemorrhage (epidural or subdural hematoma), which was found in 31/78 patient encounters (40%). Of all 78 patient encounters, 43 patients (55%) were discharged from the emergency room (ER), 17 patients (22%) were admitted for neurologic monitoring, and 18 patients (23%) were admitted solely to allow further NAT evaluation. Of those patients admitted, none experienced a neurologic decline and all had nonfocal neurologic exams on discharge. No patient returned to the ER in delayed fashion for a neurologic decline. Of all the patient encounters, no patient required surgery. Pediatric patients ≤24 months of age presenting to the ER in delayed fashion with scalp swelling after minor head trauma-who were otherwise nonfocal on examination-did not require surgical intervention and did not experience any neurologic decline. Further radiographic

  5. A study of the thermal denaturation of the S-layer protein from Lactobacillus salivarius

    Science.gov (United States)

    Lighezan, Liliana; Georgieva, Ralitsa; Neagu, Adrian

    2012-09-01

    Surface layer (S-layer) proteins display an intrinsic self-assembly property, forming monomolecular crystalline arrays, identified in outermost structures of the cell envelope in many organisms, such as bacteria and archaea. Isolated S-layer proteins also possess the ability to recrystallize into regular lattices, being used in biotechnological applications, such as controlling the architecture of biomimetic surfaces. To this end, the stability of the S-layer proteins under high-temperature conditions is very important. In this study, the S-layer protein has been isolated from Lactobacillus salivarius 16 strain of human origin, and purified by cation-exchange chromatography. Using circular dichroism (CD) spectroscopy, we have investigated the thermal denaturation of the S-layer protein. The far- and near-UV CD spectra have been collected, and the temperature dependence of the CD signal in these spectral domains has been analyzed. The variable temperature results show that the secondary and tertiary structures of the S-layer protein change irreversibly due to the heating of the sample. After the cooling of the heated protein, the secondary and tertiary structures are partially recovered. The denaturation curves show that the protein unfolding depends on the sample concentration and on the heating rate. The secondary and tertiary structures of the protein suffer changes in the same temperature range. We have also detected an intermediate state in the protein denaturation pathway. Our results on the thermal behavior of the S-layer protein may be important for the use of S-layer proteins in biotechnological applications, as well as for a better understanding of the structure and function of S-layer proteins.

  6. Helical Propensity Affects the Conformational Properties of the Denatured State of Cytochrome c'.

    Science.gov (United States)

    Danielson, Travis A; Bowler, Bruce E

    2018-01-23

    Changing the helical propensity of a polypeptide sequence might be expected to affect the conformational properties of the denatured state of a protein. To test this hypothesis, alanines at positions 83 and 87 near the center of helix 3 of cytochrome c' from Rhodopseudomonas palustris were mutated to serine to decrease the stability of this helix. A set of 13 single histidine variants in the A83S/A87S background were prepared to permit assessment of the conformational properties of the denatured state using histidine-loop formation in 3 M guanidine hydrochloride. The data are compared with previous histidine-heme loop formation data for wild-type cytochrome c'. As expected, destabilization of helix 3 decreases the global stabilities of the histidine variants in the A83S/A87S background relative to the wild-type background. Loop stability versus loop size data yields a scaling exponent of 2.1 ± 0.2, similar to the value of 2.3 ± 0.2 obtained for wild-type cytochrome c'. However, the stabilities of all histidine-heme loops, which contain the helix 3 sequence segment, are increased in the A83S/A87S background compared to the wild-type background. Rate constants for histidine-heme loop breakage are similar for the wild-type and A83S/A87S variants. However, for histidine-heme loops that contain the helix 3 sequence segment, the rate constants for loop formation increase in the A83S/A87S background compared to the wild-type background. Thus, residual helical structure appears to stiffen the polypeptide chain slowing loop formation in the denatured state. The implications of these results for protein folding mechanisms are discussed. Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.

  7. Single Molecular Level Probing of Structure and Dynamics of Papain Under Denaturation.

    Science.gov (United States)

    Sengupta, Bhaswati; Chaudhury, Apala; Das, Nilimesh; Sen, Pratik

    2017-01-01

    Papain is a cysteine protease enzyme present in papaya and known to help in digesting peptide. Thus the structure and function of the active site of papain is of interest. The objective of present study is to unveil the overall structural transformation and the local structural change around the active site of papain as a function of chemical denaturant. Papain has been tagged at Cys-25 with a thiol specific fluorescence probe N-(7- dimethylamino-4-methylcoumarin-3-yl) iodoacetamide (DACIA). Guanidine hydrochloride (GnHCl) has been used as the chemical denaturant. Steady state, time-resolved, and single molecular level fluorescence techniques was applied to map the change in the local environment. It is found that papain undergoes a two-step denaturation in the presence of GnHCl. Fluorescence correlation spectroscopic (FCS) data indicate that the size (hydrodynamic diameter) of native papain is ~36.8 Å, which steadily increases to ~53 Å in the presence of 6M GnHCl. FCS study also reveals that the conformational fluctuation time of papain is 6.3 µs in its native state, which decreased to 2.7 µs in the presence of 0.75 M GnHCl. Upon further increase in GnHCl concentration the conformational fluctuation time increase monotonically till 6 M GnHCl, where the time constant is measured as 14 µs. On the other hand, the measurement of ellipticity, hence the helical structure, by circular dichroism spectroscopy is found to be incapable to capture such structural transformation. It is concluded that in the presence of small amount of GnHCl the active site of papain takes up a more compact structure (although the overall size increases) than in the native state, which has been designated as the intermediate state. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  8. Deciphering allogeneic antibody response against native and denatured HLA epitopes in organ transplantation.

    Science.gov (United States)

    Visentin, Jonathan; Guidicelli, Gwendaline; Moreau, Jean-François; Lee, Jar-How; Taupin, Jean-Luc

    2015-07-01

    Anti-HLA donor-specific antibodies are deleterious for organ transplant survival. Class I HLA donor-specific antibodies are identified by using the Luminex single antigen beads (LSAB) assay, which also detects anti-denatured HLA antibodies (anti-dHLAs). Anti-dHLAs are thought to be unable to recognize native HLA (nHLA) on the cell surface and therefore to be clinically irrelevant. Acid denaturation of nHLA on LSAB allows anti-dHLAs to be discriminated from anti-nHLAs. We previously defined a threshold for the ratio between mean fluorescence intensity against acid-treated (D for denaturation) and nontreated (N) LSAB, D ≥ 1.2 N identifying the anti-dHLAs. However, some anti-dHLAs remained able to bind nHLA on lymphocytes in flow cytometry crossmatches, and some anti-nHLAs conserved significant reactivity toward acid-treated LSAB. After depleting serum anti-nHLA reactivity with HLA-typed cells, we analyzed the residual LSAB reactivity toward nontreated and acid-treated LSABs, and then evaluated the ability of antibodies to recognize nHLA alleles individually. We observed that sera can contain mixtures of anti-nHLAs and anti-dHLAs, or anti-nHLAs recognizing acid-resistant epitopes, all possibly targeting the same allele(s). Therefore, the anti-HLA antibody response can be highly complex and subtle, as is the accurate identification of pathogenic anti-HLA antibodies in human serum. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Stabilizing effect of biochar on soil extracellular enzymes after a denaturing stress.

    Science.gov (United States)

    Elzobair, Khalid A; Stromberger, Mary E; Ippolito, James A

    2016-01-01

    Stabilizing extracellular enzymes may maintain enzymatic activity while protecting enzymes from proteolysis and denaturation. A study determined whether a fast pyrolysis hardwood biochar (CQuest™) would reduce evaporative losses, subsequently stabilizing soil extracellular enzymes and prohibiting potential enzymatic activity loss following a denaturing stress (microwaving). Soil was incubated in the presence of biochar (0%, 1%, 2%, 5%, or 10% by wt.) for 36 days and then exposed to microwave energies (0, 400, 800, 1600, or 3200 J g(-1) soil). Soil enzymes (β-glucosidase, β-d-cellobiosidase, N-acetyl-β-glucosaminidase, phosphatase, leucine aminopeptidase, β-xylosidase) were analyzed by fluorescence-based assays. Biochar amendment reduced leucine aminopeptidase and β-xylosidase potential activity after the incubation period and prior to stress exposure. The 10% biochar rate reduced soil water loss at the lowest stress level (400 J microwave energy g(-1) soil). Enzyme stabilization was demonstrated for β-xylosidase; intermediate biochar application rates prevented a complete loss of this enzyme's potential activity after soil was exposed to 400 (1% biochar treatment) or 1600 (5% biochar treatment) J microwave energy g(-1) soil. Remaining enzyme potential activities were not affected by biochar, and activities decreased with increasing stress levels. We concluded that biochar has the potential to reduce evaporative soil water losses and stabilize certain extracellular enzymes where activity is maintained after a denaturing stress; this effect was biochar rate and enzyme dependent. While biochar may reduce the potential activity of certain soil extracellular enzymes, this phenomenon was not universal as the majority of enzymes assayed in this study were unaffected by exposure to biochar. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. On the mobility of partially denatured DNA in gel electrophoresis: a theoretical investigation

    Science.gov (United States)

    Sean, David

    There are technologies which exploit a rapid reduction of the gel electrophoretic mobility of DNA arising from partial denaturation. The underlying phenomenon behind these experiments---the mechanisms which reduce the mobility---are not very well understood. Such is the purpose of my thesis. The first chapter provides a brief introduction to the field of polymer physics. The subjects covered are carefully chosen to directly relate to the forthcoming research. There is a published semi-empirical formula used to model the rapid decrease of mobility which is largely considered to be consistent with experimental data. The second chapter of this thesis demonstrates that there is a fundamental confusion in the literature regarding the fitting parameter Lr, in the said formula. By going back to the original derivation, a physical interpretation can be given to L r. This interpretation yields theoretical values which are consistent with what has been published. However, we find that an underlying assumption---that the effect of the denaturation does not depend on its position along the DNA fragment---may systematically overestimate experimental observations of Lr. To measure the impact of this assumption, a simulation model of DNA is presented. The article presented in the third chapter reveals that indeed, the position of the denatured region affects the migration of the DNA fragment. A refined version of the formula which takes these factors into account is proposed. The simulations also reveal that, for certain fields, an unexpected conformation completely dominates during migration of the fragment. This surprising result: a squid-like conformation, is explored in chapter four.

  11. Brownian dynamics simulations of sequence-dependent duplex denaturation in dynamically superhelical DNA

    Science.gov (United States)

    Mielke, Steven P.; Grønbech-Jensen, Niels; Krishnan, V. V.; Fink, William H.; Benham, Craig J.

    2005-09-01

    The topological state of DNA in vivo is dynamically regulated by a number of processes that involve interactions with bound proteins. In one such process, the tracking of RNA polymerase along the double helix during transcription, restriction of rotational motion of the polymerase and associated structures, generates waves of overtwist downstream and undertwist upstream from the site of transcription. The resulting superhelical stress is often sufficient to drive double-stranded DNA into a denatured state at locations such as promoters and origins of replication, where sequence-specific duplex opening is a prerequisite for biological function. In this way, transcription and other events that actively supercoil the DNA provide a mechanism for dynamically coupling genetic activity with regulatory and other cellular processes. Although computer modeling has provided insight into the equilibrium dynamics of DNA supercoiling, to date no model has appeared for simulating sequence-dependent DNA strand separation under the nonequilibrium conditions imposed by the dynamic introduction of torsional stress. Here, we introduce such a model and present results from an initial set of computer simulations in which the sequences of dynamically superhelical, 147 base pair DNA circles were systematically altered in order to probe the accuracy with which the model can predict location, extent, and time of stress-induced duplex denaturation. The results agree both with well-tested statistical mechanical calculations and with available experimental information. Additionally, we find that sites susceptible to denaturation show a propensity for localizing to supercoil apices, suggesting that base sequence determines locations of strand separation not only through the energetics of interstrand interactions, but also by influencing the geometry of supercoiling.

  12. Denaturation of collagen structures and their transformation under the physical and chemical effects

    Science.gov (United States)

    Ivankin, A.; Boldirev, V.; Fadeev, G.; Baburina, M.; Kulikovskii, A.; Vostrikova, N.

    2017-11-01

    The process of denaturation of collagen structures under the influence of physical and chemical factors play an important role in the manufacture of food technology and the production of drugs for medicine and cosmetology. The paper discussed the problem of the combined effects of heat treatment, mechanical dispersion and ultrasonic action on the structural changes of the animal collagen in the presence of weak protonated organic acids. Algorithm combined effects of physical and chemical factors as a result of the formation of the technological properties of products containing collagen has been shown.

  13. Nanometer-Scale Dissection of Chromosomes by Atomic Force Microscopy Combined with Heat-Denaturing Treatment

    Science.gov (United States)

    Tsukamoto, Kazumi; Kuwazaki, Seigo; Yamamoto, Kimiko; Shichiri, Motoharu; Yoshino, Tomoyuki; Ohtani, Toshio; Sugiyama, Shigeru

    2006-03-01

    We have developed a method for dissecting chromosome fragments with a size of a few hundred nanometers by atomic force microscopy (AFM). By using this method, we demonstrated reproducible dissections of silkworm chromosomes in the pachytene phase. The dissected fragments were successfully recovered on the cantilever tips, as confirmed by fluorescent microscopy using fluorescent stained chromosomes. To recover dissected chromosome fragments from a larger chromosome, such as the human metaphase chromosome of a somatic cell, heat denaturation was found to be effective. Further improvements in this method may lead to a novel tool for isolating valuable genes and/or investigating local genome structures in the near future.

  14. Denaturing high-performance liquid chromatography mutation analysis in patients with reduced Protein S levels

    DEFF Research Database (Denmark)

    Bathum, Lise; Münster, Anna-Marie; Nybo, Mads

    2008-01-01

    diagnosis and risk estimation. The aim was to design a high-throughput genetic analysis based on denaturing high-performance liquid chromatography to identify sequence variations in the gene coding for Protein S. PATIENTS: In total, 55 patients referred to the Section of Thrombosis and Haemostasis, Odense......BACKGROUND: Patients with congenital Protein S deficiency have increased risk of venous thromboembolism. However, Protein S levels show large intra-individual variation and the biochemical assays have low accuracy and a high interlaboratory variability. Genetic analysis might aid in a more precise......, giving a precise diagnosis and subsequently a better risk estimation....

  15. DNA denaturation through a model of the partition points on a one-dimensional lattice

    International Nuclear Information System (INIS)

    Mejdani, R.; Huseini, H.

    1994-08-01

    We have shown that by using a model of the partition points gas on a one-dimensional lattice, we can study, besides the saturation curves obtained before for the enzyme kinetics, also the denaturation process, i.e. the breaking of the hydrogen bonds connecting the two strands, under treatment by heat of DNA. We think that this model, as a very simple model and mathematically transparent, can be advantageous for pedagogic goals or other theoretical investigations in chemistry or modern biology. (author). 29 refs, 4 figs

  16. Screening for mutations in the uroporphyrinogen decarboxylase gene using denaturing gradient gel electrophoresis

    DEFF Research Database (Denmark)

    Christiansen, L; Ged, C; Hombrados, I

    1999-01-01

    to exon skipping, and a 2-bp deletion (415-416delTA) resulting in a frameshift and the introduction of a premature stop codon. Heterologous expression and enzymatic studies of the mutant proteins demonstrate that the three mutations leading to shortening or truncation of the UROD protein have no residual......, confirming the heterogeneity of the underlying genetic defects of these diseases. We have established a denaturing gradient gel electrophoresis (DGGE) assay for mutation detection in the UROD gene, enabling the simultaneous screening for known and unknown mutations. The established assay has proved able...

  17. Effect of heating strategies on whey protein denaturation--Revisited by liquid chromatography quadrupole time-of-flight mass spectrometry.

    Science.gov (United States)

    Akkerman, M; Rauh, V M; Christensen, M; Johansen, L B; Hammershøj, M; Larsen, L B

    2016-01-01

    Previous standards in the area of effect of heat treatment processes on milk protein denaturation were based primarily on laboratory-scale analysis and determination of denaturation degrees by, for example, electrophoresis. In this study, whey protein denaturation was revisited by pilot-scale heating strategies and liquid chromatography quadrupole time-of-flight mass spectrometer (LC/MC Q-TOF) analysis. Skim milk was heat treated by the use of 3 heating strategies, namely plate heat exchanger (PHE), tubular heat exchanger (THE), and direct steam injection (DSI), under various heating temperatures (T) and holding times. The effect of heating strategy on the degree of denaturation of β-lactoglobulin and α-lactalbumin was determined using LC/MC Q-TOF of pH 4.5-soluble whey proteins. Furthermore, effect of heating strategy on the rennet-induced coagulation properties was studied by oscillatory rheometry. In addition, rennet-induced coagulation of heat-treated micellar casein concentrate subjected to PHE was studied. For skim milk, the whey protein denaturation increased significantly as T and holding time increased, regardless of heating method. High denaturation degrees were obtained for T >100°C using PHE and THE, whereas DSI resulted in significantly lower denaturation degrees, compared with PHE and THE. Rennet coagulation properties were impaired by increased T and holding time regardless of heating method, although DSI resulted in less impairment compared with PHE and THE. No significant difference was found between THE and PHE for effect on rennet coagulation time, whereas the curd firming rate was significantly larger for THE compared with PHE. Micellar casein concentrate possessed improved rennet coagulation properties compared with skim milk receiving equal heat treatment. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  18. The expression of miR-125b regulates angiogenesis during the recovery of heat-denatured HUVECs.

    Science.gov (United States)

    Zhou, Situo; Zhang, Pihong; Liang, Pengfei; Huang, Xiaoyuan

    2015-06-01

    In previous studies we found that miR-125b was down-regulated in denatured dermis of deep partial thickness burn patients. Moreover, miR-125b inhibited tumor-angiogenesis associated with the decrease of ERBB2 and VEGF expression in ovarian cancer cells and breast cancer cells, etc. In this study, we investigated the expression patterns and roles of miR-125b during the recovery of denatured dermis and heat-denatured human umbilical vein endothelial cells (HUVECs). Deep partial thickness burns in Sprague-Dawley rats and the heat-denatured cells (52°C, 35 s) were used for analysis. Western blot analysis and real-time PCR were applied to evaluate the expression of miR-125b and ERBB2 and VEGF. The ability of angiogenesis in heat-denatured HUVECs was analyzed by scratch wound healing and tube formation assay after pri-miR-125b or anti-miR-125b transfection. miR-125b expression was time-dependent during the recovery of heat-denatured dermis and HUVECs. Moreover, miR-125b regulated ERBB2 mRNA and Protein Expression and regulated angiogenesis association with regulating the expression of VEGF in heat-denatured HUVECs. Taken together our results show that the expression of miR-125b is time-dependent and miR-125b plays a regulatory role of angiogenesis during wound healing after burns. Copyright © 2014 Elsevier Ltd and ISBI. All rights reserved.

  19. Breast cancer: determining the genetic profile from ultrasound-guided percutaneous biopsy specimens obtained during the diagnostic workups.

    Science.gov (United States)

    López Ruiz, J A; Zabalza Estévez, I; Mieza Arana, J A

    2016-01-01

    To evaluate the possibility of determining the genetic profile of primary malignant tumors of the breast from specimens obtained by ultrasound-guided percutaneous biopsies during the diagnostic imaging workup. This is a retrospective study in 13 consecutive patients diagnosed with invasive breast cancer by B-mode ultrasound-guided 12 G core needle biopsy. After clinical indication, the pathologist decided whether the paraffin block specimens seemed suitable (on the basis of tumor size, validity of the sample, and percentage of tumor cells) before sending them for genetic analysis with the MammaPrint® platform. The size of the tumors on ultrasound ranged from 0.6cm to 5cm. In 11 patients the preserved specimen was considered valid and suitable for use in determining the genetic profile. In 1 patient (with a 1cm tumor) the pathologist decided that it was necessary to repeat the core biopsy to obtain additional samples. In 1 patient (with a 5cm tumor) the specimen was not considered valid by the genetic laboratory. The percentage of tumor cells in the samples ranged from 60% to 70%. In 11/13 cases (84.62%) it was possible to do the genetic analysis on the previously diagnosed samples. In most cases, regardless of tumor size, it is possible to obtain the genetic profile from tissue specimens obtained with ultrasound-guided 12 G core biopsy preserved in paraffin blocks. Copyright © 2015 SERAM. Published by Elsevier España, S.L.U. All rights reserved.

  20. Role of Fluorine-18-Fluorodeoxyglucose in the Work-up of Febrile AIDS Patients. Experience with Dual Head Coincidence Imaging.

    Science.gov (United States)

    Santiago, Jonas F.; Jana, Suman; Gilbert, Holly M.; Salem, Shahenda; Bellman, Paul Curtis; Hsu, Ricky K.S.; Naddaf, Sleiman; Abdel-Dayem, Hussein M.

    1999-11-01

    OBJECTIVE AND METHODS: This study was undertaken to find the role of fluorine-18-fluorodeoxyglucose (F18-FDG) in the diagnostic work-up of febrile Acquired Immune Deficiency Syndrome (AIDS) patients. Forty-seven (42 male and 5 female; mean age = 40.3 years) febrile patients with AIDS underwent imaging with F18-FDG by Dual Head Coincidence Imaging (DHCI). Findings were correlated with other imaging modalities.RESULTS: Our data show good sensitivity for scanning with F18-FDG by DHCI in determining the extent of Castleman's disease, lymphoma, Kaposi's sarcoma (KS), adenocarcinoma, and germ cell carcinoma. Various opportunistic infections also manifest with increased F18-FDG uptake.CONCLUSION: Total-body imaging can be done with F18-FDG with better resolution and a shorter procedure time compared to imaging with Gallium-67 (Ga-67). Furthermore, F18-FDG is more sensitive than Ga-67 for evaluating extent of involvement in various pathologies affecting AIDS patients. The new technology of DHCI is a good alternative for hospitals with no dedicated positron emission tomography (PET) scanner.

  1. Opportunities to deter transplant tourism exist before referral for transplantation and during the workup and management of transplant candidates.

    Science.gov (United States)

    Gill, Jagbir; Diec, Olivier; Landsberg, David N; Rose, Caren; Johnston, Olwyn; Keown, Paul A; Gill, John S

    2011-05-01

    Transplant tourism is a global issue, and physicians in the developed world may be in a position to actively deter this practice. To examine such opportunities, we identified 93 residents of British Columbia, Canada who had a kidney graft through tourism and determined their previous interactions with our transplant programs. These patients were mainly ethnic minorities (90%) who traveled to their country of origin for transplantation. Many tourists were transplanted early in their disease course, with 27 having a preemptive transplant. Among the 65 tourists referred for transplant, 33 failed to complete the evaluation. All tourists who completed an evaluation were placed on a waiting list in British Columbia and, after waiting a median of 2 years, pursued tourism. Most of these patients (62%) had a potential living donor, but none had an approved donor, with 13 donors found medically unsuitable, 8 ABO incompatible, and 12 who did not complete their evaluation. Thus, strategies to deter tourism should start before the development of end-stage renal disease and should be part of pretransplant workup and wait-list management, focusing on patients not progressing through their evaluation, those with a declined living donor, and those facing longer wait times, as these groups appear to be at higher risks for transplant tourism. Further studies are needed to identify individuals at risk for transplant tourism and to define effective strategies to deter these individuals.

  2. The role of PET in staging work-up and evaluation after therapy in patients with malignant lymphoma

    International Nuclear Information System (INIS)

    Kang, Yoon Koo; Ryoo, Baek Yeol

    1998-12-01

    The stage of disease in patients with malignant lymphoma in important in decision of treatment modality and in prediction of prognosis. The authors evaluated the usefulness of PET with F18-FDG in initial staging, in reevaluation after radical therapy and in diagnosis of recurrence or disease progression of malignant lymphoma, compared with conventional imaging studies. The efficacy of positron emission tomography (PET) with fluorine-18-fluorodeoxyglucose (F18-FDG) was evaluated in several tumors. In malignant lymphoma. It was reported that the concentration of FDG was increased in tumor tissues, and that PET with F18-FDG was more sensitive and specific in staging evaluation compared with CT scan. The visual analysis of FDG-PET would be helpful in initial staging work-up, in reevaluation of residual disease after radical therapy and in suspicious recurrence or disease progression, determining the residual disease of lymphoma after radical therapy. But more studies with larger number of cases and longer follow-up were required. The results of this study can be bases for the direction of future studies for the usefulness of PET in malignant lymphoma

  3. Clinical value of Tl per rectum scintigraphy in the work-up of patients with alcoholic liver disease

    Energy Technology Data Exchange (ETDEWEB)

    Urbain, D.; Reding, P.; Georges, B.; Thys, O.; Ham, H.R.

    1986-09-01

    The clinical value of thallium 201 per rectum scintigraphy in the work-up of patients with alcoholic liver disease was evaluated using data obtained in 104 patients. The 25th min ratio of heart to liver activities was used as an index of portal systemic shunting. This ratio was found to be normal in alcoholic patients with normal liver biopsy and also in those presenting only steatosis. It was slightly higher in patients with liver fibrosis and significantly higher values were observed in patients with liver cirrhosis. High values of the ratio were associated with a higher risk of portal systemic encephalopathy and/or gastrointestinal bleeding. The prognostic value of the test was supported by the fact that good correlations were observed between the ratio and widely accepted prognostic scores such as the Child score or the Orrego index. Moreover, high ratios were associated with an increased mortality risk at one year. We conclude that this simple test is interesting in the screening of cirrhotics at risk of encephalopathy, gastrointestinal hemorrhage, or early death.

  4. The clinical value of 201Tl per rectum scintigraphy in the work-up of patients with alcoholic liver disease

    International Nuclear Information System (INIS)

    Urbain, D.; Reding, P.; Georges, B.; Thys, O.; Ham, H.R.; Vrije Univ., Brussels

    1986-01-01

    The clinical value of thallium 201 per rectum scintigraphy in the work-up of patients with alcoholic liver disease was evaluated using data obtained in 104 patients. The 25th min ratio of heart to liver activities was used as an index of portal systemic shunting. This ratio was found to be normal in alcoholic patients with normal liver biopsy and also in those presenting only steatosis. It was slightly higher in patients with liver fibrosis and significantly higher values were observed in patients with liver cirrhosis. High values of the ratio were associated with a higher risk of portal systemic encephalopathy and/or gastrointestinal bleeding. The prognostic value of the test was supported by the fact that good correlations were observed between the ratio and widely accepted prognostic scores such as the Child score or the Orrego index. Moreover, high ratios were associated with an increased mortality risk at one year. We conclude that this simple test is interesting in the screening of cirrhotics at risk of encephalopathy, gastrointestinal hemorrhage, or early death. (orig.)

  5. Liver CT for vascular mapping during radioembolisation workup: comparison of an early and late arterial phase protocol

    Energy Technology Data Exchange (ETDEWEB)

    Hoven, Andor F. van den; Braat, Manon N.G.J.A.; Prince, Jip F.; Doormaal, Pieter J. van; Leeuwen, Maarten S. van; Lam, Marnix G.E.H.; Bosch, Maurice A.A.J. van den [University Medical Center Utrecht, Department of Radiology and Nuclear Medicine, Utrecht (Netherlands)

    2017-01-15

    To compare right gastric (RGA) and segment 4 artery (A4) origin detection rates during radioembolisation workup between early and late arterial phase liver CT protocols. 100 consecutive patients who underwent liver CT between May 2012-January 2015 with early or late arterial phase protocol (n = 50 each, 10- vs. 20-s post-threshold delay) were included. RGA/A4 origin detection rates, assessed by two raters, and contrast-to-noise ratio (CNR) of the hepatic artery relative to the portal vein were compared between the protocols. The first-second rater scored the RGA origin as visible in 58-65 % (specific proportion of agreement 82 %, κ = 0.62); A4 origin in 96-89 % (94 %, κ = 0.54). Thirty-six percent of RGA origins not detectable by DSA were identified on CT. Origin detection rates were not significantly different for early/late arterial phases. Mean CNR was higher in the early arterial phase protocol (1.7 vs. 1.2, p < 0.001). A 10-s delay arterial phase CT protocol does not significantly improve detection of small intra- and extrahepatic branches. RGA origin detection requires further optimization, whereas A4/MHA origin detection is adequate, with good inter-rater reproducibility. CT remains important for preprocedural planning, because it may reveal arterial anatomy not discernible on DSA. (orig.)

  6. Diagnostic work-up of neurological syndromes in a rural African setting: knowledge, attitudes and practices of health care providers.

    Directory of Open Access Journals (Sweden)

    Alain Mpanya

    Full Text Available BACKGROUND: Neurological disorders of infectious origin are common in rural sub-Saharan Africa and usually have serious consequences. Unfortunately, these syndromes are often poorly documented for lack of diagnostic tools. Clinical management of these diseases is a major challenge in under-equipped rural health centers and hospitals. We documented health care provider knowledge, attitudes and practices related to this syndrome in two rural health zones in Bandundu Province, Democratic Republic of Congo. METHODS: We used a qualitative research approach combining observation, in-depth interviews and focus group discussions. We observed 20 patient-provider contacts related to a neurological syndrome, conducted 12 individual interviews and 4 focus group discussions with care providers. All interviews were audiotaped and the transcripts were analyzed with the software ATLAS.ti. RESULTS: Care providers in this region usually limit their diagnostic work-up to clinical examination primarily because of the financial hurdles in this entirely out-of-pocket payment system. The patients prefer to purchase drugs rather than diagnostic tests. Moreover the general lack of diagnostic tools and the representation of the clinician as a "diviner" do not enhance any use of laboratory or other diagnostic methods. CONCLUSION: Innovation in diagnostic technology for neurological disorders is badly needed in Central-Africa, but its uptake in clinical practice will only be a success if tools are simple, affordable and embedded in a patient-centered approach.

  7. Detection of AGEs as markers for carbohydrate metabolism and protein denaturation.

    Science.gov (United States)

    Nagai, Ryoji; Shirakawa, Jun-Ichi; Fujiwara, Yukio; Ohno, Rei-Ichi; Moroishi, Narumi; Sakata, Noriyuki; Nagai, Mime

    2014-07-01

    Approximately 100 years have passed since the Maillard reaction was first reported in the field of food chemistry as a condensation reaction between reducing sugars and amino acids. This reaction is thought to progress slowly primarily from glucose with proteins in vivo. An early-stage product, called the "Amadori product", is converted into advanced glycation end products. Those accumulate in the body in accordance with age, with such accumulation being enhanced by lifestyle-related diseases that result in the denaturation of proteins. Recent studies have demonstrated that intermediate carbonyls are generated by several pathways, and rapidly generate many glycation products. However, accurate quantification of glycation products in vivo is difficult due to instability and differences in physicochemical properties. In this connection, little is known about the relationship between the structure of glycation products and pathology. Furthermore, the interaction between proteins modified by glycation and receptors for advanced glycation end products is also known to induce the production of several inflammatory cytokines. Therefore, those inhibitors have been developed over the world to prevent lifestyle-related diseases. In this review, we describe the process of protein denaturation induced by glycation and discuss the possibility of using the process as a marker of age-related diseases.

  8. Heat capacity changes in RNA folding: application of perturbation theory to hammerhead ribozyme cold denaturation.

    Science.gov (United States)

    Mikulecky, Peter J; Feig, Andrew L

    2004-01-01

    In proteins, empirical correlations have shown that changes in heat capacity (DeltaC(P)) scale linearly with the hydrophobic surface area buried upon folding. The influence of DeltaC(P) on RNA folding has been widely overlooked and is poorly understood. In addition to considerations of solvent reorganization, electrostatic effects might contribute to DeltaC(P)s of folding in polyanionic species such as RNAs. Here, we employ a perturbation method based on electrostatic theory to probe the hot and cold denaturation behavior of the hammerhead ribozyme. This treatment avoids much of the error associated with imposing two-state folding models on non-two-state systems. Ribozyme stability is perturbed across a matrix of solvent conditions by varying the concentration of NaCl and methanol co-solvent. Temperature-dependent unfolding is then monitored by circular dichroism spectroscopy. The resulting array of unfolding transitions can be used to calculate a DeltaC(P) of folding that accurately predicts the observed cold denaturation temperature. We confirm the accuracy of the calculated DeltaC(P) by using isothermal titration calorimetry, and also demonstrate a methanol-dependence of the DeltaC(P). We weigh the strengths and limitations of this method for determining DeltaC(P) values. Finally, we discuss the data in light of the physical origins of the DeltaC(P)s for RNA folding and consider their impact on biological function.

  9. Thermal and chemical denaturation of Bacillus circulans xylanase: A biophysical chemistry laboratory module.

    Science.gov (United States)

    Raabe, Richard; Gentile, Lisa

    2008-11-01

    A number of institutions have been, or are in the process of, modifying their biochemistry major to include some emphasis on the quantitative physical chemistry of biomolecules. Sometimes this is done as a replacement for part for the entire physical chemistry requirement, while at other institutions this is incorporated as a component into the traditional two-semester biochemistry series. The latter is the model used for biochemistry and molecular biology majors at the University of Richmond, whose second semester of biochemistry is a course entitled Proteins: Structure, Function, and Biophysics. What is described herein is a protein thermodynamics laboratory module, using the protein Bacillus circulans xylanase, which reinforces many lecture concepts, including: (i) the denatured (D) state ensemble of a protein can be different, depending on how it was populated; (ii) intermediate states may be detected by some spectroscopic techniques but not by others; (iii) the use and assumptions of the van't Hoff approach to calculate ΔH(o) , ΔS(o) , and ΔG(o) (T) for thermal protein unfolding transitions; and (iv) the use and assumptions of an approach that allows determination of the Gibb's free energy of a protein unfolding transition based on the linear dependence of ΔG(o) on the concentration of denaturant used. This module also requires students to design their own experimental protocols and spend time in the primary literature, both important parts of an upper division lab. Copyright © 2008 International Union of Biochemistry and Molecular Biology, Inc.

  10. Approach to analysis of single nucleotide polymorphisms by automated constant denaturant capillary electrophoresis

    International Nuclear Information System (INIS)

    Bjoerheim, Jens; Abrahamsen, Torveig Weum; Kristensen, Annette Torgunrud; Gaudernack, Gustav; Ekstroem, Per O.

    2003-01-01

    Melting gel techniques have proven to be amenable and powerful tools in point mutation and single nucleotide polymorphism (SNP) analysis. With the introduction of commercially available capillary electrophoresis instruments, a partly automated platform for denaturant capillary electrophoresis with potential for routine screening of selected target sequences has been established. The aim of this article is to demonstrate the use of automated constant denaturant capillary electrophoresis (ACDCE) in single nucleotide polymorphism analysis of various target sequences. Optimal analysis conditions for different single nucleotide polymorphisms on ACDCE are evaluated with the Poland algorithm. Laboratory procedures include only PCR and electrophoresis. For direct genotyping of individual SNPs, the samples are analyzed with an internal standard and the alleles are identified by co-migration of sample and standard peaks. In conclusion, SNPs suitable for melting gel analysis based on theoretical thermodynamics were separated by ACDCE under appropriate conditions. With this instrumentation (ABI 310 Genetic Analyzer), 48 samples could be analyzed without any intervention. Several institutions have capillary instrumentation in-house, thus making this SNP analysis method accessible to large groups of researchers without any need for instrument modification

  11. Denatured protein-coated docetaxel nanoparticles: Alterable drug state and cytosolic delivery.

    Science.gov (United States)

    Zhang, Li; Xiao, Qingqing; Wang, Yiran; Zhang, Chenshuang; He, Wei; Yin, Lifang

    2017-05-15

    Many lead compounds have a low solubility in water, which substantially hinders their clinical application. Nanosuspensions have been considered a promising strategy for the delivery of water-insoluble drugs. Here, denatured soy protein isolate (SPI)-coated docetaxel nanosuspensions (DTX-NS) were developed using an anti-solvent precipitation-ultrasonication method to improve the water-solubility of DTX, thus improving its intracellular delivery. DTX-NS, with a diameter of 150-250nm and drug-loading up to 18.18%, were successfully prepared by coating drug particles with SPI. Interestingly, the drug state of DTX-NS was alterable. Amorphous drug nanoparticles were obtained at low drug-loading, whereas at a high drug-loading, the DTX-NS drug was mainly present in the crystalline state. Moreover, DTX-NS could be internalized at high levels by cancer cells and enter the cytosol by lysosomal escape, enhancing cell cytotoxicity and apoptosis compared with free DTX. Taken together, denatured SPI has a strong stabilization effect on nanosuspensions, and the drug state in SPI-coated nanosuspensions is alterable by changing the drug-loading. Moreover, DTX-NS could achieve cytosolic delivery, generating enhanced cell cytotoxicity against cancer cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Effects of thermally induced denaturation on technological-functional properties of whey protein isolate-based films.

    Science.gov (United States)

    Schmid, M; Krimmel, B; Grupa, U; Noller, K

    2014-09-01

    This study examined how and to what extent the degree of denaturation affected the technological-functional properties of whey protein isolate (WPI)-based coatings. It was observed that denaturation affected the material properties of WPI-coated films significantly. Surface energy decreased by approximately 20% compared with native coatings. Because the surface energy of a coating should be lower than that of the substrate, this might result in enhanced wettability characteristics between WPI-based solution and substrate surface. Water vapor barrier properties increased by about 35% and oxygen barrier properties increased by approximately 33%. However, significant differences were mainly observed between coatings made of fully native WPI and ones with a degree of denaturation of 25%. Higher degrees of denaturation did not lead to further improvement of material properties. This observation offers cost-saving potential: a major share of denatured whey proteins may be replaced by fully native ones that are not exposed to energy-intensive heat treatment. Furthermore, native WPI solutions can be produced with higher dry matter content without gelatinizing. Hence, less moisture has to be removed through drying, resulting in reduced energy consumption. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  13. Ion-ion interactions in the denatured state contribute to the stabilization of CutA1 proteins.

    Science.gov (United States)

    Yutani, Katsuhide; Matsuura, Yoshinori; Naitow, Hisashi; Joti, Yasumasa

    2018-05-16

    In order to elucidate features of the denatured state ensembles that exist in equilibrium with the native state under physiological conditions, we performed 1.4-μs molecular dynamics (MD) simulations at 400 K and 450 K using the monomer subunits of three CutA1 mutants from Escherichia coli: an SH-free mutant (Ec0SH) with denaturation temperature (T d ) = 85.6 °C, a hydrophobic mutant (Ec0VV) with T d  = 113.3 °C, and an ionic mutant (Ec0VV_6) with T d  = 136.8 °C. The occupancy of salt bridges by the six substituted charged residues in Ec0VV_6 was 140.1% at 300 K and 89.5% at 450 K, indicating that even in the denatured state, salt bridge occupancy was high, approximately 60% of that at 300 K. From these results, we can infer that proteins from hyperthermophiles with a high ratio of charged residues are stabilized by a decrease in conformational entropy due to ion-ion interactions in the denatured state. The mechanism must be comparable to the stabilization conferred by disulfide bonds within a protein. This suggests that introduction of charged residues, to promote formation of salt bridges in the denatured state, would be a simple way to rationally design stability-enhanced mutants.

  14. Is thyroid hormones evaluation of clinical value in the work-up of males of infertile couples?

    Science.gov (United States)

    Lotti, F; Maseroli, E; Fralassi, N; Degl'Innocenti, S; Boni, L; Baldi, E; Maggi, M

    2016-03-01

    Is thyroid hormones (TH) evaluation of clinical value in the work-up of males of infertile couples? Our results suggest that TH evaluation is not mandatory in the work-up of male infertility. A few previous studies performed on a limited series of subjects reported a negative impact of hyper- and hypo-thyroidism on semen volume, sperm concentration, progressive motility and normal morphology. No previous study has systematically evaluated associations between TH variation, semen parameters and ultrasound characteristics of the male genital tract. Cross-sectional analysis of a consecutive series of 172 subjects seeking medical care for couple infertility from September 2010 to November 2014. Of the entire cohort, 163 men (age 38.9 ± 8.0 years) free of genetic abnormalities were studied. All subjects underwent a complete andrological and physical examination, biochemical and hormonal assessment, scrotal and transrectal colour-Doppler ultrasound (CDUS) and semen analysis (including seminal interleukin 8 levels, sIL-8) evaluation within the same day. Among the patients studied, 145 (88.9%) showed euthyroidism, 6 (3.7%) subclinical hyper- and 12 (7.4%) subclinical hypo-thyroidism. No subjects showed overt hyper- or hypo-thyroidism. At univariate analysis, no associations among thyroid-stimulating hormone (TSH) or TH levels and sperm parameters were observed. Conversely, we observed positive associations among free triiodothyronine (fT3) and free thyroxine (fT4) levels, ejaculate volume and seminal fructose levels. In a multivariate model, after adjusting for confounders such as age, body mass index, smoking habit, sexual abstinence, calculated free testosterone, prolactin and sIL-8 levels, only the associations found for fT3 levels were confirmed. When CDUS features were investigated, using the same multivariate model, we found positive associations between fT3 levels and seminal vesicles (SV) volume, both before and after ejaculation (adj. r = 0.354 and adj. r = 0

  15. Signal Peptide and Denaturing Temperature are Critical Factors for Efficient Mammalian Expression and Immunoblotting of Cannabinoid Receptors*

    Science.gov (United States)

    WANG, Chenyun; WANG, Yingying; WANG, Miao; CHEN, Jiankui; YU, Nong; SONG, Shiping; KAMINSKI, Norbert E.; ZHANG, Wei

    2013-01-01

    Summary Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports. In present study, we demonstrated cannabinoid receptor protein was not able to be properly expressed with routine mammalian expression system. This inefficient expression was rescued by endowing an exogenous signal peptide ahead of cannabinoid receptor peptide. In addition, the artificially synthesized cannabinoid receptor was found to aggregate under routine sample denaturing temperatures (i.e., ≥95°C), forming a large molecular weight band when analyzed by immunoblotting. Only denaturing temperatures ≤75°C yielded a clear band at the predicted molecular weight. Collectively, we showed that efficient mammalian expression of cannabinoid receptors need a signal peptide sequence, and described the requirement for a low sample denaturing temperature in immunoblot analysis. These findings provide very useful information for efficient mammalian expression and immunoblotting of membrane receptors. PMID:22528237

  16. Effective non-denaturing purification method for improving the solubility of recombinant actin-binding proteins produced by bacterial expression.

    Science.gov (United States)

    Chung, Jeong Min; Lee, Sangmin; Jung, Hyun Suk

    2017-05-01

    Bacterial expression is commonly used to produce recombinant and truncated mutant eukaryotic proteins. However, heterologous protein expression may render synthesized proteins insoluble. The conventional method used to express a poorly soluble protein, which involves denaturation and refolding, is time-consuming and inefficient. There are several non-denaturing approaches that can increase the solubility of recombinant proteins that include using different bacterial cell strains, altering the time of induction, lowering the incubation temperature, and employing different detergents for purification. In this study, we compared several non-denaturing protocols to express and purify two insoluble 34 kDa actin-bundling protein mutants. The solubility of the mutant proteins was not affected by any of the approaches except for treatment with the detergent sarkosyl. These results indicate that sarkosyl can effectively improve the solubility of insoluble proteins during bacterial expression. Copyright © 2016. Published by Elsevier Inc.

  17. The use of 4D-CTA in the diagnostic work-up of brain arteriovenous malformations

    Energy Technology Data Exchange (ETDEWEB)

    Willems, Peter W.A. [Toronto Western Hospital, UHN, Division of Neuroradiology, Department of Medical Imaging, Toronto, Ontario (Canada); Leiden University Medical Center, Department of Radiology, Leiden (Netherlands); Taeshineetanakul, Patamintita; Terbrugge, Karel G.; Krings, Timo [Toronto Western Hospital, UHN, Division of Neuroradiology, Department of Medical Imaging, Toronto, Ontario (Canada); Schenk, Barry; Brouwer, Patrick A. [Leiden University Medical Center, Department of Radiology, Leiden (Netherlands)

    2012-02-15

    We aimed to evaluate the use of time-resolved whole-head CT angiography (4D-CTA) in patients with an untreated arteriovenous malformation of the brain (bAVM), as demonstrated by catheter angiography (DSA). Seventeen patients with a DSA-proven bAVM were enrolled. These were subjected to 4D-CTA imaging using a 320 detector row CT scanner. Using a standardized scoring sheet, all studies were analyzed by a panel of three readers. This panel was blind to the DSA results at the time of reading the 4D-CTA. 4D-CTA detected all bAVMs. With regard to the Spetzler-Martin grade, 4D-CTA disagreed with DSA in only one case, where deep venous drainage was missed. Further discrepancies between 4D-CTA and DSA analyses included underestimation of the nidus size in small lesions (four cases), misinterpretation of a feeding vessel (one case), misinterpretation of indirect feeding through pial collaterals (three cases) and oversight of mild arterial enlargement (two cases). 4D-CTA correctly distinguished low-flow from high-flow lesions and detected dural/transosseous feeding (one case), venous narrowing (one case) and venous pouches (nine cases). In this series, 4D-CTA was able to detect all bAVMs. Although some angioarchitectural details were missed or misinterpreted when compared to DSA, 4D-CTA evaluation was sufficiently accurate to diagnose the shunt and classify it. Moreover, 4D-CTA adds cross-sectional imaging and perfusion maps, helpful in treatment planning. 4D-CTA appears to be a valuable new adjunct in the non-invasive diagnostic work-up of bAVMs and their follow-up when managed conservatively. (orig.)

  18. The use of 4D-CTA in the diagnostic work-up of brain arteriovenous malformations

    International Nuclear Information System (INIS)

    Willems, Peter W.A.; Taeshineetanakul, Patamintita; Terbrugge, Karel G.; Krings, Timo; Schenk, Barry; Brouwer, Patrick A.

    2012-01-01

    We aimed to evaluate the use of time-resolved whole-head CT angiography (4D-CTA) in patients with an untreated arteriovenous malformation of the brain (bAVM), as demonstrated by catheter angiography (DSA). Seventeen patients with a DSA-proven bAVM were enrolled. These were subjected to 4D-CTA imaging using a 320 detector row CT scanner. Using a standardized scoring sheet, all studies were analyzed by a panel of three readers. This panel was blind to the DSA results at the time of reading the 4D-CTA. 4D-CTA detected all bAVMs. With regard to the Spetzler-Martin grade, 4D-CTA disagreed with DSA in only one case, where deep venous drainage was missed. Further discrepancies between 4D-CTA and DSA analyses included underestimation of the nidus size in small lesions (four cases), misinterpretation of a feeding vessel (one case), misinterpretation of indirect feeding through pial collaterals (three cases) and oversight of mild arterial enlargement (two cases). 4D-CTA correctly distinguished low-flow from high-flow lesions and detected dural/transosseous feeding (one case), venous narrowing (one case) and venous pouches (nine cases). In this series, 4D-CTA was able to detect all bAVMs. Although some angioarchitectural details were missed or misinterpreted when compared to DSA, 4D-CTA evaluation was sufficiently accurate to diagnose the shunt and classify it. Moreover, 4D-CTA adds cross-sectional imaging and perfusion maps, helpful in treatment planning. 4D-CTA appears to be a valuable new adjunct in the non-invasive diagnostic work-up of bAVMs and their follow-up when managed conservatively. (orig.)

  19. No Value for Routine Chest Radiography in the Work-Up of Early Stage Cervical Cancer Patients

    Science.gov (United States)

    Hoogendam, Jacob P.; Zweemer, Ronald P.; Verkooijen, Helena M.; de Jong, Pim A.; van den Bosch, Maurice A. A. J.; Verheijen, René H. M.; Veldhuis, Wouter B.

    2015-01-01

    Aim Evidence supporting the recommendation to include chest radiography in the work-up of all cervical cancer patients is limited. We investigated the diagnostic value of routine chest radiography in cervical cancer staging. Methods All consecutive cervical cancer patients who presented at our tertiary referral center in the Netherlands (January 2006 – September 2013), and for whom ≥6 months follow-up was available, were included. As part of the staging procedure, patients underwent a routine two-directional digital chest radiograph. Findings were compared to a composite reference standard consisting of all imaging studies and histology obtained during the 6 months following radiography. Results Of the 402 women who presented with cervical cancer, 288 (71.6%) underwent chest radiography and had ≥6 months follow-up. Early clinical stage (I/II) cervical cancer was present in 244/288 (84.7%) women, while 44 (15.3%) presented with advanced disease (stage III/IV). The chest radiograph of 1 woman – with advanced pre-radiograph stage (IVA) disease – showed findings consistent with pulmonary metastases. Radiographs of 7 other women – 4 early, 3 advanced stage disease – were suspicious for pulmonary metastases which was confirmed by additional imaging in only 1 woman (with pre-radiograph advanced stage (IIIB) disease) and excluded in 6 cases, including all women with early stage disease. In none of the 288 women were thoracic skeletal metastases identified on imaging or during 6 months follow up. Radiography was unremarkable in 76.4% of the study population, and showed findings unrelated to the cervical carcinoma in 21.2%. Conclusion Routine chest radiography was of no value for any of the early stage cervical cancer patients presenting at our tertiary center over a period of 7.7 years. PMID:26135733

  20. Male infertility workup needs additional testing of expressed prostatic secretion and/or post-massage urine.

    Directory of Open Access Journals (Sweden)

    Margus Punab

    Full Text Available The male factor accounts for almost 50% of infertility cases. Inflammation may reduce semen quality via several pathways, including oxidative stress (OxS. As male infertility routinely is assessed using semen analysis only, the possible presence of non-leukocytospermic asymptomatic inflammatory prostatitis may be overlooked. We compared local and systemic OxS levels in male partners of infertile couples with different inflammation patterns in their genital tract and/or oligospermia. Subjects (n=143 were grouped according to inflammation in their semen, expressed prostatic secretion (EPS, and/or post-massage urine (post-M. Systemic (8-isoprostanes in urine and local (diene conjugates and total antioxidant capacity in seminal plasma OxS was measured The levels of OxS markers were significantly elevated in both severe inflammation groups--leukocytospermic men and subjects whose inflammation was limited only to EPS and/or post-M. Comparison between oligospermic and non-oligospermic men with genital tract inflammation, and oligozoospermic men with or without inflammation in the genital tract indicated that inflammation but not oligospermia status had significant impact on the measured OxS markers. Hence, a high leukocyte count in prostate-specific materials (EPS, post-M, even in absence of clear leukocytopsermia, is an important source of local and systemic OxS that may be associated with male infertility and affect general health. We suggest including the tests for detection of inflammation of the prostate into the workup of infertile men as was suggested in the WHO 1993 recommendation.

  1. Validating the Modified Drug Adherence Work-Up (M-DRAW) Tool to Identify and Address Barriers to Medication Adherence.

    Science.gov (United States)

    Lee, Sun; Bae, Yuna H; Worley, Marcia; Law, Anandi

    2017-09-08

    Barriers to medication adherence stem from multiple factors. An effective and convenient tool is needed to identify these barriers so that clinicians can provide a tailored, patient-centered consultation with patients. The Modified Drug Adherence Work-up Tool (M-DRAW) was developed as a 13-item checklist questionnaire to identify barriers to medication adherence. The response scale was a 4-point Likert scale of frequency of occurrence (1 = never to 4 = often). The checklist was accompanied by a GUIDE that provided corresponding motivational interview-based intervention strategies for each identified barrier. The current pilot study examined the psychometric properties of the M-DRAW checklist (reliability, responsiveness and discriminant validity) in patients taking one or more prescription medication(s) for chronic conditions. A cross-sectional sample of 26 patients was recruited between December 2015 and March 2016 at an academic medical center pharmacy in Southern California. A priming question that assessed self-reported adherence was used to separate participants into the control group of 17 "adherers" (65.4%), and into the intervention group of nine "unintentional and intentional non-adherers" (34.6%). Comparable baseline characteristics were observed between the two groups. The M-DRAW checklist showed acceptable reliability (13 item; alpha = 0.74) for identifying factors and barriers leading to medication non-adherence. Discriminant validity of the tool and the priming question was established by the four-fold number of barriers to adherence identified within the self-selected intervention group compared to the control group (4.4 versus 1.2 barriers, p tool will include construct validation.

  2. Validating the Modified Drug Adherence Work-Up (M-DRAW Tool to Identify and Address Barriers to Medication Adherence

    Directory of Open Access Journals (Sweden)

    Sun Lee

    2017-09-01

    Full Text Available Barriers to medication adherence stem from multiple factors. An effective and convenient tool is needed to identify these barriers so that clinicians can provide a tailored, patient-centered consultation with patients. The Modified Drug Adherence Work-up Tool (M-DRAW was developed as a 13-item checklist questionnaire to identify barriers to medication adherence. The response scale was a 4-point Likert scale of frequency of occurrence (1 = never to 4 = often. The checklist was accompanied by a GUIDE that provided corresponding motivational interview-based intervention strategies for each identified barrier. The current pilot study examined the psychometric properties of the M-DRAW checklist (reliability, responsiveness and discriminant validity in patients taking one or more prescription medication(s for chronic conditions. A cross-sectional sample of 26 patients was recruited between December 2015 and March 2016 at an academic medical center pharmacy in Southern California. A priming question that assessed self-reported adherence was used to separate participants into the control group of 17 “adherers” (65.4%, and into the intervention group of nine “unintentional and intentional non-adherers” (34.6%. Comparable baseline characteristics were observed between the two groups. The M-DRAW checklist showed acceptable reliability (13 item; alpha = 0.74 for identifying factors and barriers leading to medication non-adherence. Discriminant validity of the tool and the priming question was established by the four-fold number of barriers to adherence identified within the self-selected intervention group compared to the control group (4.4 versus 1.2 barriers, p < 0.05. The current study did not investigate construct validity due to small sample size and challenges on follow-up with patients. Future testing of the tool will include construct validation.

  3. A calorimetric study of the interactions in the aqueous solutions of lysozyme in the presence of denaturing cosolvents

    International Nuclear Information System (INIS)

    Castronuovo, Giuseppina; Niccoli, Marcella

    2012-01-01

    Highlights: ► A thermodynamic method is reported to monitor the chemical denaturation of lysozyme. ► The enthalpic interaction coefficients are very useful parameters to gain information about the mechanism through which two hydrated molecules interact in solution. ► Hypotheses are proposed about the mechanism underlying the denaturation of lysozyme induced by high concentrations of urea or ethanol. - Abstract: A thermodynamic method is reported to monitor the chemical denaturation of lysozyme. Heats of dilution of the protein in concentrated aqueous solutions of urea or ethanol have been determined at 298.15 K by flow microcalorimetry. The pairwise enthalpic interaction coefficients of the protein in the different solvent media are derived. These parameters allow to gain information about the influence of the cosolvents on the interactions acting between two interacting hydrated molecules of lysozyme, hence on the denaturation process. At increasing urea concentration, up to about 6 mol kg −1 , the values of the interaction coefficients are large and negative and remain almost unaltered. The invariance of the coefficients underlines that, even in highly concentrated urea, the hydration shell of the protein is such to maintain essentially unaltered the native conformation. At higher urea concentrations, a sudden change in the sign of the coefficients monitors the variation in the interactions between two hydrated denatured protein molecules. The same trend is found when ethanol is the cosolvent. At increasing concentration of the cosolvent, coefficients are, at first, almost invariant. After that, denaturation occurs, detected as a jump toward much more negative values. The results obtained are rationalized on the basis of those previously found for small model molecules in concentrated solutions of urea or ethanol. The thermodynamic framework allows useful comments to be made on the possible mode of action of the two cosolvents on the stability of proteins

  4. A calorimetric study of the interactions in the aqueous solutions of lysozyme in the presence of denaturing cosolvents

    Energy Technology Data Exchange (ETDEWEB)

    Castronuovo, Giuseppina, E-mail: giuseppina.castronuovo@unina.it [Department of Chemistry, University Federico II of Naples, Complesso Universitario Monte S. Angelo, via Cintia, 80126 Naples (Italy); Niccoli, Marcella [Department of Chemistry, University Federico II of Naples, Complesso Universitario Monte S. Angelo, via Cintia, 80126 Naples (Italy)

    2012-09-10

    Highlights: Black-Right-Pointing-Pointer A thermodynamic method is reported to monitor the chemical denaturation of lysozyme. Black-Right-Pointing-Pointer The enthalpic interaction coefficients are very useful parameters to gain information about the mechanism through which two hydrated molecules interact in solution. Black-Right-Pointing-Pointer Hypotheses are proposed about the mechanism underlying the denaturation of lysozyme induced by high concentrations of urea or ethanol. - Abstract: A thermodynamic method is reported to monitor the chemical denaturation of lysozyme. Heats of dilution of the protein in concentrated aqueous solutions of urea or ethanol have been determined at 298.15 K by flow microcalorimetry. The pairwise enthalpic interaction coefficients of the protein in the different solvent media are derived. These parameters allow to gain information about the influence of the cosolvents on the interactions acting between two interacting hydrated molecules of lysozyme, hence on the denaturation process. At increasing urea concentration, up to about 6 mol kg{sup -1}, the values of the interaction coefficients are large and negative and remain almost unaltered. The invariance of the coefficients underlines that, even in highly concentrated urea, the hydration shell of the protein is such to maintain essentially unaltered the native conformation. At higher urea concentrations, a sudden change in the sign of the coefficients monitors the variation in the interactions between two hydrated denatured protein molecules. The same trend is found when ethanol is the cosolvent. At increasing concentration of the cosolvent, coefficients are, at first, almost invariant. After that, denaturation occurs, detected as a jump toward much more negative values. The results obtained are rationalized on the basis of those previously found for small model molecules in concentrated solutions of urea or ethanol. The thermodynamic framework allows useful comments to be made on

  5. Chemical denaturation of globular proteins at the air/water interface: an x-ray and neutron reflectometry study

    International Nuclear Information System (INIS)

    Perriman, A.W.; Henderson, M.J.; White, J.W.

    2003-01-01

    Full text: X-ray and neutron reflectometry has been used to probe the equilibrium surface structure of hen egg white lysozyme (lysozyme) and bovine β -lactoglobulin (β -lactoglobulin) under denaturing conditions at the air-water interface. This was achieved by performing experiments on 10 mg mL -1 protein solutions containing increasing concentrations of the chemical denaturant guanidinium hydrochloride (G.HCl). For solutions containing no G.HCl, the surface structure of the proteins was represented by a two-layer model with total thicknesses of 48 Angstroms and 38 Angstroms for lysozyme and β -lactoglobulin, respectively. The total volume of a single protein molecule and the associated water molecules was evaluated to be approximately 45 (0.3) nm 3 for lysozyme, and 60 (0.3) nm 3 for β-lactoglobulin. The thickness dimensions and the total volumes compared favourably with the crystal dimensions of 45 x 30 x 30 Angstroms (40.5 nm 3 ),1 and 36 x 36 x 36 Angstroms (47 nm 3 ) 2 for lysozyme and β -lactoglobulin, respectively. This comparison suggests that when no denaturant was present, the structures of lysozyme and β -lactoglobulin were near to their native conformations at the air-water interface. The response to the presence of the chemical denaturant was different for each protein. The surface layer of β-lactoglobulin expanded at very low concentrations (0.2 mol dm -3 ) of G.HCl. In contrast, the lysozyme layer contracted. At higher concentrations, unfolding of both the proteins led to the formation of a third diffuse layer. In general, lysozyme appeared to be less responsive to the chemical denaturant, which is most likely a result of the higher disulfide content of lysozyme. A protocol allowing quantitative thermodynamic analysis of the contribution from the air-water interface to the chemical denaturation of a protein was developed

  6. Pragmatic approach to the clinical work-up of patients with putative allergic disease to metallic orthopaedic implants before and after surgery

    DEFF Research Database (Denmark)

    Thyssen, J P; Menné, T; Schalock, P C

    2011-01-01

    on in the work-up of patients with putative allergic complications following surgery. Few studies have investigated whether subjects with metal contact allergy have increased risk of developing complications following orthopaedic implant insertion. Metal allergy might in a minority increase the risk...... testing prior to surgery unless the patient has already had implant surgery with complications suspected to be allergic or has a history of clinical metal intolerance of sufficient magnitude to be of concern to the patient or a health provider. The clinical work-up of a patient suspected of having......Allergic complications following insertion of metallic orthopaedic implants include allergic dermatitis reactions but also extracutaneous complications. As metal-allergic patients and/or surgeons may ask dermatologists and allergologists for advice prior to planned orthopaedic implant surgery...

  7. Can value for money be improved by changing the sequence of our donor work-up in the living kidney donor programme?

    DEFF Research Database (Denmark)

    Larsen, J.; Sorensen, S.S.; Feldt-Rasmussen, B.

    2009-01-01

    and December 2006 at our department. The cost in euro (euro) for the programme was estimated using the Danish diagnosis-related group-system (DRG). The donor work-up programme was described. One hundred and thirty-three potential donors were identified; 66 male- and 67 female subjects, median age of 52 years......The aim of the study was to identify procedures of maximum importance for acceptance or rejection of kidney donation from a living donor as well as making the process more cost-effective. We identified all potential living related donors who were examined during the period between January 2002...... was the procedure identifying most subjects who were unsuited for kidney donation. A rearrangement of the present donor work-up programme could potentially reduce the costs from euro6911 to euro5292 per donor--saving 23% of the costs. By changing the sequence of examinations, it might be possible to cut down...

  8. Biological function evaluation and effects of laser micro-pore burn-denatured acellular dermal matrix.

    Science.gov (United States)

    Zhang, Youlai; Zeng, Yuanlin; Xin, Guohua; Zou, Lijin; Ding, Yuewei; Duyin, Jiang

    2018-03-01

    In the field of burns repairs, many problems exist in the shortage of donor skin, the expense of allograft or xenograft skin, temporary substitution and unsatisfactory extremity function after wound healing. Previous studies showed that burn-denatured skin could return to normal dermis formation and function. This study investigates the application of laser micro-pore burn-denatured acellular dermis matrix (DADM) from an escharotomy in the repair of burn wounds and evaluates the biological properties and wound repair effects of DADM in implantation experiments in Kunming mice. Specific-pathogen-free (SPF) Kunming mice were used in this study. A deep II° burn wound was created on the dorsum of the mice by an electric heated water bath. The full-thickness wound tissue was harvested. The necrotic tissue and subcutaneous tissue were removed. The denatured dermis was preserved and treated with 0.25% trypsin, 0.5% Triton X-100. The DADM was drilled by laser micro-pore. The biological properties and grafting effects of laser micro-pore burn-DADM were evaluated by morphology, cytokine expression levels and subcutaneous implantation experiments in Kunming mice. We found statistical significance (Ppore burn-DADM (experimental group) compared to the control group (no laser micro-pore burn-DADM). Cytokine expression level was different in the dermal matrixes harvested at various time points after burn (24h, 48h, 72h and infected wound group). Comparing the dermal matrix from 24h burn tissue to infected wound tissue, the expression level of IL-6, MMP-24, VE-cadherin and VEGF were decreased. We found no inflammatory cells infiltration in the dermal matrix were observed in both experimental and control groups (24h burn group), while the obviously vascular infiltration and fiber fusion were observed in the experimental group after subcutaneous implantation experiments. There was better bio-performance, low immunogenicity and better dermal incorporation after treated by laser

  9. Glutamate Induced Thermal Equilibrium Intermediate and Counteracting Effect on Chemical Denaturation of Proteins.

    Science.gov (United States)

    Anumalla, Bramhini; Prabhu, N Prakash

    2018-01-25

    When organisms are subjected to stress conditions, one of their adaptive responses is accumulation of small organic molecules called osmolytes. These osmolytes affect the structure and stability of the biological macromolecules including proteins. The present study examines the effect of a negatively charged amino acid osmolyte, glutamate (Glu), on two model proteins, ribonuclease A (RNase A) and α-lactalbumin (α-LA), which have positive and negative surface charges at pH 7, respectively. These proteins follow two-state unfolding transitions during both heat and chemical induced denaturation processes. The addition of Glu stabilizes the proteins against temperature and induces an early equilibrium intermediate during unfolding. The stability is found to be enthalpy-driven, and the free energy of stabilization is more for α-LA compared to RNase A. The decrease in the partial molar volume and compressibility of both of the proteins in the presence of Glu suggests that the proteins attain a more compact state through surface hydration which could provide a more stable conformation. This is also supported by molecule dynamic simulation studies which demonstrate that the water density around the proteins is increased upon the addition of Glu. Further, the intermediates could be completely destabilized by lower concentrations (∼0.5 M) of guanidinium chloride and salt. However, urea subverts the Glu-induced intermediate formed by α-LA, whereas it only slightly destabilizes in the case of RNase A which has a positive surface charge and could possess charge-charge interactions with Glu. This suggests that, apart from hydration, columbic interactions might also contribute to the stability of the intermediate. Gdm-induced denaturation of RNase A and α-LA in the absence and the presence of Glu at different temperatures was carried out. These results also show the Glu-induced stabilization of both of the proteins; however, all of the unfolding transitions followed two

  10. Preparation of denatured sup(99m)Tc labeled HSA aerosols of different median diameters for various imaging studies

    International Nuclear Information System (INIS)

    Raghunath, B.; Kotrappa, P.; Soni, P.S.; Ganatra, R.D.

    1982-01-01

    The preparation of denatured sup(99m)Tc-labelled human serum albumin (HSA) aerosols of different median diameters is described using the BARC (Bhabha Atomic Research Centre) dry aerosol generation and delivery system. The applications of these radioactive aerosols are demonstrated in aerosol scintigraphy of lungs, mucociliary movement studies and lymphoscintigraphy in rabbits. It is concluded that the BARC system gives a simplified, rapid and versatile procedure for generation of denatured volume tagged HSA aerosols for a variety of clinical applications. (U.K.)

  11. Dynamics of Ionic Liquid-Assisted Refolding of Denatured Cytochrome c: A Study of Preferential Interactions toward Renaturation.

    Science.gov (United States)

    Singh, Upendra Kumar; Patel, Rajan

    2018-05-25

    In vitro refolding of denatured protein and the influence of the alkyl chain on the refolding of a protein were tested using long chain imidazolium chloride salts, 1-methyl-3-octylimidazolium chloride [C 8 mim][Cl], and 1-decyl-3-methylimidazolium chloride [C 10 mim][Cl]. The horse heart cytochrome c (h-cyt c) was denatured by urea and guanidinium hydrochloride (GdnHCl), as well as by base-induced denaturation at pH 13, to provide a broad overview of the overall refolding behavior. The variation in the alkyl chain of the ionic liquids (ILs) showed a profound effect on the refolding of denatured h-cyt c. The ligand-induced refolding was correlated to understand the mechanism of the conformational stability of proteins in aqueous solutions of ILs. The results showed that the long chain ILs having the [C 8 mim] + and [C 10 mim] + cations promote the refolding of alkali-denatured h-cyt c. The IL having the [C 10 mim] + cation efficiently refolded the alkali-denatured h-cyt c with the formation of the MG state, whereas the IL having the [C 8 mim] + cation, which is known to be compatible for protein stability, shows slight refolding and forms a different transition state. The lifetime results show successful refolding of alkaline-denatured h-cyt c by both of the ILs, however, more refolding was observed in the case of [C 10 mim][Cl], and this was correlated with the fast and medium lifetimes (τ 1 and τ 2 ) obtained, which show an increase accompanied by an increase in secondary structure. The hydrophobic interactions plays an important role in the refolding of chemically and alkali-denatured h-cyt c by long chain imidazolium ILs. The formation of the MG state by [C 10 mim][Cl] was also confirmed, as some regular structure exists far below the CMC of IL. The overall results suggested that the [C 10 mim] + cation bound to the unfolded h-cyt c triggers its refolding by electrostatic and hydrophobic interactions that stabilize the MG state.

  12. Fuel utilization improvement in PWRs using the denatured 233U-Th cycle

    International Nuclear Information System (INIS)

    Jones, H.M.; Schwenk, G.A.; Toops, E.C.; Yotinen, V.O.

    1980-06-01

    A number of changes in PWR core design and/or operating strategy were evaluated to assess the fuel utilization improvement achievable by their implementation in a PWR using thorium-based fuel and operating in a recycle mode. The reference PWR for this study was identical to the B and W Standard Plant except that the fuel pellets were of denatured ( 233 U/ 238 U-Th)O 2 . An initial scoping study identified the three most promising improvement concepts as (1) a very tight lattice, (2) thorium blankets, and (3) ThO 2 rods placed in available guide tubes. A conceptual core design incorporating these changes was then developed, and the fuel utilization of this modified design was compared with that of the reference case

  13. Investigating the fermentation of cocoa by correlating denaturing gradient gel electrophoresis profiles and near infrared spectra

    DEFF Research Database (Denmark)

    Nielsen, Dennis Sandris; Snitkjær, Pia; van der Berg, Franciscus Winfried J

    2008-01-01

    demonstrating the microbial succession taking place during the fermentation. Subsequently the DGGE spectra were correlated to the NIR spectra using Partial Least Squares regression models (PLS2). Correlations of 0.87 (bacterial derived DGGE spectra) and 0.81 (yeast derived DGGE spectra) were obtained indicating......Raw cocoa has an astringent, unpleasant taste and flavour, and has to be fermented, dried and roasted in order to obtain the characteristic cocoa flavour and taste. During the fermentation microbial activity outside the cocoa beans induces biochemical and physical changes inside the beans...... of the beans and the chemical processes inside the beans have been carried out previously. Recently it has been shown that Denaturing Gradient Gel Electrophoresis (DGGE) offers an efficient tool for monitoring the microbiological changes taking place during the fermentation of cocoa. Near Infrared (NIR...

  14. Protective role of salt in catalysis and maintaining structure of halophilic proteins against denaturation

    Science.gov (United States)

    Sinha, Rajeshwari; Khare, Sunil K.

    2014-01-01

    Search for new industrial enzymes having novel properties continues to be a desirable pursuit in enzyme research. The halophilic organisms inhabiting under saline/ hypersaline conditions are considered as promising source of useful enzymes. Their enzymes are structurally adapted to perform efficient catalysis under saline environment wherein n0n-halophilic enzymes often lose their structure and activity. Haloenzymes have been documented to be polyextremophilic and withstand high temperature, pH, organic solvents, and chaotropic agents. However, this stability is modulated by salt. Although vast amount of information have been generated on salt mediated protection and structure function relationship in halophilic proteins, their clear understanding and correct perspective still remain incoherent. Furthermore, understanding their protein architecture may give better clue for engineering stable enzymes which can withstand harsh industrial conditions. The article encompasses the current level of understanding about haloadaptations and analyzes structural basis of their enzyme stability against classical denaturants. PMID:24782853

  15. Fluorescence lifetime components reveal kinetic intermediate states upon equilibrium denaturation of carbonic anhydrase II.

    Science.gov (United States)

    Nemtseva, Elena V; Lashchuk, Olesya O; Gerasimova, Marina A; Melnik, Tatiana N; Nagibina, Galina S; Melnik, Bogdan S

    2017-12-21

    In most cases, intermediate states of multistage folding proteins are not 'visible' under equilibrium conditions but are revealed in kinetic experiments. Time-resolved fluorescence spectroscopy was used in equilibrium denaturation studies. The technique allows for detecting changes in the conformation and environment of tryptophan residues in different structural elements of carbonic anhydrase II which in its turn has made it possible to study the intermediate states of carbonic anhydrase II under equilibrium conditions. The results of equilibrium and kinetic experiments using wild-type bovine carbonic anhydrase II and its mutant form with the substitution of leucine for alanine at position 139 (L139A) were compared. The obtained lifetime components of intrinsic tryptophan fluorescence allowed for revealing that, the same as in kinetic experiments, under equilibrium conditions the unfolding of carbonic anhydrase II ensues through formation of intermediate states.

  16. Functionality screen of streptavidin mutants by non-denaturing SDS-PAGE using biotin-4-fluorescein.

    Science.gov (United States)

    Humbert, Nicolas; Ward, Thomas R

    2008-01-01

    Site-directed mutagenesis or directed evolution of proteins often leads to the production of inactive mutants. For streptavidin and related proteins, mutations may lead to the loss of their biotin-binding properties. With high-throughput screening methodologies in mind, it is imperative to detect, prior to the high-density protein production, the bacteria that produce non-functional streptavidin isoforms. Based on the incorporation of biotin-4-fluorescein in streptavidin mutants present in Escherichia coli bacterial extracts, we detail a functional screen that allows the identification of biotin-binding streptavidin variants. Bacteria are cultivated in a small volume, followed by a rapid treatment of the cells; biotin-4-fluorescein is added to the bacterial extract and loaded on an Sodium Dodecyl Sulfate Poly-Acrylamide Gel Electrophoresis (SDS-PAGE) under non-denaturing conditions. Revealing is performed using a UV transilluminator. This screen is thus easy to implement, cheap and requires only readily available equipment.

  17. 125I-labeled cortisol radioimmunoassay in which serum binding protein are enzymatically denatured

    International Nuclear Information System (INIS)

    Hasler, M.J.; Painter, K.; Niswender, G.D.

    1976-01-01

    We report an iodine-125 radioimmunoassay for cortisol in biological fluids, in which interfering binding proteins are enzymatically denatured. An antiserum to cortisol-3-carboxymethyloxime-bovine serum albumin, extremely low cross-reacting with other corticosteroids, was raised in rabbits. A cortisol-3-carboxymethyloxime tyrosine methyl ester derivative was synthesized and labeled with iodine-125 by standard radioiodination techniques. To eliminate the need for extraction and recovery procedures, we digested interfering binding with a proteolytic enzyme, which then was heat-inactivated before adding the labeled derivative and the premixed, preincubated antiserum complex. There was quantitative analytical recovery of esogenous cortisol added to sera from a normal man, a normal woman, and a pregnant woman. Values for the same samples agreed after extraction and chromatographic purification and agreed well with values obtained by other techniques by independent reference laboratories. The five-step assay can be done in 6 h or less

  18. Thermal, chemical and pH induced unfolding of turmeric root lectin: modes of denaturation.

    Directory of Open Access Journals (Sweden)

    Himadri Biswas

    Full Text Available Curcuma longa rhizome lectin, of non-seed origin having antifungal, antibacterial and α-glucosidase inhibitory activities, forms a homodimer with high thermal stability as well as acid tolerance. Size exclusion chromatography and dynamic light scattering show it to be a dimer at pH 7, but it converts to a monomer near pH 2. Circular dichroism spectra and fluorescence emission maxima are virtually indistinguishable from pH 7 to 2, indicating secondary and tertiary structures remain the same in dimer and monomer within experimental error. The tryptophan environment as probed by acrylamide quenching data yielded very similar data at pH 2 and pH 7, implying very similar folding for monomer and dimer. Differential scanning calorimetry shows a transition at 350.3 K for dimer and at 327.0 K for monomer. Thermal unfolding and chemical unfolding induced by guanidinium chloride for dimer are both reversible and can be described by two-state models. The temperatures and the denaturant concentrations at which one-half of the protein molecules are unfolded, are protein concentration-dependent for dimer but protein concentration-independent for monomer. The free energy of unfolding at 298 K was found to be 5.23 Kcal mol-1 and 14.90 Kcal mol-1 for the monomer and dimer respectively. The value of change in excess heat capacity upon protein denaturation (ΔCp is 3.42 Kcal mol-1 K-1 for dimer. The small ΔCp for unfolding of CLA reflects a buried hydrophobic core in the folded dimeric protein. These unfolding experiments, temperature dependent circular dichroism and dynamic light scattering for the dimer at pH 7 indicate its higher stability than for the monomer at pH 2. This difference in stability of dimeric and monomeric forms highlights the contribution of inter-subunit interactions in the former.

  19. Aqueous Solutions of the Ionic Liquid 1-butyl-3-methylimidazolium Chloride Denature Proteins

    International Nuclear Information System (INIS)

    Baker, Gary A.; Heller, William T.

    2009-01-01

    As we advance our understanding, ionic liquids (ILs) are finding ever broader scope within the chemical sciences including, most recently, pharmaceutical, enzymatic, and bioanalytical applications. With examples of enzymatic activity reported in both neat ILs and in IL/water mixtures, enzymes are frequently assumed to adopt a quasi-native conformation, even if little work has been carried out to date toward characterizing the conformation, dynamics, active-site perturbation, cooperativity of unfolding transitions, free energy of stabilization, or aggregation/oligomerization state of enzymes in the presence of an IL solvent component. In this study, human serum albumin and equine heart cytochrome c were characterized in aqueous solutions of the fully water-miscible IL 1-butyl-3-methylimidazolium chloride, (bmim)Cl, by small-angle neutron and X-ray scattering. At (bmim)Cl concentrations up to 25 vol.%, these two proteins were found to largely retain their higher-order structures whereas both proteins become highly denatured at the highest IL concentration studied here (i.e., 50 vol.% (bmim)Cl). The response of these proteins to (bmim)Cl is analogous to their behavior in the widely studied denaturants guanidine hydrochloride and urea which similarly lead to random coil conformations at excessive molar concentrations. Interestingly, human serum albumin dimerizes in response to (bmim)Cl, whereas cytochrome c remains predominantly in monomeric form. These results have important implications for enzymatic studies in aqueous IL media, as they suggest a facile pathway through which biocatalytic activity can be altered in these nascent and potentially green electrolyte systems

  20. Mammographic findings of women recalled for diagnostic work-up in digital versus screen-film mammography in a population-based screening program

    International Nuclear Information System (INIS)

    Lipasti, Seppo; Pamilo, Martti; Anttila, Ahti

    2010-01-01

    Background: Limited information is available concerning differences in the radiological findings of women recalled for diagnostic work-up in digital mammography (DM) versus screen-film mammography (SFM) screening. Purpose: To compare the radiological findings, their positive predictive values (PPVs) for cancer and other process indicators of DM screening performed by computed radiography (CR) technology and SFM screening in a population-based program. Material and Methods: The material consisted of women, 50-59 years of age, who were invited for screening: 30 153 women with DM in 2007-2008 and 32 939 women with SFM in 1999-2000. The attendance rate was 77.7% (23 440) in the DM arm and 83.8% (27 593) in the SFM arm. In the DM arm, 1.71% of those screened (401) and in the SFM arm 1.59% (438) were recalled for further work-up. The images resulting in the recall were classified as: 1) tumor-like mass, 2) parenchymal distortion/asymmetry, 3) calcifications, and 4) combination of mass and calcifications. The distributions of the various radiological findings and their PPVs for cancer were compared in both study groups. The recall rates, cancer detection rates, test specificities, and PPVs of the DM and SFM groups were also compared. Results: Women were recalled for diagnostic work-up most often due to tumor-like mass. It was more common in SFM (1.08% per woman screened) than in DM (0.93%). The second most common finding was parenchymal distortion and asymmetry, more often in DM (0.58%) than in SFM (0.37%). Calcifications were the third most common finding. DM exposed calcifications more often (0.49%) than SFM (0.26%). The PPVs for cancer of the recalls were higher in DM than in SFM in all subgroups of radiological findings. The test specificities were similar (DM 98.9%, SFM 98.8%). Significantly more cancers were detected by DM (cancer detection rate 0.623% per woman screened, n=146) than by SFM (cancer detection rate 0.406% per woman screened, n=112). The PPVs for

  1. Should there be a standardised approach to the diagnostic workup of suspected adult encephalitis? a case series from Australia

    Directory of Open Access Journals (Sweden)

    Williams David

    2010-12-01

    assessment is necessary to exclude treatable agents and identify pathogens warranting public health interventions, such as those transmitted by mosquitoes and those that are vaccine preventable. An algorithm and guidelines for the diagnostic workup of encephalitis cases would assist in optimising laboratory testing so that clinical management can be best tailored to the pathogen, and appropriate public health measures implemented.

  2. T1ρ is superior to T2 mapping for the evaluation of articular cartilage denaturalization with osteoarthritis: radiological-pathological correlation after total knee arthroplasty.

    Science.gov (United States)

    Takayama, Yukihisa; Hatakenaka, Masamitsu; Tsushima, Hidetoshi; Okazaki, Ken; Yoshiura, Takashi; Yonezawa, Masato; Nishikawa, Kei; Iwamoto, Yukihide; Honda, Hiroshi

    2013-04-01

    We compared the diagnostic performance of T1ρ and T2 mappings in the evaluation of denatured articular cartilage with osteoarthritis of the knee. 2D-Sagittal T1ρ and T2 mappings of the knee were obtained from 16 patients before total knee arthroplasty. After surgery, specimens of the femur and tibia were regionally segmented according to a 5-point scale of the severity of denaturalization. The T1ρ and T2 values in the full thickness of the articular cartilage in each region were measured by two observers. The two mappings were compared for their ability to differentiate between normal and denatured articular cartilage and also for their usefulness in grading the severity of the denaturalization using the area under receiver operating characteristic curves (Az). A pT2 mapping for the differentiation between normal and denatured articular cartilage (pT2 mapping could not. However, there were no significant differences between the two mappings in the discrimination of mild versus moderate denaturalization or of moderate versus severe denaturalization. The two observers showed good agreement in the results (intraclass correlation coefficient=0.81 for T1ρ and 0.92 for T2). T1ρ mapping is superior to T2 mapping for the evaluation of denatured articular cartilage with osteoarthritis of the knee. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  3. Laboratory Exercise for Studying the Morphology of Heat-Denatured and Amyloid Aggregates of Lysozyme by Atomic Force Microscopy

    Science.gov (United States)

    Gokalp, Sumeyra; Horton, William; Jónsdóttir-Lewis, Elfa B.; Foster, Michelle; Török, Marianna

    2018-01-01

    To facilitate learning advanced instrumental techniques, essential tools for visualizing biomaterials, a simple and versatile laboratory exercise demonstrating the use of Atomic Force Microscopy (AFM) in biomedical applications was developed. In this experiment, the morphology of heat-denatured and amyloid-type aggregates formed from a low-cost…

  4. Evaluation of several techniques to modify denatured muscle tissue to obtain a scaffold for peripheral nerve regeneration

    NARCIS (Netherlands)

    Meek, MF; den Dunnen, WFA; Schakenraad, JM; Robinson, PH

    The aim of this study was to (1) evaluate the effect of several preparation techniques of denatured muscle tissue to obtain an open three-dimensional structure, and (2) test if this scaffold is suitable for peripheral nerve regeneration. Four samples (A-D) of muscle tissue specimens were evaluated

  5. Sequential Events in the Irreversible Thermal Denaturation of Human Brain-Type Creatine Kinase by Spectroscopic Methods

    Directory of Open Access Journals (Sweden)

    Yan-Song Gao

    2010-06-01

    Full Text Available The non-cooperative or sequential events which occur during protein thermal denaturation are closely correlated with protein folding, stability, and physiological functions. In this research, the sequential events of human brain-type creatine kinase (hBBCK thermal denaturation were studied by differential scanning calorimetry (DSC, CD, and intrinsic fluorescence spectroscopy. DSC experiments revealed that the thermal denaturation of hBBCK was calorimetrically irreversible. The existence of several endothermic peaks suggested that the denaturation involved stepwise conformational changes, which were further verified by the discrepancy in the transition curves obtained from various spectroscopic probes. During heating, the disruption of the active site structure occurred prior to the secondary and tertiary structural changes. The thermal unfolding and aggregation of hBBCK was found to occur through sequential events. This is quite different from that of muscle-type CK (MMCK. The results herein suggest that BBCK and MMCK undergo quite dissimilar thermal unfolding pathways, although they are highly conserved in the primary and tertiary structures. A minor difference in structure might endow the isoenzymes dissimilar local stabilities in structure, which further contribute to isoenzyme-specific thermal stabilities.

  6. New comprehensive denaturing-gradient-gel-electrophoresis assay for KRAS mutation detection applied to paraffin-embedded tumours

    NARCIS (Netherlands)

    Hayes, VM; Westra, JL; Verlind, E; Bleeker, W; Plukker, JT; Hofstra, RMW; Buys, CHCM

    2000-01-01

    A comprehensive mutation detection assay is presented for the entire coding region and all splice site junctions of the KRAS oncogene. The assay is based on denaturing gradient gel electrophoresis and applicable to archival paraffin-embedded tumour material. All KRAS amplicons are analysed within

  7. Denaturation and in Vitro Gastric Digestion of Heat-Treated Quinoa Protein Isolates Obtained at Various Extraction pH

    NARCIS (Netherlands)

    Ruiz, Geraldine Avila; Opazo-Navarrete, Mauricio; Meurs, Marlon; Minor, Marcel; Sala, Guido; Boekel, van Tiny; Stieger, Markus; Janssen, Anja E.M.

    2016-01-01

    The aim of this study was to determine the influence of heat processing on denaturation and digestibility properties of protein isolates obtained from sweet quinoa (Chenopodium quinoa Willd) at various extraction pH values (8, 9, 10 and 11). Pretreatment of suspensions of protein isolates at 60,

  8. The effect of Na+ and K+ on the thermal denaturation of Na+ and + K+-dependent ATPase.

    Science.gov (United States)

    Fischer, T H

    1983-01-01

    To increase our understanding of the physical nature of the Na+ and K+ forms of the Na+ + K+-dependent ATPase, thermal-denaturation studies were conducted in different types of ionic media. Thermal-denaturation measurements were performed by measuring the regeneration of ATPase activity after slow pulse exposure to elevated temperatures. Two types of experiments were performed. First, the dependence of the thermal-denaturation rate on Na+ and K+ concentrations was examined. It was found that both cations stabilized the pump protein. Also, K+ was a more effective stabilizer of the native state than was Na+. Secondly, a set of thermodynamic parameters was obtained by measuring the temperature-dependence of the thermal-denaturation rate under three ionic conditions: 60 mM-K+, 150 mM-Na+ and no Na+ or K+. It was found that ion-mediated stabilization of the pump protein was accompanied by substantial increases in activation enthalpy and entropy, the net effect being a less-pronounced increase in activation free energy. PMID:6309139

  9. Pragmatic approach to the clinical work-up of patients with putative allergic disease to metallic orthopaedic implants before and after surgery.

    Science.gov (United States)

    Thyssen, J P; Menné, T; Schalock, P C; Taylor, J S; Maibach, H I

    2011-03-01

    Allergic complications following insertion of metallic orthopaedic implants include allergic dermatitis reactions but also extracutaneous complications. As metal-allergic patients and/or surgeons may ask dermatologists and allergologists for advice prior to planned orthopaedic implant surgery, and as surgeons may refer patients with complications following total joint arthroplasty for diagnostic work-up, there is a continuous need for updated guidelines. This review presents published evidence for patch testing prior to surgery and proposes tentative diagnostic criteria which clinicians can rely on in the work-up of patients with putative allergic complications following surgery. Few studies have investigated whether subjects with metal contact allergy have increased risk of developing complications following orthopaedic implant insertion. Metal allergy might in a minority increase the risk of complications caused by a delayed-type hypersensitivity reaction. At present, we do not know how to identify the subgroups of metal contact allergic patients with a potentially increased risk of complications following insertion of a metal implant. We recommend that clinicians should refrain from routine patch testing prior to surgery unless the patient has already had implant surgery with complications suspected to be allergic or has a history of clinical metal intolerance of sufficient magnitude to be of concern to the patient or a health provider. The clinical work-up of a patient suspected of having an allergic reaction to a metal implant should include patch testing and possibly in vitro testing. We propose diagnostic criteria for allergic dermatitis reactions as well as noneczematous complications caused by metal implants. © 2011 The Authors. BJD © 2011 British Association of Dermatologists.

  10. The Added Diagnostic Value of 18F-Fluorodihydroxyphenylalanine PET/CT in the Preoperative Work-Up of Small Bowel Neuroendocrine Tumors.

    Science.gov (United States)

    Addeo, Pietro; Poncet, Gilles; Goichot, Bernard; Leclerc, Loic; Brigand, Cécile; Mutter, Didier; Romain, Benoit; Namer, Izzie-Jacques; Bachellier, Philippe; Imperiale, Alessio

    2018-04-01

    The precise localization of the primary tumor and/or the identification of multiple primary tumors improves the preoperative work-up in patients with small bowel (SB) neuroendocrine tumor (NET). The present study assesses the diagnostic value of 18 F-fluorodihydroxyphenylalanine ( 18 F-FDOPA) positron emission tomography/computed tomography (PET/CT) during the preoperative wok-up of SB NETs. Between January 2010 and June 2017, all consecutive patients with SB NETs undergoing preoperative 18 F-FDOPA PET/CT and successive resection were analyzed. Preoperative work-up included computed tomography (CT), somatostatin receptor scintigraphy (SRS), and 18 F-FDOPA PET/CT. Sensitivity and accuracy ratio for primary and multiple tumor detection were compared with data from surgery and pathology. There were 17 consecutive patients with SB NETs undergoing surgery. Nine patients (53%) had multiple tumors, 15 (88%) metastatic lymph nodes, 3 (18%) peritoneal carcinomatosis, and 9 patients (53%) liver metastases. A total of 70 SB NETs were found by pathology. Surgery identified the primary in 17/17 (100%) patients and recognized seven of 9 patients (78%) with multiple synchronous SB. Preoperatively, 18 F-FDOPA PET/CT displayed a statistically significant higher sensitivity for primary tumor localization (100 vs. 23.5 vs. 29.5%) and multiple tumor detection (78 vs. 22 vs. 11%) over SRS and CT. Compared with pathology, 18 F-FDOPA PET/CT displayed the highest accuracy ratio for number of tumor detected over CT and SRS (2.0 ± 2.2 vs. 0.4 ± 0.7 vs. 0.6 ± 1.5, p = 0.0003). 18 F-FDOPA PET/CT significantly increased the sensitivity and accuracy for primary and multiple SB NET identification. 18 F-FDOPA PET/CT should be included systematically in the preoperative work-up of SB NET.

  11. Histologic work-up of non-palpable breast lesions classified as probably benign at initial mammography and/or ultrasound (BI-RADS category 3).

    Science.gov (United States)

    Gruber, R; Jaromi, S; Rudas, M; Pfarl, G; Riedl, C C; Flöry, D; Graf, O; Sickles, E A; Helbich, T H

    2013-03-01

    To determine the accuracy of a probably benign assessment of non-palpable breast lesions (BI-RADS category 3) at mammography and/or ultrasound with immediate histological work-up. Stereotactic or ultrasound guided core needle breast biopsy (NBB) was performed to evaluate 288 lesions, which were prospectively assessed as BI-RADS category 3. Imaging findings included 195 masses, 73 calcification cases, 16 focal asymmetries, and four architectural distortion cases. After NBB, patients underwent either open surgical biopsy (OSB) (n=204) or mammographic follow-up (n=84) for at least 24 months. Histological results of NBB were compared with those of OSB. Three of the 288 lesions (1.0%) proved to be malignant at histological work-up, two of them were ductal carcinoma in situ (DCIS) and one of them was an invasive carcinoma. NBB revealed invasive carcinoma in 1/288 (0.35%) and atypical ductal hyperplasia (ADH) in 13/288 (4.5%) lesions. OSB revealed DCIS in 2/204 (1%) and invasive carcinoma in 1/204 (0.5%) lesions. The two DCIS were underestimated as ADH by NBB. The remaining 285 (99%) lesions proved to be benign at OSB or remained stable during follow-up. Confirmed by tissue diagnosis, the low likelihood of malignancy of prospectively assessed probably benign lesions is below the 2% threshold established for BI-RADS category 3. Imaging follow-up is a safe and effective alternative to immediate histological work-up for such lesions. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  12. Structural and dynamical study about denatured states of yeast phosphoglycerate kinase by neutrons scattering and X-rays; Etude structurale et dynamique des etats denatures de la phosphoglycerate kinase de levure par diffusion des neutrons et des rayons X

    Energy Technology Data Exchange (ETDEWEB)

    Receveur, V

    1997-04-28

    During a long time, the neutron scattering and X-rays techniques have not been used for the studies bearing on the folding of proteins. The compactness and the globularness of a protein are two structural characteristics describing the denatured states and the intermediate states of folding, and the neutrons and x-rays scattering are probably the two techniques the most appropriate to give this kind of information; they are sensible to the spatial extent and to the molecules compactness, and to their general shape. For these three or four last years, the works using these techniques are increasing, giving precious knowledge on the different steps of folding and on the interactions stabilizing the denatured or intermediate states. This thesis falls into this category. (N.C.).

  13. Pu Denaturing by Transmutation of MA in FBR Multi-cycle

    Energy Technology Data Exchange (ETDEWEB)

    Meiliza, Yoshitalia; Saito, Masaki; Sagara, Hiroshi [Tokyo Institute of Technology, 2-12-1-N1-1 Ookayama, Meguro-ku, Tokyo, 1528550 (Japan)

    2009-06-15

    Pu accumulation and its recycling is important in the term of energy resources, however one of the most sensitive issues is non-proliferation in the future fuel cycle based on fast breeder reactor (FBR). The present paper utilizes Protected Pu Production (P{sup 3}) concept for the production of {sup 238}Pu and {sup 242}Pu by Minor Actinides (MA) transmutation to enhance the proliferation resistance of Pu in the fuel. Increase in the {sup 238}Pu and {sup 242}Pu isotopic fraction creates a high rate of internal heat generation by alpha decay (DH) and/or a high neutron source of spontaneous fission (SFN) in Pu that would be encountered during manufacturing and maintaining of nuclear explosive device. The feasibility of denaturing of Pu by MA transmutation in medium size FBR has been studied from the viewpoint of even-mass number Pu accumulation during multi-cycle of Pu and MA. The proliferation resistance property of Pu is also evaluated based on the specific decay heat and spontaneous fission neutron, compared with the reference criteria. In present paper, the P{sup 3} technology based on multi-recycled Pu and MA is compared with the conventional technology based on multi-recycled Pu only. The detail of mass balance behavior is, however, beyond the scope of the present paper. (authors)

  14. Insight into the effect mechanism of urea-induced protein denaturation by dielectric spectroscopy.

    Science.gov (United States)

    Zhang, Cancan; Yang, Man; Zhao, Kongshuang

    2017-12-06

    Dielectric relaxation spectroscopy was applied to study how urea affects the phase transition of a thermosensitive polymer, poly(N-isopropylacrylamide) (PNIPAM), which has been widely used as a protein model. It was found that there is a pronounced relaxation near 10 GHz for the ternary system of PNIPAM in urea aqueous solution. The temperature dependence of dielectric parameters indicates that urea can reduce the lower critical solution temperature (LCST) of PNIPAM, i.e., stabilize the globule state of PNIPAM and collapse the PNIPAM chains. Based on our results, the interaction mechanism of urea on the conformational transition of PNIPAM was presented: urea replaces water molecules directly bonding with PNIPAM and acts as the bridging agent for the adjacent side chains of PNIPAM. Accordingly, the mechanism with which urea denatures protein was deduced. In addition, it is worth mentioning that, from the temperature dependence of the dielectric parameters obtained in the presence of urea, an interesting phenomenon was found in which the effect of urea on PNIPAM seems to take 2 M as a unit. This result may be the reason why urea and TMAO exit marine fishes at a specific ratio of 2 : 1.

  15. Precise determination of protein extinction coefficients under native and denaturing conditions using SV-AUC.

    Science.gov (United States)

    Hoffmann, Andreas; Grassl, Kerstin; Gommert, Janine; Schlesak, Christian; Bepperling, Alexander

    2018-04-17

    The accurate determination of protein concentration is an important though non-trivial task during the development of a biopharmaceutical. The fundamental prerequisite for this is the availability of an accurate extinction coefficient. Common approaches for the determination of an extinction coefficient for a given protein are either based on the theoretical prediction utilizing the amino acid sequence or the photometric determination combined with a measurement of absolute protein concentration. Here, we report on an improved SV-AUC based method utilizing an analytical ultracentrifuge equipped with absorbance and Rayleigh interference optics. Global fitting of datasets helped to overcome some of the obstacles encountered with the traditional method employing synthetic boundary cells. Careful calculation of dn/dc values taking glycosylation and solvent composition into account allowed the determination of the extinction coefficients of monoclonal antibodies and an Fc-fusion protein under native as well as under denaturing conditions. An intra-assay precision of 0.9% and an accuracy of 1.8% compared to the theoretical value was achieved for monoclonal antibodies. Due to the large number of data points of a single dataset, no meaningful difference between the ProteomeLab XL-I and the new Optima AUC platform could be observed. Thus, the AUC-based approach offers a precise, convenient and versatile alternative to conventional methods like total amino acid analysis (AAA).

  16. Detection of urea-induced internal denaturation of dsDNA using solid-state nanopores.

    Science.gov (United States)

    Singer, Alon; Kuhn, Heiko; Frank-Kamenetskii, Maxim; Meller, Amit

    2010-11-17

    The ability to detect and measure dsDNA thermal fluctuations is of immense importance in understanding the underlying mechanisms responsible for transcription and replication regulation. We describe here the ability of solid-state nanopores to detect sub-nanometer changes in DNA structure as a result of chemically enhanced thermal fluctuations. In this study, we investigate the subtle changes in the mean effective diameter of a dsDNA molecule with 3-5 nm solid-state nanopores as a function of urea concentration and the DNA's AT content. Our studies reveal an increase in the mean effective diameter of a DNA molecule of approximately 0.6 nm at 8.7 M urea. In agreement with the mechanism of DNA local denaturation, we observe a sigmoid dependence of these effects on urea concentration. We find that the translocation times in urea are markedly slower than would be expected if the dynamics were governed primarily by viscous effects. Furthermore, we find that the sensitivity of the nanopore is sufficient to statistically differentiate between DNA molecules of nearly identical lengths differing only in sequence and AT content when placed in 3.5 M urea. Our results demonstrate that nanopores can detect subtle structural changes and are thus a valuable tool for detecting differences in biomolecules' environment.

  17. Detection of urea-induced internal denaturation of dsDNA using solid-state nanopores

    International Nuclear Information System (INIS)

    Singer, Alon; Kuhn, Heiko; Frank-Kamenetskii, Maxim; Meller, Amit

    2010-01-01

    The ability to detect and measure dsDNA thermal fluctuations is of immense importance in understanding the underlying mechanisms responsible for transcription and replication regulation. We describe here the ability of solid-state nanopores to detect sub-nanometer changes in DNA structure as a result of chemically enhanced thermal fluctuations. In this study, we investigate the subtle changes in the mean effective diameter of a dsDNA molecule with 3-5 nm solid-state nanopores as a function of urea concentration and the DNA's AT content. Our studies reveal an increase in the mean effective diameter of a DNA molecule of approximately 0.6 nm at 8.7 M urea. In agreement with the mechanism of DNA local denaturation, we observe a sigmoid dependence of these effects on urea concentration. We find that the translocation times in urea are markedly slower than would be expected if the dynamics were governed primarily by viscous effects. Furthermore, we find that the sensitivity of the nanopore is sufficient to statistically differentiate between DNA molecules of nearly identical lengths differing only in sequence and AT content when placed in 3.5 M urea. Our results demonstrate that nanopores can detect subtle structural changes and are thus a valuable tool for detecting differences in biomolecules' environment.

  18. CalFitter: a web server for analysis of protein thermal denaturation data.

    Science.gov (United States)

    Mazurenko, Stanislav; Stourac, Jan; Kunka, Antonin; Nedeljkovic, Sava; Bednar, David; Prokop, Zbynek; Damborsky, Jiri

    2018-05-14

    Despite significant advances in the understanding of protein structure-function relationships, revealing protein folding pathways still poses a challenge due to a limited number of relevant experimental tools. Widely-used experimental techniques, such as calorimetry or spectroscopy, critically depend on a proper data analysis. Currently, there are only separate data analysis tools available for each type of experiment with a limited model selection. To address this problem, we have developed the CalFitter web server to be a unified platform for comprehensive data fitting and analysis of protein thermal denaturation data. The server allows simultaneous global data fitting using any combination of input data types and offers 12 protein unfolding pathway models for selection, including irreversible transitions often missing from other tools. The data fitting produces optimal parameter values, their confidence intervals, and statistical information to define unfolding pathways. The server provides an interactive and easy-to-use interface that allows users to directly analyse input datasets and simulate modelled output based on the model parameters. CalFitter web server is available free at https://loschmidt.chemi.muni.cz/calfitter/.

  19. Formation of Polyphenol-Denatured Protein Flocs in Alcohol Beverages Sweetened with Refined Cane Sugars.

    Science.gov (United States)

    Eggleston, Gillian; Triplett, Alexa

    2017-11-08

    The sporadic appearance of floc from refined, white cane sugars in alcohol beverages remains a technical problem for both beverage manufacturers and sugar refiners. Cane invert sugars mixed with 60% pure alcohol and water increased light scattering by up to ∼1000-fold. Insoluble and soluble starch, fat, inorganic ash, oligosaccharides, Brix, and pH were not involved in the prevailing floc-formation mechanism. Strong polynomial correlations existed between the haze floc and indicator values (IVs) (color at 420 nm pH 9.0/color at pH 4.0-an indirect measure of polyphenolic and flavonoid colorants) (R 2 = 0.815) and protein (R 2 = 0.819) content of the invert sugars. Ethanol-induced denaturation of the protein exposed hydrophobic polyphenol-binding sites that were further exposed when heated to 80 °C. A tentative mechanism for floc formation was advanced by molecular probing with a haze (floc) active protein and polyphenol as well as polar, nonpolar, and ionic solvents.

  20. Modulation of the Extent of Cooperative Structural Change During Protein Folding by Chemical Denaturant.

    Science.gov (United States)

    Jethva, Prashant N; Udgaonkar, Jayant B

    2017-09-07

    Protein folding and unfolding reactions invariably appear to be highly cooperative reactions, but the structural and sequence determinants of cooperativity are poorly understood. Importantly, it is not known whether cooperative structural change occurs throughout the protein, or whether some parts change cooperatively and other parts change noncooperatively. In the current study, hydrogen exchange mass spectrometry has been used to show that the mechanism of unfolding of the PI3K SH3 domain is similar in the absence and presence of 5 M urea. The data are well described by a four state N ↔ I N ↔ I 2 ↔ U model, in which structural changes occur noncooperatively during the N ↔ I N and I N ↔ I 2 transitions, and occur cooperatively during the I 2 ↔ U transition. The nSrc-loop and RT-loop, as well as β strands 4 and 5 undergo noncooperative unfolding, while β strands 1, 2, and 3 unfold cooperatively in the absence of urea. However, in the presence of 5 M urea, the unfolding of β strand 4 switches to become cooperative, leading to an increase in the extent of cooperative structural change. The current study highlights the relationship between protein stability and cooperativity, by showing how the extent of cooperativity can be varied, using chemical denaturant to alter protein stability.

  1. Effect of free cysteine on the denaturation and aggregation of holo α-lactalbumin

    DEFF Research Database (Denmark)

    Nielsen, Line R.; Lund, Marianne N.; Davies, Michael J.

    2018-01-01

    α-Lactalbumin (α-LA) is a key commercial whey protein for nutritional purposes. The holo protein (calcium saturated) is considered the most heat stable whey protein, capable of refolding from unfolded states under many conditions. This is due to the absence of free thiols (cysteine residues......) that are typically involved in thermal aggregation and thiol–disulphide exchange reactions of other whey proteins. Heating (0–120 min at 90 °C, pH 7.0) holo α-LA generates free thiols through thermal cleavage of disulphide bonds, resulting in aggregates comprising unfolded α-LA species. The addition of free cysteine...... promotes the formation of soluble aggregates, effectively decreasing the holding time required to reach a particular aggregate size in a dose-dependent manner (0.35–1.4 mM cysteine). Excess cysteine (≥14 mM) causes a destabilisation of α-LA, shown by decreased denaturation temperature and gel formation...

  2. Size distribution of DNA molecules recovered from non-denaturing filter elution

    International Nuclear Information System (INIS)

    Bloecher, D.; Iliakis, G.

    1991-01-01

    DNA fragments removed from the filter during non-denaturing filter elution were collected and loaded on top of neutral sucrose gradients. Their size distribution was determined by low-speed centrifugation in neutral sucrose gradients. The average size of eluted DNA was found to be approximately 110 S; the average size of DNA collected after short elution times was found to be slightly larger than after long elution times. It is concluded that the size of eluted DNA fragments is not correlated with elution rate, and it is proposed that shear forces generated at the filter pores cause degradation of the DNA. Comparison of sedimentation profiles of carefully prepared cellular DNA before and after elution revealed that generated shear forces during elution break down DNA to an extent equivalent to around 20 000 DNA double-strand breaks (dsb) per G 1 cell. The size of DNA fragments decreased with increasing radiation dose; five times more dsb were found than expected after exposure to radiation alone. It is proposed that excess of dsb may derive from the transformation of other radiation-induced lesions to dsb under the action of shear forces generated during elution. (author)

  3. Programmable self-assembly of carbon nanotubes assisted by reversible denaturation of a protein

    International Nuclear Information System (INIS)

    Nithiyasri, P; Parthasarathy, M; Balaji, K; Brindha, P

    2012-01-01

    Self-assembly of pristine multi-walled carbon nanotubes (CNTs) in aqueous dispersion using a protein, bovine serum albumin (BSA), has been demonstrated. Step-wise conformational changes in BSA as a function of temperature have been deployed to direct the assembly of nanotubes. More specifically, CNTs distributed randomly in native BSA at 35 °C as well as completely denatured BSA solution at 80 °C self-assemble in the intermediate temperature range of 45–65 °C, as evident from scanning and transmission electron microscopy. Fourier transform infrared (FTIR) and fluorescence studies indicate significant changes in the α-helical content of the protein with respect to the amide I and II bands and tryptophan emission intensity, respectively. The stability of CNT dispersion in BSA solution has been attributed to the hydrophobic interaction between nanotubes and the protein molecule by adding sodium cholate to the dispersion. Moreover, a mechanism based on electrostatic repulsion between BSA-bound CNTs has been proposed for the thermally reversible assembly of CNTs in BSA solution based on evidence from zeta potential measurements and FTIR spectroscopy. Thus the present report demonstrates bio-mimetic self-assembly of as-synthesized CNTs using changes in surface charge and conformation of an unfolding protein for biomedical applications and nanobiotechnology. (paper)

  4. Protein Denaturation on p-T Axes--Thermodynamics and Analysis.

    Science.gov (United States)

    Smeller, László

    2015-01-01

    Proteins are essential players in the vast majority of molecular level life processes. Since their structure is in most cases substantial for their correct function, study of their structural changes attracted great interest in the past decades. The three dimensional structure of proteins is influenced by several factors including temperature, pH, presence of chaotropic and cosmotropic agents, or presence of denaturants. Although pressure is an equally important thermodynamic parameter as temperature, pressure studies are considerably less frequent in the literature, probably due to the technical difficulties associated to the pressure studies. Although the first steps in the high-pressure protein study have been done 100 years ago with Bridgman's ground breaking work, the field was silent until the modern spectroscopic techniques allowed the characterization of the protein structural changes, while the protein was under pressure. Recently a number of proteins were studied under pressure, and complete pressure-temperature phase diagrams were determined for several of them. This review summarizes the thermodynamic background of the typical elliptic p-T phase diagram, its limitations and the possible reasons for deviations of the experimental diagrams from the theoretical one. Finally we show some examples of experimentally determined pressure-temperature phase diagrams.

  5. The quaternary structure of the recombinant bovine odorant-binding protein is modulated by chemical denaturants.

    Directory of Open Access Journals (Sweden)

    Olga V Stepanenko

    Full Text Available A large group of odorant-binding proteins (OBPs has attracted great scientific interest as promising building blocks in constructing optical biosensors for dangerous substances, such as toxic and explosive molecules. Native tissue-extracted bovine OBP (bOBP has a unique dimer folding pattern that involves crossing the α-helical domain in each monomer over the other monomer's β-barrel. In contrast, recombinant bOBP maintaining the high level of stability inherent to native tissue bOBP is produced in a stable native-like state with a decreased tendency for dimerization and is a mixture of monomers and dimers in a buffered solution. This work is focused on the study of the quaternary structure and the folding-unfolding processes of the recombinant bOBP in the absence and in the presence of guanidine hydrochloride (GdnHCl. Our results show that the recombinant bOBP native dimer is only formed at elevated GdnHCl concentrations (1.5 M. This process requires re-organizing the protein structure by progressing through the formation of an intermediate state. The bOBP dimerization process appears to be irreversible and it occurs before the protein unfolds. Though the observed structural changes for recombinant bOBP at pre-denaturing GdnHCl concentrations show a local character and the overall protein structure is maintained, such changes should be considered where the protein is used as a sensitive element in a biosensor system.

  6. Potential use of gradient denaturing gel electrophoresis in obtaining mutational spectra from human cells

    International Nuclear Information System (INIS)

    Thilly, W.G.

    1985-01-01

    A method is described to isolate mutations in DNA in human cells. When a double-stranded DNA migrates through an electric field on an electrophoretic gel, it is compact hydrodynamic structure relative to the same material in a melted form. Normally the solution in electrophoretic gels is uniform, but a way has been devised to set up a stable gradient of increasing solute concentration in the direction of DNA motion. Thus, as a double-stranded DNA molecule is drawn by the electric field into higher and higher concentrations of urea/formamide, it will eventually reach a point at which the concentration is high enough to melt the lower-melting-point region. The melting results in an essentially immobile structure within the gel so that the position at which the DNA molecule stops on the gradient gel is determined by its melting point, which is uniquely determined by its nucleotide sequence. A single base pair substitution within a low melting point sequence of some 100 base pairs changed the expected melting point by 0.4 0 C and resulted in about a 2-cm displacement under appropriate denaturing gel conditions. This expectation leads to the idea that if a mixture of DNA sequences derived from point mutations within the same restriction fragment were permitted to anneal with a complementary wild-type sequence, the melting point of each type of heteroduplex would differ depending on the kind and position of each mutation

  7. Use of denaturing gradient gel electrophoresis to detect Actinobacteria associated with the human faecal microbiota.

    Science.gov (United States)

    Hoyles, Lesley; Clear, Jessica A; McCartney, Anne L

    2013-08-01

    With the exceptions of the bifidobacteria, propionibacteria and coriobacteria, the Actinobacteria associated with the human gastrointestinal tract have received little attention. This has been due to the seeming absence of these bacteria from most clone libraries. In addition, many of these bacteria have fastidious growth and atmospheric requirements. A recent cultivation-based study has shown that the Actinobacteria of the human gut may be more diverse than previously thought. The aim of this study was to develop a denaturing gradient gel electrophoresis (DGGE) approach for characterizing Actinobacteria present in faecal samples. Amount of DNA added to the Actinobacteria-specific PCR used to generate strong PCR products of equal intensity from faecal samples of five infants, nine adults and eight elderly adults was anti-correlated with counts of bacteria obtained using fluorescence in situ hybridization probe HGC69A. A nested PCR using Actinobacteria-specific and universal PCR-DGGE primers was used to generate profiles for the Actinobacteria. Cloning of sequences from the DGGE bands confirmed the specificity of the Actinobacteria-specific primers. In addition to members of the genus Bifidobacterium, species belonging to the genera Propionibacterium, Microbacterium, Brevibacterium, Actinomyces and Corynebacterium were found to be part of the faecal microbiota of healthy humans. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. A Comparison Between Denaturing Gradient Gel Electrophoresis and Denaturing High Performance Liquid Chromatography in Detecting Mutations in Genes Associated with Hereditary Non-Polyposis Colorectal Cancer (HNPCC and the Identification of 9 New Mutations Previously Unidentified by DGGE

    Directory of Open Access Journals (Sweden)

    Meldrum Cliff J

    2003-12-01

    Full Text Available Abstract Denaturing high performance liquid chromatography is a relatively new method by which heteroduplex structures formed during the PCR amplification of heterozygote samples can be rapidly identified. The use of this technology for mutation detection in hereditary non-polyposis colorectal cancer (HNPCC has the potential to appreciably shorten the time it takes to analyze genes associated with this disorder. Prior to acceptance of this method for screening genes associated with HNPCC, assessment of the reliability of this method should be performed. In this report we have compared mutation and polymorphism detection by denaturing gradient gel electrophoresis (DGGE with denaturing high performance liquid chromatography (DHPLC in a set of 130 families. All mutations/polymorphisms representing base substitutions, deletions, insertions and a 23 base pair inversion were detected by DHPLC whereas DGGE failed to identify four single base substitutions and a single base pair deletion. In addition, we show that DHPLC has been used for the identification of 5 different mutations in exon 7 of hMSH2 that could not be detected by DGGE. From this study we conclude that DHPLC is a more effective and rapid alternative to the detection of mutations in hMSH2 and hMLH1 with the same or better accuracy than DGGE. Furthermore, this technique offers opportunities for automation, which have not been realised for the majority of other methods of gene analysis.

  9. Structural and dynamical study about denatured states of yeast phosphoglycerate kinase by neutrons scattering and X-rays

    International Nuclear Information System (INIS)

    Receveur, V.

    1997-01-01

    During a long time, the neutron scattering and X-rays techniques have not been used for the studies bearing on the folding of proteins. The compactness and the globularness of a protein are two structural characteristics describing the denatured states and the intermediate states of folding, and the neutrons and x-rays scattering are probably the two techniques the most appropriate to give this kind of information; they are sensible to the spatial extent and to the molecules compactness, and to their general shape. For these three or four last years, the works using these techniques are increasing, giving precious knowledge on the different steps of folding and on the interactions stabilizing the denatured or intermediate states. This thesis falls into this category. (N.C.)

  10. Non-invasive diagnostic workup of patients with suspected stable angina by combined computed tomography coronary angiography and magnetic resonance perfusion imaging

    International Nuclear Information System (INIS)

    Kirschbaum, S.W.; Nieman, K.; Springeling, T.

    2011-01-01

    The background of this study was to evaluate additional adenosine magnetic resonance perfusion (MRP) imaging in the diagnostic workup of patients with suspected stable angina with computed tomography coronary angiography (CTCA) as first-line diagnostic modality. Two hundred and thirty symptomatic patients (male, 52%; age, 56 year) with suspected stable angina underwent CTCA. In patients with a stenosis of >50% as visually assessed, MRP was performed and the quantitative myocardial perfusion reserve index (MPRI) was calculated. Coronary flow reserve (CFR) using invasive coronary flow measurements served as the standard of reference. CTCA showed non-significant coronary artery disease (CAD) in 151/230 (66%) patients and significant CAD in 79/230 patients (34%), of whom 50 subsequently underwent MRP and CFR. MRP showed reduced perfusion in 32 patients (64%), which was confirmed by CFR in 27 (84%). All 18 cases of normal MRP (36%) were confirmed by CFR. The positive likelihood ratio of MRP for the presence of functional significant disease in patients with a lesion on CTCA was 4.49 (95% confidence interval [CI] 2.12-9.99). The negative likelihood ratio was 0.05 (95% CI 0.01-0.34). CTCA as first-line diagnostic modality excluded coronary artery disease in a high percentage of patients referred for diagnostic workup of suspected stable angina. MRP made a significant contribution to the detection of functional significant lesions in patients with a positive CTCA. (author)

  11. International recommendation for a comprehensive neuropathologic workup of epilepsy surgery brain tissue: A consensus Task Force report from the ILAE Commission on Diagnostic Methods.

    Science.gov (United States)

    Blümcke, Ingmar; Aronica, Eleonora; Miyata, Hajime; Sarnat, Harvey B; Thom, Maria; Roessler, Karl; Rydenhag, Bertil; Jehi, Lara; Krsek, Pavel; Wiebe, Samuel; Spreafico, Roberto

    2016-03-01

    Epilepsy surgery is an effective treatment in many patients with drug-resistant focal epilepsies. An early decision for surgical therapy is facilitated by a magnetic resonance imaging (MRI)-visible brain lesion congruent with the electrophysiologically abnormal brain region. Recent advances in the pathologic diagnosis and classification of epileptogenic brain lesions are helpful for clinical correlation, outcome stratification, and patient management. However, application of international consensus classification systems to common epileptic pathologies (e.g., focal cortical dysplasia [FCD] and hippocampal sclerosis [HS]) necessitates standardized protocols for neuropathologic workup of epilepsy surgery specimens. To this end, the Task Force of Neuropathology from the International League Against Epilepsy (ILAE) Commission on Diagnostic Methods developed a consensus standard operational procedure for tissue inspection, distribution, and processing. The aims are to provide a systematic framework for histopathologic workup, meeting minimal standards and maximizing current and future opportunities for morphofunctional correlations and molecular studies for both clinical care and research. Whenever feasible, anatomically intact surgical specimens are desirable to enable systematic analysis in selective hippocampectomies, temporal lobe resections, and lesional or nonlesional neocortical samples. Correct orientation of sample and the sample's relation to neurophysiologically aberrant sites requires good communication between pathology and neurosurgical teams. Systematic tissue sampling of 5-mm slabs along a defined anatomic axis and application of a limited immunohistochemical panel will ensure a reliable differential diagnosis of main pathologies encountered in epilepsy surgery. Wiley Periodicals, Inc. © 2016 International League Against Epilepsy.

  12. Can value for money be improved by changing the sequence of our donor work-up in the living kidney donor programme?

    Science.gov (United States)

    Larsen, Jesper; Sørensen, Søren Schwartz; Feldt-Rasmussen, Bo

    2009-08-01

    The aim of the study was to identify procedures of maximum importance for acceptance or rejection of kidney donation from a living donor as well as making the process more cost-effective. We identified all potential living related donors who were examined during the period between January 2002 and December 2006 at our department. The cost in euro (euro) for the programme was estimated using the Danish diagnosis-related group-system (DRG). The donor work-up programme was described. One hundred and thirty-three potential donors were identified; 66 male- and 67 female subjects, median age of 52 years (range 22-69). Sixty-four participants were rejected as donors. Abdominal CT-scan with angiography and urography ruled out 22 of the above 64 potential organ donors; thus, 48% of the volunteers for living kidney donation were unsuited for donation. Abdominal CT-scan with angiography and urography was the procedure identifying most subjects who were unsuited for kidney donation. A rearrangement of the present donor work-up programme could potentially reduce the costs from euro6911 to euro5292 per donor--saving 23% of the costs. By changing the sequence of examinations, it might be possible to cut down on time spent and number of tests needed for approving or rejecting subjects for living kidney donation.

  13. Protective role of microRNA-29a in denatured dermis and skin fibroblast cells after thermal injury

    Directory of Open Access Journals (Sweden)

    Jie Zhou

    2016-03-01

    Full Text Available Our previous study has suggested that downregulated microRNA (miR-29a in denatured dermis might be involved in burn wound healing. However, the exact role of miR-29a in healing of burn injury still remains unclear. Here, we found that expression of miR-29a was notably upregulated in denatured dermis tissues and skin fibroblast cells after thermal injury, and thereafter gradually downregulated compared with control group. By contrast, the expression of collagen, type I, alpha 2 (COL1A2 and vascular endothelial growth factor (VEGF-A were first reduced and subsequently upregulated in denatured dermis tissues and skin fibroblast cells after thermal injury. We further identified COL1A2 as a novel target of miR-29a, which is involved in type I collagen synthesis, and showed that miR-29a negatively regulated the expression level of COL1A2 in skin fibroblast cells. In addition, VEGF-A, another target gene of miR-29a, was also negatively mediated by miR-29a in skin fibroblast cells. Inhibition of miR-29a expression significantly promoted the proliferation and migration of skin fibroblast cells after thermal injury, and knockdown of COL1A2 and VEGF-A reversed the effects of miR-29a on the proliferation and migration of skin fibroblast cells. Furthermore, we found that Notch2/Jagged2 signaling was involved in miR-29a response to burn wound healing. Our findings suggest that downregulated miR-29a in denatured dermis may help burn wound healing in the later phase, probably via upregulation of COL1A2 and VEGF-A expression, which can further enhance type I collagen synthesis and angiogenesis.

  14. Counteraction of urea-induced protein denaturation by trimethylamine N-oxide: A chemical chaperone at atomic resolution

    OpenAIRE

    Bennion, Brian J.; Daggett, Valerie

    2004-01-01

    Proteins are very sensitive to their solvent environments. Urea is a common chemical denaturant of proteins, yet some animals contain high concentrations of urea. These animals have evolved an interesting mechanism to counteract the effects of urea by using trimethylamine N-oxide (TMAO). The molecular basis for the ability of TMAO to act as a chemical chaperone remains unknown. Here, we describe molecular dynamics simulations of a small globular protein, chymotrypsin inhibitor 2, in 8 M urea ...

  15. Preparation of denatured protein bone sterilized with gamma radiation; Preparacion de hueso desproteinizado esterilizado con radiacion gamma

    Energy Technology Data Exchange (ETDEWEB)

    Luna Z, D [ININ, 52045 Ocoyoacac, Estado de Mexico (Mexico)

    2005-07-01

    The bone is one of the tissues more transplanted in the entire world by that the bone necessity for transplant every day becomes bigger. In the Bank of tissues Radio sterilized of the ININ the amnion and the pig skin are routinely processed. The tissue with which will be continued is with bone. Due to that in our country it doesn't have enough bone of human origin for the necessities required in the bone transplant, an option is the bone of bovine. Of this bone one can obtain denatured protein bone, with the same characteristics of the denatured protein human bone, the one which has been proven that it has good acceptance and incorporation in the human body when is transplanted. The method for the obtaining of the denatured protein bone of bovine, with the confirmation of the final product by means of X-ray diffraction is described. The radiosterilization of this bone with gamma rays and the determination of the lead content. (Author)

  16. Interchange reaction of disulfides and denaturation of oxytocin by copper(II)/ascorbic acid/O2 system.

    Science.gov (United States)

    Inoue, H; Hirobe, M

    1987-05-29

    The interchange reaction of disulfides was caused by the copper(II)/ascorbic acid/O2 system. The incubation of two symmetric disulfides, L-cystinyl-bis-L-phenylalanine (PP) and L-cystinyl-bis-L-tyrosine (TT), with L-ascorbic acid and CuSO4 in potassium phosphate buffer (pH 7.2, 50 mM) resulted in the formation of an asymmetric disulfide, L-cystinyl-L-phenylalanine-L-tyrosine (PT), and the final ratio of PP:PT:TT was 1:2:1. As the reaction was inhibited by catalase and DMSO only at the initial time, hydroxyl radical generated by the copper(II)/ascorbic acid/O2 system seemed to be responsible for the initiation of the reaction. Oxytocin and insulin were denatured by this system, and catalase and DMSO similarly inhibited these denaturations. As the composition of amino acids was unchanged after the reaction, hydroxyl radical was thought to cause the cleavage and/or interchange reaction of disulfides to denature the peptides.

  17. A Proposed Mechanism for the Thermal Denaturation of a Recombinant Bacillus Halmapalus Alpha-amylase - the Effect of Calcium Ions

    Science.gov (United States)

    Nielsen, Anders D.; Pusey, Marc L.; Fuglsang, Claus C.; Westh, Peter

    2003-01-01

    The thermal stability of a recombinant alpha-amylase from Bacillus halmapalus alpha-amylase (BHA) has been investigated using circular dichroism spectroscopy (CD) and differential scanning calorimetry (DSC). This alpha-amylase is homologous to other Bacillus alpha-amylases where previous crystallographic studies have identified the existence of 3 calcium binding sites in the structure. Denaturation of BHA is irreversible with a Tm of approximately 89 C, and DSC thermograms can be described using a one-step irreversible model. A 5 C increase in T(sub m) in the presence of 10 fold excess CaCl2 was observed. However, a concomitant increase in the tendency to aggregate was also observed. The presence of 30-40 fold excess calcium chelator (EDTA or EGTA) results in a large destabilization of BHA corresponding to about 40 C lower T(sub m), as determined by both CD and DSC. Ten fold excess EGTA reveals complex DSC thermograms corresponding to both reversible and irreversible transitions, which possibly originate from different populations of BHA:calcium complexes. The observations in the present study have, in combination with structural information of homologous alpha-amylases, provided the basis for the proposal of a simple denaturation mechanism of BHA. The proposed mechanism describes the irreversible thermal denaturation of different BHA:calcium complexes and the calcium binding equilibrium involved. Furthermore, the model accounts for a temperature induced reversible structural change associated with calcium binding.

  18. Selection for Protein Kinetic Stability Connects Denaturation Temperatures to Organismal Temperatures and Provides Clues to Archaean Life.

    Directory of Open Access Journals (Sweden)

    M Luisa Romero-Romero

    Full Text Available The relationship between the denaturation temperatures of proteins (Tm values and the living temperatures of their host organisms (environmental temperatures: TENV values is poorly understood. Since different proteins in the same organism may show widely different Tm's, no simple universal relationship between Tm and TENV should hold, other than Tm≥TENV. Yet, when analyzing a set of homologous proteins from different hosts, Tm's are oftentimes found to correlate with TENV's but this correlation is shifted upward on the Tm axis. Supporting this trend, we recently reported Tm's for resurrected Precambrian thioredoxins that mirror a proposed environmental cooling over long geological time, while remaining a shocking ~50°C above the proposed ancestral ocean temperatures. Here, we show that natural selection for protein kinetic stability (denaturation rate can produce a Tm↔TENV correlation with a large upward shift in Tm. A model for protein stability evolution suggests a link between the Tm shift and the in vivo lifetime of a protein and, more specifically, allows us to estimate ancestral environmental temperatures from experimental denaturation rates for resurrected Precambrian thioredoxins. The TENV values thus obtained match the proposed ancestral ocean cooling, support comparatively high Archaean temperatures, and are consistent with a recent proposal for the environmental temperature (above 75°C that hosted the last universal common ancestor. More generally, this work provides a framework for understanding how features of protein stability reflect the environmental temperatures of the host organisms.

  19. NF2 tumor suppressor gene: a comprehensive and efficient detection of somatic mutations by denaturing HPLC and microarray-CGH.

    Science.gov (United States)

    Szijan, Irene; Rochefort, Daniel; Bruder, Carl; Surace, Ezequiel; Machiavelli, Gloria; Dalamon, Viviana; Cotignola, Javier; Ferreiro, Veronica; Campero, Alvaro; Basso, Armando; Dumanski, Jan P; Rouleau, Guy A

    2003-01-01

    The NF2 tumor suppressor gene, located in chromosome 22q12, is involved in the development of multiple tumors of the nervous system, either associated with neurofibromatosis 2 or sporadic ones, mainly schwannomas and meningiomas. In order to evaluate the role of the NF2 gene in sporadic central nervous system (CNS) tumors, we analyzed NF2 mutations in 26 specimens: 14 meningiomas, 4 schwannomas, 4 metastases, and 4 other histopathological types of neoplasms. Denaturing high performance liquid chromatography (denaturing HPLC) and comparative genomic hybridization on a DNA microarray (microarray- CGH) were used as scanning methods for small mutations and gross rearrangements respectively. Small mutations were identified in six out of seventeen meningiomas and schwannomas, one mutation was novel. Large deletions were detected in six meningiomas. All mutations were predicted to result in truncated protein or in the absence of a large protein domain. No NF2 mutations were found in other histopathological types of CNS tumors. These results provide additional evidence that mutations in the NF2 gene play an important role in the development of sporadic meningiomas and schwannomas. Denaturing HPLC analysis of small mutations and microarray-CGH of large deletions are complementary, fast, and efficient methods for the detection of mutations in tumor tissues.

  20. Preparation of denatured protein bone sterilized with gamma radiation; Preparacion de hueso desproteinizado esterilizado con radiacion gamma

    Energy Technology Data Exchange (ETDEWEB)

    Luna Z, D. [ININ, 52045 Ocoyoacac, Estado de Mexico (Mexico)]. e-mail: dlz@nuclear.inin.mx

    2005-07-01

    The bone is one of the tissues more transplanted in the entire world by that the bone necessity for transplant every day becomes bigger. In the Bank of tissues Radio sterilized of the ININ the amnion and the pig skin are routinely processed. The tissue with which will be continued is with bone. Due to that in our country it doesn't have enough bone of human origin for the necessities required in the bone transplant, an option is the bone of bovine. Of this bone one can obtain denatured protein bone, with the same characteristics of the denatured protein human bone, the one which has been proven that it has good acceptance and incorporation in the human body when is transplanted. The method for the obtaining of the denatured protein bone of bovine, with the confirmation of the final product by means of X-ray diffraction is described. The radiosterilization of this bone with gamma rays and the determination of the lead content. (Author)

  1. Transition from uranium to denatured uranium/thorium fuel in an existing PWR

    International Nuclear Information System (INIS)

    Walters, M.A.

    1982-01-01

    The purpose of this research was to determine whether it is possible to make a gradual transition from uranium to denatured uranium/thorium (DUTH) fuel in an existing PWR by adding DUTH assemblies during each scheduled refueling and, if the transition is possible, to develop a general procedure for making it. The feasibility of the transition was established by identifying acceptable refueling schemes for a series of transition cores, and in the process, a method for identifying acceptable schemes evolved. The utility of the method was then demonstrated by applying it to a standard reactor operating under normal conditions. The vehicle used to examine proposed fuel mixtures and to select acceptable ones was a set of one-dimensional computer codes. The core was modeled as a set of five concentric fuel zones with a reflector. Fuel mixtures were proposed and the computer codes were used to determine whether a mixture was acceptable, i.e., whether it had the desired k-effective and flux and power distributions. The parameters allowed to vary in selection of proposed fuel mixtures were enrichment of fresh fuel assemblies, number of uranium and DUTH assemblies added during each refueling, and distribution of fuel in the core. Results of the research showed that a gradual transition is possible. Furthermore, there is a method that allows the identification of fuel mixtures that are likely to be acceptable. It requires the calculation of K-infinity for the entire proposed core and for some of its regions. These values of K-infinity and relationships developed in this research can be used to predict the flux distribution and the final k-effective for the proposed fuel mixture

  2. Polymerase chain reaction-based denaturing gradient gel electrophoresis in the evaluation of oral microbiota.

    Science.gov (United States)

    Li, Y; Saxena, D; Barnes, V M; Trivedi, H M; Ge, Y; Xu, T

    2006-10-01

    Clinical evaluation of oral microbial reduction after a standard prophylactic treatment has traditionally been based on bacterial cultivation methods. However, not all microbes in saliva or dental plaque can be cultivated. Polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE) is a cultivation-independent molecular fingerprinting technique that allows the assessment of the predominant bacterial species present in the oral cavity. This study sought to evaluate the oral microbial changes that occurred after a standard prophylactic treatment with a conventional oral care product using PCR-DGGE. Twelve healthy adults participated in the study. Pooled plaque samples were collected at baseline, 24 h after prophylaxis (T1), and 4 days after toothbrushing with fluoride toothpaste (T4). The total microbial genomic DNA of the plaque was isolated. PCR was performed with a set of universal bacterial 16S rDNA primers. The PCR-amplified 16S rDNA fragments were separated by DGGE. The effects of the treatment and of dental brushing were assessed by comparing the PCR-DGGE fingerprinting profiles. The mean numbers of detected PCR amplicons were 22.3 +/- 6.1 for the baseline group, 13.0 +/- 3.1 for the T1 group, and 13.5 +/- 4.3 for the T4 group; the differences among the three groups were statistically significant (P < 0.01). The study also found a significant difference in the mean similarities of microbial profiles between the baseline and the treatment groups (P < 0.001). PCR-based DGGE has been shown to be an excellent means of rapidly and accurately assessing oral microbial changes in this clinical study.

  3. An improved method for detecting genetic variation in DNA using denaturing gradient gel electrophoresis

    International Nuclear Information System (INIS)

    Takahashi, Norio; Hiyama, Keiko; Kodaira, Mieko; Satoh, Chiyoko.

    1990-05-01

    We have examined the feasibility of denaturing gradient gel electrophoresis (DGGE) of RNA:DNA duplexes to detect variations in genomic and cloned DNAs. The result has demonstrated that use of RNA:DNA duplexes makes DGGE much more practical for screening a large number of samples than use of DNA:DNA heteroduplexes, because preparation of RNA probes is easier than that of DNA probes. Three different 32 P-labeled RNA probes were produced. Genomic or cloned DNAs were digested with restriction enzymes and hybridized to labeled RNA probes, and resulting RNA:DNA duplexes were examined by DGGE. The presence of a mismatch(es) was detected as a difference in the mobility of bands on the gel. The experimental conditions were determined using DNA segments from cloned normal and three thalassemic human β-globin genes. The results from experiments on the cloned DNAs suggest that DGGE of RNA:DNA duplexes will detect nucleotide substitutions and deletions in DNA. In the course of these studies, a polymorphism due to a single-base substitution at position 666 of IVS2 (IVS2-666) of the human β-globin gene was directly identified using genomic DNA samples. A study of 59 unrelated Japanese from Hiroshima was undertaken in which the frequency of the allele with C at IVS2-666 was 0.48 and that of the allele with T was 0.52. This approach was found to be very effective for detecting heritable variation and should be a powerful tool for detecting fresh mutations in DNA, which occur outside the known restriction sites. (author)

  4. Nonlinear acoustic properties of ex vivo bovine liver and the effects of temperature and denaturation

    International Nuclear Information System (INIS)

    Jackson, E J; Coussios, C-C; Cleveland, R O

    2014-01-01

    Thermal ablation by high intensity focused ultrasound (HIFU) has a great potential for the non-invasive treatment of solid tumours. Due to the high pressure amplitudes involved, nonlinear acoustic effects must be understood and the relevant medium property is the parameter of nonlinearity B/A. Here, B/A was measured in ex vivo bovine liver, over a heating/cooling cycle replicating temperatures reached during HIFU ablation, adapting a finite amplitude insertion technique, which also allowed for measurement of sound-speed and attenuation. The method measures the nonlinear progression of a plane wave through liver and B/A was chosen so that numerical simulations matched the measured waveforms. To create plane-wave conditions, sinusoidal bursts were transmitted by a 100 mm diameter 1.125 MHz unfocused transducer and measured using a 15 mm diameter 2.25 MHz broadband transducer in the near field. Attenuation and sound-speed were calculated using a reflected pulse from the smaller transducer using the larger transducer as the reflecting interface. Results showed that attenuation initially decreased with heating then increased after denaturation, the sound-speed initially increased with temperature and then decreased, and B/A showed an increase with temperature but no significant post-heating change. The B/A data disagree with other reports that show a significant change and we suggest that any nonlinear enhancement in the received ultrasound signal post-treatment is likely due to acoustic cavitation rather than changes in tissue nonlinearity. (paper)

  5. A differential scanning calorimetric study of the effects of metal ions, substrate/product, substrate analogues and chaotropic anions on the thermal denaturation of yeast enolase 1.

    Science.gov (United States)

    Brewer, J M; Wampler, J E

    2001-03-14

    The thermal denaturation of yeast enolase 1 was studied by differential scanning calorimetry (DSC) under conditions of subunit association/dissociation, enzymatic activity or substrate binding without turnover and substrate analogue binding. Subunit association stabilizes the enzyme, that is, the enzyme dissociates before denaturing. The conformational change produced by conformational metal ion binding increases thermal stability by reducing subunit dissociation. 'Substrate' or analogue binding additionally stabilizes the enzyme, irrespective of whether turnover is occurring, perhaps in part by the same mechanism. More strongly bound metal ions also stabilize the enzyme more, which we interpret as consistent with metal ion loss before denaturation, though possibly the denaturation pathway is different in the absence of metal ion. We suggest that some of the stabilization by 'substrate' and analogue binding is owing to the closure of moveable polypeptide loops about the active site, producing a more 'closed' and hence thermostable conformation.

  6. Achieving continuous manufacturing: technologies and approaches for synthesis, workup, and isolation of drug substance. May 20-21, 2014 Continuous Manufacturing Symposium.

    Science.gov (United States)

    Baxendale, Ian R; Braatz, Richard D; Hodnett, Benjamin K; Jensen, Klavs F; Johnson, Martin D; Sharratt, Paul; Sherlock, Jon-Paul; Florence, Alastair J

    2015-03-01

    This whitepaper highlights current challenges and opportunities associated with continuous synthesis, workup, and crystallization of active pharmaceutical ingredients (drug substances). We describe the technologies and requirements at each stage and emphasize the different considerations for developing continuous processes compared with batch. In addition to the specific sequence of operations required to deliver the necessary chemical and physical transformations for continuous drug substance manufacture, consideration is also given to how adoption of continuous technologies may impact different manufacturing stages in development from discovery, process development, through scale-up and into full scale production. The impact of continuous manufacture on drug substance quality and the associated challenges for control and for process safety are also emphasized. In addition to the technology and operational considerations necessary for the adoption of continuous manufacturing (CM), this whitepaper also addresses the cultural, as well as skills and training, challenges that will need to be met by support from organizations in order to accommodate the new work flows. Specific action items for industry leaders are: Develop flow chemistry toolboxes, exploiting the advantages of flow processing and including highly selective chemistries that allow use of simple and effective continuous workup technologies. Availability of modular or plug and play type equipment especially for workup to assist in straightforward deployment in the laboratory. As with learning from other industries, standardization is highly desirable and will require cooperation across industry and academia to develop and implement. Implement and exploit process analytical technologies (PAT) for real-time dynamic control of continuous processes. Develop modeling and simulation techniques to support continuous process development and control. Progress is required in multiphase systems such as

  7. The Challenge of Prenatal Diagnostic Work-Up of Maternally Inherited X-Linked Opitz G/BBB: Case Report and Literature Review

    Directory of Open Access Journals (Sweden)

    Marialuigia Spinelli

    2015-01-01

    Full Text Available Background. Prenatal diagnosis of Optiz G/BBB syndrome (OS is challenging because the characteristic clinical features, such as facial and genitourinary anomalies, may be subtle at sonography and rather unspecific. Furthermore, molecular testing of the disease gene is not routinely performed, unless a specific diagnosis is suggested. Method. Both familial and ultrasound data were used to achieve the diagnosis of X-linked OS (XLOS, which was confirmed by molecular testing of MID1 gene (Xp22.3 at birth. Results. Sequencing of MID1 gene disclosed the nucleotide change c.1285 +1 G>T, previously associated with XLOS. Conclusions. This case illustrates current challenges of the prenatal diagnostic work-up of XLOS and exemplifies how clinical investigation, including family history, and accurate US foetal investigations can lead to the correct diagnosis.

  8. Multidetector computed tomography angiography in clinically suspected hyperacute ischemic stroke in the anterior circulation: an etiological workup in a cohort of Brazilian patients

    Directory of Open Access Journals (Sweden)

    Felipe Torres Pacheco

    2015-05-01

    Full Text Available Objective The potential of computed tomography angiography (CTA was assessed for early determination of stroke subtypes in a Brazilian cohort of patients with stroke. Method From July 2011 to July 2013, we selected patients with suspected hyperacute stroke (< 6 hours. Intracranial and cervical arteries were scrutinized on CTA and their imaging features were correlated with concurrent subtype of stroke. Results Stroke was documented in 50/106 selected patients (47.2% based on both clinical grounds and imaging follow-up (stroke group, with statistically significant arterial stenosis and vulnerable plaques on CTA. Intracranial large artery disease was demonstrated in 34% of patients in the stroke group. Partial territorial infarct prevailed (86% while artery-to-artery embolization was the most common stroke mechanism (52%. Conclusion Multidetector CTA was useful for the etiologic work-up of hyperacute ischemic stroke and facilitated the knowledge about the topographic pattern of brain infarct in accordance with its causative mechanism.

  9. Value of computed tomography arthrography with delayed acquisitions in the work-up of ganglion cysts of the tarsal tunnel: report of three cases

    International Nuclear Information System (INIS)

    Omoumi, Patrick; Gheldere, Antoine de; Leemrijse, Thibaut; Galant, Christine; Van den Bergh, Peter; Malghem, Jacques; Simoni, Paolo; Berg, Bruno C.V.; Lecouvet, Frederic E.

    2010-01-01

    Ganglion cysts are a common cause of tarsal tunnel syndrome. As in other locations, these cysts are believed to communicate with neighboring joints. The positive diagnosis and preoperative work-up of these cysts require identification and location of the cyst pedicles so that they may be excised and the risk of recurrence decreased. This can be challenging with ultrasonography and magnetic resonance (MR) imaging. We present three cases of symptomatic ganglion cysts of the tarsal tunnel, diagnosed by MR imaging, where computed tomography (CT) arthrography with delayed acquisitions helped to confirm the diagnosis and identify precisely the topography of the communication with the subtalar joint. These cases provide new evidence of the articular origin of ganglion cysts developing in the tarsal tunnel. (orig.)

  10. Can value for money be improved by changing the sequence of our donor work-up in the living kidney donor programme?

    DEFF Research Database (Denmark)

    Larsen, Jesper; Sørensen, Søren Schwartz; Feldt-Rasmussen, Bo

    2009-01-01

    (range 22-69). Sixty-four participants were rejected as donors. Abdominal CT-scan with angiography and urography ruled out 22 of the above 64 potential organ donors; thus, 48% of the volunteers for living kidney donation were unsuited for donation. Abdominal CT-scan with angiography and urography......The aim of the study was to identify procedures of maximum importance for acceptance or rejection of kidney donation from a living donor as well as making the process more cost-effective. We identified all potential living related donors who were examined during the period between January 2002...... was the procedure identifying most subjects who were unsuited for kidney donation. A rearrangement of the present donor work-up programme could potentially reduce the costs from euro6911 to euro5292 per donor--saving 23% of the costs. By changing the sequence of examinations, it might be possible to cut down...

  11. Is bias in the eye of the beholder? A vignette study to assess recognition of cognitive biases in clinical case workups.

    Science.gov (United States)

    Zwaan, Laura; Monteiro, Sandra; Sherbino, Jonathan; Ilgen, Jonathan; Howey, Betty; Norman, Geoffrey

    2017-02-01

    Many authors have implicated cognitive biases as a primary cause of diagnostic error. If this is so, then physicians already familiar with common cognitive biases should consistently identify biases present in a clinical workup. The aim of this paper is to determine whether physicians agree on the presence or absence of particular biases in a clinical case workup and how case outcome knowledge affects bias identification. We conducted a web survey of 37 physicians. Each participant read eight cases and listed which biases were present from a list provided. In half the cases the outcome implied a correct diagnosis; in the other half, it implied an incorrect diagnosis. We compared the number of biases identified when the outcome implied a correct or incorrect primary diagnosis. Additionally, the agreement among participants about presence or absence of specific biases was assessed. When the case outcome implied a correct diagnosis, an average of 1.75 cognitive biases were reported; when incorrect, 3.45 biases (F=71.3, p<0.00001). Individual biases were reported from 73% to 125% more often when an incorrect diagnosis was implied. There was no agreement on presence or absence of individual biases, with κ ranging from 0.000 to 0.044. Individual physicians are unable to agree on the presence or absence of individual cognitive biases. Their judgements are heavily influenced by hindsight bias; when the outcome implies a diagnostic error, twice as many biases are identified. The results present challenges for current error reduction strategies based on identification of cognitive biases. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  12. The role of whole body spiral CT in the primary work-up of polytrauma patients - comparison with conventional radiography and abdominal sonography

    International Nuclear Information System (INIS)

    Albrecht, T.; Schlippenbach, J. von; Wolf, K.J.; Stahel, P.F.; Ertel, W.

    2004-01-01

    Purpose: To evaluate the role of routine 'whole body spiral CT' in the primary work-up of polytrauma patients for injuries of the thorax, abdomen and spine, and to compare the results with those of conventional radiography of the chest and spine and abdominal ultrasound. Materials and Methods: Fifty consecutive polytrauma patients underwent contrast-enhanced single slice spiral CT (5 mm collimation) from the vertex to the floor of the pelvis as part of the primary work-up after emergency room admission. Overlapping high resolution sections and sagittal reformations of the spine were obtained. Reports of additional chest radiographs (n=43), abdominal ultrasound examinations (n=47) and spine radiographs (n=36) performed in the emergency room were available for retrospective comparison. The 'final diagnoses', which served as the standard of reference, were taken from the patients' records using all information that became available until discharge or death, such as findings from further imaging, surgery and autopsy. Results: CT showed 109 (97%) of 112 thoracic and abdominal soft-tissue injuries. Relevant injuries missed were an early splenic laceration and an early pelvic hematoma, both of which became clinically apparent several hours later. There were 4 false positive CT findings. Conventional chest radiography demonstrated only 20% of thoracic and sonography 22% of abdominal injuries. Chest radiography and sonography produced 2 false-positive findings each. CT showed 66 (87%) of 76 vertebral fractures including all 19 unstable ones. CT missed 5 anterior vertebral body and 5 spinous/transverse process fractures. Conventional radiography found 71% of vertebral fractures including only 50% of the unstable one. (orig.)

  13. Contrast-enhanced spectral mammography as work-up tool in patients recalled from breast cancer screening has low risks and might hold clinical benefits.

    Science.gov (United States)

    Houben, I P L; Van de Voorde, P; Jeukens, C R L P N; Wildberger, J E; Kooreman, L F; Smidt, M L; Lobbes, M B I

    2017-09-01

    Contrast-enhanced spectral mammography (CESM) is a reliable problem solving tool in the work-up of women recalled from breast cancer screening. We evaluated additional findings caused by CESM alone and outweighed them against the disadvantages of this technique. From December 2012 to December 2015, all women recalled from screening who underwent CESM were considered for this study. Radiation exposure and number of adverse contrast reactions were analysed. An experienced breast radiologist reviewed all exams and identified cases with lesions detected by CESM alone and scored their conspicuity. From these cases, data on breast density and final diagnosis were collected. For malignant cases, tumour grade and receptor characteristics were also collected. During this study, 839 women underwent CESM after a screening recall, in which five minor adverse contrast reactions were observed. Median radiation dose per exam was 6.0mGy (0.9-23.4mGy). Seventy CESM-only lesions were detected in 65 patients. Of these 70 lesions, 54.3% proved to be malignant, most commonly invasive ductal carcinomas. The remaining CESM-only lesions were benign, predominantly fibroadenomas. No complications were observed during biopsy of these lesions. Retrospectively, the majority of the lesions were either occult or a 'minimal sign' on low-energy CESM images or the screening mammogram. Using CESM as a work-up tool for women recalled from screening carries low risk for the patient, while additionally detected tumour foci might hold important clinical implications which need to be further studied in large, randomized controlled trials. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Prioritizing molecular markers to test for in the initial workup of advanced non-small cell lung cancer: wants versus needs.

    Science.gov (United States)

    West, Howard

    2017-09-01

    The current standard of care for molecular marker testing in patients with advanced non-small cell lung cancer (NSCLC) has been evolving over several years and is a product of the quality of the evidence supporting a targeted therapy for a specific molecular marker, the pre-test probability of that marker in the population, and the magnitude of benefit seen with that treatment. Among the markers that have one or more matched targeted therapies, only a few are in the subset for which they should be considered as most clearly worthy of prioritizing to detect in the first line setting in order to have them supplant other first line alternatives, and in only a subset of patients, as defined currently by NSCLC histology. Specifically, this currently includes testing for an activating epidermal growth factor receptor ( EGFR ) mutation or an anaplastic lymphoma kinase ( ALK ) or ROS1 rearrangement. This article reviews the history and data supporting the prioritization of these markers in patients with non-squamous NSCLC, a histologically selected population in whom the probability of these markers combined with the anticipated efficacy of targeted therapies against them is high enough to favor these treatments in the first line setting. In reviewing the evidence supporting this very limited core subset of most valuable molecular markers to detect in the initial workup of such patients, we can also see the criteria by which other actionable markers need to reach in order to be widely recognized as reliably valuable enough to warrant prioritization to detect in the initial workup of advanced NSCLC as well.

  15. Autistic Siblings with Novel Mutations in Two Different Genes: Insight for Genetic Workups of Autistic Siblings and Connection to Mitochondrial Dysfunction

    Directory of Open Access Journals (Sweden)

    Barrett J. Burger

    2017-10-01

    Full Text Available The prevalence of autism spectrum disorder (ASD is high, yet the etiology of this disorder is still uncertain. Advancements in genetic analysis have provided the ability to identify potential genetic changes that may contribute to ASD. Interestingly, several genetic syndromes have been linked to metabolic dysfunction, suggesting an avenue for treatment. In this case study, we report siblings with ASD who had similar initial phenotypic presentations. Whole exome sequencing (WES revealed a novel c.795delT mutation in the WDR45 gene affecting the girl, which was consistent with her eventual progression to a Rett-like syndrome phenotype including seizures along with a stereotypical cyclic breathing pattern. Interestingly, WES identified that the brother harbored a novel heterozygous Y1546H variant in the DEP domain-containing protein 5 (DEPDC5 gene, consistent with his presentation. Both siblings underwent a metabolic workup that demonstrated different patterns of mitochondrial dysfunction. The girl demonstrated statistically significant elevations in mitochondrial activity of complex I + III in both muscle and fibroblasts and increased respiration in peripheral blood mononuclear cells (PBMCs on Seahorse Extracellular Flux analysis. The boy demonstrates a statistically significant decrease in complex IV activity in buccal epithelium and decreased respiration in PBMCs. These cases highlight the differences in genetic abnormalities even in siblings with ASD phenotypes as well as highlights the individual role of novel mutations in the WDR45 and DEPDC5 genes. These cases demonstrate the importance of advanced genetic testing combined with metabolic evaluations in the workup of children with ASD.

  16. An audit to investigate the impact of false positive breast screening results and diagnostic work-up on re-engagement with subsequent routine screening

    International Nuclear Information System (INIS)

    Nightingale, Julie M.; Borgen, Rita; Porter-Bennett, Lisa; Szczepura, Katy

    2015-01-01

    Introduction: Women attending breast screening may have suspicious mammographic findings that are subsequently found at assessment clinic to be normal (false positive, FP). A false positive diagnosis is not harmless, with short and long term negative psychosocial consequences reported. Women are at increased relative risk of breast cancer therefore their attendance at subsequent screening is essential. Aims: To assess the impact of FP breast screening diagnosis and diagnostic work-up on re-attendance rates across four consecutive screening rounds at a typical breast screening centre. Method: Diagnostic interventions and screening re-attendance rates at one prior and two consecutive rounds were analysed for women receiving an FP diagnosis between 2004 and 2006. Results: 397 women (5.57%) were referred for further assessment, including 228 (57.43%) false positives. 34 eligible women failed to re-attend routine screening (+3 years), with 17 failing to re-attend subsequently (+6 years). 70.6% (24/34) of non-attenders had attended at least two screening rounds prior to FP assessment. 75% of FP women had an imaging-only assessment with 17.5% (30/171) failing to re-attend, and 25% received a biopsy, with 7% (4/57) failing to re-attend subsequently. Conclusion: This study is unique as it follows FP women through four consecutive screening rounds. FP non-attendance rates were considerably lower compared to the general screening population, with diagnostic work-up having limited influence. FP non-attendance may appear insignificant in comparison to total screened population, but these women are at greater risk of subsequent cancer so should be actively encouraged to re-engage with the screening programme

  17. Cost-effectiveness of substituting dual-energy CT for SPECT in the assessment of myocardial perfusion for the workup of coronary artery disease

    Energy Technology Data Exchange (ETDEWEB)

    Meyer, Mathias [Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Institute of Clinical Radiology and Nuclear Medicine, University Medical Center Mannheim, Medical Faculty Mannheim–Heidelberg University, Mannheim (Germany); Nance, John W. [Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Schoepf, U. Joseph, E-mail: schoepf@musc.edu [Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Department of Medicine, Division of Cardiology, Medical University of South Carolina, Charleston, SC (United States); Moscariello, Antonio [Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Department of Bioimaging and Radiological Sciences, Catholic University of Sacred Heart, “A. Gemelli” Hospital, Rome (Italy); Weininger, Markus; Rowe, Garrett W.; Ruzsics, Balazs [Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Kang, Doo Kyoung [Department of Radiology and Radiological Science, Medical University of South Carolina, Charleston, SC (United States); Department of Radiology, Ajou University School of Medicine, Suwon (Korea, Republic of); Chiaramida, Salvatore A. [Department of Medicine, Division of Cardiology, Medical University of South Carolina, Charleston, SC (United States); Schoenberg, Stefan O.; Fink, Christian [Institute of Clinical Radiology and Nuclear Medicine, University Medical Center Mannheim, Medical Faculty Mannheim–Heidelberg University, Mannheim (Germany); and others

    2012-12-15

    Purpose: We compared cost-effectiveness and potential lifetime benefits of using dual-energy computed tomography (DECT) for myocardial perfusion assessment instead of single photon emission computed tomography (SPECT) for the workup of coronary artery disease (CAD). Materials and methods: A decision and simulation model was developed to estimate cost and health effects of using DECT myocardial perfusion imaging instead of SPECT for identifying patients in need of invasive imaging and possible revascularization. The model was based on the performance indices of stress/rest DECT compared with stress/rest SPECT for detecting myocardial perfusion deficits in 50 patients (mean age 61 ± 10 years) with CAD. Stress/rest perfusion and delayed enhancement cardiac MRI served as reference standard. For DECT a reimbursement of US$1700 was assumed but costs of cardiac MRI were not included in the model. All other actual healthcare costs in these patients were derived from MUSC's hospital billing system. Results: Compared with cardiac MRI, DECT (versus SPECT) had 90% (85%) sensitivity and 71% (58%) specificity for identifying patients with obstructive CAD. Compared with the no imaging and no treatment strategy, routine SPECT gained 13.49 quality-adjusted life-years (QALYs) with an incremental cost-effectiveness ratio (ICER) of US$3557 (in 2010) per QALY. In comparison, DECT ICER was lower (US$3.191 per QALY, p = 0.0002) and an additional 0.64 QALYs was obtained (total of 14.13 QALYs) if compared with the SPECT strategy as well as the no imaging and no treatment strategy. Conclusion: Using DECT as the first-line imaging test for myocardial perfusion for the workup of patients with CAD has the potential to provide gains in QALYs, while lowering costs if compared to routine myocardial perfusion SPECT.

  18. Cost-effectiveness of substituting dual-energy CT for SPECT in the assessment of myocardial perfusion for the workup of coronary artery disease

    International Nuclear Information System (INIS)

    Meyer, Mathias; Nance, John W.; Schoepf, U. Joseph; Moscariello, Antonio; Weininger, Markus; Rowe, Garrett W.; Ruzsics, Balazs; Kang, Doo Kyoung; Chiaramida, Salvatore A.; Schoenberg, Stefan O.; Fink, Christian

    2012-01-01

    Purpose: We compared cost-effectiveness and potential lifetime benefits of using dual-energy computed tomography (DECT) for myocardial perfusion assessment instead of single photon emission computed tomography (SPECT) for the workup of coronary artery disease (CAD). Materials and methods: A decision and simulation model was developed to estimate cost and health effects of using DECT myocardial perfusion imaging instead of SPECT for identifying patients in need of invasive imaging and possible revascularization. The model was based on the performance indices of stress/rest DECT compared with stress/rest SPECT for detecting myocardial perfusion deficits in 50 patients (mean age 61 ± 10 years) with CAD. Stress/rest perfusion and delayed enhancement cardiac MRI served as reference standard. For DECT a reimbursement of US$1700 was assumed but costs of cardiac MRI were not included in the model. All other actual healthcare costs in these patients were derived from MUSC's hospital billing system. Results: Compared with cardiac MRI, DECT (versus SPECT) had 90% (85%) sensitivity and 71% (58%) specificity for identifying patients with obstructive CAD. Compared with the no imaging and no treatment strategy, routine SPECT gained 13.49 quality-adjusted life-years (QALYs) with an incremental cost-effectiveness ratio (ICER) of US$3557 (in 2010) per QALY. In comparison, DECT ICER was lower (US$3.191 per QALY, p = 0.0002) and an additional 0.64 QALYs was obtained (total of 14.13 QALYs) if compared with the SPECT strategy as well as the no imaging and no treatment strategy. Conclusion: Using DECT as the first-line imaging test for myocardial perfusion for the workup of patients with CAD has the potential to provide gains in QALYs, while lowering costs if compared to routine myocardial perfusion SPECT.

  19. INTERACTION OF IRON(II MIXED-LIGAND COMPLEXES WITH DNA: BASE-PAIR SPECIFICITY AND THERMAL DENATURATION STUDIES

    Directory of Open Access Journals (Sweden)

    Mudasir Mudasir

    2010-06-01

    Full Text Available A research about base-pair specificity of the DNA binding of [Fe(phen3]2+, [Fe(phen2(dip]2+ and [Fe(phen(dip2]2+ complexes and the effect of calf-thymus DNA (ct-DNA binding of these metal complexes on thermal denaturation of ct-DNA has been carried out. This research is intended to evaluate the preferential binding of the complexes to the sequence of DNA (A-T or G-C sequence and to investigate the binding strength and mode upon their interaction with DNA. Base-pair specificity of the DNA binding of the complexes was determined by comparing the equilibrium binding constant (Kb of each complex to polysynthetic DNA that contain only A-T or G-C sequence. The Kb value of the interaction was determined by spectrophotometric titration and thermal denaturation temperature (Tm was determined by monitoring the absorbance of the mixture solution of each complex and ct-DNA at λ =260 nm as temperature was elevated in the range of 25 - 100 oC. Results of the study show that in general all iron(II complexes studied exhibit a base-pair specificity in their DNA binding to prefer the relatively facile A-T sequence as compared to the G-C one. The thermal denaturation experiments have demonstrated that Fe(phen3]2+ and [Fe(phen2(dip]2+ interact weakly with double helical DNA via electrostatic interaction as indicated by insignificant changes in melting temperature, whereas [Fe(phen2(dip]2+  most probably binds to DNA in mixed modes of interaction, i.e.: intercalation and electrostatic interaction. This conclusion is based on the fact that the binding of [Fe(phen2(dip]2+ to ct-DNA moderately increase the Tm value of ct- DNA   Keywords: DNA Binding, mixed-ligand complexes

  20. Denatured states of yeast cytochrome c induced by heat and guanidinium chloride are structurally and thermodynamically different.

    Science.gov (United States)

    Zaidi, Sobia; Haque, Md Anzarul; Ubaid-Ullah, Shah; Prakash, Amresh; Hassan, Md Imtaiyaz; Islam, Asimul; Batra, Janendra K; Ahmad, Faizan

    2017-05-01

    A sequence alignment of mammalian cytochromes c with yeast iso-1-cytochrome c (y-cyt-c) shows that the yeast protein contains five extra N-terminal residues. We have been interested in understanding the question: What is the role of these five extra N-terminal residues in folding and stability of the protein? To answer this question we have prepared five deletants of y-cyt-c by sequentially removing these extra residues. During our studies on the wild type (WT) protein and its deletants, we observed that the amount of secondary structure in the guanidinium chloride (GdmCl)-induced denatured (D) state of each protein is different from that of the heat-induced denatured (H) state. This finding is confirmed by the observation of an additional cooperative transition curve of optical properties between H and D states on the addition of different concentrations of GdmCl to the already heat denatured WT y-cyt-c and its deletants at pH 6.0 and 68°C. For each protein, analysis of transition curves representing processes, native (N) state ↔ D state, N state ↔ H state, and H state ↔ D state, was done to obtain Gibbs free energy changes associated with all the three processes. This analysis showed that, for each protein, thermodynamic cycle accommodates Gibbs free energies associated with transitions between N and D states, N and H states, and H and D states, the characteristics required for a thermodynamic function. All these experimental observations have been supported by our molecular dynamics simulation studies.

  1. Fourier transform infrared spectroscopic studies of the secondary structure and thermal denaturation of CaATPase from rabbit skeletal muscle

    Science.gov (United States)

    Jaworsky, Mark; Brauner, Joseph W.; Mendelsohn, Richard

    Fourier transform i.r. spectroscopy has been used to monitor structural alterations induced by thermal denaturation of the intrinsic membrane protein CaATPase in aqueous media. The protein has been isolated, purified and studied in five forms: (i) In its native lipid environment after isolation from rabbit sarcoplasmic reticulum, both in H 2O and D 2O suspensions. (ii) After both mild and extensive tryptic digestion has cleaved those residues external to the membrane bilayer. (iii) Reconstituted in vesicle form with bovine brain sphingomyelin. Fourier deconvolution techniques have been used to enhance the resolution of the intrinsically overlapped Amide I and Amide II spectral regions. Large spectral alterations apparent in the deconvoluted spectra occur in these regions upon thermal denaturation of the protein which are consistent with the formation of a large proportion of β-antiparallel sheet form. The alteration parallels the loss in ATPase activity. A mild tryptic digestion increases slightly the proportion of α-helix and/or random coil secondary structure. A thermal transition to a form containing a high proportion of β structure is still evident. Extensive tryptic digestion nearly abolishes the alpha helical plus random coil secondary structure, while producing a high proportion of β form which is resistant to further thermally induced structural alterations. Studies of CaATPase reconstituted into vesicles with bovine brain sphingomyelin reveal a higher proportion of β structure than the native enzyme, with further introduction of β structure on thermal denaturation. Both the utility of deconvolution techniques and the necessity for caution in their application are apparent from the current experiments.

  2. Heterogeneity of equilibrium molten globule state of cytochrome c induced by weak salt denaturants under physiological condition.

    Directory of Open Access Journals (Sweden)

    Hamidur Rahaman

    Full Text Available While many proteins are recognized to undergo folding via intermediate(s, the heterogeneity of equilibrium folding intermediate(s along the folding pathway is less understood. In our present study, FTIR spectroscopy, far- and near-UV circular dichroism (CD, ANS and tryptophan fluorescence, near IR absorbance spectroscopy and dynamic light scattering (DLS were used to study the structural and thermodynamic characteristics of the native (N, denatured (D and intermediate state (X of goat cytochorme c (cyt-c induced by weak salt denaturants (LiBr, LiCl and LiClO4 at pH 6.0 and 25°C. The LiBr-induced denaturation of cyt-c measured by Soret absorption (Δε400 and CD ([θ]409, is a three-step process, N ↔ X ↔ D. It is observed that the X state obtained along the denaturation pathway of cyt-c possesses common structural and thermodynamic characteristics of the molten globule (MG state. The MG state of cyt-c induced by LiBr is compared for its structural and thermodynamic parameters with those found in other solvent conditions such as LiCl, LiClO4 and acidic pH. Our observations suggest: (1 that the LiBr-induced MG state of cyt-c retains the native Met80-Fe(III axial bond and Trp59-propionate interactions; (2 that LiBr-induced MG state of cyt-c is more compact retaining the hydrophobic interactions in comparison to the MG states induced by LiCl, LiClO4 and 0.5 M NaCl at pH 2.0; and (3 that there exists heterogeneity of equilibrium intermediates along the unfolding pathway of cyt-c as highly ordered (X1, classical (X2 and disordered (X3, i.e., D ↔ X3 ↔ X2 ↔ X1 ↔ N.

  3. Pressure-assisted cold denaturation of hen egg white lysozyme: the influence of co-solvents probed by hydrogen exchange nuclear magnetic resonance.

    Science.gov (United States)

    Vogtt, K; Winter, R

    2005-08-01

    COSY proton nuclear magnetic resonance was used to measure the exchange rates of amide protons of hen egg white lysozyme (HEWL) in the pressure-assisted cold-denatured state and in the heat-denatured state. After dissolving lysozyme in deuterium oxide buffer, labile protons exchange for deuterons in such a way that exposed protons are substituted rapidly, whereas "protected" protons within structured parts of the protein are substituted slowly. The exchange rates k obs were determined for HEWL under heat treatment (80 degrees C) and under high pressure conditions at low temperature (3.75 kbar, -13 degrees C). Moreover, the influence of co-solvents (sorbitol, urea) on the exchange rate was examined under pressure-assisted cold denaturation conditions, and the corresponding protection factors, P, were determined. The exchange kinetics upon heat treatment was found to be a two-step process with initial slow exchange followed by a fast one, showing residual protection in the slow-exchange state and P-factors in the random-coil-like range for the final temperature-denatured state. Addition of sorbitol (500 mM) led to an increase of P-factors for the pressure-assisted cold denatured state, but not for the heat-denatured state. The presence of 2 M urea resulted in a drastic decrease of the P-factors of the pressure-assisted cold denatured state. For both types of co-solvents, the effect they exert appears to be cooperative, i.e., no particular regions within the protein can be identified with significantly diverse changes of P-factors.

  4. Pressure-assisted cold denaturation of hen egg white lysozyme: the influence of co-solvents probed by hydrogen exchange nuclear magnetic resonance

    Directory of Open Access Journals (Sweden)

    K. Vogtt

    2005-08-01

    Full Text Available COSY proton nuclear magnetic resonance was used to measure the exchange rates of amide protons of hen egg white lysozyme (HEWL in the pressure-assisted cold-denatured state and in the heat-denatured state. After dissolving lysozyme in deuterium oxide buffer, labile protons exchange for deuterons in such a way that exposed protons are substituted rapidly, whereas "protected" protons within structured parts of the protein are substituted slowly. The exchange rates k obs were determined for HEWL under heat treatment (80ºC and under high pressure conditions at low temperature (3.75 kbar, -13ºC. Moreover, the influence of co-solvents (sorbitol, urea on the exchange rate was examined under pressure-assisted cold denaturation conditions, and the corresponding protection factors, P, were determined. The exchange kinetics upon heat treatment was found to be a two-step process with initial slow exchange followed by a fast one, showing residual protection in the slow-exchange state and P-factors in the random-coil-like range for the final temperature-denatured state. Addition of sorbitol (500 mM led to an increase of P-factors for the pressure-assisted cold denatured state, but not for the heat-denatured state. The presence of 2 M urea resulted in a drastic decrease of the P-factors of the pressure-assisted cold denatured state. For both types of co-solvents, the effect they exert appears to be cooperative, i.e., no particular regions within the protein can be identified with significantly diverse changes of P-factors.

  5. Detection by denaturing gradient gel electrophoresis of ammonia-oxidizing bacteria in microcosms of crude oil-contaminated mangrove sediments.

    Science.gov (United States)

    dos Santos, A C F; Marques, E L S; Gross, E; Souza, S S; Dias, J C T; Brendel, M; Rezende, R P

    2012-01-27

    Currently, the effect of crude oil on ammonia-oxidizing bacterium communities from mangrove sediments is little understood. We studied the diversity of ammonia-oxidizing bacteria in mangrove microcosm experiments using mangrove sediments contaminated with 0.1, 0.5, 1, 2, and 5% crude oil as well as non-contaminated control and landfarm soil from near an oil refinery in Camamu Bay in Bahia, Brazil. The evolution of CO(2) production in all crude oil-contaminated microcosms showed potential for mineralization. Cluster analysis of denaturing gradient gel electrophoresis-derived samples generated with primers for gene amoA, which encodes the functional enzyme ammonia monooxygenase, showed differences in the sample contaminated with 5% compared to the other samples. Principal component analysis showed divergence of the non-contaminated samples from the 5% crude oil-contaminated sediment. A Venn diagram generated from the banding pattern of PCR-denaturing gradient gel electrophoresis was used to look for operational taxonomic units (OTUs) in common. Eight OTUs were found in non-contaminated sediments and in samples contaminated with 0.5, 1, or 2% crude oil. A Jaccard similarity index of 50% was found for samples contaminated with 0.1, 0.5, 1, and 2% crude oil. This is the first study that focuses on the impact of crude oil on the ammonia-oxidizing bacterium community in mangrove sediments from Camamu Bay.

  6. Evidence of β-sheet structure induced kinetic stability of papain upon thermal and sodium dodecyl sulphate denaturation

    Directory of Open Access Journals (Sweden)

    Rašković Brankica

    2015-01-01

    Full Text Available Papain is a protease that consists of α-helical and β-sheet domains which unfold almost independently. Both, papain considerable thermal stability and sodium dodecyl sulphate (SDS resistance have been shown. However, the ability of each domain to unfold upon thermal and SDS denaturation has never been studied. This work shows that fruit papain has slightly higher thermal inactivation resistance when it is compared to stem papain with rather high activation energy (Ea of 223 ± 16 kJmol-1 and Tm50 value of 79 ± 2 °C. SDS resistance of fruit papain was estimated by SDS-PAGE analysis and activity staining. It has been noted that, in the presence of SDS, unless heat energy was applied in order to unfold papain, the protein remained active. Furthermore, it has been proven via Fourier transform infrared spectroscopy (FT-IR that α-helical domain of fruit papain is more prone to unfolding at elevated temperatures and in the presence of SDS then β-sheet rich domain. Thermal denaturation of papain without detergent present led to accelerated formation of aggregation specific intermolecular β-sheets as compared to native protein. Presented results are both, of fundamental and application importance. [Projekat Ministarstva nauke Republike Srbije, br. 172049

  7. Purification of inclusion bodies using PEG precipitation under denaturing conditions to produce recombinant therapeutic proteins from Escherichia coli.

    Science.gov (United States)

    Chen, Huanhuan; Li, Ninghuan; Xie, Yueqing; Jiang, Hua; Yang, Xiaoyi; Cagliero, Cedric; Shi, Siwei; Zhu, Chencen; Luo, Han; Chen, Junsheng; Zhang, Lei; Zhao, Menglin; Feng, Lei; Lu, Huili; Zhu, Jianwei

    2017-07-01

    It has been documented that the purification of inclusion bodies from Escherichia coli by size exclusion chromatography (SEC) may benefit subsequent refolding and recovery of recombinant proteins. However, loading volume and the high cost of the column limits its application in large-scale manufacturing of biopharmaceutical proteins. We report a novel process using polyethylene glycol (PEG) precipitation under denaturing conditions to replace SEC for rapid purification of inclusion bodies containing recombinant therapeutic proteins. Using recombinant human interleukin 15 (rhIL-15) as an example, inclusion bodies of rhIL-15 were solubilized in 7 M guanidine hydrochloride, and rhIL-15 was precipitated by the addition of PEG 6000. A final concentration of 5% (w/v) PEG 6000 was found to be optimal to precipitate target proteins and enhance recovery and purity. Compared to the previously reported S-200 size exclusion purification method, PEG precipitation was easier to scale up and achieved the same protein yields and quality of the product. PEG precipitation also reduced manufacturing time by about 50 and 95% of material costs. After refolding and further purification, the rhIL-15 product was highly pure and demonstrated a comparable bioactivity with a rhIL-15 reference standard. Our studies demonstrated that PEG precipitation of inclusion bodies under denaturing conditions holds significant potential as a manufacturing process for biopharmaceuticals from E. coli protein expression systems.

  8. Effect of quencher, denaturants, temperature and pH on the fluorescent properties of BSA protected gold nanoclusters

    Energy Technology Data Exchange (ETDEWEB)

    Chib, Rahul, E-mail: Rahul.chib@live.unthsc.edu [Department of Cell Biology and Immunology, Center for Fluorescence Technologies and Nanomedicine, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States); Butler, Susan [Department of Cell Biology and Immunology, Center for Fluorescence Technologies and Nanomedicine, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States); Raut, Sangram [Department of Cell Biology and Immunology, Center for Fluorescence Technologies and Nanomedicine, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States); Department of Physics and Astronomy, Texas Christian University, Fort Worth, TX 76129 (United States); Shah, Sunil; Borejdo, Julian [Department of Cell Biology and Immunology, Center for Fluorescence Technologies and Nanomedicine, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States); Gryczynski, Zygmunt [Department of Cell Biology and Immunology, Center for Fluorescence Technologies and Nanomedicine, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States); Department of Physics and Astronomy, Texas Christian University, Fort Worth, TX 76129 (United States); Gryczynski, Ignacy, E-mail: ignacy.gryczynski@unthsc.edu [Department of Cell Biology and Immunology, Center for Fluorescence Technologies and Nanomedicine, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States)

    2015-12-15

    In this paper, we have synthesized BSA protected gold nanoclusters (BSA Au nanocluster) and studied the effect of quencher, protein denaturant, pH and temperature on the fluorescence properties of the tryptophan molecule of the BSA Au nanocluster and native BSA. We have also studied their effect on the peak emission of BSA Au nanoclusters (650 nm). The photophysical characterization of a newly developed fluorophore in different environments is absolutely necessary to futher develop their biomedical and analytical applications. It was observed from our experiments that the tryptophan in BSA Au nanoclusters is better shielded from the polar environment. Tryptophan in native BSA showed a red shift in its peak emission wavelength position. Tryptophan is a highly polarity sensitive dye and a minimal change in its microenvironment can be easily observed in its photophysical properties. - Highlights: • Tryptophan is easily accessible in native BSA compared to BSA Au nanoclusters. • Guanidine HCL denatures native BSA more compared to BSA Au nanoclusters. • High temperature decreases the quantum yield of tryptophan and BSA Au nanocluster. • Emission wavelength of BSA Au nanoclusters remains constant with increasing pH. • BSA Au nanoclusters are robust to the changes in their environments.

  9. Effect of quencher, denaturants, temperature and pH on the fluorescent properties of BSA protected gold nanoclusters

    International Nuclear Information System (INIS)

    Chib, Rahul; Butler, Susan; Raut, Sangram; Shah, Sunil; Borejdo, Julian; Gryczynski, Zygmunt; Gryczynski, Ignacy

    2015-01-01

    In this paper, we have synthesized BSA protected gold nanoclusters (BSA Au nanocluster) and studied the effect of quencher, protein denaturant, pH and temperature on the fluorescence properties of the tryptophan molecule of the BSA Au nanocluster and native BSA. We have also studied their effect on the peak emission of BSA Au nanoclusters (650 nm). The photophysical characterization of a newly developed fluorophore in different environments is absolutely necessary to futher develop their biomedical and analytical applications. It was observed from our experiments that the tryptophan in BSA Au nanoclusters is better shielded from the polar environment. Tryptophan in native BSA showed a red shift in its peak emission wavelength position. Tryptophan is a highly polarity sensitive dye and a minimal change in its microenvironment can be easily observed in its photophysical properties. - Highlights: • Tryptophan is easily accessible in native BSA compared to BSA Au nanoclusters. • Guanidine HCL denatures native BSA more compared to BSA Au nanoclusters. • High temperature decreases the quantum yield of tryptophan and BSA Au nanocluster. • Emission wavelength of BSA Au nanoclusters remains constant with increasing pH. • BSA Au nanoclusters are robust to the changes in their environments.

  10. Clinical utility of (18)F-FDG positron emission tomography/computed tomography scan vs. (99m)Tc-HMPAO white blood cell single-photon emission computed tomography in extra-cardiac work-up of infective endocarditis

    DEFF Research Database (Denmark)

    Lauridsen, Trine K; Iversen, Kasper K; Ihlemann, Nikolaj

    2017-01-01

    The extra-cardiac work-up in infective endocarditis (IE) comprises a search for primary and secondary infective foci. Whether18FDG-PET/CT or WBC-SPECT/CT is superior in detection of clinically relevant extra-cardiac manifestations in IE is unexplored. The objectives of this study were to identify...

  11. Should PET/CT be implemented in the routine imaging work-up of locally advanced head and neck squamous cell carcinoma? A prospective analysis

    Energy Technology Data Exchange (ETDEWEB)

    Cacicedo, Jon; Bilbao, Pedro [Cruces University Hospital, Radiation Oncology Department, Barakaldo, Bizkaia (Basque Country) (Spain); BioCruces Health Research Institute, Bizkaia, Basque Country (Spain); Fernandez, Iratxe [Cruces University Hospital, Nuclear Medicine Department, Barakaldo (Spain); Hoyo, Olga del; Hortelano, Eduardo [Cruces University Hospital, Radiation Oncology Department, Barakaldo, Bizkaia (Basque Country) (Spain); Dolado, Ainara [Cruces University Hospital, Radiodiagnostic and Medical Imaging Department, Barakaldo (Spain); Gomez-Suarez, Javier [Cruces University Hospital, Otolaryngology Department, Barakaldo (Spain); Sancho, Aintzane [Cruces University Hospital, Medical Oncology Department, Barakaldo (Spain); Pijoan, Jose I. [BioCruces Health Research Institute, Bizkaia, Basque Country (Spain); Cruces University Hospital, Clinical Epidemiology Unit, Barakaldo (Spain); CIBER de Epidemiologia y Salud Publica (CIBERESP), Madrid (Spain); Alvarez, Julio [Cruces University Hospital, Oral and Maxillofacial Surgery Department, Barakaldo (Spain); Espinosa, Jose M. [Cruces University Hospital, Medical Physics Department, Barakaldo (Spain); Gaafar, Ayman [Cruces University Hospital, Department of Pathology, Barakaldo (Spain)

    2015-08-15

    The objective of this study was to determine the incremental staging information provided by positron emission tomography/computed tomography (PET/CT) and its impact on management plans in patients with untreated stage III-IV head and neck squamous cell carcinoma (HNSCC). We prospectively studied, between September 2011 and February 2013, 84 consecutive patients [median age 63.5 years (39-84); 73 men] with histologically confirmed HNSCC. First, based on a conventional work-up (physical examination, CT imaging of the head, neck and chest), the multidisciplinary Head and Neck Tumour Board documented the TNM stage and a management plan for each patient, outlining the modalities to be used, including surgery, radiation therapy (RT), chemotherapy or a combination. After release of the PET/CT results, new TNM staging and management plans were agreed on by the multidisciplinary Tumour Board. Any changes in stage or intended management due to the PET/CT findings were then analysed. The impact on patient management was classified as: low (treatment modality, delivery and intent unchanged), moderate (change within the same treatment modality: type of surgery, radiation technique/dose) or high (change in treatment intent and/or treatment modality → curative to palliative, or surgery to chemoradiation or detection of unknown primary tumour or a synchronous second primary tumour). TNM stage was validated by histopathological analysis, additional imaging or follow-up. Accuracy of the conventional and PET/CT-based staging was compared using McNemar's test. Conventional and PET/CT stages were discordant in 32/84 (38 %) cases: the T stage in 2/32 (6.2 %), the N stage in 21/32 (65.7 %) and the M stage 9/32 (28.1 %). Patient management was altered in 22/84 (26 %) patients, with a moderate impact in 8 (9.5 %) patients and high impact in 14 (16.6 %) patients. PET/CT TNM classification was significantly more accurate (92.5 vs 73.7 %) than conventional staging with a p value < 0

  12. Is Whole-Body Computed Tomography the Standard Work-up for Severely-Injured Children? Results of a Survey among German Trauma Centers.

    Science.gov (United States)

    Bayer, J; Reising, K; Kuminack, K; Südkamp, N P; Strohm, P C

    2015-01-01

    Whole-body computed tomography is accepted as the standard procedure in the primary diagnostic of polytraumatised adults in the emergency room. Up to now there is still controversial discussion about the same algorithm in the primary diagnostic of children. The aim of this study was to survey the participation of German trauma-centres in the care of polytraumatised children and the hospital dependant use of whole-body computed tomography for initial patient work-up. A questionnaire was mailed to every Department of Traumatology registered in the DGU (German Trauma Society) databank. We received 60,32% of the initially sent questionnaires and after applying exclusion criteria 269 (53,91%) were applicable to statistical analysis. In the three-tiered German hospital system no statistical difference was seen in the general participation of children polytrauma care between hospitals of different tiers (p = 0.315). Even at the lowest hospital level 69,47% of hospitals stated to participate in polytrauma care for children, at the intermediate and highest level hospitals 91,89% and 95,24% stated to be involved in children polytrauma care, respectively. Children suspicious of multiple injuries or polytrauma received significantly fewer primary whole-body CTs in lowest level compared to intermediate level hospitals (36,07% vs. 56,57%; p = 0.015) and lowest level compared to highest level hospitals (36,07% vs. 68,42%; p = 0.001). Comparing the use of whole-body CT in intermediate to highest level hospitals a not significant increase in its use could be seen in highest level hospitals (56,57% vs. 68,42%; p = 0.174). According to our survey, taking care of polytraumatised children in Germany is not limited to specialised hospitals or a defined hospital level-of-care. Additionally, there is no established radiologic standard in work-up of the polytraumatised child. However, in higher hospital care -levels a higher percentage of hospitals employs whole-body CTs for primary

  13. Should PET/CT be implemented in the routine imaging work-up of locally advanced head and neck squamous cell carcinoma? A prospective analysis

    International Nuclear Information System (INIS)

    Cacicedo, Jon; Bilbao, Pedro; Fernandez, Iratxe; Hoyo, Olga del; Hortelano, Eduardo; Dolado, Ainara; Gomez-Suarez, Javier; Sancho, Aintzane; Pijoan, Jose I.; Alvarez, Julio; Espinosa, Jose M.; Gaafar, Ayman

    2015-01-01

    The objective of this study was to determine the incremental staging information provided by positron emission tomography/computed tomography (PET/CT) and its impact on management plans in patients with untreated stage III-IV head and neck squamous cell carcinoma (HNSCC). We prospectively studied, between September 2011 and February 2013, 84 consecutive patients [median age 63.5 years (39-84); 73 men] with histologically confirmed HNSCC. First, based on a conventional work-up (physical examination, CT imaging of the head, neck and chest), the multidisciplinary Head and Neck Tumour Board documented the TNM stage and a management plan for each patient, outlining the modalities to be used, including surgery, radiation therapy (RT), chemotherapy or a combination. After release of the PET/CT results, new TNM staging and management plans were agreed on by the multidisciplinary Tumour Board. Any changes in stage or intended management due to the PET/CT findings were then analysed. The impact on patient management was classified as: low (treatment modality, delivery and intent unchanged), moderate (change within the same treatment modality: type of surgery, radiation technique/dose) or high (change in treatment intent and/or treatment modality → curative to palliative, or surgery to chemoradiation or detection of unknown primary tumour or a synchronous second primary tumour). TNM stage was validated by histopathological analysis, additional imaging or follow-up. Accuracy of the conventional and PET/CT-based staging was compared using McNemar's test. Conventional and PET/CT stages were discordant in 32/84 (38 %) cases: the T stage in 2/32 (6.2 %), the N stage in 21/32 (65.7 %) and the M stage 9/32 (28.1 %). Patient management was altered in 22/84 (26 %) patients, with a moderate impact in 8 (9.5 %) patients and high impact in 14 (16.6 %) patients. PET/CT TNM classification was significantly more accurate (92.5 vs 73.7 %) than conventional staging with a p value < 0

  14. Time-efficient and convenient synthesis of [18F]altanserin for human PET imaging by a new work-up procedure

    International Nuclear Information System (INIS)

    Massarweh, G.; Kovacevic, M.; Rosa-Neto, P.; Evans, A.C.; Diksic, M.; Schirrmacher, R.

    2009-01-01

    [ 18 F]Altanserin, an important PET radioligand for the in vivo imaging of the 5-HT 2A receptor, was synthesized from its precursor nitro-altanserin in DMF or DMSO at high temperatures of 150 deg. C in an overall radiochemical yield (EOB) of 23-25% after 75 min. A new solid phase work-up procedure involving the acidification of the crude reaction mixture and a C18-SepPak-solid phase separation preceded the final HPLC purification. This led to a significantly reduced synthesis time as a result of a stable and early elution from the HPLC column using improved HPLC conditions (MeOH/THF/NaOAc 0.05 N pH 5: 27/18/55, flow: 5 mL/min, Symetry Prep 7 μm C18 (Waters)). The synthesis was performed semi-automatically in a modified GE TracerLab synthesis module using an in-house-developed program. The synthesized [ 18 F]altanserin was used in our ongoing human and animal PET imaging studies.

  15. Time-efficient and convenient synthesis of [{sup 18}F]altanserin for human PET imaging by a new work-up procedure

    Energy Technology Data Exchange (ETDEWEB)

    Massarweh, G. [McConnell Brain Imaging Centre, Montreal Neurological Institute, McGill University, 3801 University Street, Montreal, Quebec (Canada)], E-mail: gassan.wassarweh@mcgill.ca; Kovacevic, M.; Rosa-Neto, P.; Evans, A.C.; Diksic, M. [McConnell Brain Imaging Centre, Montreal Neurological Institute, McGill University, 3801 University Street, Montreal, Quebec (Canada); Schirrmacher, R. [McConnell Brain Imaging Centre, Montreal Neurological Institute, McGill University, 3801 University Street, Montreal, Quebec (Canada)], E-mail: ralf.schirrmacher@mcgill.ca

    2009-11-15

    [{sup 18}F]Altanserin, an important PET radioligand for the in vivo imaging of the 5-HT{sub 2A} receptor, was synthesized from its precursor nitro-altanserin in DMF or DMSO at high temperatures of 150 deg. C in an overall radiochemical yield (EOB) of 23-25% after 75 min. A new solid phase work-up procedure involving the acidification of the crude reaction mixture and a C18-SepPak-solid phase separation preceded the final HPLC purification. This led to a significantly reduced synthesis time as a result of a stable and early elution from the HPLC column using improved HPLC conditions (MeOH/THF/NaOAc 0.05 N pH 5: 27/18/55, flow: 5 mL/min, Symetry Prep 7 {mu}m C18 (Waters)). The synthesis was performed semi-automatically in a modified GE TracerLab synthesis module using an in-house-developed program. The synthesized [{sup 18}F]altanserin was used in our ongoing human and animal PET imaging studies.

  16. Can the Diagnostics of Triangular Fibrocartilage Complex Lesions Be Improved by MRI-Based Soft-Tissue Reconstruction? An Imaging-Based Workup and Case Presentation.

    Science.gov (United States)

    Hammer, Niels; Hirschfeld, Ulrich; Strunz, Hendrik; Werner, Michael; Wolfskämpf, Thomas; Löffler, Sabine

    2017-01-01

    Introduction . The triangular fibrocartilage complex (TFCC) provides both mobility and stability of the radiocarpal joint. TFCC lesions are difficult to diagnose due to the complex anatomy. The standard treatment for TFCC lesions is arthroscopy, posing surgery-related risks onto the patients. This feasibility study aimed at developing a workup for soft-tissue reconstruction using clinical imaging, to verify these results in retrospective patient data. Methods . Microcomputed tomography ( μ -CT), 3 T magnetic resonance imaging (MRI), and plastination were used to visualize the TFCC in cadaveric specimens applying segmentation-based 3D reconstruction. This approach further trialed the MRI dataset of a patient with minor radiological TFCC alterations but persistent pain. Results . TFCC reconstruction was impossible using μ -CT only but feasible using MRI, resulting in an appreciation of its substructures, as seen in the plastinates. Applying this approach allowed for visualizing a Palmer 2C lesion in a patient, confirming ex postum the arthroscopy findings, being markedly different from MRI (Palmer 1B). Discussion . This preliminary study showed that image-based TFCC reconstruction may help to identify pathologies invisible in standard MRI. The combined approach of μ -CT, MRI, and plastination allowed for a three-dimensional appreciation of the TFCC. Image quality and time expenditure limit the approach's usefulness as a diagnostic tool.

  17. Can the Diagnostics of Triangular Fibrocartilage Complex Lesions Be Improved by MRI-Based Soft-Tissue Reconstruction? An Imaging-Based Workup and Case Presentation

    Directory of Open Access Journals (Sweden)

    Niels Hammer

    2017-01-01

    Full Text Available Introduction. The triangular fibrocartilage complex (TFCC provides both mobility and stability of the radiocarpal joint. TFCC lesions are difficult to diagnose due to the complex anatomy. The standard treatment for TFCC lesions is arthroscopy, posing surgery-related risks onto the patients. This feasibility study aimed at developing a workup for soft-tissue reconstruction using clinical imaging, to verify these results in retrospective patient data. Methods. Microcomputed tomography (μ-CT, 3 T magnetic resonance imaging (MRI, and plastination were used to visualize the TFCC in cadaveric specimens applying segmentation-based 3D reconstruction. This approach further trialed the MRI dataset of a patient with minor radiological TFCC alterations but persistent pain. Results. TFCC reconstruction was impossible using μ-CT only but feasible using MRI, resulting in an appreciation of its substructures, as seen in the plastinates. Applying this approach allowed for visualizing a Palmer 2C lesion in a patient, confirming ex postum the arthroscopy findings, being markedly different from MRI (Palmer 1B. Discussion. This preliminary study showed that image-based TFCC reconstruction may help to identify pathologies invisible in standard MRI. The combined approach of μ-CT, MRI, and plastination allowed for a three-dimensional appreciation of the TFCC. Image quality and time expenditure limit the approach’s usefulness as a diagnostic tool.

  18. High resolution MRI for preoperative work-up of neonates with an anorectal malformation: a direct comparison with distal pressure colostography/fistulography

    Energy Technology Data Exchange (ETDEWEB)

    Thomeer, Maarten G. [Erasmus MC, Department of Radiology, Rotterdam (Netherlands); Devos, Annick; Lequin, Maarten; Graaf, Nanko de; Meradji, Morteza [Erasmus MC, Department of Pediatric Radiology, Rotterdam (Netherlands); Meeussen, Conny J.H.M.; Blaauw, Ivo de; Sloots, Cornelius E.J. [Erasmus MC, Department of Pediatric Surgery, Rotterdam (Netherlands)

    2015-12-15

    To compare MRI and colostography/fistulography in neonates with anorectal malformations (ARM), using surgery as reference standard. Thirty-three neonates (22 boys) with ARM were included. All patients underwent both preoperative high-resolution MRI (without sedation or contrast instillation) and colostography/fistulography. The Krickenbeck classification was used to classify anorectal malformations, and the level of the rectal ending in relation to the levator muscle was evaluated. Subjects included nine patients with a bulbar recto-urethral fistula, six with a prostatic recto-urethral fistula, five with a vestibular fistula, five with a cloacal malformation, four without fistula, one with a H-type fistula, one with anal stenosis, one with a rectoperineal fistula and one with a bladderneck fistula. MRI and colostography/fistulography predicted anatomy in 88 % (29/33) and 61 % (20/33) of cases, respectively (p = 0.012). The distal end of the rectal pouch was correctly predicted in 88 % (29/33) and 67 % (22/33) of cases, respectively (p = 0.065). The length of the common channel in cloacal malformation was predicted with MRI in all (100 %, 5/5) and in 80 % of cases (4/5) with colostography/fistulography. Two bowel perforations occurred during colostography/fistulography. MRI provides the most accurate evaluation of ARM and should be considered a serious alternative to colostography/fistulography during preoperative work-up. (orig.)

  19. Splenic scintigraphy using Tc-99m-labeled heat-denatured red blood cells in pediatric patients: concise communication

    International Nuclear Information System (INIS)

    Ehrlich, C.P.; Papanicolaou, N.; Treves, S.; Hurwitz, R.A.; Richards, P.

    1982-01-01

    Ten children underwent splenic imaging with heat-denatured red blood cells labeled with technetium-99m (Tc-99m DRBC). The presenting problems included the heterotaxia syndrome, recurrent idiopathic thrombocytopenic purpura following splenectomy, mass in the left posterior hemithorax, and blunt abdominal trauma. In nine patients, the presence or absence of splenic tissue was established. A splenic hematoma was identified in the tenth patient. All patients were initially scanned with Tc-99m sulfur colloid (Tc-99m SC), and were selected for Tc-99m DRBC scintigraphy only after the results of the SC scans failed to establish the clinical problem beyond doubt. The availability of kits containing stannous ions, essential for efficient and stable labeling of red blood cells with Tc-99m and requiring only a small volume of blood, make splenic scintigraphy in children a relatively simple and definitive diagnostic procedure, when identification of splenic tissue is of clinical importance

  20. Splenic scintigraphy using Tc-99m-labeled heat-denatured red blood cells in pediatric patients: concise communication

    Energy Technology Data Exchange (ETDEWEB)

    Ehrlich, C.P.; Papanicolaou, N.; Treves, S.; Hurwitz, R.A.; Richards, P.

    1982-03-01

    Ten children underwent splenic imaging with heat-denatured red blood cells labeled with technetium-99m (Tc-99m DRBC). The presenting problems included the heterotaxia syndrome, recurrent idiopathic thrombocytopenic purpura following splenectomy, mass in the left posterior hemithorax, and blunt abdominal trauma. In nine patients, the presence or absence of splenic tissue was established. A splenic hematoma was identified in the tenth patient. All patients were initially scanned with Tc-99m sulfur colloid (Tc-99m SC), and were selected for Tc-99m DRBC scintigraphy only after the results of the SC scans failed to establish the clinical problem beyond doubt. The availability of kits containing stannous ions, essential for efficient and stable labeling of red blood cells with Tc-99m and requiring only a small volume of blood, make splenic scintigraphy in children a relatively simple and definitive diagnostic procedure, when identification of splenic tissue is of clinical importance.

  1. Antibodies to uv light denatured DNA in systemic lupus erythematosus: detection by filter radioimmunoassay and clinical correlations

    Energy Technology Data Exchange (ETDEWEB)

    Davis, P; Russell, A S; Percy, J S

    1976-12-01

    Antibodies to ultraviolet light denatured DNA (UV DNA) have been measured in patients with systemic lupus erythematosus (SLE) and normal subjects, using a millipore filter radioimmunoassay. High levels of UV DNA binding were only found in patients with SLE. The presence of UV DNA antibodies correlated well with the presence of native DNA antibodies, although immunodiffusion studies and inhibition techniques showed these antibodies to be immunologically distinct in many cases. Forty-one percent of the SLE patients had had photosensitivity at some stage of their disease, but there was a poor correlation between this symptom and the presence of UV DNA antibodies. Although UV DNA is known to be a potent immunogen, none of the results from this study suggests that antibodies to UV DNA are more than another example of the broad spectrum of antinuclear antibodies seen in SLE.

  2. The detection of Mycoplasma (formerly Eperythrozoon) wenyonii by 16S rDNA PCR and denaturing gradient gel electrophoresis.

    Science.gov (United States)

    McAuliffe, Laura; Lawes, Joanna; Bell, Suzanna; Barlow, Alex; Ayling, Roger; Nicholas, Robin

    2006-10-31

    Although the role of Mycoplasma wenyonii in disease is still subject to some debate, infections have been reported to result in parasitaemia, anaemia, scrotal and hind limb oedema, tachycardia, pyrexia, infertility, swollen teats, prefemoral lymphadenopathy and decreased milk production. Previously, diagnosis of M. wenyonii has been based on blood smears but is not specific for M. wenyonii and can be difficult to interpret. We have previously described the use of PCR and denaturing gradient gel electrophoresis (DGGE) for the detection and differentiation of Mycoplasma species. DGGE enables the rapid and specific identification of Mycoplasma species and is ideally suited to detecting both mixed infections and new and unusual species. In this study, we have used DGGE with universal primers to detect M. wenyonii DNA from blood samples. DGGE can be used on blood samples as a rapid and specific test for M. wenyonii and can also be used as a screening test for other blood borne pathogens.

  3. The effects of urea and n-propanol on collagen denaturation: using DSC, circular dicroism and viscosity

    International Nuclear Information System (INIS)

    Usha, R.; Ramasami, T.

    2004-01-01

    The effect of urea and n-propanol on circular dichroism (CD) and viscosity of purified type1 collagen solution at various temperatures and differential scanning calorimetry (DSC) of rat-tail tendon (RTT) collagen fibre have been studied. CD reveals a spectrum with a positive peak at around 220 nm and a negative peak at 200 nm characteristics of collagen triple helix. The molar ellipticity decreases as the concentration of urea increases up to particular concentration (collagen solution treated with 265 μM of urea) and after that it increases (collagen solution treated with 500 μM of urea). There is a linear decrease in molar ellipticity as the concentration of n-propanol increases. Denaturation temperature of urea and n-propanol treated with purified collagen solution has been studied using viscosity method. Additives such as urea and n-propanol decrease the thermal stability of collagen triple helix in solution and in RTT collagen fibre. Thermal helix to coil transition of urea and n-propanol treated collagen depends on the degree of hydration and the concentration of these additives. Thermodynamic parameters such as the peak temperature, enthalpy of activation, and energy of activation for collagen-gelatin transition for native, urea and n-propanol treated RTT collagen fibre has been calculated using DSC. The change in the thermodynamic parameters has been observed for native, urea and n-propanol treated RTT collagen fibres. The experimental results show that the change in the water structure, dehydration and desolvation induced by different additives such as urea and n-propanol on RTT may vary with the type of denaturation

  4. Effects of sterilization, packaging, and storage on vitamin C degradation, protein denaturation, and glycation in fortified milks.

    Science.gov (United States)

    Gliguem, H; Birlouez-Aragon, I

    2005-03-01

    Monitoring the nutritional quality of dietetic milk throughout its shelf life is particularly important due to the high susceptibility of some vitamins to oxidation, and the continuous development of the Maillard reaction during storage. The objective of this paper was to evaluate the vitamin C content and protein modification by denaturation and glycation on fortified milk samples (growth milks) destined for 1- to 3-yr-old children. The influences of the sterilization process, formulation, packaging, and storage duration at ambient temperature in the dark were studied. Vitamin C degradation was particularly influenced by type of packaging. The use of a 3-layered opaque bottle was associated with complete oxidation of vitamin C after 1 mo of storage, whereas in the 6-layered opaque bottle, which has an oxygen barrier, the vitamin C content slowly decreased to reach 25% of the initial concentration after 4 mo of storage. However, no significant effect of vitamin C degradation during storage could be observed in terms of Maillard reaction, despite the fact that a probable impact occurred during sterilization. Furosine content and the FAST (fluorescence of advanced Maillard products and soluble tryptophan) index-indicators of the early and advanced Maillard reaction, respectively-were significantly higher in the in-bottle sterilized milk samples compared with UHT samples, and in fortified milk samples compared with cow milk. However, after 1 mo, the impact of storage was predominant, increasing the furosine level and the FAST index at similar levels for the differently processed samples. The early Maillard reaction developed continuously throughout the storage period.In conclusion, only packaging comprising an oxygen and light barrier is compatible with vitamin C fortification of milk. Furthermore, short storage time or low storage temperature is needed to retard vitamin C degradation, protein denaturation, and development of the Maillard reaction.

  5. Application of denaturing high-performance liquid chromatography for monitoring sulfate-reducing bacteria in oil fields.

    Science.gov (United States)

    Priha, Outi; Nyyssönen, Mari; Bomberg, Malin; Laitila, Arja; Simell, Jaakko; Kapanen, Anu; Juvonen, Riikka

    2013-09-01

    Sulfate-reducing bacteria (SRB) participate in microbially induced corrosion (MIC) of equipment and H2S-driven reservoir souring in oil field sites. Successful management of industrial processes requires methods that allow robust monitoring of microbial communities. This study investigated the applicability of denaturing high-performance liquid chromatography (DHPLC) targeting the dissimilatory sulfite reductase ß-subunit (dsrB) gene for monitoring SRB communities in oil field samples from the North Sea, the United States, and Brazil. Fifteen of the 28 screened samples gave a positive result in real-time PCR assays, containing 9 × 10(1) to 6 × 10(5) dsrB gene copies ml(-1). DHPLC and denaturing gradient gel electrophoresis (DGGE) community profiles of the PCR-positive samples shared an overall similarity; both methods revealed the same samples to have the lowest and highest diversity. The SRB communities were diverse, and different dsrB compositions were detected at different geographical locations. The identified dsrB gene sequences belonged to several phylogenetic groups, such as Desulfovibrio, Desulfococcus, Desulfomicrobium, Desulfobulbus, Desulfotignum, Desulfonatronovibrio, and Desulfonauticus. DHPLC showed an advantage over DGGE in that the community profiles were very reproducible from run to run, and the resolved gene fragments could be collected using an automated fraction collector and sequenced without a further purification step. DGGE, on the other hand, included casting of gradient gels, and several rounds of rerunning, excising, and reamplification of bands were needed for successful sequencing. In summary, DHPLC proved to be a suitable tool for routine monitoring of the diversity of SRB communities in oil field samples.

  6. Changes in the diversity of pig ileal lactobacilli around weaning determined by means of 16S rRNA gene amplification and denaturing gradient gel electrophoresis

    NARCIS (Netherlands)

    Janzcyk, P.; Pieper, R.; Smidt, H.; Souffrant, W.B.

    2007-01-01

    Our study aimed to provide a comprehensive characterization of changes in porcine intestinal Lactobacillus populations around the time of weaning based on 16S rRNA gene amplification and denaturing gradient gel electrophoresis (DGGE). DNA was extracted from the ileal contents of piglets at weaning

  7. Evaluation of functional nerve recovery after reconstruction with a poly (DL-lactide-epsilon-caprolactone) nerve guide, filled with modified denatured muscle tissue

    NARCIS (Netherlands)

    Meek, MF; Den Dunnen, WFA; Schakenraad, JM; Robinson, PH

    1996-01-01

    The aim of this study was to compare the speed of functional nerve recovery after reconstruction with a biodegradable p(DLLA-epsilon -CL) nerve guide, as filled with either modified denatured muscle tissue (MDMT) or phosphate-buffered saline (PBS). To evaluate both motor and sensory nerve recovery,

  8. Development and application of a selective pcr-denaturing gradient gel electrophoresis approach to detect a recently cultivated Bacillus group predominant in soil

    NARCIS (Netherlands)

    Tzeneva, V.A.; Li, Y.; Felske, A.; Vos, de W.M.; Akkermans, A.D.L.; Vaughan, E.E.; Smidt, H.

    2004-01-01

    The worldwide presence of a hitherto-nondescribed group of predominant soil microorganisms related to Bacillus benzoevorans was analyzed after development of two sets of selective primers targeting 16S rRNA genes in combination with denaturing gradient gel electrophoresis (DGGE). The high abundance

  9. Lysis solution composition and non-linear dose-response to ionizing radiation in the non-denaturing DNA filter elution technique

    International Nuclear Information System (INIS)

    Radford, I.R.

    1990-01-01

    The suggestion by Okayasu and Iliakis (1989) that the non-linear dose-response curve, obtained with the non-denaturing filter elution technique for mammalian cells exposed to low-LET radiation, is the result of a technical artefact, was not confirmed. (author)

  10. The recognition of adsorbed and denatured proteins of different topographies by β2 integrins and effects on leukocyte adhesion and activation

    DEFF Research Database (Denmark)

    Brevig, T.; Holst, B.; Ademovic, Z.

    2005-01-01

    Leukocyte beta(2) integrins Mac-1 and p150,95 are promiscuous cell-surface receptors that recognise and mediate cell adhesion to a variety of adsorbed and denatured proteins. We used albumin as a model protein to study whether leukocyte adhesion and activation depended on the nm-scale topography...

  11. Electronmicroscopical evaluation of short-term nerve regeneration through a thin-walled biodegradable poly(DLLA-epsilon-CL) nerve guide filled with modified denatured muscle tissue

    NARCIS (Netherlands)

    Meek, MF; Robinson, PH; Stokroos, [No Value; Blaauw, EH; Kors, G; den Dunnen, WFA

    The aim of this study was to evaluate short-term peripheral nerve regeneration across a 15-mm gap in the sciatic nerve of the rat, using a thin-walled biodegradable poly(DL-lactide-epsilon -caprolactone) nerve guide filled with modified denatured muscle tissue (MDMT). The evaluation was performed

  12. With the Advent of Tomosynthesis in the Workup of Mammographic Abnormality, is Spot Compression Mammography Now Obsolete? An Initial Clinical Experience.

    Science.gov (United States)

    Ni Mhuircheartaigh, Neasa; Coffey, Louise; Fleming, Hannah; O' Doherty, Ann; McNally, Sorcha

    2017-09-01

    To determine if the routine use of spot compression mammography is now obsolete in the assessment of screen detected masses, asymmetries and architectural distortion since the availability of digital breast tomosynthesis. We introduced breast tomosynthesis in the workup of screen detected abnormalities in our screening center in January 2015. During an initial learning period with tomosynthesis standard spot compression views were also performed. Three consultant breast radiologists retrospectively reviewed all screening mammograms recalled for assessment over the first 6-month period. We assessed retrospectively whether there was any additional diagnostic information obtained from spot compression views not already apparent on tomography. All cases were also reviewed for any additional lesions detected by tomosynthesis, not detected on routine 2-view screening mammography. 548 women screened with standard 2-view digital screening mammography were recalled for assessment in the selected period and a total of 565 lesions were assessed. 341 lesions were assessed by both tomosynthesis and routine spot compression mammography. The spot compression view was considered more helpful than tomosynthesis in only one patient. This was because the breast was inadequately positioned for tomosynthesis and the area in question was not adequately imaged. Apart from this technical error there was no asymmetry, distortion or mass where spot compression provided more diagnostic information than tomosynthesis alone. We detected three additional cancers on tomosynthesis, not detected by routine screening mammography. From our initial experience with tomosynthesis we conclude that spot compression mammography is now obsolete in the assessment of screen detected masses, asymmetries and distortions where tomosynthesis is available. © 2017 Wiley Periodicals, Inc.

  13. Mandatory imaging cuts costs and reduces the rate of unnecessary surgeries in the diagnostic work-up of patients suspected of having appendicitis

    Energy Technology Data Exchange (ETDEWEB)

    Lahaye, M.J.; Lambregts, D.M.J.; Mutsaers, E.; Beets-Tan, R.G.H. [Maastricht University Medical Centre, Department of Radiology, Maastricht (Netherlands); Essers, B.A.B. [Maastricht University Medical Centre, Department of Epidemiology and Medical Technology, Maastricht (Netherlands); Breukink, S.; Beets, G.L. [Maastricht University Medical Centre, Department of Surgery, Maastricht (Netherlands); Cappendijk, V.C. [Jeroen Bosch Hospital, Department of Radiology, ' s Hertogenbosch (Netherlands)

    2015-05-01

    To evaluate whether mandatory imaging is an effective strategy in suspected appendicitis for reducing unnecessary surgery and costs. In 2010, guidelines were implemented in The Netherlands recommending the mandatory use of preoperative imaging to confirm/refute clinically suspected appendicitis. This retrospective study included 1,556 consecutive patients with clinically suspected appendicitis in 2008-2009 (756 patients/group I) and 2011-2012 (800 patients/group II). Imaging use (none/US/CT and/or MRI) was recorded. Additional parameters were: complications, medical costs, surgical and histopathological findings. The primary study endpoint was the number of unnecessary surgeries before and after guideline implementation. After clinical examination by a surgeon, 509/756 patients in group I and 540/800 patients in group II were still suspected of having appendicitis. In group I, 58.5% received preoperative imaging (42% US/12.8% CT/3.7% both), compared with 98.7% after the guidelines (61.6% US/4.4% CT/ 32.6% both). The percentage of unnecessary surgeries before the guidelines was 22.9%. After implementation, it dropped significantly to 6.2% (p<0.001). The surgical complication rate dropped from 19.9% to 14.2%. The average cost-per-patient decreased by 594 EUR from 2,482 to 1,888 EUR (CL:-1081; -143). Increased use of imaging in the diagnostic work-up of patients with clinically suspected appendicitis reduced the rate of negative appendectomies, surgical complications and costs. (orig.)

  14. Raman spectral markers of collagen denaturation and hydration in human cortical bone tissue are affected by radiation sterilization and high cycle fatigue damage.

    Science.gov (United States)

    Flanagan, Christopher D; Unal, Mustafa; Akkus, Ozan; Rimnac, Clare M

    2017-11-01

    Thermal denaturation and monotonic mechanical damage alter the organic and water-related compartments of cortical bone. These changes can be detected using Raman spectroscopy. However, less is known regarding Raman sensitivity to detect the effects of cyclic fatigue damage and allograft sterilization doses of gamma radiation. To determine if Raman spectroscopic biomarkers of collagen denaturation and hydration are sensitive to the effects of (a) high cycle fatigue damage and (b) 25kGy irradiation. Unirradiated and gamma-radiation sterilized human cortical bone specimens previously tested in vitro under high-cycle (> 100,000 cycles) fatigue conditions at 15MPa, 25MPa, 35MPa, 45MPa, and 55MPa cyclic stress levels were studied. Cortical bone Raman spectral profiles from wavenumber ranges of 800-1750cm -1 and 2700-3800cm -1 were obtained and compared from: a) non-fatigue vs fatigue fracture sites and b) radiated vs. unirradiated states. Raman biomarker ratios 1670/1640 and 3220/2949, which reflect collagen denaturation and organic matrix (mainly collagen)-bound water, respectively, were assessed. One- and two-way ANOVA analyses were utilized to identify differences between groups along with interaction effects between cyclic fatigue and radiation-induced damage. Cyclic fatigue damage resulted in increases in collagen denaturation (1670/1640: 1.517 ± 0.043 vs 1.579 ± 0.021, p Raman spectroscopy can detect the effects of cyclic fatigue damage and 25kGy irradiation via increases in organic matrix (mainly collagen)-bound water. A Raman measure of collagen denaturation was sensitive to cyclic fatigue damage but not 25kGy irradiation. Collagen denaturation was correlated with organic matrix-bound water, suggesting that denaturation of collagen to gelatinous form may expose more binding sites to water by unwinding the triple alpha chains. This research may eventually be useful to help identify allograft quality and more appropriately match donors to recipients. Copyright

  15. Whole-body-MR imaging including DWIBS in the work-up of patients with head and neck squamous cell carcinoma: A feasibility study

    Energy Technology Data Exchange (ETDEWEB)

    Noij, Daniel P., E-mail: d.noij@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Boerhout, Els J., E-mail: e.boerhout@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Pieters-van den Bos, Indra C., E-mail: i.pieters@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Comans, Emile F., E-mail: efi.comans@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Oprea-Lager, Daniela, E-mail: d.oprea-lager@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Reinhard, Rinze, E-mail: r.reinhard@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Hoekstra, Otto S., E-mail: os.hoekstra@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Bree, Remco de, E-mail: r.debree@vumc.nl [Department Otolaryngology/Head and Neck Surgery, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Graaf, Pim de, E-mail: p.degraaf@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands); Castelijns, Jonas A., E-mail: j.castelijns@vumc.nl [Department of Radiology and Nuclear Medicine, VU University Medical Center, De Boelelaan 1117, PO Box 7057, 1007 MB Amsterdam (Netherlands)

    2014-07-15

    Objectives: To assess the feasibility of whole-body magnetic resonance imaging (WB-MRI) including diffusion-weighted whole-body imaging with background-body-signal-suppression (DWIBS) for the evaluation of distant malignancies in head and neck squamous cell carcinoma (HNSCC); and to compare WB-MRI findings with {sup 18}F-fluorodeoxyglucose positron emission tomography/computed tomography ({sup 18}F-FDG-PET/CT) and chest-CT. Methods: Thirty-three patients with high risk for metastatic spread (26 males; range 48–79 years, mean age 63 ± 7.9 years (mean ± standard deviation) years) were prospectively included with a follow-up of six months. WB-MRI protocol included short-TI inversion recovery and T1-weighted sequences in the coronal plane and half-fourier acquisition single-shot turbo spin-echo T2 and contrast-enhanced-T1-weighted sequences in the axial plane. Axial DWIBS was reformatted in the coronal plane. Interobserver variability was assessed using weighted kappa and the proportion specific agreement (PA). Results: Two second primary tumors and one metastasis were detected on WB-MRI. WB-MRI yielded seven clinically indeterminate lesions which did not progress at follow-up. The metastasis and one second primary tumor were found when combining {sup 18}F-FDG-PET/CT and chest-CT findings. Interobserver variability for WB-MRI was κ = 0.91 with PA ranging from 0.82 to 1.00. For {sup 18}F-FDG-PET/CT κ could not be calculated due to a constant variable in the table and PA ranged from 0.40 to 0.99. Conclusions: Our WB-MRI protocol with DWIBS is feasible in the work-up of HNSCC patients for detection and characterization of distant pathology. WB-MRI can be complementary to {sup 18}F-FDG-PET/CT, especially in the detection of non {sup 18}F-FDG avid second primary tumors.

  16. Dynamic and structural study of neocarzinostatin native and denatured states, by differential microcalorimetry, optical spectroscopies and X-ray and neutron scattering

    International Nuclear Information System (INIS)

    Russo, Daniela

    2000-01-01

    A structural and dynamic characterization of proteins denatured states is essential to the understanding of mechanisms which control proteins folding. It is in this framework that this study has been undertaken in taking as model the neocarzinostatin globular protein. It is formed with seven cell-layers which form a barrel pattern maintained by two bi-sulfur bonds. A full characterization of native and denatured states, both from structural and dynamic point of view, has been implemented with several techniques able to bring data at different levels. During the experiments, ncs has been stabilized by temperature and by the use of a chaotropic agent: the guanidinium chloride (gdmcl). Small angle x-ray and neutron scattering have allowed us to obtain data on the variation of the protein compactness in terms of gdmcl temperature and concentration. The diffusion spectra show that ncs loses completely its globular structure above 80 C or in presence of about 5 m of gdmcl. Temperature and concentration of half denaturation are tm= 70 C and cm=3.5 m (in heavy water), respectively. Spectra analysis of strongly denatured protein has allowed us to obtain values of its chain length and of its persistence length which are in agreement with those theoretically estimated. Experiments have been carried out too to measure the radius of gyration to zero concentration and the second virial coefficient of the solution in order to estimate the interactions between the molecules. A full characterization has been performed in terms of gdmcl temperature and concentration by fluorescence and circular dichroism. These two techniques reveal the variations of the local three-dimensional structure and secondary structure of the protein respectively. Microcalorimetry measurements have shown that thermal denaturation of ncs is completely reversible and has been used to measure the enthalpy variation during the transition. At last, it has been possible to study ncs intramolecular dynamics in

  17. Variations among Japanese of the factor IX gene (F9) detected by PCR-denaturing gradient gel electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Satoh, Chiyoko; Takahashi, Norio; Asakawa, Junichi; Hiyama, Keiko; Kodaira, Meiko (Radiation Effects Research Foundation, Hiroshima (Japan))

    1993-01-01

    In the course of feasibility studies to examine the efficiencies and practicalities of various techniques for screening for genetic variations, the human coagulation factor IX (F9) genes of 63 Japanese families were examined by PCR-denaturing gradient gel electrophoresis (PCR-DGGE). Four target sequences with lengths of 983-2,891 bp from the F9 genes of 126 unrelated individuals from Hiroshima and their 100 children were amplified by PCR, digested with restriction enzymes to approximately 500-bp fragments, and examined by DGGE - a total of 6,724 bp being examined per individual. GC-rich sequences (GC-clamps) of 40 bp were attached to both ends of the target sequences, as far as was feasible. Eleven types of new nucleotide substitutions were detected in the population, none of which produced RFLPs or caused hemophilia B. By examining two target sequences in a single lane, approximately 8,000 bp in a diploid individual could be examined. This approach is very effective for the detection of variations in DNA and is applicable to large-scale population studies. 46 refs., 3 figs., 1 tab.

  18. Denaturing gradient gel electrophoresis and barcoded pyrosequencing reveal unprecedented archaeal diversity in mangrove sediment and rhizosphere samples.

    Science.gov (United States)

    Pires, Ana C C; Cleary, Daniel F R; Almeida, Adelaide; Cunha, Angela; Dealtry, Simone; Mendonça-Hagler, Leda C S; Smalla, Kornelia; Gomes, Newton C M

    2012-08-01

    Mangroves are complex ecosystems that regulate nutrient and sediment fluxes to the open sea. The importance of bacteria and fungi in regulating nutrient cycles has led to an interest in their diversity and composition in mangroves. However, very few studies have assessed Archaea in mangroves, and virtually nothing is known about whether mangrove rhizospheres affect archaeal diversity and composition. Here, we studied the diversity and composition of Archaea in mangrove bulk sediment and the rhizospheres of two mangrove trees, Rhizophora mangle and Laguncularia racemosa, using denaturing gradient gel electrophoresis (DGGE) and pyrosequencing of archaeal 16S rRNA genes with a nested-amplification approach. DGGE profiles revealed significant structural differences between bulk sediment and rhizosphere samples, suggesting that roots of both mangrove species influence the sediment archaeal community. Nearly all of the detected sequences obtained with pyrosequencing were identified as Archaea, but most were unclassified at the level of phylum or below. Archaeal richness was, furthermore, the highest in the L. racemosa rhizosphere, intermediate in bulk sediment, and the lowest in the R. mangle rhizosphere. This study shows that rhizosphere microhabitats of R. mangle and L. racemosa, common plants in subtropical mangroves located in Rio de Janeiro, Brazil, hosted distinct archaeal assemblages.

  19. Robust Denaturation of Villin Headpiece by MoS2 Nanosheet: Potential Molecular Origin of the Nanotoxicity

    Science.gov (United States)

    Gu, Zonglin; Yang, Zaixing; Kang, Seung-Gu; Yang, Jerry R.; Luo, Judong; Zhou, Ruhong

    2016-06-01

    MoS2 nanosheet, a new two-dimensional transition metal dichalcogenides nanomaterial, has attracted significant attentions lately due to many potential promising biomedical applications. Meanwhile, there is also a growing concern on its biocompatibility, with little known on its interactions with various biomolecules such as proteins. In this study, we use all-atom molecular dynamics simulations to investigate the interaction of a MoS2 nanosheet with Villin Headpiece (HP35), a model protein widely used in protein folding studies. We find that MoS2 exhibits robust denaturing capability to HP35, with its secondary structures severely destroyed within hundreds of nanosecond simulations. Both aromatic and basic residues are critical for the protein anchoring onto MoS2 surface, which then triggers the successive protein unfolding process. The main driving force behind the adsorption process is the dispersion interaction between protein and MoS2 monolayer. Moreover, water molecules at the interface between some key hydrophobic residues (e.g. Trp-64) and MoS2 surface also help to accelerate the process driven by nanoscale drying, which provides a strong hydrophobic force. These findings might have shed new light on the potential nanotoxicity of MoS2 to proteins with atomic details, which should be helpful in guiding future biomedical applications of MoS2 with its nanotoxicity mitigated.

  20. Bacteria community study of combined periodontal-endodontic lesions using denaturing gradient gel electrophoresis and sequencing analysis.

    Science.gov (United States)

    Li, Hong; Guan, Rui; Sun, Jinghua; Hou, Benxiang

    2014-10-01

    The entire microbial population and predominant microflora of root canals (RCs) and adjacent periodontal pockets (PPs) from teeth with combined periodontal-endodontic lesions were determined and compared. Pooled RC and PP samples were collected from the molars of 20 patients diagnosed with combined periodontal-endodontic lesions. DNA was extracted for polymerase chain reaction-based denaturing gradient gel electrophoresis (PCR-DGGE), cloning, and sequence analysis. A coefficient of similarity (Cs) was used to determine the similarity of the bacterial profiles from RCs and PPs. Significantly fewer bands were produced by PCR-DGGE from RCs (5.9 ± 1.7) than from PPs (8.0 ± 1.8) (P bacteria in both the RC and PP samples were (in descending order) Filifactor alocis, Parvimonas micra, Porphyromonas gingivalis, and Tannerella forsythia. The high similarity in the sets of organisms present in both RC and PP samples in this study suggests that the pocket could be a source of RC infection. The data also demonstrate that combined periodontal-endodontic lesions consist of a diverse and complex microbial community.

  1. Bacterial analysis of combined periodontal-endodontic lesions by polymerase chain reaction-denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Xia, Minghui; Qi, Qingguo

    2013-01-01

    We used denaturing gradient gel electrophoresis (DGGE) to compare bacterial profiles in periodontium and root canals of teeth with combined periodontal-endodontic lesions. Samples of dental plaque and necrotic pulp were collected from thirteen extracted teeth with advanced periodontitis. Genomic DNA was extracted for polymerase chain reaction (PCR) analysis using universal bacterial primers. The PCR products were then loaded onto DGGE gels to obtain fractionated bands. Characteristic DGGE bands were excised and DNA was cloned and sequenced. The number of bands, which indicates the number of bacterial species, was compared between dental plaques and necrotic pulp tissues from the same tooth. Although the difference was statistically significant (P bacteria species were present in both the periodontal pockets and root canals of the same tooth; however, periodontal bacteria did not always invade the root canals, and some bacteria in root canals were not present in periodontal pockets of the same tooth. In some teeth, unique bacteria in root canals had not passed from periodontal pockets. A basic local alignment search tool (BLAST) sequence search in Genbank indicated that new bacteria species were present in periodontal pockets and root canals. Their characteristics must thus be further analyzed.

  2. 16S rRNA PCR-Denaturing Gradient Gel Electrophoresis of Oral Lactobacillus casei Group and Their Phenotypic Appearances.

    Science.gov (United States)

    Piwat, S; Teanpaisan, R

    2013-01-01

    This study aimed to develop a 16S rRNA PCR-denaturing gradient gel electrophoresis (DGGE) to identify the species level of Lactobacillus casei group and to investigate their characteristics of acid production and inhibitory effect. PCR-DGGE has been developed based on the 16S rRNA gene, and a set of HDA-1-GC and HDA-2, designed at V2-V3 region, and another set of CARP-1-GC and CARP-2, designed at V1 region, have been used. The bacterial strains included L. casei ATCC 393, L. paracasei CCUG 32212, L. rhamnosus ATCC 7469, L. zeae CCUG 35515, and 46 clinical strains of L. casei/paracasei/rhamnosus. Inhibitory effect against Streptococcus mutans and acid production were examined. Results revealed that each type species strain and identified clinical isolate showed its own unique DGGE pattern using CARP1-GC and CARP2 primers. HDA1-GC and HDA2 primers could distinguish the strains of L. paracasei from L. casei. It was found that inhibitory effect of L. paracasei was stronger than L. casei and L. rhamnosus. The acid production of L. paracasei was lower than L. casei and L. rhamnosus. In conclusion, the technique has been proven to be able to differentiate between closely related species in L. casei group and thus provide reliable information of their phenotypic appearances.

  3. Monitoring of the microbial communities involved in the soy sauce manufacturing process by PCR-denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Tanaka, Yasushi; Watanabe, Jun; Mogi, Yoshinobu

    2012-08-01

    Soy sauce is a traditional seasoning produced through the fermentation of soybeans and wheat using microbes. In this study, the microbial communities involved in the soy sauce manufacturing process were analyzed by PCR-Denaturing Gradient Gel Electrophoresis (PCR-DGGE). The bacterial DGGE profile indicated that the bacterial microbes in the koji were Weissella cibaria (Weissella confusa, Weissella kimchii, Weissella salipiscis, Lactobacillus fermentum, Lactobacillus plantarum, Lactobacillus iners, or Streptococcus thermophilus), Staphylococcus gallinarum (or Staphylococcus xylosus), and Staphylococcus kloosii. In addition to these bacteria, Tetragenococcus halophilus was also detected in the mash during lactic acid fermentation. The fungal DGGE profile indicated that the fungal microbes in the koji were not only Aspergillus oryzae but also several yeasts. In the mash, Zygosaccharomyces rouxii appeared in the early fermentation stage, Candida etchellsii (or Candida nodaensis) and Candida versatilis were detected at the middle fermentation stage, and Candida etchellsii was detected at the mature fermentation stage. These results suggest that the microbial communities present during the soy sauce manufacturing process change drastically throughout its production. This is the first report to reveal the microbial communities involved in the soy sauce manufacturing process using a culture-independent method. Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.

  4. Analysis of microbial diversity on deli slicers using polymerase chain reaction and denaturing gradient gel electrophoresis technologies.

    Science.gov (United States)

    Koo, O K; Mertz, A W; Akins, E L; Sirsat, S A; Neal, J A; Morawicki, R; Crandall, P G; Ricke, S C

    2013-02-01

    Cross-contamination of pathogenic and spoilage bacteria from food-contact surfaces to food products is a serious public health issue. Bacteria may survive and attach to food-contact surfaces by residual food components and/or background bacteria which may subsequently transfer to other food products. Deli slicers, generally used for slicing ready-to-eat products, can serve as potential sources for considerable bacterial transfer. The objective of this study was to assess the extent and distribution of microbial diversity of deli slicers by identification of pathogenic and background bacteria. Slicer-swab samples were collected from restaurants in Arkansas and Texas in the United States. Ten surface areas for each slicer were swabbed using sterile sponges. Denaturing gradient gel electrophoresis (DGGE) was applied to investigate the fingerprint of samples, and each band was further identified by sequence analysis. Pseudomonads were identified as the dominant bacteria followed by Enterobacteriaceae family, and lactic acid bacteria such as Lactococcus lactis and Streptococcus thermophilus were also found. Bacterial distribution was similar for all surface areas, while the blade guard exhibited the greatest diversity. This study provides a profile of the microbial ecology of slicers using DGGE to develop more specific sanitation practices and to reduce cross-contamination during slicing. © 2012 The Society for Applied Microbiology.

  5. Denaturing gradient gel electrophoresis (DGGE as a powerful novel alternative for differentiation of epizootic ISA virus variants.

    Directory of Open Access Journals (Sweden)

    Marisela Carmona

    Full Text Available Infectious Salmon Anemia is a devastating disease critically affecting world-wide salmon production. Chile has been particularly stricken by this disease which in all cases has been directly related with its causative agent, a novel orthomyxovirus which presents specific and distinctive infective features. Among these, two molecular markers have been directly associated with pathogenicity in two of the eight RNA sub genomic coding units of the virus: an insertion hot spot region present in viral segment 5 and a Highly Polymorphic Region (HPR located in viral segment 6. Here we report the successful adaptation of a PCR-dependent denaturing gel electrophoresis technique (DGGE, which enables differentiation of selected reported HPR epizootic variants detected in Chile. At the same time, the technique allows us to distinguish one nucleotide differences in sequences associated with the intriguing, and still not well-understood, insertion events which tend to occur on RNA Segment 5. Thus, the versatility of the technique opens new opportunities for improved understanding of the complex biology of all ISA variants as well as possible applications to other highly variable pathogens.

  6. Heat-denaturation and aggregation of quinoa (Chenopodium quinoa) globulins as affected by the pH value.

    Science.gov (United States)

    Mäkinen, Outi E; Zannini, Emanuele; Koehler, Peter; Arendt, Elke K

    2016-04-01

    The influence of heating (100 °C; 0-15 min) on the relative molecular mass, protein unfolding and secondary structure of quinoa globulins was studied at pH 6.5 (low solubility), 8.5 and 10.5 (high solubility). The patterns of denaturation and aggregation varied with pH. Heating triggered the disruption of the disulfide bonds connecting the acidic and basic chains of the chenopodin subunits at pH 8.5 and 10.5, but not at pH 6.5. Large aggregates unable to enter a 4% SDS-PAGE gel were formed at pH 6.5 and 8.5, which became soluble under reducing conditions. Heating at pH 10.5 lead to a rapid dissociation of the native chenopodin and to the disruption of the subunits, but no SDS-insoluble aggregates were formed. No major changes in secondary structure occurred during a 15 min heating, but an increase in hydrophobicity indicated unfolding of the tertiary structure in all samples. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Counteraction of urea-induced protein denaturation by trimethylamine N-oxide: a chemical chaperone at atomic resolution.

    Science.gov (United States)

    Bennion, Brian J; Daggett, Valerie

    2004-04-27

    Proteins are very sensitive to their solvent environments. Urea is a common chemical denaturant of proteins, yet some animals contain high concentrations of urea. These animals have evolved an interesting mechanism to counteract the effects of urea by using trimethylamine N-oxide (TMAO). The molecular basis for the ability of TMAO to act as a chemical chaperone remains unknown. Here, we describe molecular dynamics simulations of a small globular protein, chymotrypsin inhibitor 2, in 8 M urea and 4 M TMAO/8 M urea solutions, in addition to other control simulations, to investigate this effect at the atomic level. In 8 M urea, the protein unfolds, and urea acts in both a direct and indirect manner to achieve this effect. In contrast, introduction of 4 M TMAO counteracts the effect of urea and the protein remains well structured. TMAO makes few direct interactions with the protein. Instead, it prevents unfolding of the protein by structuring the solvent. In particular, TMAO orders the solvent and discourages it from competing with intraprotein H bonds and breaking up the hydrophobic core of the protein.

  8. Application of the denaturing gradient gel electrophoresis (DGGE) technique as an efficient diagnostic tool for ciliate communities in soil.

    Science.gov (United States)

    Jousset, Alexandre; Lara, Enrique; Nikolausz, Marcell; Harms, Hauke; Chatzinotas, Antonis

    2010-02-01

    Ciliates (or Ciliophora) are ubiquitous organisms which can be widely used as bioindicators in ecosystems exposed to anthropogenic and industrial influences. The evaluation of the environmental impact on soil ciliate communities with methods relying on morphology-based identification may be hampered by the large number of samples usually required for a statistically supported, reliable conclusion. Cultivation-independent molecular-biological diagnostic tools are a promising alternative to greatly simplify and accelerate such studies. In this present work a ciliate-specific fingerprint method based on the amplification of a phylogenetic marker gene (i.e. the 18S ribosomal RNA gene) with subsequent analysis by denaturing gradient gel electrophoresis (DGGE) was developed and used to monitor community shifts in a polycyclic aromatic hydrocarbon (PAH) polluted soil. The semi-nested approach generated ciliate-specific amplification products from all soil samples and allowed to distinguish community profiles from a PAH-polluted and a non-polluted control soil. Subsequent sequence analysis of excised bands provided evidence that polluted soil samples are dominated by organisms belonging to the class Colpodea. The general DGGE approach presented in this study might thus in principle serve as a fast and reproducible diagnostic tool, complementing and facilitating future ecological and ecotoxicological monitoring of ciliates in polluted habitats. Copyright 2009 Elsevier B.V. All rights reserved.

  9. A Fast Silver Staining Protocol Enabling Simple and Efficient Detection of SSR Markers using a Non-denaturing Polyacrylamide Gel.

    Science.gov (United States)

    Huang, Ling; Deng, Xiaohui; Li, Ronghua; Xia, Yanshi; Bai, Guihua; Siddique, Kadambot H M; Guo, Peiguo

    2018-04-20

    Simple Sequence Repeat (SSR) is one of the most effective markers used in plant and animal genetic research and molecular breeding programs. Silver staining is a widely used method for the detection of SSR markers in a polyacrylamide gel. However, conventional protocols for silver staining are technically demanding and time-consuming. Like many other biological laboratory techniques, silver staining protocols have been steadily optimized to improve detection efficiency. Here, we report a simplified silver staining method that significantly reduces reagent costs and enhances the detection resolution and picture clarity. The new method requires two major steps (impregnation and development) and three reagents (silver nitrate, sodium hydroxide, and formaldehyde), and only 7 min of processing for a non-denaturing polyacrylamide gel. Compared to previously reported protocols, this new method is easier, quicker and uses fewer chemical reagents for SSR detection. Therefore, this simple, low-cost, and effective silver staining protocol will benefit genetic mapping and marker-assisted breeding by a quick generation of SSR marker data.

  10. Behavior of variable V3 region from 16S rDNA of lactic acid bacteria in denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Ercolini, D; Moschetti, G; Blaiotta, G; Coppola, S

    2001-03-01

    Separation of amplified V3 region from 16S rDNA by denaturing gradient gel electrophoresis (DGGE) was tested as a tool for differentiation of lactic acid bacteria commonly isolated from food. Variable V3 regions of 21 reference strains and 34 wild strains referred to species belonging to the genera Pediococcus, Enterococcus, Lactococcus, Lactobacillus, Leuconostoc, Weissella, and Streptococcus were analyzed. DGGE profiles obtained were species-specific for most of the cultures tested. Moreover, it was possible to group the remaining LAB reference strains according to the migration of their 16S V3 region in the denaturing gel. The results are discussed with reference to their potential in the analysis of LAB communities in food, besides shedding light on taxonomic aspects.

  11. Purification of correctly oxidized MHC class I heavy-chain molecules under denaturing conditions: a novel strategy exploiting disulfide assisted protein folding

    DEFF Research Database (Denmark)

    Ferré, Henrik; Ruffet, Emmanuel; Blicher, Thomas

    2003-01-01

    The aim of this study has been to develop a strategy for purifying correctly oxidized denatured major histocompability complex class I (MHC-I) heavy-chain molecules, which on dilution, fold efficiently and become functional. Expression of heavy-chain molecules in bacteria results in the formation...... of insoluble cellular inclusion bodies, which must be solubilized under denaturing conditions. Their subsequent purification and refolding is complicated by the fact that (1). correct folding can only take place in combined presence of beta(2)-microglobulin and a binding peptide; and (2). optimal in vitro...... conditions for disulfide bond formation ( approximately pH 8) and peptide binding ( approximately pH 6.6) are far from complementary. Here we present a two-step strategy, which relies on uncoupling the events of disulfide bond formation and peptide binding. In the first phase, heavy-chain molecules...

  12. Purification of correctly oxidized MHC class I heavy-chain molecules under denaturing conditions: A novel strategy exploiting disulfide assisted protein folding

    DEFF Research Database (Denmark)

    Ferré, Henrik; Ruffet, E.; Blicher, T.

    2003-01-01

    The aim of this study has been to develop a strategy for purifying correctly oxidized denatured major histocompability complex class I (MHC-I) heavy-chain molecules, which on dilution, fold efficiently and become functional. Expression of heavy-chain molecules in bacteria results in the formation...... of insoluble cellular inclusion bodies, which must be solubilized under denaturing conditions. Their subsequent purification and refolding is complicated by the fact that (1) correct folding can only take place in combined presence of beta(2)-microglobulin and a binding peptide; and (2) optimal in vitro...... conditions for disulfide bond formation (similar topH 8) and peptide binding (similar topH 6.6) are far from complementary. Here we present a two-step strategy, which relies on uncoupling the events of disulfide bond formation and peptide binding. In the first phase, heavy-chain molecules with correct...

  13. 1H, 15N and 13C assignments of domain 5 of Dictyostelium discoideum gelation factor (ABP-120) in its native and 8M urea-denatured states.

    Science.gov (United States)

    Hsu, Shang-Te Danny; Cabrita, Lisa D; Christodoulou, John; Dobson, Christopher M

    2009-06-01

    The gelation factor from Dictyostelium discoideum (ABP-120) is an actin binding protein consisting of six immunoglobulin (Ig) domains in the C-terminal rod domain. We have recently used the pair of domains 5 and 6 of ABP-120 as a model system for studying multi-domain nascent chain folding on the ribosome. Here we present the NMR assignments of domain 5 in its native and 8M urea-denatured states.

  14. Sampling the Denatured State of Polypeptides in Water, Urea, and Guanidine Chloride to Strict Equilibrium Conditions with the Help of Massively Parallel Computers.

    Science.gov (United States)

    Meloni, Roberto; Camilloni, Carlo; Tiana, Guido

    2014-02-11

    The denatured state of polypeptides and proteins, stabilized by chemical denaturants like urea and guanidine chloride, displays residual secondary structure when studied by nuclear-magnetic-resonance spectroscopy. However, these experimental techniques are weakly sensitive, and thus molecular-dynamics simulations can be useful to complement the experimental findings. To sample the denatured state, we made use of massively-parallel computers and of a variant of the replica exchange algorithm, in which the different branches, connected with unbiased replicas, favor the formation and disruption of local secondary structure. The algorithm is applied to the second hairpin of GB1 in water, in urea, and in guanidine chloride. We show with the help of different criteria that the simulations converge to equilibrium. It results that urea and guanidine chloride, besides inducing some polyproline-II structure, have different effect on the hairpin. Urea disrupts completely the native region and stabilizes a state which resembles a random coil, while guanidine chloride has a milder effect.

  15. Diagnostic work-up of pulmonary nodules. Management of pulmonary nodules detected with low-dose CT screening; Abklaerung von Lungenrundherden. Management durch Frueherkennungsuntersuchungen detektierter pulmonaler Rundherde

    Energy Technology Data Exchange (ETDEWEB)

    Wormanns, D. [Evangelische Lungenklinik Berlin, Berlin (Germany)

    2016-09-15

    Pulmonary nodules are the most frequent pathological finding in low-dose computed tomography (CT) scanning for early detection of lung cancer. Early stages of lung cancer are often manifested as pulmonary nodules; however, the very commonly occurring small nodules are predominantly benign. These benign nodules are responsible for the high percentage of false positive test results in screening studies. Appropriate diagnostic algorithms are necessary to reduce false positive screening results and to improve the specificity of lung cancer screening. Such algorithms are based on some of the basic principles comprehensively described in this article. Firstly, the diameter of nodules allows a differentiation between large (>8 mm) probably malignant and small (<8 mm) probably benign nodules. Secondly, some morphological features of pulmonary nodules in CT can prove their benign nature. Thirdly, growth of small nodules is the best non-invasive predictor of malignancy and is utilized as a trigger for further diagnostic work-up. Non-invasive testing using positron emission tomography (PET) and contrast enhancement as well as invasive diagnostic tests (e.g. various procedures for cytological and histological diagnostics) are briefly described in this article. Different nodule morphology using CT (e.g. solid and semisolid nodules) is associated with different biological behavior and different algorithms for follow-up are required. Currently, no obligatory algorithm is available in German-speaking countries for the management of pulmonary nodules, which reflects the current state of knowledge. The main features of some international and American recommendations are briefly presented in this article from which conclusions for the daily clinical use are derived. (orig.) [German] Lungenrundherde sind die haeufigsten pathologischen Befunde bei Untersuchungen mit der Niedrigdosis-CT zur Lungenkrebsfrueherkennung. Fruehstadien des Lungenkarzinoms manifestieren sich meist als Rundherd

  16. Mapping Proteoforms and Protein Complexes From King Cobra Venom Using Both Denaturing and Native Top-down Proteomics.

    Science.gov (United States)

    Melani, Rafael D; Skinner, Owen S; Fornelli, Luca; Domont, Gilberto B; Compton, Philip D; Kelleher, Neil L

    2016-07-01

    Characterizing whole proteins by top-down proteomics avoids a step of inference encountered in the dominant bottom-up methodology when peptides are assembled computationally into proteins for identification. The direct interrogation of whole proteins and protein complexes from the venom of Ophiophagus hannah (king cobra) provides a sharply clarified view of toxin sequence variation, transit peptide cleavage sites and post-translational modifications (PTMs) likely critical for venom lethality. A tube-gel format for electrophoresis (called GELFrEE) and solution isoelectric focusing were used for protein fractionation prior to LC-MS/MS analysis resulting in 131 protein identifications (18 more than bottom-up) and a total of 184 proteoforms characterized from 14 protein toxin families. Operating both GELFrEE and mass spectrometry to preserve non-covalent interactions generated detailed information about two of the largest venom glycoprotein complexes: the homodimeric l-amino acid oxidase (∼130 kDa) and the multichain toxin cobra venom factor (∼147 kDa). The l-amino acid oxidase complex exhibited two clusters of multiproteoform complexes corresponding to the presence of 5 or 6 N-glycans moieties, each consistent with a distribution of N-acetyl hexosamines. Employing top-down proteomics in both native and denaturing modes provides unprecedented characterization of venom proteoforms and their complexes. A precise molecular inventory of venom proteins will propel the study of snake toxin variation and the targeted development of new antivenoms or other biotherapeutics. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Mechanism of Protein Denaturation: Partial Unfolding of the P22 Coat Protein I-Domain by Urea Binding

    Science.gov (United States)

    Newcomer, Rebecca L.; Fraser, LaTasha C.R.; Teschke, Carolyn M.; Alexandrescu, Andrei T.

    2015-01-01

    The I-domain is an insertion domain of the bacteriophage P22 coat protein that drives rapid folding and accounts for over half of the stability of the full-length protein. We sought to determine the role of hydrogen bonds (H-bonds) in the unfolding of the I-domain by examining 3JNC’ couplings transmitted through H-bonds, the temperature and urea-concentration dependence of 1HN and 15N chemical shifts, and native-state hydrogen exchange at urea concentrations where the domain is predominantly folded. The native-state hydrogen-exchange data suggest that the six-stranded β-barrel core of the I-domain is more stable against unfolding than a smaller subdomain comprised of a short α-helix and three-stranded β-sheet. H-bonds, separately determined from solvent protection and 3JNC’ H-bond couplings, are identified with an accuracy of 90% by 1HN temperature coefficients. The accuracy is improved to 95% when 15N temperature coefficients are also included. In contrast, the urea dependence of 1HN and 15N chemical shifts is unrelated to H-bonding. The protein segments with the largest chemical-shift changes in the presence of urea show curved or sigmoidal titration curves suggestive of direct urea binding. Nuclear Overhauser effects to urea for these segments are also consistent with specific urea-binding sites in the I-domain. Taken together, the results support a mechanism of urea unfolding in which denaturant binds to distinct sites in the I-domain. Disordered segments bind urea more readily than regions in stable secondary structure. The locations of the putative urea-binding sites correlate with the lower stability of the structure against solvent exchange, suggesting that partial unfolding of the structure is related to urea accessibility. PMID:26682823

  18. Effects of Metal Ions, Temperature, and a Denaturant on the Oxidative Folding Pathways of Bovine α-Lactalbumin

    Directory of Open Access Journals (Sweden)

    Reina Shinozaki

    2017-09-01

    Full Text Available Bovine α-lactalbumin (αLA has four disulfide (SS bonds in the native form (N. On the oxidative folding pathways of this protein, two specific SS folding intermediates, i.e., (61–77, 73–91 and des[6–120], which have two and three native SS bonds, respectively, accumulate predominantly in the presence of Ca2+. In this study, we reinvestigated the pathways using a water-soluble cyclic selenoxide reagent, trans-3,4-dihydroxyselenolane oxide (DHSox, as a strong and quantitative oxidant to oxidize the fully reduced form (R. In the presence of ethylenediaminetetraacetic acid (EDTA (under a metal-free condition, SS formation randomly proceeded, and N did not regenerate. On the other hand, two specific SS intermediates transiently generated in the presence of Ca2+. These intermediates could be assigned to (61–77, 73–91 and des[6–120] having two common SS bonds, i.e., Cys61-Cys77 and Cys73-Cys91, near the calcium binding pocket of the β-sheet domain. Much faster folding to N was observed in the presence of Mn2+, whereas Na+, K+, Mg2+, and Zn2+ did not affect the pathways. The two key intermediates were susceptible to temperature and a denaturant. The oxidative folding pathways revealed were significantly different from those of hen egg white lysozyme, which has the same SS-bonding pattern as αLA, suggesting that the folding pathways of SS-containing proteins can alter depending on the amino acid sequence and other factors, even when the SS-bond topologies are similar to each other.

  19. Weak and saturable protein-surfactant interactions in the denaturation of apo-alpha-lactalbumin by acidic and lactonic sophorolipid

    Directory of Open Access Journals (Sweden)

    Kell K Andersen

    2016-11-01

    Full Text Available Biosurfactants are of growing interest as sustainable alternatives to fossil-fuel-derived chemical surfactants, particularly for the detergent industry. To realize this potential, it is necessary to understand how they affect proteins which they may encounter in their applications. However knowledge of such interactions is limited. Here we present a study of the interactions between the model protein apo-alpha-lactalbumin and the biosurfactant sophorolipid (SL produced by the yeast Starmerella bombicola. SL occurs both as an acidic and a lactonic form; the lactonic form (lactSL is sparingly soluble and has a lower critical micelle concentration than the acidic form (acidSL. We show that acidSL affects apo-aLA in a similar way to the related glycolipid biosurfactant rhamnolipid (RL, with the important difference that RL is also active below the cmc in contrast to acidSL. Using isothermal titration calorimetry data, we show that acidSL has weak and saturable interactions with apo-aLA at low concentrations; due to the relatively low cmc of acidSL (which means that the monomer concentration is limited to ca. 0-1 mM SL, it is only possible to observe interactions with monomeric acidSL at high apo-aLA concentrations. However, the denaturation kinetics of apo-aLA in the presence of acidSL are consistent with a collaboration between monomeric and micellar surfactant species, similar to RL and nonionic or zwitterionic surfactants. Inclusion of lactSL as mixed micelles with acidSL lowers the cmc and this effectively reduces the rate of unfolding, emphasizing that SL like other biosurfactants is a gentle anionic surfactant. Our data highlight the potential of these biosurfactants for future use in the detergent industry.

  20. Origins of protein denatured state compactness and hydrophobic clustering in aqueous urea: inferences from nonpolar potentials of mean force.

    Science.gov (United States)

    Shimizu, Seishi; Chan, Hue Sun

    2002-12-01

    Free energies of pairwise hydrophobic association are simulated in aqueous solutions of urea at concentrations ranging from 0-8 M. Consistent with the expectation that hydrophobic interactions are weakened by urea, the association of relatively large nonpolar solutes is destabilized by urea. However, the association of two small methane-sized nonpolar solutes in water has the opposite tendency of being slightly strengthened by the addition of urea. Such size effects and the dependence of urea-induced stability changes on the configuration of nonpolar solutes are not predicted by solvent accessible surface area approaches based on energetic parameters derived from bulk-phase solubilities of model compounds. Thus, to understand hydrophobic interactions in proteins, it is not sufficient to rely solely on transfer experiment data that effectively characterize a single nonpolar solute in an aqueous environment but not the solvent-mediated interactions among two or more nonpolar solutes. We find that the m-values for the rate of change of two-methane association free energy with respect to urea concentration is a dramatically nonmonotonic function of the spatial separation between the two methanes, with a distance-dependent profile similar to the corresponding two-methane heat capacity of association in pure water. Our results rationalize the persistence of residual hydrophobic contacts in some proteins at high urea concentrations and explain why the heat capacity signature (DeltaC(P)) of a compact denatured state can be similar to DeltaC(P) values calculated by assuming an open random-coil-like unfolded state. Copyright 2002 Wiley-Liss, Inc.

  1. Is there still a role for thyroid scintigraphy in the workup of a thyroid nodule in the era of fine needle aspiration cytology and molecular testing? [version 1; referees: 3 approved

    Directory of Open Access Journals (Sweden)

    Rodrigo Moreno-Reyes

    2016-04-01

    Full Text Available Thyroid scintigraphy is now rarely used in the work-up of a thyroid nodule except in the presence of a low TSH value. Therefore, autonomously functioning thyroid nodules (AFTNs with a normal TSH value are diagnosed only in the rare medical centers that continue to use thyroid scan systematically in the presence of a thyroid nodule. In this review, we discuss the prevalence of AFTN with a normal TSH level and the possible consequences of performing fine needle aspiration cytology (FNAC in an undiagnosed AFTN. We also discuss the risk of malignant AFTN which may be higher than previously stated.

  2. Biological effects of DNA repair, including mutagenesis. Progress report, June 15, 1980-May 1, 1981

    International Nuclear Information System (INIS)

    Hutchinson, F.

    1981-01-01

    A mutagen of interest is used to create mutations in the cI gene of lambda phage, producing a clear plaque phenotype. Genetic mapping is used to determine which restriction fragment in the gene contains the mutation. The fragment is then isolated and sequenced by the Maxam-Gilbert technique. Thus, the actual changes in base sequence produced by the action of the mutagen may be determined and also the base sequence surrounding the mutation. Such studies will be useful in understanding the mechanisms by which a mutation occurs, since any proposed mechanism must be consistent with the observations. The results also give information on the specificity of mutagens

  3. Guanidine hydrochloride denaturation of human serum albumin originates by local unfolding of some stable loops in domain III.

    Science.gov (United States)

    Ahmad, Basir; Ahmed, Md Zulfazal; Haq, Soghra Khatun; Khan, Rizwan Hasan

    2005-06-15

    The effect of guanidine hydrochloride (GnHCl) on the global stability of human serum albumin (HSA) has been studied by fluorescence and circular dichroism spectroscopic measurements. The differential stability of native conformation of three HSA domains were explored by using domain-specific ligands, hemin (domain I), chloroform (domain II), bilirubin (at domain I/domain II interface) and diazepam (domain III). GnHCl induced unfolding transition curves as monitored by probes for secondary and tertiary structures were cooperative but noncoincidental. A strong ANS binding to the protein was observed around 1.8 M GnHCl, suggesting existence of intermediate states in the unfolding pathway of HSA. A gradual decrease (in the GnHCl concentration range 0.0-1.8 M) in the binding of diazepam indicates that domain III is the most labile to GnHCl denaturation. A significant increase in the binding of bilirubin up to 1.4 M GnHCl and decrease thereafter leading to complete abolishment of bilirubin binding at around 2.0 M GnHCl suggest favorable rearrangement and separation of domains I and II at 1.4 and 2.0 M GnHCl concentration, respectively. Above 1.6 M GnHCl, decrease of the binding of hemin, a ligand for domain I, chloroform, which binds in domain II and lone tryptophanyl fluorescence (Trp-214 located in domain II) indicate that at higher concentration of GnHCl domains I and II start unfolding simultaneously but the stability of domain I (7.4 Kcal/mol) is much more than domain II (4.3 Kcal/mol). A pictorial model for the unfolding of HSA domains, consistent with all these results, has been formulated, suggesting that domain III is the most labile followed by domain II while domain I is the most stable. A molten globule like state of domain III around 1.8 M GnHCl has also been identified and characterized.

  4. Monomorphism in humans and sequence differences among higher primates for a sequence tagged site (STS) in homeo box cluster 2 as assayed by denaturing gradient electrophoresis

    Energy Technology Data Exchange (ETDEWEB)

    Ruano, G.; Ruddle, F.H.; Kidd, K.K. (Yale Univ., New Haven, CT (United States)); Gray, M.R. (Tufts Univ., Boston, MA (United States)); Miki, Tetsuro (Osaka Univ. (Japan)); Ferguson-Smith, A.C. (Inst. of Animal Physiology and Genetics Research, Cambridge (United Kingdom))

    1990-03-11

    The human homeo box cluster 2 (HOX2) contains genes coding for DNA binding proteins involved in developmental control and is highly conserved between mouse and man. The authors have applied in concert the Polymerase Chain Reaction (PCR) and Denaturing Gradient Electrophoresis (DGE) to amplify defined primate HOX2 segments and to detect sequence differences among them. They have sequenced a PstI fragment 4 kb upstream from HOX 2.2 and synthesized primers delimiting both halves of 630 bp segment within it PCR on various unrelated humans and SC-PCR on chimpanzee, gorilla, orangutan and gibbon yielded products of the same length for each primer pair.

  5. Essential roles of protein-solvent many-body correlation in solvent-entropy effect on protein folding and denaturation: Comparison between hard-sphere solvent and water

    International Nuclear Information System (INIS)

    Oshima, Hiraku; Kinoshita, Masahiro

    2015-01-01

    In earlier works, we showed that the entropic effect originating from the translational displacement of water molecules plays the pivotal role in protein folding and denaturation. The two different solvent models, hard-sphere solvent and model water, were employed in theoretical methods wherein the entropic effect was treated as an essential factor. However, there were similarities and differences in the results obtained from the two solvent models. In the present work, to unveil the physical origins of the similarities and differences, we simultaneously consider structural transition, cold denaturation, and pressure denaturation for the same protein by employing the two solvent models and considering three different thermodynamic states for each solvent model. The solvent-entropy change upon protein folding/unfolding is decomposed into the protein-solvent pair (PA) and many-body (MB) correlation components using the integral equation theories. Each component is further decomposed into the excluded-volume (EV) and solvent-accessible surface (SAS) terms by applying the morphometric approach. The four physically insightful constituents, (PA, EV), (PA, SAS), (MB, EV), and (MB, SAS), are thus obtained. Moreover, (MB, SAS) is discussed by dividing it into two factors. This all-inclusive investigation leads to the following results: (1) the protein-water many-body correlation always plays critical roles in a variety of folding/unfolding processes; (2) the hard-sphere solvent model fails when it does not correctly reproduce the protein-water many-body correlation; (3) the hard-sphere solvent model becomes problematic when the dependence of the many-body correlation on the solvent number density and temperature is essential: it is not quite suited to studies on cold and pressure denaturating of a protein; (4) when the temperature and solvent number density are limited to the ambient values, the hard-sphere solvent model is usually successful; and (5) even at the ambient

  6. 99 mTc-sulphur-colloid and heat-denatured 99mTc-labelled red cell scans demonstrating a giant intrapelvic spleen in a girl after splenectomy

    International Nuclear Information System (INIS)

    Kao, P.F.; Tzen, K.Y.; Tsai, M.F.; Lin, J.N.

    2001-01-01

    A 17 x 12 x 5-cm giant intrapelvic mass in a 14-year-old girl is reported. This mass developed 6 years after a splenectomy for splenic torsion. The heat-denatured 99 m Tc-labelled red cell scan and 99 m Tc- sulphur-colloid scan confirmed the specific red cell sequestration function and reticuloendothelial activity in the giant intrapelvic spleen. The size and development of the giant intrapelvic spleen are unusual. The usefulness of functional images to diagnosis the nature of the intrapelvic mass is well demonstrated. (orig.)

  7. The H1 histone-specific proteinase is associated with nuclear matrix and stimulated by DNA containing breaks of denatured sites

    International Nuclear Information System (INIS)

    Gaziev, A.I.; Kutsyj, M.P.

    1988-01-01

    Discovery of proteinase in nuclear matrix specific of H1 histone and dependent presence of breaks or denatured sites in DNA permits to assume that the given enzyme, obviously, participates in replication and DNA repair, in regulation of genes expression. Removal of H1 histone by proteinase is, probably, necessary for procedure of these processes, and, obviously, this proteinase suffers conformational changes in the composition of the DNA-histone complex. H1 histone disintegration in nucleohistone containing damaged sites of DNA by specific proteinase, probably, represents one of the mechanisms for providing DNA repair in cells of higher organisms

  8. A Comprehensive Quality Evaluation System for Complex Herbal Medicine Using PacBio Sequencing, PCR-Denaturing Gradient Gel Electrophoresis, and Several Chemical Approaches

    Science.gov (United States)

    Zheng, Xiasheng; Zhang, Peng; Liao, Baosheng; Li, Jing; Liu, Xingyun; Shi, Yuhua; Cheng, Jinle; Lai, Zhitian; Xu, Jiang; Chen, Shilin

    2017-01-01

    Herbal medicine is a major component of complementary and alternative medicine, contributing significantly to the health of many people and communities. Quality control of herbal medicine is crucial to ensure that it is safe and sound for use. Here, we investigated a comprehensive quality evaluation system for a classic herbal medicine, Danggui Buxue Formula, by applying genetic-based and analytical chemistry approaches to authenticate and evaluate the quality of its samples. For authenticity, we successfully applied two novel technologies, third-generation sequencing and PCR-DGGE (denaturing gradient gel electrophoresis), to analyze the ingredient composition of the tested samples. For quality evaluation, we used high performance liquid chromatography assays to determine the content of chemical markers to help estimate the dosage relationship between its two raw materials, plant roots of Huangqi and Danggui. A series of surveys were then conducted against several exogenous contaminations, aiming to further access the efficacy and safety of the samples. In conclusion, the quality evaluation system demonstrated here can potentially address the authenticity, quality, and safety of herbal medicines, thus providing novel insight for enhancing their overall quality control. Highlight: We established a comprehensive quality evaluation system for herbal medicine, by combining two genetic-based approaches third-generation sequencing and DGGE (denaturing gradient gel electrophoresis) with analytical chemistry approaches to achieve the authentication and quality connotation of the samples. PMID:28955365

  9. A Comprehensive Quality Evaluation System for Complex Herbal Medicine Using PacBio Sequencing, PCR-Denaturing Gradient Gel Electrophoresis, and Several Chemical Approaches

    Directory of Open Access Journals (Sweden)

    Xiasheng Zheng

    2017-09-01

    Full Text Available Herbal medicine is a major component of complementary and alternative medicine, contributing significantly to the health of many people and communities. Quality control of herbal medicine is crucial to ensure that it is safe and sound for use. Here, we investigated a comprehensive quality evaluation system for a classic herbal medicine, Danggui Buxue Formula, by applying genetic-based and analytical chemistry approaches to authenticate and evaluate the quality of its samples. For authenticity, we successfully applied two novel technologies, third-generation sequencing and PCR-DGGE (denaturing gradient gel electrophoresis, to analyze the ingredient composition of the tested samples. For quality evaluation, we used high performance liquid chromatography assays to determine the content of chemical markers to help estimate the dosage relationship between its two raw materials, plant roots of Huangqi and Danggui. A series of surveys were then conducted against several exogenous contaminations, aiming to further access the efficacy and safety of the samples. In conclusion, the quality evaluation system demonstrated here can potentially address the authenticity, quality, and safety of herbal medicines, thus providing novel insight for enhancing their overall quality control.Highlight: We established a comprehensive quality evaluation system for herbal medicine, by combining two genetic-based approaches third-generation sequencing and DGGE (denaturing gradient gel electrophoresis with analytical chemistry approaches to achieve the authentication and quality connotation of the samples.

  10. A Comprehensive Quality Evaluation System for Complex Herbal Medicine Using PacBio Sequencing, PCR-Denaturing Gradient Gel Electrophoresis, and Several Chemical Approaches.

    Science.gov (United States)

    Zheng, Xiasheng; Zhang, Peng; Liao, Baosheng; Li, Jing; Liu, Xingyun; Shi, Yuhua; Cheng, Jinle; Lai, Zhitian; Xu, Jiang; Chen, Shilin

    2017-01-01

    Herbal medicine is a major component of complementary and alternative medicine, contributing significantly to the health of many people and communities. Quality control of herbal medicine is crucial to ensure that it is safe and sound for use. Here, we investigated a comprehensive quality evaluation system for a classic herbal medicine, Danggui Buxue Formula, by applying genetic-based and analytical chemistry approaches to authenticate and evaluate the quality of its samples. For authenticity, we successfully applied two novel technologies, third-generation sequencing and PCR-DGGE (denaturing gradient gel electrophoresis), to analyze the ingredient composition of the tested samples. For quality evaluation, we used high performance liquid chromatography assays to determine the content of chemical markers to help estimate the dosage relationship between its two raw materials, plant roots of Huangqi and Danggui. A series of surveys were then conducted against several exogenous contaminations, aiming to further access the efficacy and safety of the samples. In conclusion, the quality evaluation system demonstrated here can potentially address the authenticity, quality, and safety of herbal medicines, thus providing novel insight for enhancing their overall quality control. Highlight : We established a comprehensive quality evaluation system for herbal medicine, by combining two genetic-based approaches third-generation sequencing and DGGE (denaturing gradient gel electrophoresis) with analytical chemistry approaches to achieve the authentication and quality connotation of the samples.

  11. Resonance assignment of disordered protein with repetitive and overlapping sequence using combinatorial approach reveals initial structural propensities and local restrictions in the denatured state

    Energy Technology Data Exchange (ETDEWEB)

    Malik, Nikita; Kumar, Ashutosh, E-mail: askutoshk@iitb.ac.in [Indian Institute of Technology Bombay, Department of Bioscience and Bioengineering (India)

    2016-09-15

    NMR resonance assignment of intrinsically disordered proteins poses a challenge because of the limited dispersion of amide proton chemical shifts. This becomes even more complex with the increase in the size of the system. Residue specific selective labeling/unlabeling experiments have been used to resolve the overlap, but require multiple sample preparations. Here, we demonstrate an assignment strategy requiring only a single sample of uniformly labeled {sup 13}C,{sup 15}N-protein. We have used a combinatorial approach, involving 3D-HNN, CC(CO)NH and 2D-MUSIC, which allowed us to assign a denatured centromeric protein Cse4 of 229 residues. Further, we show that even the less sensitive experiments, when used in an efficient manner can lead to the complete assignment of a complex system without the use of specialized probes in a relatively short time frame. The assignment of the amino acids discloses the presence of local structural propensities even in the denatured state accompanied by restricted motion in certain regions that provides insights into the early folding events of the protein.

  12. Resonance assignment of disordered protein with repetitive and overlapping sequence using combinatorial approach reveals initial structural propensities and local restrictions in the denatured state

    International Nuclear Information System (INIS)

    Malik, Nikita; Kumar, Ashutosh

    2016-01-01

    NMR resonance assignment of intrinsically disordered proteins poses a challenge because of the limited dispersion of amide proton chemical shifts. This becomes even more complex with the increase in the size of the system. Residue specific selective labeling/unlabeling experiments have been used to resolve the overlap, but require multiple sample preparations. Here, we demonstrate an assignment strategy requiring only a single sample of uniformly labeled "1"3C,"1"5N-protein. We have used a combinatorial approach, involving 3D-HNN, CC(CO)NH and 2D-MUSIC, which allowed us to assign a denatured centromeric protein Cse4 of 229 residues. Further, we show that even the less sensitive experiments, when used in an efficient manner can lead to the complete assignment of a complex system without the use of specialized probes in a relatively short time frame. The assignment of the amino acids discloses the presence of local structural propensities even in the denatured state accompanied by restricted motion in certain regions that provides insights into the early folding events of the protein.

  13. pH dependence of cyanide binding to the ferric heme domain of the direct oxygen sensor from Escherichia coli and the effect of alkaline denaturation.

    Science.gov (United States)

    Bidwai, Anil K; Ok, Esther Y; Erman, James E

    2008-09-30

    The spectrum of the ferric heme domain of the direct oxygen sensor protein from Escherichia coli ( EcDosH) has been measured between pH 3.0 and 12.6. EcDosH undergoes acid denaturation with an apparent p K a of 4.24 +/- 0.05 and a Hill coefficient of 3.1 +/- 0.6 and reversible alkaline denaturation with a p K a of 9.86 +/- 0.04 and a Hill coefficient of 1.1 +/- 0.1. Cyanide binding to EcDosH has been investigated between pH 4 and 11. The EcDosH-cyanide complex is most stable at pH 9 with a K D of 0.29 +/- 0.06 microM. The kinetics of cyanide binding are monophasic between pH 4 and 8. At pH >or=8.5, the reaction is biphasic with the fast phase dependent upon the cyanide concentration and the slow phase independent of cyanide. The slow phase is attributed to conversion of denatured EcDosH to the native state, with a pH-independent rate of 0.052 +/- 0.006 s (-1). The apparent association rate constant for cyanide binding to EcDosH increases from 3.6 +/- 0.1 M (-1) s (-1) at pH 4 to 520 +/- 20 M (-1) s (-1) at pH 11. The dissociation rate constant averages (8.6 +/- 1.3) x 10 (-5) s (-1) between pH 5 and 9, increasing to (1.4 +/- 0.1) x 10 (-3) s (-1) at pH 4 and (2.5 +/- 0.1) x 10 (-3) s (-1) at pH 12.2. The mechanism of cyanide binding is consistent with preferential binding of the cyanide anion to native EcDosH. The reactions of imidazole and H 2O 2 with ferric EcDosH were also investigated and show little reactivity.

  14. High pressure inactivation of relevant target microorganisms in poultry meat products and the evaluation of pressure-induced protein denaturation of marinated poultry under different high pressure treatments

    Science.gov (United States)

    Schmidgall, Johanna; Hertel, Christian; Bindrich, Ute; Heinz, Volker; Toepfl, Stefan

    2011-03-01

    In this study, the possibility of extending shelf life of marinated poultry meat products by high pressure processing was evaluated. Relevant spoilage and pathogenic strains were selected and used as target microorganisms (MOs) for challenge experiments. Meat and brine were inoculated with MOs and treated at 450 MPa, 4 °C for 3 min. The results of inactivation show a decreasing pressure tolerance in the series Lactobacillus > Arcobacter > Carnobacterium > Bacillus cereus > Brochothrix thermosphacta > Listeria monocytogenes. Leuconostoc gelidum exhibited the highest pressure tolerance in meat. A protective effect of poultry meat was found for L. sakei and L. gelidum. In parallel, the influence of different marinade formulations (pH, carbonates, citrates) on protein structure changes during a pressure treatment was investigated. Addition of sodium carbonate shows a protection against denaturation of myofibrillar proteins and provides a maximum water-holding capacity. Caustic marinades allowed a higher retention of product characteristics than low-pH marinades.

  15. Application of denaturing high-performance liquid chromatography (DHPLC) for the identification of fish: a new way to determine the composition of processed food containing multiple species.

    Science.gov (United States)

    Le Fresne, Sophie; Popova, Milena; Le Vacon, Françoise; Carton, Thomas

    2011-12-14

    The identification of fish species in transformed food products is difficult because the existing methods are not adapted to heat-processed products containing more than one species. Using a common to all vertebrates region of the cytochrome b gene, we have developed a denaturing high-performance liquid chromatography (DHPLC) fingerprinting method, which allowed us to identify most of the species in commercial crab sticks. Whole fish and fillets were used for the creation of a library of referent DHPLC profiles. Crab sticks generated complex DHPLC profiles in which the number of contained fish species can be estimated by the number of major fluorescence peaks. The identity of some of the species was predicted by comparison of the peaks with the referent profiles, and others were identified after collection of the peak fractions, reamplification, and sequencing. DHPLC appears to be a quick and efficient method to analyze the species composition of complex heat-processed fish products.

  16. In vitro study of prebiotic properties of levan-type exopolysaccharides from Lactobacilli and non-digestible carbohydrates using denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Bello, F D; Walter, J; Hertel, C; Hammes, W P

    2001-07-01

    Batch cultures inoculated with human faeces were used to study the prebiotic properties of levan-type exopolysaccharides (EPS) from Lactobacillus sanfranciscensis as well as levan, inulin, and fructooligosaccharide (FOS). Denaturing gradient gel electrophoresis of 16S rDNA fragments generated by PCR with universal primers was used to analyse the cultures. Characteristic changes were revealed in the composition of the gut bacteria during fermentation of the carbohydrates. An enrichment of Bifidobacterium spp. was found for the EPS and inulin but not for levan and FOS. The bifidogenic effect of the EPS was confirmed by culturing on selective medium. In addition, the use of EPS and FOS resulted in enhanced growth of Eubacterium biforme and Clostridium perfringens, respectively.

  17. Culture-based and denaturing gradient gel electrophoresis analysis of the bacterial community from Chungkookjang, a traditional Korean fermented soybean food.

    Science.gov (United States)

    Hong, Sung Wook; Choi, Jae Young; Chung, Kun Sub

    2012-10-01

    The bacterial community of Chungkookjang and raw rice-straw collected from various areas in South Korea was investigated using both culture-dependent and culture-independent methods. Pure cultures were isolated from Chungkookjang and raw rice-straw on tryptic soy agar plates with 72 to 121 colonies and identified by 16S rDNA gene sequence analysis, respectively. The traditional culture-based method and denaturing gradient gel electrophoresis analysis of PCR-amplified 16S rDNA confirmed that Pantoea agglomerans and B. subtilis were identified as predominant in the raw rice-straw and Chungkookjang, respectively, from Iljuk district of Gyeonggi province, P. ananatis and B. licheniformis were identified as predominant in the raw rice-straw and Chungkookjang from Wonju district of Gangwon province, and Microbacterium sp. and B. licheniformis were identified as predominant in the raw rice-straw and Chungkookjang from Sunchang district of Jeolla province. Other strains, such as Bacillus, Enterococcus, Pseudomonas, Rhodococcus, and uncultured bacteria were also present in raw rice-straw and Chungkookjang. A comprehensive analysis of these microorganisms would provide a more detailed understanding of the biologically active components of Chungkookjang and help improve its quality. Polymerase chain reaction-denaturing gradient gel electrophoresis analysis can be successfully applied to a fermented food to detect unculturable or more species than the culture-dependent method. This technique is an effective and convenient culture-independent method for studying the bacterial community in Chungkookjang. In this study, the bacterial community of Chungkookjang collected from various areas in South Korea was investigated using both culture-dependent and culture-independent methods. © 2012 Institute of Food Technologists®

  18. Classical Hodgkin's lymphoma in adults: guidelines of the Italian Society of Hematology, the Italian Society of Experimental Hematology, and the Italian Group for Bone Marrow Transplantation on initial work-up, management, and follow-up.

    Science.gov (United States)

    Brusamolino, Ercole; Bacigalupo, Andrea; Barosi, Giovanni; Biti, Giampaolo; Gobbi, Paolo G; Levis, Alessandro; Marchetti, Monia; Santoro, Armando; Zinzani, Pier Luigi; Tura, Sante

    2009-04-01

    The Italian Society of Hematology (SIE), the Italian Society of Experimental Haematology (SIES) and the Italian Group for Bone Marrow Transplantation (GITMO) commissioned a project to develop practice guidelines for the initial work-up, therapy and follow-up of classical Hodgkin's lymphoma. Key questions to the clinical evaluation and treatment of this disease were formulated by an Advisory Committee, discussed and approved by an Expert Panel (EP) composed of senior hematologists and one radiotherapist. After a comprehensive and systematic literature review, the EP recommendations were graded according to their supporting evidence. An explicit approach to consensus methodologies was used for evidence interpretation and for producing recommendations in the absence of a strong evidence. The EP decided that the target domain of the guidelines should include only classical Hodgkin's lymphoma, as defined by the WHO classification, and exclude lymphocyte predominant histology. Distinct recommendations were produced for initial work-up, first-line therapy of early and advanced stage disease, monitoring procedures and salvage therapy, including hemopoietic stem cell transplant. Separate recommendations were formulated for elderly patients. Pre-treatment volumetric CT scan of the neck, thorax, abdomen, and pelvis is mandatory, while FDG-PET is recommended. As to the therapy of early stage disease, a combined modality approach is still recommended with ABVD followed by involved-field radiotherapy; the number of courses of ABVD will depend on the patient risk category (favorable or unfavorable). Full-term chemotherapy with ABVD is recommended in advanced stage disease; adjuvant radiotherapy in patients without initial bulk who achieved a complete remission is not recommended. In the elderly, chemotherapy regimens more intensive than ABVD are not recommended. Early evaluation of response with FDG-PET scan is suggested. Relapsed or refractory patients should receive high

  19. The effect of a non-denaturing detergent and a guanidinium-based inactivation agent on the viability of Ebola virus in mock clinical serum samples.

    Science.gov (United States)

    Burton, J E; Easterbrook, L; Pitman, J; Anderson, D; Roddy, S; Bailey, D; Vipond, R; Bruce, C B; Roberts, A D

    2017-12-01

    The 2014 Ebola outbreak in West Africa required the rapid testing of clinical material for the presence of potentially high titre Ebola virus (EBOV). Safe, fast and effective methods for the inactivation of such clinical samples are required so that rapid diagnostic tests including downstream analysis by RT-qPCR or nucleotide sequencing can be carried out. One of the most commonly used guanidinium - based denaturing agents, AVL (Qiagen) has been shown to fully inactivate EBOV once ethanol is added, however this is not compatible with the use of automated nucleic acid extraction systems. Additional inactivation agents need to be identified that can be used in automated systems. A candidate inactivation agent is Triton X-100, a non-denaturing detergent that is frequently used in clinical nucleic acid extraction procedures and has previously been used for inactivation of EBOV. In this study the effect of 0.1% and 1.0% Triton X-100 (final concentration 0.08% and 0.8% respectively) alone and in combination with AVL on the viability of EBOV (10 6 TCID 50 /ml) spiked into commercially available pooled negative human serum was tested. The presence of viable EBOV in the treated samples was assessed by carrying out three serial passages of the samples in Vero E6 cells (37°C, 5% CO 2 , 1 week for each passage). At the end of each passage the cells were observed for evidence of cytopathic effect and samples were taken for rRT-PCR analysis for the presence of EBOV RNA. Before cell culture cytotoxic components of AVL and Triton X-100 were removed from the samples using size exclusion spin column technology or a hydrophobic adsorbent resin. The results of this study showed that EBOV spiked into human serum was not fully inactivated when treated with either 0.1% (v/v) Triton X-100 for 10 mins or 1.0% (v/v) Triton X-100 for 20 mins (final concentrations 0.08% and 0.8% Triton X-100 respectively). AVL alone also did not consistently provide complete inactivation. Samples treated

  20. Insights into signal transduction by a hybrid FixL: Denaturation study of on and off states of a multi-domain oxygen sensor.

    Science.gov (United States)

    Guimarães, Wellinson G; Gondim, Ana C S; Costa, Pedro Mikael da Silva; Gilles-Gonzalez, Marie-Alda; Lopes, Luiz G F; Carepo, Marta S P; Sousa, Eduardo H S

    2017-07-01

    FixL from Rhizobium etli (ReFixL) is a hybrid oxygen sensor protein. Signal transduction in ReFixL is effected by a switch off of the kinase activity on binding of an oxygen molecule to ferrous heme iron in another domain. Cyanide can also inhibit the kinase activity upon binding to the heme iron in the ferric state. The unfolding by urea of the purified full-length ReFixL in both active pentacoordinate form, met-FixL(Fe III ) and inactive cyanomet-FixL (Fe III -CN - ) form was monitored by UV-visible absorption spectroscopy, circular dichroism (CD) and fluorescence spectroscopy. The CD and UV-visible absorption spectroscopy revealed two states during unfolding, whereas fluorescence spectroscopy identified a three-state unfolding mechanism. The unfolding mechanism was not altered for the active compared to the inactive state; however, differences in the ΔG H2O were observed. According to the CD results, compared to cyanomet-FixL, met-FixL was more stable towards chemical denaturation by urea (7.2 vs 4.8kJmol -1 ). By contrast, electronic spectroscopy monitoring of the Soret band showed cyanomet-FixL to be more stable than met-FixL (18.5 versus 36.2kJmol -1 ). For the three-state mechanism exhibited by fluorescence, the ΔG H2O for both denaturation steps were higher for the active-state met-FixL than for cyanomet-FixL. The overall stability of met-FixL is higher in comparison to cyanomet-FixL suggesting a more compact protein in the active form. Nonetheless, hydrogen bonding by bound cyanide in the inactive state promotes the stability of the heme domain. This work supports a model of signal transduction by FixL that is likely shared by other heme-based sensors. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Diagnostic work-up in cardiomyopathies

    DEFF Research Database (Denmark)

    Rapezzi, Claudio; Arbustini, Eloisa; Caforio, Alida L P

    2013-01-01

    a framework for the clinical approach to diagnosis in cardiomyopathies based on the recognition of diagnostic 'red flags' that can be used to guide rational selection of specialized tests including genetic analysis. The basic premise is that the adoption of a cardiomyopathy-specific mindset which combines...

  2. Evaluation of the antibody-detection ELISA using plates precoated with denatured T. congolense and T. vivax antigens for monitoring tsetse and trypanosomosis control in Zambia

    International Nuclear Information System (INIS)

    Machila, N.; Sinyangwe, L.; Eisler, M.C.

    2000-01-01

    An evaluation of the indirect enzyme-linked immunosorbent assay (ELISA) for the detection of anti-trypanosomal antibodies in bovine serum was conducted using ELISA plates pre-coated with denatured T. congolense and T. vivax antigen. The study was conducted on 280 samples from a parasitologically positive cattle population and 200 samples from a negative cattle population. The overwhelming majority of trypanosome infections in the parasitologically positive cattle were T. congolense. The Ab-ELISA was able to discriminate between the negative and positive cattle populations, at an optimum cut-off point of 40% positivity (T. congolense antigen), with 89% sensitivity and 86% specificity; and for the T. vivax antigen, the optimum cut-off point was at 30%, with 81% sensitivity and 70% specificity. However, the optical densities (OD) and percentage positivity (PP) values for sera from both the reference positive and negative cattle populations were unacceptably high particularly in the ELISA using T. congolense antigen. Furthermore the quality control sera used in the assay appear to have inappropriately low OD and PP values by comparison to typical sera from the reference positive and negative cattle populations. It is suggested that these ELISA's be re-titrated using more appropriate quality assurance sera. This should result in OD and PP values for sera from the reference populations failing within acceptable ranges. (author)

  3. Detection in a Japanese population of a length polymorphism in the 5' flanking region of the human β-globin gene with denaturing gradient gel electrophoresis

    International Nuclear Information System (INIS)

    Takahashi, Noria; Hiyama, Keiko; Kodaira, Mieko; Satoh, Chiyoko

    1992-10-01

    An analysis of the ATTTT repeat polymorphism located approximately 1,400 base pairs upstream from the β-globin structural gene was carried out by denaturing gradient gel electrophoresis (DGGE) of RNA:DNA duplexes. Genomic or cloned DNAs were digested with restriction enzymes and hybridized with 32 P-labeled RNA probes, and resulting RNA:DNA duplexes were examined by DGGE. A difference in the number of repeat units was recognized by differences in duplex mobility on the DGGE gel. In this study of 81 unrelated Japanese from Hiroshima, a sequence heteromorphism was observed at this site. Alleles with 5 and 6 repeats of the ATTTT unit, which had already been reported, were found in polymorphic proportions. In addition, two unreported alleles, one having 7 repeats and the other having an A-to-G nucleotide substitution in the 5th repeat, were detected. Family study data showed that the segregation of these four types of variants is consistent with an autosomal codominant mode of inheritance. This study also demonstrated that DGGE of RNA:DNA duplexes is a sensitive tool for detecting variations in DNA. (author)

  4. Abundance and Diversity of Hydrogenotrophic Microorganisms in the Infant Gut before the Weaning Period Assessed by Denaturing Gradient Gel Electrophoresis and Quantitative PCR

    Directory of Open Access Journals (Sweden)

    Valeria Sagheddu

    2017-06-01

    Full Text Available Delivery mode (natural vs. cesarean and feeding type (breast vs. formula feeding are relevant factors for neonatal gut colonization. Biomolecular methods have shown that the ecological structure of infant microbiota is more complex than previously proposed, suggesting a relevant presence of unculturable bacteria. It has also been postulated that among unculturable bacteria, hydrogenotrophic populations might play a key role in infant health. Sulfate-reducing bacteria (SRB, acetogens, and methanogenic archaea use hydrogenotrophic pathways within the human colon. However, to date, few studies have reported detection of hydrogenotrophic microorganisms in newborns, possibly because of limitations on available group-specific, culture-independent quantification procedures. In the present work, we analyzed 16 fecal samples of healthy babies aged 1–6 months by means of quantitative PCR (qPCR targeting the 16S rRNA or metabolic functional genes and by denaturing gradient gel electrophoresis (DGGE. qPCR data showed quantifiable levels of methanogens, SRB, and acetogens in all samples, indicating that the relative abundances of these microbial groups were not affected by delivery mode (natural vs. caesarian. DGGE revealed a high prevalence of the Blautia genus within the acetogenic bacteria despite strong interindividual variability. Our preliminary results suggest that hydrogenotrophic microorganisms, which have been a neglected group to date, should be included in future ecological and metabolic studies evaluating the infant intestinal microbiota.

  5. Behavior of Heat-Denatured Whey: Buttermilk Protein Aggregates during the Yogurt-Making Process and Their Influence on Set-Type Yogurt Properties

    Directory of Open Access Journals (Sweden)

    Maxime Saffon

    2013-09-01

    Full Text Available The objective of this study was to assess the impact of using heat-denatured whey:buttermilk protein aggregate in acid-set type yogurt production. Whey and buttermilk (25:75 protein concentrate was adjusted to pH 4.6, heated at 90 °C for 5 min, homogenized and freeze-dried. Set-type yogurts were prepared from skim milk standardized to 15% (w/v total solids and 4.2% (w/v protein using different levels of powdered skim milk or freeze-dried protein aggregate. The use of the protein aggregate significantly modified yogurt texture, but did not affect the water-holding capacity of the gel. Confocal laser-scanning microscope images showed the presence of large particles in milk enriched with protein aggregate, which directly affected the homogeneity of the clusters within the protein matrix. Thiol groups were freed during heating of the protein aggregate suspended in water, suggesting that the aggregates could interact with milk proteins during heating.

  6. Hand-held optical sensor using denatured antibody coated electro-active polymer for ultra-trace detection of copper in blood serum and environmental samples.

    Science.gov (United States)

    Chandra, Sutapa; Dhawangale, Arvind; Mukherji, Soumyo

    2018-07-01

    An optimum copper concentration in environment is highly desired for all forms of life. We have developed an ultrasensitive copper sensor which functions from femto to micro molar concentration accurately (R 2 = 0.98). The sensor is based on denatured antibody immunoglobulin G (IgG), immobilized on polyaniline (PAni) which in turn is the coating on the core of an optical fiber. The sensing relies on changes in evanescent wave absorbance in the presence of the analyte. The sensor showed excellent selectivity towards Cu (II) ions over all other metal ions. The sensor was tested with lake and marine water samples to determine unknown concentrations of copper ions and the recovery results were within 90-115%, indicating reasonable accuracy. We further integrated the fiber-optic sensor with a miniaturized hand-held instrumentation platform to develop an accurate and field deployable device which can broadly be applicable to determine Cu (II) concentration in a wide range of systems - natural water bodies, soil as well as blood serum. Copyright © 2018 Elsevier B.V. All rights reserved.

  7. Succession of bacterial and fungal communities during a traditional pot fermentation of rice vinegar assessed by PCR-mediated denaturing gradient gel electrophoresis.

    Science.gov (United States)

    Haruta, Shin; Ueno, Shintaro; Egawa, Isao; Hashiguchi, Kazunori; Fujii, Akira; Nagano, Masanobu; Ishii, Masaharu; Igarashi, Yasuo

    2006-05-25

    Denaturing gradient gel electrophoresis (DGGE) based on small subunit rRNA gene was applied to a traditional rice vinegar fermentation process in which the conversion of rice starch into acetic acid proceeded in a pot. The fungal DGGE profile indicated that the transition from Aspergillus oryzae to Saccharomyces sp. took place at the initial stage at which alcohol production was observed. The early stage was characterized by the coexistence of Saccharomyces sp. and lactic acid bacteria. Almost all of the bacterial DGGE bands related to lactic acid bacteria were replaced by bands derived from Lactobacillus acetotolerance and Acetobacter pasteurianus at the stage at which acetic acid started to accumulate. The microbial succession, tested in three different pots, was found to be essentially identical. Among the bacteria isolated at the early stage, some species differed from those detected by DGGE. This is the first report to reveal the microbial community succession that occurs during a unique vinegar fermentation process, as determined by a culture-independent method.

  8. Dynamics of Vaginal Bacterial Communities in Women Developing Bacterial Vaginosis, Candidiasis, or No Infection, Analyzed by PCR-Denaturing Gradient Gel Electrophoresis and Real-Time PCR▿

    Science.gov (United States)

    Vitali, Beatrice; Pugliese, Ciro; Biagi, Elena; Candela, Marco; Turroni, Silvia; Bellen, Gert; Donders, Gilbert G. G.; Brigidi, Patrizia

    2007-01-01

    The microbial flora of the vagina plays a major role in preventing genital infections, including bacterial vaginosis (BV) and candidiasis (CA). An integrated approach based on PCR-denaturing gradient gel electrophoresis (PCR-DGGE) and real-time PCR was used to study the structure and dynamics of bacterial communities in vaginal fluids of healthy women and patients developing BV and CA. Universal eubacterial primers and Lactobacillus genus-specific primers, both targeted at 16S rRNA genes, were used in DGGE and real-time PCR analysis, respectively. The DGGE profiles revealed that the vaginal flora was dominated by Lactobacillus species under healthy conditions, whereas several potentially pathogenic bacteria were present in the flora of women with BV. Lactobacilli were the predominant bacterial population in the vagina for patients affected by CA, but changes in the composition of Lactobacillus species were observed. Real-time PCR analysis allowed the quantitative estimation of variations in lactobacilli associated with BV and CA diseases. A statistically significant decrease in the relative abundance of lactobacilli was found in vaginal fluids of patients with BV compared to the relative abundance of lactobacilli in the vaginal fluids of healthy women and patients with CA. PMID:17644631

  9. Dynamics of vaginal bacterial communities in women developing bacterial vaginosis, candidiasis, or no infection, analyzed by PCR-denaturing gradient gel electrophoresis and real-time PCR.

    Science.gov (United States)

    Vitali, Beatrice; Pugliese, Ciro; Biagi, Elena; Candela, Marco; Turroni, Silvia; Bellen, Gert; Donders, Gilbert G G; Brigidi, Patrizia

    2007-09-01

    The microbial flora of the vagina plays a major role in preventing genital infections, including bacterial vaginosis (BV) and candidiasis (CA). An integrated approach based on PCR-denaturing gradient gel electrophoresis (PCR-DGGE) and real-time PCR was used to study the structure and dynamics of bacterial communities in vaginal fluids of healthy women and patients developing BV and CA. Universal eubacterial primers and Lactobacillus genus-specific primers, both targeted at 16S rRNA genes, were used in DGGE and real-time PCR analysis, respectively. The DGGE profiles revealed that the vaginal flora was dominated by Lactobacillus species under healthy conditions, whereas several potentially pathogenic bacteria were present in the flora of women with BV. Lactobacilli were the predominant bacterial population in the vagina for patients affected by CA, but changes in the composition of Lactobacillus species were observed. Real-time PCR analysis allowed the quantitative estimation of variations in lactobacilli associated with BV and CA diseases. A statistically significant decrease in the relative abundance of lactobacilli was found in vaginal fluids of patients with BV compared to the relative abundance of lactobacilli in the vaginal fluids of healthy women and patients with CA.

  10. Bacterial Diversity in the Digestive Tracts of Four Indian Air-Breathing Fish Species Investigated by PCR Based Denaturing Gradient Gel Electrophoresis

    Directory of Open Access Journals (Sweden)

    Suxu He

    Full Text Available ABSTRACT An investigation was conducted to identify the allochthonous microbiota (entire intestine and the autochthonous microbiota in proximal intestine (PI and distal intestine (DI of four species of Indian air-breathing fish (climbing perch; Anabas testudineus, murrel; Channa punctatus, walking catfish; Clarias batrachus and stinging catfish; Heteropneustes fossilis by PCR based denaturing gradient gel electrophoresis (DGGE. High similarities of the allochthonous microbiota were observed between climbing perch and murrel, walking catfish and stinging catfish, indicating similar food behavior. The autochthonous microbiota of PI and DI from climbing perch and murrel revealed more similarity, than the result obtained from walking catfish and stinging catfish. The autochthonous microbiota of climbing perch and murrel were similar with regard to the allochthonous microbiota, but no such similarity was observed in case of walking catfish and stinging catfish. The fish genotype and intestinal bacteria are well matched and show co-evolutionary relationship. Three fish species has its unique bacteria; autochthonous Enterobacter cloacae, Edwardsiella tarda and Sphingobium sp. in DI of climbing perch, Pseudomonas sp.; allochthonous and autochthonous in PI of walking catfish and uncultured bacterium (EU697160.1, uncultured bacterium (JF018065.1 and uncultured bacterium (EU697160.1 for stinging catfish. In murrel, no unique bacteria were detected.

  11. Efficient detection of factor IX mutations by denaturing high-performance liquid chromatography in Taiwanese hemophilia B patients, and the identification of two novel mutations

    Directory of Open Access Journals (Sweden)

    Pei-Chin Lin

    2014-04-01

    Full Text Available Hemophilia B (HB is an X-linked recessive disorder characterized by mutations in the clotting factor IX (FIX gene that result in FIX deficiency. Previous studies have shown a wide variation of FIX gene mutations in HB. Although the quality of life in HB has greatly improved mainly because of prophylactic replacement therapy with FIX concentrates, there exists a significant burden on affected families and the medical care system. Accurate detection of FIX gene mutations is critical for genetic counseling and disease prevention in HB. In this study, we used denaturing high-performance liquid chromatography (DHPLC, which has proved to be a highly informative and practical means of detecting mutations, for the molecular diagnosis of our patients with HB. Ten Taiwanese families affected by HB were enrolled. We used the DHPLC technique followed by direct sequencing of suspected segments to detect FIX gene mutations. In all, 11 FIX gene mutations (8 point mutations, 2 small deletions/insertions, and 1 large deletion, including two novel mutations (exon6 c.687–695, del 9 mer and c.460–461, ins T were found. According to the HB pedigrees, 25% and 75% of our patients were defined as familial and sporadic HB cases, respectively. We show that DHPLC is a highly sensitive and cost-effective method for FIX gene analysis and can be used as a convenient system for disease prevention.

  12. Chromatin structure influence of DNA damage measurements by four assays: pulsed- and constant-field gel electrophoresis, DNA precipitation and non-denaturing filter elution

    International Nuclear Information System (INIS)

    Wlodek, D.; Olive, P.L.

    1996-01-01

    The of elution of DNA during non-denaturing filter elution (NFE) often correlates with cell sensitivity to radiation. The elution rate is influenced by two cellular factors: chromatin structure and the number of DNA-strand breaks (DSBs) produced in an intact cell by ionizing radiation. To determine which of the above factors is relevant to cell radiosensitivity, four assays were used to measure induction of DNA damage in three cell lines varying in radiosensitivity (V79, CHO, and L5178Y-R). Each of the assays, neutral filter elution (NFE), DNA precipitation, constant (CFGE) and pulsed field gel electrophoresis (PFGE) have different physical basis for DNA damage measurement and might be differently affected by chromatin structure. Three of the methods used to measure DNA double-strand breaks gave different results: NFE was dependent on cell type and location of DNA relative to the replication fork, gel electrophoresis was independent of cell type but was affected by proximity to the replication fork, and the precipitation assay was independent of both cell type and replication status. Pulsed field gel electrophoresis produced the same results and constant field gel electrophoresis for 3 cell lines examined. Only NFE showed differences in sensitivity which correlated with cell survival following irradiation. The results suggest that three is the same initial amount of DSBs in cells from all three lines and that the sensitivity to radiation is determined by some additional factors, probably chromatin structure. (author). 18 refs, 5 figs

  13. Bacterial community structure in Apis florea larvae analyzed by denaturing gradient gel electrophoresis and 16S rRNA gene sequencing.

    Science.gov (United States)

    Saraithong, Prakaimuk; Li, Yihong; Saenphet, Kanokporn; Chen, Zhou; Chantawannakul, Panuwan

    2015-10-01

    This study characterizes the colonization and composition of bacterial flora in dwarf Asian honeybee (Apis florea) larvae and compares bacterial diversity and distribution among different sampling locations. A. florea larvae were collected from 3 locations in Chiang Mai province, Thailand. Bacterial DNA was extracted from each larva using the phenol-chloroform method. Denaturing gradient gel electrophoresis was performed, and the dominant bands were excised from the gels, cloned, and sequenced for bacterial species identification. The result revealed similarities of bacterial community profiles in each individual colony, but differences between colonies from the same and different locations. A. florea larvae harbor bacteria belonging to 2 phyla (Firmicutes and Proteobacteria), 5 classes (Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Bacilli, and Clostridia), 6 genera (Clostridium, Gilliamella, Melissococcus, Lactobacillus, Saccharibacter, and Snodgrassella), and an unknown genus from uncultured bacterial species. The classes with the highest abundance of bacteria were Alphaproteobacteria (34%), Bacilli (25%), Betaproteobacteria (11%), Gammaproteobacteria (10%), and Clostridia (8%), respectively. Similarly, uncultured bacterial species were identified (12%). Environmental bacterial species, such as Saccharibacter floricola, were also found. This is the first study in which sequences closely related to Melissococcus plutonius, the causal pathogen responsible for European foulbrood, have been identified in Thai A. florea larvae. © 2014 Institute of Zoology, Chinese Academy of Sciences.

  14. Parallel characterization of anaerobic toluene- and ethylbenzene-degrading microbial consortia by PCR-denaturing gradient gel electrophoresis, RNA-DNA membrane hybridization, and DNA microarray technology

    Science.gov (United States)

    Koizumi, Yoshikazu; Kelly, John J.; Nakagawa, Tatsunori; Urakawa, Hidetoshi; El-Fantroussi, Said; Al-Muzaini, Saleh; Fukui, Manabu; Urushigawa, Yoshikuni; Stahl, David A.

    2002-01-01

    A mesophilic toluene-degrading consortium (TDC) and an ethylbenzene-degrading consortium (EDC) were established under sulfate-reducing conditions. These consortia were first characterized by denaturing gradient gel electrophoresis (DGGE) fingerprinting of PCR-amplified 16S rRNA gene fragments, followed by sequencing. The sequences of the major bands (T-1 and E-2) belonging to TDC and EDC, respectively, were affiliated with the family Desulfobacteriaceae. Another major band from EDC (E-1) was related to an uncultured non-sulfate-reducing soil bacterium. Oligonucleotide probes specific for the 16S rRNAs of target organisms corresponding to T-1, E-1, and E-2 were designed, and hybridization conditions were optimized for two analytical formats, membrane and DNA microarray hybridization. Both formats were used to characterize the TDC and EDC, and the results of both were consistent with DGGE analysis. In order to assess the utility of the microarray format for analysis of environmental samples, oil-contaminated sediments from the coast of Kuwait were analyzed. The DNA microarray successfully detected bacterial nucleic acids from these samples, but probes targeting specific groups of sulfate-reducing bacteria did not give positive signals. The results of this study demonstrate the limitations and the potential utility of DNA microarrays for microbial community analysis.

  15. Comparative effectiveness of coronary CT angiography vs stress cardiac imaging in patients following hospital admission for chest pain work-up: The Prospective First Evaluation in Chest Pain (PERFECT) Trial.

    Science.gov (United States)

    Uretsky, Seth; Argulian, Edgar; Supariwala, Azhar; Agarwal, Shiv K; El-Hayek, Georges; Chavez, Patricia; Awan, Hira; Jagarlamudi, Ashadevi; Puppala, Siva P; Cohen, Randy; Rozanski, Alan

    2017-08-01

    Because the frequency of cardiac event rates is low among chest pain patients following either performance of coronary CT angiography (CCTA) or stress testing, there is a need to better assess how these tests influence the central management decisions that follow from cardiac testing. The present study was performed to assess the relative impact of CCTA vs stress testing on medical therapies and downstream resource utilization among patients admitted for the work-up of chest pain. The admitted patients were randomized in a 1:1 ratio to either cardiac imaging stress test or CCTA. Primary outcomes were time to discharge, change in medication usage, and frequency of downstream testing, cardiac interventions, and cardiovascular re-hospitalizations. We randomized 411 patients, 205 to stress testing, and 206 to CCTA. There were no differences in time to discharge or initiation of new cardiac medications at discharge. At 1 year follow-up, there was no difference in the number of patients who underwent cardiovascular downstream tests in the CCTA vs stress test patients (21% vs 15%, P = .1) or cardiovascular hospitalizations (14% vs 16%, P = .5). However, there was a higher frequency of invasive angiography in the CCTA group (11% vs 2%, P = .001) and percutaneous coronary interventions (6% vs 0%, P work-up to either CCTA or to stress testing resulted in similar discharge times, change in medical therapies at discharge, frequency of downstream noninvasive testing, and repeat hospitalizations. However, a higher frequency of invasive coronary angiography and revascularization procedures were performed in the CCTA arm. (ClinicalTrials.gov number, NCT01604655.).

  16. Preoperative workup to assess indication for laparoscopic treatment in gastroesophageal reflux disease Aseguramiento preoperatorio en la indicación de tratamiento laparoscópico en la enfermedad por reflujo gastroesofágico

    Directory of Open Access Journals (Sweden)

    S. Pérez-Holanda

    2008-07-01

    Full Text Available Introduction and objectives: antireflux surgery performed by an experienced surgeon is a maintenance option for patients with well-documented gastroesophageal reflux disease (GERD. Well-documented GERD is difficult to find, as GERD is a multifactorial disease in which the gastroesophageal junction, with its special anatomical and functional components, is important. In order to examine patient preoperative workups, and their indication for surgical treatment in GERD, we retrospectively studied patients who underwent a laparoscopic antireflux procedure. Methods: preoperative workups in patients from our health care area who underwent a laparoscopic antireflux procedure from December 1997 to February 2007 were retrospectively analyzed. Data related to epidemiological findings, symptoms, morphologic and functional evaluation, medical therapy, and indication for surgical treatment were recorded and statistically analyzed by means of a bivariate test. Differences were significant when the p value was equal to or less than 0.05. Results: 100 patients (50 % female, 51.31 ± 13.53 years of age underwent a laparoscopic antireflux surgery after 56.47 ± 61.33 months with symptoms. Ninety-five percent of patients had an anatomical abnormality. The pH monitoring test diagnosed three quarters of cases. The most frequent indication for GERD treatment was persistent or recurrent esophagitis despite adequate medical treatment (52 cases. Conclusions: based on our preoperative workup, as described, 100 percent of subjects were well documented and diagnosed with GERD (both non-erosive reflux disease and erosive reflux disease, and their indication for laparoscopic treatment was retrospectively assessed in 94% of cases.Introducción y objetivos: la cirugía antirreflujo realizada por un cirujano experto es una opción para el tratamiento de mantenimiento para el paciente con enfermedad por reflujo gastroesofágico (ERGE bien documentada. La "buena documentaci

  17. 15NH/D-SOLEXSY experiment for accurate measurement of amide solvent exchange rates: application to denatured drkN SH3

    International Nuclear Information System (INIS)

    Chevelkov, Veniamin; Xue, Yi; Krishna Rao, D.; Forman-Kay, Julie D.; Skrynnikov, Nikolai R.

    2010-01-01

    Amide solvent exchange rates are regarded as a valuable source of information on structure/dynamics of unfolded (disordered) proteins. Proton-based saturation transfer experiments, normally used to measure solvent exchange, are known to meet some serious difficulties. The problems mainly arise from the need to (1) manipulate water magnetization and (2) discriminate between multiple magnetization transfer pathways that occur within the proton pool. Some of these issues are specific to unfolded proteins. For example, the compensation scheme used to cancel the Overhauser effect in the popular CLEANEX experiment is not designed for use with unfolded proteins. In this report we describe an alternative experimental strategy, where amide 15 N is used as a probe of solvent exchange. The experiment is performed in 50% H 2 O-50% D 2 O solvent and is based on the (HACACO)NH pulse sequence. The resulting spectral map is fully equivalent to the conventional HSQC. To fulfill its purpose, the experiment monitors the conversion of deuterated species, 15 N D , into protonated species, 15 N H , as effected by the solvent exchange. Conceptually, this experiment is similar to EXSY which prompted the name of 15 N H/D -SOLEXSY (SOLvent EXchange SpectroscopY). Of note, our experimental scheme, which relies on nitrogen rather than proton to monitor solvent exchange, is free of the complications described above. The developed pulse sequence was used to measure solvent exchange rates in the chemically denatured state of the drkN SH3 domain. The results were found to correlate well with the CLEANEX-PM data, r = 0.97, thus providing a measure of validation for both techniques. When the experimentally measured exchange rates are converted into protection factors, most of the values fall in the range 0.5-2, consistent with random-coil behavior. However, elevated values, ca. 5, are obtained for residues R38 and A39, as well as the side-chain indole of W36. This is surprising, given that high

  18. Denaturing Gradient Gel Electrophoresis-Polymerase Chain Reaction Comparison of Chitosan Effects on Anaerobic Cultures of Broiler Cecal Bacteria and Salmonella Typhimurium.

    Science.gov (United States)

    Hume, Michael; Sohail, Muhammad Umar

    2018-04-01

    Enteropathogen colonization and product contamination are major poultry industry problems. The emergence of antibiotic resistance, and associated risks to human health, is limiting the use of antibiotics as first-line defense against enteropathogens in poultry. The chitin derivative, chitosan, has drawn substantial attention for its bactericidal properties. Different molecular weight (MW) chitosans can have varied effects against different bacteria in monoculture. In the current study, cecal contents from each of three market-age broilers and Salmonella Typhimurium, as indicator enteropathogen, were exposed to in vitro anaerobic culture to three chitosan preparations (0.08%, wt/vol), low (LMW), medium (MMW), and coarse (CMW). Effects of chitosan and the carrier solvent acetic acid, on cecal bacteria and Salmonella, were examined by denaturing gradient gel electrophoresis (DGGE) and Salmonella enumeration. Bacterial profiles for the three cecal contents were shown by DGGE to be very different. Each of the three cecal contents grown in the presence of 0.08% acetic acid was very different from the same contents grown without the chitosan solvent. Culturing cecal contents in the presence of chitosan altered the bacterial DGGE profiles from the control and acetic acid-only cultures. The DGGE chitosan-treated profiles for all three cecal sources were identical to each other regardless of the MW chitosan in the culture medium. Compared with Salmonella in monoculture, Salmonella decreased (p < 0.05) by about 1.5 log CFU/mL when grown in mixed culture with cecal contents. Salmonella monocultures in the presence of 0.08% of the chitosan solvent acetic acid decreased (p < 0.05) counts by almost 3.5 log CFU/mL. Combining acetic acid and cecal contents reduced (p < 0.05) Salmonella by 7 log CFU/mL. Adding the chitosan preparations to the mixtures reduced (p < 0.05) Salmonella by 8 log CFU/mL.

  19. Improving the reversibility of thermal denaturation and catalytic efficiency of Bacillus licheniformis α-amylase through stabilizing a long loop in domain B.

    Directory of Open Access Journals (Sweden)

    Zhu Li

    Full Text Available The reversibility of thermal denaturation and catalytic efficiency of Bacillus licheniformis α-amylase were improved through site-directed mutagenesis. By using multiple sequence alignment and PoPMuSiC algorithm, Ser187 and Asn188, which located within a long loop in Domain B of Bacillus licheniformis α-amylase, were selected for mutation. In addition, Ala269, which is adjacent to Ser187 and Asn188, was also investigated. Seven mutants carrying the mutations S187D, N188T, N188S, A269K, A269K/S187D, S187D/N188T, and A269K/S187D/N188T were generated and characterized. The most thermostable mutant, A269K/S187D/N188T, exhibited a 9-fold improvement in half-life at 95°C and pH 5.5, compared with that of the wild-type enzyme. Mutant A269K/S187D/N188T also exhibited improved catalytic efficiency. The catalytic efficiency of mutant A269K/S187D/N188T reached 5.87×103±0.17 g·L-1·s-1 at pH 5.5, which is 1.84-fold larger than the corresponding value determined for the wild-type enzyme. Furthermore, the structure analysis showed that immobilization of the loop containing Ser187 and Asn188 plays a significant role in developing the properties of Bacillus licheniformis α-amylase.

  20. Improving the reversibility of thermal denaturation and catalytic efficiency of Bacillus licheniformis α-amylase through stabilizing a long loop in domain B.

    Science.gov (United States)

    Li, Zhu; Duan, Xuguo; Chen, Sheng; Wu, Jing

    2017-01-01

    The reversibility of thermal denaturation and catalytic efficiency of Bacillus licheniformis α-amylase were improved through site-directed mutagenesis. By using multiple sequence alignment and PoPMuSiC algorithm, Ser187 and Asn188, which located within a long loop in Domain B of Bacillus licheniformis α-amylase, were selected for mutation. In addition, Ala269, which is adjacent to Ser187 and Asn188, was also investigated. Seven mutants carrying the mutations S187D, N188T, N188S, A269K, A269K/S187D, S187D/N188T, and A269K/S187D/N188T were generated and characterized. The most thermostable mutant, A269K/S187D/N188T, exhibited a 9-fold improvement in half-life at 95°C and pH 5.5, compared with that of the wild-type enzyme. Mutant A269K/S187D/N188T also exhibited improved catalytic efficiency. The catalytic efficiency of mutant A269K/S187D/N188T reached 5.87×103±0.17 g·L-1·s-1 at pH 5.5, which is 1.84-fold larger than the corresponding value determined for the wild-type enzyme. Furthermore, the structure analysis showed that immobilization of the loop containing Ser187 and Asn188 plays a significant role in developing the properties of Bacillus licheniformis α-amylase.

  1. Application of PCR-denaturing-gradient gel electrophoresis (DGGE) method to examine microbial community structure in asparagus fields with growth inhibition due to continuous cropping.

    Science.gov (United States)

    Urashima, Yasufumi; Sonoda, Takahiro; Fujita, Yuko; Uragami, Atsuko

    2012-01-01

    Growth inhibition due to continuous cropping of asparagus is a major problem; the yield of asparagus in replanted fields is low compared to that in new fields, and missing plants occur among young seedlings. Although soil-borne disease and allelochemicals are considered to be involved in this effect, this is still controversial. We aimed to develop a technique for the biological field diagnosis of growth inhibition due to continuous cropping. Therefore, in this study, fungal community structure and Fusarium community structure in continuously cropped fields of asparagus were analyzed by polymerase chain reaction/denaturing-gradient gel electrophoresis (PCR-DGGE). Soil samples were collected from the Aizu region of Fukushima Prefecture, Japan. Soil samples were taken from both continuously cropped fields of asparagus with growth inhibition and healthy neighboring fields of asparagus. The soil samples were collected from the fields of 5 sets in 2008 and 4 sets in 2009. We were able to distinguish between pathogenic and non-pathogenic Fusarium by using Alfie1 and Alfie2GC as the second PCR primers and PCR-DGGE. Fungal community structure was not greatly involved in the growth inhibition of asparagus due to continuous cropping. By contrast, the band ratios of Fusarium oxysporum f. sp. asparagi in growth-inhibited fields were higher than those in neighboring healthy fields. In addition, there was a positive correlation between the band ratios of Fusarium oxysporum f. sp. asparagi and the ratios of missing asparagus plants. We showed the potential of biological field diagnosis of growth inhibition due to continuous cropping of asparagus using PCR-DGGE.

  2. Identification and characterization of single nucleotide polymorphisms in 6 growth-correlated genes in porcine by denaturing high performance liquid chromatography.

    Science.gov (United States)

    Liu, Dewu; Zhang, Yushan; Du, Yinjun; Yang, Guanfu; Zhang, Xiquan

    2007-06-01

    The growth-correlated genes that are part of the neuroendocrine growth axis play crucial roles in the regulation of growth and development of pig. The identification of genetic polymorphisms in these genes will enable the scientist to evaluate the biological relevance of such polymorphisms and to gain a better understanding of quantitative traits like growth. In the present study, seven pairs of primers were designed to obtain unknown sequences of growth-correlated genes, and other 25 pairs of primers were designed to identify single nucleotide polymorphisms (SNP) using the denaturing high-performance liquid chromatography (DHPLC) technology in four pig breeds (Duroc, Landrace, Lantang and Wuzhishan), significantly differing in growth and development characteristics. A total of 101 polymorphisms were discovered in 10,707 base pairs (bp) from six genes of the ghrelin (GHRL), leptin (LEP), insulin-like growth factor II (IGF-II), insulin-like growth factor binding protein 2 (IGFBP-2), insulin-like growth factor binding protein 3 (IGFBP-3), and somatostatin (SS). The observed average distances between the SNP in the 5'UTR, coding regions, introns and 3'UTR were 134, 521, 81 and 92 bp, respectively. Four SNPs were found in the coding regions of IGF-II, IGFBP-2 and LEP, respectively. Two synonymous mutations were obtained in IGF-II and LEP genes respectively, and two non-synonymous were found in IGFBP-2 and LEP genes, respectively. Seven other mutations were also observed. Thirty-two PCR-RFLP markers were found among 101 polymorphisms of the six genes. The SNP discovered in this study would provide suitable markers for association studies of candidate genes with growth related traits in pig.

  3. [Effects of redox state of disulfide bonds on the intrinsic fluorescence and denaturation of Trx-fused gibberellin-induced cysteine-rich protein from Gymnadnia conopsea].

    Science.gov (United States)

    Zhang, Teng; Feng, Juan; Li, Yang; Chen, Rui; Tang, Li-Xia; Pang, Xiao-Feng; Ren, Zheng-Long

    2010-02-01

    In the present paper, thioredoxin-fused gibberellin-induced cysteine-rich protein from Gymnadnia conopsea, desigated as Trx-GcGASA and expressed prokaryotically, was purified and identified by using Ni(2+) -NTA affinity chromatography column and SDS-PAGE, and then its intrinsic fluorescence was investigated in the absence and presence of dithiothreitol (DTT), oxidized glutathione (GSSG), peroxide and guanidine hydrochloride (GdnHCl) by means of steady-state fluorescence spectroscopic methods. It was found that (1) at the neutral pH Trx-GcGASA had maximum fluorescence emission at 305 nm following excitation at different wavelengths varying from 250 to 280 nm, which was ascribed to the fluorescence emission from tyrosine residues. (2) The reduction of disulphide bonds lead to the changes in the relative fluorescence intensity between tyrosine and tryptophan residues from 0.7 to 1.8. (3) Both Tyr and Trp residues underwent 12%-21% decrease in fluorescence intensity with the addition of 0.5 mmol x L(-1) GSSG or 5 mmol x L(-1) peroxide. The latter was roughly consistent with the antioxidative activity reported in vivo. (4) No matter whether 1 mmol x L(-1) DTT was absent or present, the fusion protein could not be fully unfolded with lambda(max) Trx-GcGASA experienced GdnHCl-induced denaturation process, and the unfolding equilibrium curve could be well fitted by using two-state model, giving the Gibbs free energy change (deltaG) of 3.7 kJ x mol(-1). However, it was not the case for reduced Trx-GcGASA protein. The aforementioned experimental results will not only provide some guides to investigate the effects of fusion partner Trx on the unfolding thermodynamics, kinetics and refolding process of Trx-GcGASA, but also will be useful for further studies on the strucuture of GA-induced cysteine-rich protein with the help of spectroscopic methods.

  4. Use of pyrosequencing and denaturing gradient gel electrophoresis to examine the effects of probiotics and essential oil blends on digestive microflora in broilers under mixed Eimeria infection.

    Science.gov (United States)

    Hume, Michael E; Barbosa, Nei A; Dowd, Scot E; Sakomura, Nilva K; Nalian, Armen G; Martynova-Van Kley, Alexandra; Oviedo-Rondón, Edgar O

    2011-11-01

    A protective digestive microflora helps prevent and reduce broiler infection and colonization by enteropathogens. In the current experiment, broilers fed diets supplemented with probiotics and essential oil (EO) blends were infected with a standard mixed Eimeria spp. to determine effects of performance enhancers on ileal and cecal microbial communities (MCs). Eight treatment groups included four controls (uninfected-unmedicated [UU], unmedicated-infected, the antibiotic BMD plus the ionophore Coban as positive control, and the ionophore as negative control), and four treatments (probiotics BC-30 and Calsporin; and EO, Crina Poultry Plus, and Crina PoultryAF). Day-old broilers were raised to 14 days in floor pens on used litter and then were moved to Petersime batteries and inoculated at 15 days with mixed Eimeria spp. Ileal and cecal samples were collected at 14 days and 7 days postinfection. Digesta DNA was subjected to pyrosequencing for sequencing of individual cecal bacteria and denaturing gradient gel electrophoresis (DGGE) for determination of changes in ileal and cecal MC according to percentage similarity coefficient (%SC). Pyrosequencing is very sensitive detecting shifts in individual bacterial sequences, whereas DGGE is able to detect gross shifts in entire MC. These combined techniques offer versatility toward identifying feed additive and mild Eimeria infection modulation of broiler MC. Pyrosequencing detected 147 bacterial species sequences. Additionally, pyrosequencing revealed the presence of relatively low levels of the potential human enteropathogens Campylobacter sp. and four Shigella spp. as well as the potential poultry pathogen Clostridiun perfringens. Pre- and postinfection changes in ileal (56%SC) and cecal (78.5%SC) DGGE profiles resulted from the coccidia infection and with increased broiler age. Probiotics and EO changed MC from those seen in UU ilea and ceca. Results potentially reflect the performance enhancement above expectations in

  5. Comparison of automated ribosomal intergenic spacer analysis (ARISA) and denaturing gradient gel electrophoresis (DGGE) techniques for analysing the influence of diet on ruminal bacterial diversity.

    Science.gov (United States)

    Saro, Cristina; Molina-Alcaide, Eduarda; Abecia, Leticia; Ranilla, María José; Carro, María Dolores

    2018-04-01

    The objective of this study was to compare the automated ribosomal intergenic spacer analysis (ARISA) and the denaturing gradient gel electrophoresis (DGGE) techniques for analysing the effects of diet on diversity in bacterial pellets isolated from the liquid (liquid-associated bacteria (LAB)) and solid (solid-associated bacteria (SAB)) phase of the rumen. The four experimental diets contained forage to concentrate ratios of 70:30 or 30:70 and had either alfalfa hay or grass hay as forage. Four rumen-fistulated animals (two sheep and two goats) received the diets in a Latin square design. Bacterial pellets (LAB and SAB) were isolated at 2 h post-feeding for DNA extraction and analysed by ARISA and DGGE. The number of peaks in individual samples ranged from 48 to 99 for LAB and from 41 to 95 for SAB with ARISA, and values of DGGE-bands ranged from 27 to 50 for LAB and from 18 to 45 for SAB. The LAB samples from high concentrate-fed animals tended (p forage-fed animals with ARISA, but no differences were identified with DGGE. The SAB samples from high concentrate-fed animals had lower (p forage diets with ARISA, but only a trend was noticed for these parameters with DGGE (p forage type on LAB diversity was detected by any technique. In this study, ARISA detected some changes in ruminal bacterial communities that were not detected by DGGE, and therefore ARISA was considered more appropriate for assessing bacterial diversity of ruminal bacterial pellets. The results highlight the impact of the fingerprinting technique used to draw conclusions on dietary factors affecting bacterial diversity in ruminal bacterial pellets.

  6. 18F-Fluorocholine PET/CT Complementing the Role of Dynamic Contrast-Enhanced MRI for Providing Comprehensive Diagnostic Workup in Prostate Cancer Patients With Suspected Relapse Following Radical Prostatectomy.

    Science.gov (United States)

    Vadi, Shelvin Kumar; Singh, Baljinder; Basher, Rajender K; Watts, Ankit; Sood, Ashwani K; Lal, Anupam; Kakkar, Nandita; Singh, S K

    2017-08-01

    The aim of this study was to compare the diagnostic performance of F-fluorocholine (FCH) PET/CT and dynamic contrast-enhanced MRI (DCE-MRI) of pelvis in restaging prostate cancer (PC) patients with biochemical recurrence (BCR) following radical prostatectomy (RP). Twenty PC patients who had undergone RP and had BCR were recruited in this study. All the patients underwent whole-body FCH PET/CT and DCE-MRI of the pelvis. An overall pattern of recurrent disease was analyzed, and diagnostic accuracy for the detection of pelvic disease recurrence by the 2 modalities was evaluated by taking histopathologic analysis as the criterion standard. The whole-body FCH PET/CT images were also analyzed separately for the presence of any extra lesion(s). The initial mean Gleason score was 6.3 ± 1.53 (range, 4-9). The mean prostate-specific antigen levels at the time of relapse were 1.9 ± 2.87 ng/mL (range, 0.24-13.2 ng/mL). MRI findings were positive for primary tumor recurrence in the prostate bed in 6 patients (6/20 [30.0%]), pelvic lymph node metastases in 4 patients (4/20 [20.0%]), and for pelvic skeletal metastases in 2 patients (2/20 [10.0%]), respectively. On the other hand, FCH PET/CT results were positive in the corresponding sites in 7 (7/20 [35.0%]), 9 (9/20 [45.0%]), and 2 patients (2/20 [10.0%]), respectively. F-fluorocholine PET/CT and MRI showed comparable results in terms of sensitivity, specificity, and positive and negative predictive values for PC characterization. The whole-body FCH PET/CT was found to be useful in identifying unknown distant metastases in a significant proportion of patients. The correlative whole-body FCH PET/CT and pelvic DCE-MRI offer a complementary and comprehensive diagnostic workup for better management of PC patients with BCR following RP.

  7. Residue-specific description of non-native transient structures in the ensemble of acid-denatured structures of the all-beta protein c-src SH3

    DEFF Research Database (Denmark)

    Rösner, Heike I; Poulsen, Flemming Martin

    2010-01-01

    -src loop to the third beta-strand, exhibited an apparent helicity of nearly 45%. Furthermore, the RT loop and the diverging turn appeared to adopt non-native-like helical conformations. Interestingly, none of the residues found in transient helical conformations exhibited significant varphi-values [Riddle......Secondary chemical shift analysis has been used to characterize the unfolded state of acid-denatured c-src SH3. Even though native c-src SH3 adopts an all-beta fold, we found evidence of transient helicity in regions corresponding to native loops. In particular, residues 40-46, connecting the n...

  8. {sup 99} {sup m}Tc-sulphur-colloid and heat-denatured {sup 99} {sup m}Tc-labelled red cell scans demonstrating a giant intrapelvic spleen in a girl after splenectomy

    Energy Technology Data Exchange (ETDEWEB)

    Kao, P.F. [Dept. of Nuclear Medicine, Chang Gung Memorial Hospital and Chang Gung University School of Medicine, Tauyuan, Taiwan (Taiwan); Dept. of Nuclear Medicine, Chang Gung Memorial Hospital, Taipei, Taiwan (Taiwan); Tzen, K.Y.; Tsai, M.F. [Dept. of Nuclear Medicine, Chang Gung Memorial Hospital and Chang Gung University School of Medicine, Tauyuan, Taiwan (Taiwan); Lin, J.N. [Dept. of Paediatric Surgery, Chang Gung Childrens Hospital and Chang Gung University School of Medicine, Tauyuan, Taiwan (Taiwan)

    2001-04-01

    A 17 x 12 x 5-cm giant intrapelvic mass in a 14-year-old girl is reported. This mass developed 6 years after a splenectomy for splenic torsion. The heat-denatured {sup 99} {sup m}Tc-labelled red cell scan and {sup 99} {sup m}Tc- sulphur-colloid scan confirmed the specific red cell sequestration function and reticuloendothelial activity in the giant intrapelvic spleen. The size and development of the giant intrapelvic spleen are unusual. The usefulness of functional images to diagnosis the nature of the intrapelvic mass is well demonstrated. (orig.)

  9. Evaluation of different PCR primers for denaturing gradient gel electrophoresis (DGGE) analysis of fungal community structure in traditional fermentation starters used for Hong Qu glutinous rice wine.

    Science.gov (United States)

    Lv, Xu-Cong; Jiang, Ya-Jun; Liu, Jie; Guo, Wei-Ling; Liu, Zhi-Bin; Zhang, Wen; Rao, Ping-Fan; Ni, Li

    2017-08-16

    Denaturing gradient gel electrophoresis (DGGE) has become a widely used tool to examine microbial community structure. However, when DGGE is applied to evaluate the fungal community of traditional fermentation starters, the choice of hypervariable ribosomal RNA gene regions is still controversial. In the current study, several previously published fungal PCR primer sets were compared and evaluated using PCR-DGGE, with the purpose of screening a suitable primer set to study the fungal community of traditional fermentation starters for Hong Qu glutinous rice wine. Firstly, different primer sets were used to amplify different hypervariable regions from pure fungal cultures. Except NS1/FR1+ and ITS1fGC/ITS4, other primer sets (NL1+/LS2R, NL3A/NL4GC, FF390/FR1+, NS1/GCFung, NS3+/YM951r and ITS1fGC/ITS2r) amplified the target DNA sequences successfully. Secondly, the selected primer sets were further evaluated based on their resolution to distinguish different fungal cultures through DGGE fingerprints. Three primer sets (NL1+/LS2R, NS1/GCFung and ITS1fGC/ITS2r) were finally selected for investigating the fungal community structure of different traditional fermentation starters for Hong Qu glutinous rice wine. The internal transcribed spacer (ITS) region amplified by ITS1fGC/ITS2r, which is more hypervariable than the 18S rRNA gene and 26S rRNA gene, provides an excellent tool to separate amplification products of different fungal species. Results indicated that PCR-DGGE profile using ITS1fGC/ITS2r showed more abundant fungal species than that using NL1+/LS2R and NS1/GCFung. Therefore, ITS1fGC/ITS2r is the most suitable primer set for PCR-DGGE analysis of fungal community structure in traditional fermentation starters for Hong Qu glutinous rice wine. DGGE profiles based on ITS1fGC/ITS2r revealed the presence of twenty-four fungal species in traditional fermentation starter. A significant difference of fungal community can be observed directly from DGGE fingerprints and

  10. Enrichment of methylated molecules using enhanced-ice-co-amplification at lower denaturation temperature-PCR (E-ice-COLD-PCR) for the sensitive detection of disease-related hypermethylation.

    Science.gov (United States)

    Mauger, Florence; Kernaleguen, Magali; Lallemand, Céline; Kristensen, Vessela N; Deleuze, Jean-François; Tost, Jörg

    2018-05-01

    The detection of specific DNA methylation patterns bears great promise as biomarker for personalized management of cancer patients. Co-amplification at lower denaturation temperature-PCR (COLD-PCR) assays are sensitive methods, but have previously only been able to analyze loss of DNA methylation. Enhanced (E)-ice-COLD-PCR reactions starting from 2 ng of bisulfite-converted DNA were developed to analyze methylation patterns in two promoters with locked nucleic acid (LNA) probes blocking amplification of unmethylated CpGs. The enrichment of methylated molecules was compared to quantitative (q)PCR and quantified using serial dilutions. E-ice-COLD-PCR allowed the multiplexed enrichment and quantification of methylated DNA. Assays were validated in primary breast cancer specimens and circulating cell-free DNA from cancer patients. E-ice-COLD-PCR could prove a useful tool in the context of DNA methylation analysis for personalized medicine.

  11. Comparative carbon-13 nuclear magnetic resonance study at 67. 9 MHz on lysozyme (human and egg-white) and. cap alpha. -lactalbumin (human and bovine) in their native and denatured state

    Energy Technology Data Exchange (ETDEWEB)

    van Binst, G; Biesemans, M [Brussels Univ. (Belgium). Faculte des Sciences

    1975-01-01

    A first detailed comparison of the /sup 13/C spectra at high field of two lysozymes (human and egg-white) and two ..cap alpha..-lactalbumins (human and bovine milk) is presented. Assignments were made on most of the resonance peaks in the aliphatic and aromatic regions of the denatured proteins. The relative peak intensities clearly demonstrate the differences in the amino acid composition of the related proteins. The broadening and the complexity of the spectra of the native proteins reflect the non equivalence of the chemical groups in the folded conformation. The usefulness of /sup 13/C nmr spectroscopy in the study of the interaction between small molecules and proteins was tested on N-acteyl-glucosamine in the presence of lysozyme.

  12. A novel polymerase chain reaction (PCR) - denaturing gradient gel electrophoresis (DGGE) for the identification of Micrococcaceae strains involved in meat fermentations. Its application to naturally fermented Italian sausages.

    Science.gov (United States)

    Cocolin, L; Manzano, M; Aggio, D; Cantoni, C; Comi, G

    2001-05-01

    A new molecular method consisting of polymerase chain reaction (PCR) amplification and denaturing gradient gel electrophoresis (DGGE) of a small fragment from the 16S rRNA gene identified the Micrococcaceae strains isolated from natural fermented Italian sausages. Lactic acid bacteria, total aerobic mesophilic flora, Enterobacteriaceae and faecal enterococci were also monitored. Micrococcaceaea control strains from international collections were used to optimise the method and 90 strains, isolated from fermented sausages, were identified by biochemical tests and PCR-DGGE. No differences were observed between the methods used. The results reported in this paper prove that Staphylococcus xylosus is the main bacterium involved in fermented sausage production, representing, from the tenth day of ripening, the only Micrococcaceaea species isolated.

  13. Heat shock protein 70 negatively regulates the heat-shock-induced suppression of the IκB/NF-κB cascade by facilitating IκB kinase renaturation and blocking its further denaturation

    International Nuclear Information System (INIS)

    Lee, Kyoung-Hee; Lee, Choon-Taek; Kim, Young Whan; Han, Sung Koo; Shim, Young-Soo; Yoo, Chul-Gyu

    2005-01-01

    Heat shock (HS) treatment has been previously shown to suppress the IκB/nuclear factor-κB (NF-κB) cascade by denaturing, and thus inactivating IκB kinase (IKK). HS is characterized by the induction of a group of heat shock proteins (HSPs). However, their role in the HS-induced suppression of the IκB/NF-κB cascade is unclear. Adenovirus-mediated HSP70 overexpression was found not to suppress the TNF-α-induced activation of the IκB/NF-κB pathway, thus suggesting that HSP70 is unlikely to suppress this pathway. When TNF-α-induced activation of the IκB/NF-κB pathway was regained 24 h after HS, HSP70 was found to be highly up-regulated. Moreover, blocking HSP70 induction delayed TNF-α-induced IκBα degradation and the resolubilization of IKK. In addition, HSP70 associated physically with IKK, suggesting that HSP70 is involved in the recovery process via molecular chaperone effect. Adenovirus-mediated HSP70 overexpression prior to HS blocked the IκBα stabilizing effect of HS by suppressing IKK insolubilization. Moreover, the up-regulation of endogenous HSP70 by preheating, suppressed this subsequent HS-induced IKK insolubilization, and this effect was abrogated by blocking HSP70 induction. These findings indicate that HSP70 accumulates during HS and negatively regulates the HS-induced suppression of the IκB/NF-κB cascade by facilitating the renaturation of IKK and blocking its further denaturation

  14. The role of whole body spiral CT in the primary work-up of polytrauma patients - comparison with conventional radiography and abdominal sonography; Die Rolle der Ganzkoerper-Spiral-CT bei der Primaerdiagnostik polytraumatisierter Patienten - Vergleich mit konventioneller Radiographie und Abdomensonographie

    Energy Technology Data Exchange (ETDEWEB)

    Albrecht, T.; Schlippenbach, J. von; Wolf, K.J. [Klinik und Poliklinik fuer Radiologie und Nuklearmedizin, Charite - Campus Benjamin Franklin (Germany); Stahel, P.F.; Ertel, W. [Klinik fuer Unfall- und Wiederherstellungschirurgie, Charite - Campus Benjamin Franklin (Germany)

    2004-08-01

    Purpose: To evaluate the role of routine 'whole body spiral CT' in the primary work-up of polytrauma patients for injuries of the thorax, abdomen and spine, and to compare the results with those of conventional radiography of the chest and spine and abdominal ultrasound. Materials and Methods: Fifty consecutive polytrauma patients underwent contrast-enhanced single slice spiral CT (5 mm collimation) from the vertex to the floor of the pelvis as part of the primary work-up after emergency room admission. Overlapping high resolution sections and sagittal reformations of the spine were obtained. Reports of additional chest radiographs (n=43), abdominal ultrasound examinations (n=47) and spine radiographs (n=36) performed in the emergency room were available for retrospective comparison. The 'final diagnoses', which served as the standard of reference, were taken from the patients' records using all information that became available until discharge or death, such as findings from further imaging, surgery and autopsy. Results: CT showed 109 (97%) of 112 thoracic and abdominal soft-tissue injuries. Relevant injuries missed were an early splenic laceration and an early pelvic hematoma, both of which became clinically apparent several hours later. There were 4 false positive CT findings. Conventional chest radiography demonstrated only 20% of thoracic and sonography 22% of abdominal injuries. Chest radiography and sonography produced 2 false-positive findings each. CT showed 66 (87%) of 76 vertebral fractures including all 19 unstable ones. CT missed 5 anterior vertebral body and 5 spinous/transverse process fractures. Conventional radiography found 71% of vertebral fractures including only 50% of the unstable one. (orig.)

  15. Final report of the safety assessment of Alcohol Denat., including SD Alcohol 3-A, SD Alcohol 30, SD Alcohol 39, SD Alcohol 39-B, SD Alcohol 39-C, SD Alcohol 40, SD Alcohol 40-B, and SD Alcohol 40-C, and the denaturants, Quassin, Brucine Sulfate/Brucine, and Denatonium Benzoate.

    Science.gov (United States)

    2008-01-01

    Alcohol Denat. is the generic term used by the cosmetics industry to describe denatured alcohol. Alcohol Denat. and various specially denatured (SD) alcohols are used as cosmetic ingredients in a wide variety of products. Many denaturants have been previously considered, on an individual basis, as cosmetic ingredients by the Cosmetic Ingredient Review (CIR) Expert Panel, whereas others, including Brucine and Brucine Sulfate, Denatonium Benzoate, and Quassin, have not previously been evaluated. Quassin is a bitter alkaloid obtained from the wood of Quassia amara. Quassin has been used as an insect antifeedant and insecticide and several studies demonstrate its effectiveness. At oral doses up to 1000 mg/kg using rats, Quassin was not toxic in acute and short-term tests, but some reversible piloerection, decrease in motor activity, and a partial loss of righting reflex were found in mice at 500 mg/kg. At 1000 mg/kg given intraperitoneally (i.p.), all mice died within 24 h of receiving treatment. In a cytotoxicity test with brine shrimp, 1 mg/ml of Quassin did not possess any cytotoxic or antiplasmodial activity. Quassin administered to rat Leydig cells in vitro at concentrations of 5-25 ng/ml inhibited both the basal and luteinizing hormone (LH)-stimulated testosterone secretion in a dose-related fashion. Quassin at doses up to 2.0 g/kg in drinking water using rats produced no significant effect on the body weights, but the mean weights of the testes, seminal vesicles, and epididymides were significantly reduced, and the weights of the anterior pituitary glands were significantly increased. The sperm counts and levels of LH, follicle-stimulating hormone (FSH), and testosterone were significantly lower in groups treated with Quassin. Brucine is a derivative of 2-hydroxystrychnine. Swiss-Webster mice given Brucine base, 30 ml/kg, had an acute oral LD(50) of 150 mg/kg, with central nervous system depression followed by convulsions and seizures in some cases. In those

  16. A novel technique using DNA denaturation to detect multiply induced single-strand breaks in a hydrated plasmid DNA molecule by X-ray and 4He2+ ion irradiation

    International Nuclear Information System (INIS)

    Yokoya, A.; Shikazono, N.; Fujii, K.; Noguchi, M.; Urushibara, A.

    2011-01-01

    To detect multiple single-strand breaks (SSBs) produced in plasmid DNA molecules by direct energy deposition from radiation tracks, we have developed a novel technique using DNA denaturation by which irradiated DNA is analysed as single-strand DNA (SS-DNA). The multiple SSBs that arise in both strands of DNA, but do not induce a double-strand break, are quantified as loss of SS-DNA using agarose gel electrophoresis. We have applied this method to X-ray and 4 He 2+ ion-irradiated samples of fully hydrated pUC18 plasmid DNA. The fractions of both SS-DNA and closed circular DNA (CC-DNA) exponentially decrease with the increasing dose of X rays and 4 He 2+ ions. The efficiency of the loss of SS-DNA was half that of CC-DNA for both types of irradiation, indicating that one of two strands in DNA is not broken when one SSB is produced in CC-DNA by irradiation. Contrary to our initial expectation, these results indicate that SSBs are not multiply induced even by high linear energy transfer radiation distributed in both strands. (authors)

  17. An X-band Co2+ EPR study of Zn1-xCoxO (x=0.005-0.1) nanoparticles prepared by chemical hydrolysis methods using diethylene glycol and denaturated alcohol at 5 K

    Science.gov (United States)

    Misra, Sushil K.; Andronenko, S. I.; Srinivasa Rao, S.; Chess, Jordan; Punnoose, A.

    2015-11-01

    EPR investigations on two types of dilute magnetic semiconductor (DMS) ZnO nanoparticles doped with 0.5-10% Co2+ ions, prepared by two chemical hydrolysis methods, using: (i) diethylene glycol ((CH2CH2OH)2O) (NC-rod-like samples), and (ii) denatured ethanol (CH3CH2OH) solutions (QC-spherical samples), were carried out at X-band (9.5 GHz) at 5 K. The analysis of EPR data for NC samples revealed the presence of several types of EPR lines: (i) two types, intense and weak, of high-spin Co2+ ions in the samples with Co concentration >0.5%; (ii) surface oxygen vacancies, and (iii) a ferromagnetic resonance (FMR) line. QC samples exhibit an intense FMR line and an EPR line due to high-spin Co2+ ions. FMR line is more intense, than the corresponding line exhibited by NC samples. These EPR spectra varied for sample with different doping concentrations. The magnetic states of these samples as revealed by EPR spectra, as well as the origin of ferromagnetism DMS samples are discussed.

  18. Utilidad del estudio de las heces para el diagnóstico y manejo de lactantes y prescolares con diarrea aguda Stool work-up protocol in infants and preschool children with acute diarrhea: is it useful for diagnosis and treatment

    Directory of Open Access Journals (Sweden)

    Alfredo Larrosa-Haro

    2002-07-01

    analyze the results of a stool work-up protocol in a series of infants and preschoolers with acute diarrhea. Material and Methods. A cross-sectional descriptive study was conducted between April 1999 and March 2000, among 288 children seen at a pediatric office in Guadalajara, Mexico. The mean age (±1SD was 23.1±13.9 months; 43% were females. Data were collected on demographic and clinical characteristics. The stool work-up consisted of fresh smear and methylene blue and Kinyoun smears, as well as determination of pH and reducing substances. Stool culture was performed in samples with ³ 3 leukocytes/microscopic field and rotavirus antigen detection only in selected cases. Data were analyzed using descriptive statistics, chi², odds ratios, and 95% confidence intervals. Results. Enterophatogens (% identified were: rotavirus 47.1, Campylobacter jejuni 27.4, Salmonella spp. 5.1, Shigella spp. 4.3, Cryptosporidium parvum 2.8, Giardia lamblia 2.4, Blastocystis hominis 1.4, Entamoeba histolytica 0.7. An OR of 5.7 was obtained for isolation of enteroinvasive bacteria in the presence of fecal leukocytes. Lactose intolerance was detected in 19.1%. The frequencies of rotavirus antigen identification and lactose intolerance were significantly higher in infants; the OR for lactose intolerance in infants with rotavirus was 21. Mucus and blood in the stools were associated to enteroinvasive bacteria and Cryptosporidium parvum. Conclusions. The current stool work-up protocol allowed the identification of enteropathogenic parasites, rotavirus and lactose intolerance in a short period of time. Leukocytes in stools were associated to the isolation of enteroinvasive bacteria. The frequency of agents associated to diarrheal disease was similar to that from other national studies. This stool work-up protocol could be useful as a tool to limit the unnecessary prescription of drugs and to follow universal recommendations for dietary management of these patients.

  19. Raman spectroscopy of DNA-metal complexes. II. The thermal denaturation of DNA in the presence of Sr2+, Ba2+, Mg2+, Ca2+, Mn2+, Co2+, Ni2+, and Cd2+.

    Science.gov (United States)

    Duguid, J G; Bloomfield, V A; Benevides, J M; Thomas, G J

    1995-12-01

    Differential scanning calorimetry, laser Raman spectroscopy, optical densitometry, and pH potentiometry have been used to investigate DNA melting profiles in the presence of the chloride salts of Ba2+, Sr2+, Mg2+, Ca2+, Mn2+, Co2+, Ni2+, and Cd2+. Metal-DNA interactions have been observed for the molar ratio [M2+]/[PO2-] = 0.6 in aqueous solutions containing 5% by weight of 160 bp mononucleosomal calf thymus DNA. All of the alkaline earth metals, plus Mn2+, elevate the melting temperature of DNA (Tm > 75.5 degrees C), whereas the transition metals Co2+, Ni2+, and Cd2+ lower Tm. Calorimetric (delta Hcal) and van't Hoff (delta HVH) enthalpies of melting range from 6.2-8.7 kcal/mol bp and 75.6-188.6 kcal/mol cooperative unit, respectively, and entropies from 17.5 to 24.7 cal/K mol bp. The average number of base pairs in a cooperative melting unit () varied from 11.3 to 28.1. No dichotomy was observed between alkaline earth and transition DNA-metal complexes for any of the thermodynamic parameters other than their effects on Tm. These results complement Raman difference spectra, which reveal decreases in backbone order, base unstacking, distortion of glycosyl torsion angles, and rupture of hydrogen bonds, which occur after thermal denaturation. Raman difference spectroscopy shows that transition metals interact with the N7 atom of guanine in duplex DNA. A broader range of interaction sites with single-stranded DNA includes ionic phosphates, the N1 and N7 atoms of purines, and the N3 atom of pyrimidines. For alkaline earth metals, very little interaction was observed with duplex DNA, whereas spectra of single-stranded complexes are very similar to those of melted DNA without metal. However, difference spectra reveal some metal-specific perturbations at 1092 cm-1 (nPO2-), 1258 cm-1 (dC, dA), and 1668 cm-1 (nC==O, dNH2 dT, dG, dC). Increased spectral intensity could also be observed near 1335 cm-1 (dA, dG) for CaDNA. Optical densitometry, employed to detect DNA

  20. Discrimination of three mutational events that result in a disruption of the R122 primary autolysis site of the human cationic trypsinogen (PRSS1 by denaturing high performance liquid chromatography

    Directory of Open Access Journals (Sweden)

    Férec Claude

    2001-11-01

    Full Text Available Abstract Background R122, the primary autolysis site of the human cationic trypsinogen (PRSS1, constitutes an important "self-destruct" or "fail-safe" defensive mechanism against premature trypsin activation within the pancreas. Disruption of this site by a missense mutation, R122H, was found to cause hereditary pancreatitis. In addition to a c.365G>A (CGC>CAC single nucleotide substitution, a c.365~366GC>AT (CGC>CAT gene conversion event in exon 3 of PRSS1 was also found to result in a R122H mutation. This imposes a serious concern on the genotyping of pancreatitis by a widely used polymerase chain reaction-restriction fragment length polymorphism assay, which could only detect the commonest c.365G>A variant. Materials and methods DNA samples containing either the known c.365G>A or c.365~366GC>AT variant in exon 3 of PRSS1 were used as positive controls to establish a denaturing high performance liquid chromatography (DHPLC assay. Results DHPLC could readily discriminate the two known different mutational events resulting in the R122H mutation. More importantly, under the same experimental conditions, it identified a further mutational event that also occurs in the R122 primary autolysis site but results in a different amino acid substitution: c.364C>T (CGC>TGC; R122C. Conclusions A rapid, simple, and low-cost assay for detecting both the known and new mutations occuring in the R122 primary autolysis site of PRSS1 was established. In addition, the newly found R122C variant represents a likely pancreatitis-predisposing mutation.

  1. The Diagnostic Work-Up of Eating Disorders

    DEFF Research Database (Denmark)

    Sjögren, Magnus

    2015-01-01

    Background: The mortality of Anorexia Nervosa (AN) is the highest among all psychiatric disorders, and Eating Disorders (ED) overall pose serious health threats to a significant proportion of the population. In spite of an increasing recognition of the clinical impact of ED, often, the general re...

  2. Stitch in the side: causes, workup, and solutions.

    Science.gov (United States)

    Eichner, E Randy

    2006-12-01

    Side stitch is an acute, localized, sharp, transient pain that occurs during exercise, most often in runners or swimmers, but also in those participating in team sports, and less often in cyclists. The pain is vexing and performance-limiting, but fleeting and benign. Three theories compete to explain it: diaphragmatic ischemia, stress on peritoneal ligaments, and irritation of the parietal peritoneum. The differential diagnosis seems broad, but the classic side stitch is so unique in its features and setting that a reasonable working diagnosis can be made from a careful history and physical examination. Practical tips, although anecdotal, offer lasting solutions.

  3. MDCT diagnostic work-up of patients with subarachnoid haemorrhage

    International Nuclear Information System (INIS)

    Phanis, S.; Nikolopoulos, P.; Veliou, K.; Tsampoulas, C.; Efremidis, S.

    2006-01-01

    Full text: Our presentation is intended to analyze our experience over the last two years, in the expeditious work up of patients with subarachnoid haemorrhage, as well as patients whose clinical symptoms combined with radiological findings are highly suspicious of an aneurysm, acute arterial occlusion, cerebral vascular malformation, venous occlusion, dissection of the internal carotid artery or vertebral artery. 276 CT Angiographies of the brain have been performed and recorded over the last two years in our Department, on a 16 Detector Philips MX8000IDT Computed Tomograph. The patients are referred to our Department from the Emergency Department of our Hospital, as well as from Hospitals all over Northwestern Greece. The CT Angiographies are performed with examination protocols, which we will analyze thoroughly in our presentation. The primary data are then studied in workstations away from the main console of the Computed Tomograph. The study of the primary data consists in multiplanar reconstructions with the Maximum Intensity Algorithm, as well as Volume Rendering and three Dimensional Reconstruction. The stages we follow for these procedures will be presented as well as the procedures for the removal of bone structures, with the intention of the best possible angiographic results. Of the 276 patients 40 were diagnosed with an aneurysm,19 with a venous angioma, 2 with a vascular malformation, 3 with venous occlusion and 2 with dissection, a total of 66/276 with aneurysms and vascular malformations. These patients were subsequently referred to the Neurosurgery Clinic or to the Department of Interventional Radiology for further treatment. The remaining 210 patients, underwent in the following days of their hospitalization a Digital Subtraction Angiography, and in 16 (7.6%) of them a small aneurysm, 3-5 mm was diagnosed. The aneurysm was detected in a second reading of the CT Angiography in 8 patients (3.8%). The CT Angiography in patients with subarachnoid haemorrhage, performed in the new MDCT Computed Tomographs (16 Detectors), is a fast, minimally invasive, well tolerated, and a diagnostic method gradually gaining international recognition by radiologists as well as by clinical doctors, especially as the experience of radiologists in the method is increasing. Digital Subtraction Angiography still remains the Gold Standard method of examination, the trend though appears to be in favor for MDCT Angiography. We are anxiously awaiting for the primary results from the newest 64 Detector and 256 Detector Computed Tomographs

  4. Work-up of Pediatric Urinary Tract Infection.

    Science.gov (United States)

    Schmidt, Bogdana; Copp, Hillary L

    2015-11-01

    Pediatric urinary tract infection (UTI) costs the health care system more than $180 million annually, and accounts for more than 1.5 million clinician visits per year. Accurate and timely diagnosis of these infections is important for determining appropriate treatment and preventing long-term complications such as renal scarring, hypertension, and end-stage renal disease. After the first 12 months, girls are more likely to be diagnosed with a UTI. About half of boys with UTI are diagnosed within the first 12 months of life. Diagnosis of UTI is made based on history and examination findings and confirmed by urine testing. Copyright © 2015 Elsevier Inc. All rights reserved.

  5. Is hysterosalpingography still relevant in workup of infertility? A ...

    African Journals Online (AJOL)

    Objective: To examine the current role of Hysterosalpingography in the investigation of the infertile couple in view of the increasing use of Laparoscopy in the evaluation and management of infertility. Materials: Electronic search of published literature was done using the Pubmed, Cochrane libraries and Google search ...

  6. Idiopathic interstitial pneumonias. From classification to diagnostic work-up

    International Nuclear Information System (INIS)

    Mueller-Mang, C.; Bankier, A.; Stiebellehner, L.; Schmid, K.

    2007-01-01

    Idiopathic interstitial pneumonias (IIP) comprise seven entities with distinct histologic patterns. In their idiopathic form IIP are rare diseases. They are, nevertheless, considered prototypes of the much more common secondary interstitial pneumonias. The advent of high-resolution computed tomography (HRCT) has had a profound impact on the imaging of IIP, because the detailed delineation of the lung anatomy allows a close correlation between the histologic patterns of IIP and the CT features. On the basis of CT morphology and in the correct clinical context, the radiologist can achieve an accurate diagnosis in many cases. However, due to overlap between the various entities, complementary lung biopsy is recommended in virtually all cases. This article reviews the CT pattern of IIP and offers relevant clinical and histological information for the purpose of enabling the radiologist to understand and participate in the multidisciplinary concept of IIP. (orig.) [de

  7. [Denaturation of egg antigens by cooking].

    Science.gov (United States)

    Watanabe, Hiroko; Akaboshi, Chie; Sekido, Haruko; Tanaka, Kouki; Tanaka, Kazuko; Shimojo, Naoki

    2012-01-01

    Changes in egg protein contents by cooking were measured with an ELISA kit using Tris-HCl buffer in model foods including cake, meatballs, pasta and pudding made with whole egg, egg-white and egg-yolk. The egg protein contents were lowest in the deep-fried model foods of cakes and meatballs. Ovalbumin (OVA) was undetectable (meatballs, suggesting that processing temperature and uniform heat-treatment affect the detection of egg protein. Furthermore, egg protein contents were below 6 µg/g in the pouched meatballs and pasta made with egg-yolk, and OVA and OVM were not detected by Western blotting analysis with human IgE from patients' serum. On the other hand, processed egg proteins were detected with an ELISA kit using a surfactant and reductant in the extract buffer.

  8. 27 CFR 21.151 - List of denaturants authorized for denatured spirits.

    Science.gov (United States)

    2010-04-01

    ..., liquid S.D.A. 36. Cedar leaf oil, U.S.P.XIII S.D.A. 38-B. Chloroform S.D.A. 20. Chlorothymol, N.F.XII S.D... (Glycerol), U.S.P S.D.A. 31-A. Green soap, U.S.P S.D.A. 27-B. Guaiacol, N.F.X S.D.A. 38-B. Heptane C.D.A. 18...) S.D.A. 45. Soap, hard, N.F.XI S.D.A. 31-A. Sodium iodide, U.S.P S.D.A. 25, 25-A. Sodium, metallic S...

  9. 78 FR 38628 - Reclassification of Specially Denatured Spirits and Completely Denatured Alcohol Formulas and...

    Science.gov (United States)

    2013-06-27

    ... this proposed rulemaking: Rum is any spirit produced from sugar cane products and distilled at less...--with ethanol in the resulting formulations. Thus, it would be more difficult to separate the ethanol...

  10. DNA-directed alkylating ligands as potential antitumor agents: sequence specificity of alkylation by intercalating aniline mustards.

    Science.gov (United States)

    Prakash, A S; Denny, W A; Gourdie, T A; Valu, K K; Woodgate, P D; Wakelin, L P

    1990-10-23

    The sequence preferences for alkylation of a series of novel parasubstituted aniline mustards linked to the DNA-intercalating chromophore 9-aminoacridine by an alkyl chain of variable length were studied by using procedures analogous to Maxam-Gilbert reactions. The compounds alkylate DNA at both guanine and adenine sites. For mustards linked to the acridine by a short alkyl chain through a para O- or S-link group, 5'-GT sequences are the most preferred sites at which N7-guanine alkylation occurs. For analogues with longer chain lengths, the preference of 5'-GT sequences diminishes in favor of N7-adenine alkylation at the complementary 5'-AC sequence. Magnesium ions are shown to selectively inhibit alkylation at the N7 of adenine (in the major groove) by these compounds but not the alkylation at the N3 of adenine (in the minor groove) by the antitumor antibiotic CC-1065. Effects of chromophore variation were also studied by using aniline mustards linked to quinazoline and sterically hindered tert-butyl-9-aminoacridine chromophores. The results demonstrate that in this series of DNA-directed mustards the noncovalent interactions of the carrier chromophores with DNA significantly modify the sequence selectivity of alkylation by the mustard. Relationships between the DNA alkylation patterns of these compounds and their biological activities are discussed.

  11. Reaction of single-standard DNA with hydroxyl radical generated by iron(II)-ethylenediaminetetraacetic acid

    International Nuclear Information System (INIS)

    Prigodich, R.V.; Martin, C.T.

    1990-01-01

    This study demonstrates that the reaction of Fe(II)-EDTA and hydrogen peroxide with the single-stranded nucleic acids d(pT) 70 and a 29-base sequence containing a mixture of bases results in substantial damage which is not directly detected by gel electrophoresis. Cleavage of the DNA sugar backbone is enhanced significantly after the samples are incubated at 90 degree C in the presence of piperidine. The latter reaction is used in traditional Maxam-Gilbert DNA sequencing to detect base damage, and the current results are consistent with reaction of the hydroxyl radical with the bases in single-stranded DNA (although reaction with sugar may also produce adducts that are uncleaved but labile to cleavage by piperidine). We the authors propose that hydroxyl radicals may react preferentially with the nucleic acid bases in ssDNA and that reaction of the sugars in dsDNA is dominant because the bases are sequestered within the double helix. These results have implications both for the study of single-stranded DNA binding protein binding sites and for the interpretation of experiments using the hydroxyl radical to probe DNA structure or to footprint double-stranded DNA binding protein binding sites

  12. An X-band Co{sup 2+} EPR study of Zn{sub 1−x}Co{sub x}O (x=0.005–0.1) nanoparticles prepared by chemical hydrolysis methods using diethylene glycol and denaturated alcohol at 5 K

    Energy Technology Data Exchange (ETDEWEB)

    Misra, Sushil K., E-mail: skmisra@alcor.concordia.ca [Physics Department, Concordia University, Montreal, QC, Canada H3G 1M8 (Canada); Andronenko, S.I. [Physics Institute, Kazan Federal University, Kazan 420008 (Russian Federation); Srinivasa Rao, S.; Chess, Jordan; Punnoose, A. [Department of Physics, Boise State University, Boise, ID 83725-1570 (United States)

    2015-11-15

    EPR investigations on two types of dilute magnetic semiconductor (DMS) ZnO nanoparticles doped with 0.5–10% Co{sup 2+} ions, prepared by two chemical hydrolysis methods, using: (i) diethylene glycol ((CH{sub 2}CH{sub 2}OH){sub 2}O) (NC-rod-like samples), and (ii) denatured ethanol (CH{sub 3}CH{sub 2}OH) solutions (QC-spherical samples), were carried out at X-band (9.5 GHz) at 5 K. The analysis of EPR data for NC samples revealed the presence of several types of EPR lines: (i) two types, intense and weak, of high-spin Co{sup 2+} ions in the samples with Co concentration >0.5%; (ii) surface oxygen vacancies, and (iii) a ferromagnetic resonance (FMR) line. QC samples exhibit an intense FMR line and an EPR line due to high-spin Co{sup 2+} ions. FMR line is more intense, than the corresponding line exhibited by NC samples. These EPR spectra varied for sample with different doping concentrations. The magnetic states of these samples as revealed by EPR spectra, as well as the origin of ferromagnetism DMS samples are discussed. - Highlights: • 5 K X band Co{sup 2+} EPR investigations on QC and NC ZnO dilute magnetic semiconductor nanoparticles. • NC and QC samples exhibited high-spin Co{sup 2+} EPR lines and ferromagnetic resonance line. • NC sample also exhibit line due surface oxygen vacancies. • FMR line is more intense in QC than that in NC samples. • Magnetic states and the origin of ferromagnetism are discussed.

  13. Native and denatured enzyme enterokinase determined by electrochemical methods

    Czech Academy of Sciences Publication Activity Database

    Janovjáková, A.; Gál, M.; Krahulec, J.; Sokolová, Romana; Naumowicz, M.; Híveš, J.

    2017-01-01

    Roč. 148, č. 3 (2017), s. 549-553 ISSN 0026-9247 Institutional support: RVO:61388955 Keywords : enterokinase * proteins conformation * chronopotentiometry Subject RIV: CG - Electrochemistry OBOR OECD: Physical chemistry Impact factor: 1.282, year: 2016

  14. The Denaturation of Environmental Education: Exploring the Role of Ecotechnologies

    Science.gov (United States)

    Gough, Annette; Gough, Noel

    2016-01-01

    This article explores the changing ways "environment" has been represented in the discourses of environmental education and education for sustainable development (ESD) in United Nations (and related) publications since the 1970s. It draws on the writings of Jean-Luc Nancy and discusses the increasingly dominant view of the environment as…

  15. The application of polymerase chain reaction-denaturing gradient ...

    African Journals Online (AJOL)

    Jane

    2011-05-23

    May 23, 2011 ... dominance in microbial ecology if the corresponding environment samples had been provided. This ... yeast peptone dextrose; PCR, polymerase chain reaction. method, DGGE method ..... Two nuclear mutations that block.

  16. Evaluation of denatured thorium fuel cycles in pressurized water reactors

    International Nuclear Information System (INIS)

    Matzie, R.A.; Rec, J.R.; Terney, A.N.

    1977-01-01

    A developing national energy policy that is based in part on a substantial expansion of the LWR-based electrical generating capacity with deferment of the LMFBR has prompted a re-evaluation of our nuclear fuel resources and their utilization. The ancillary policy of minimizing nuclear weapons proliferation through diversion of bred fissile material has left in doubt the viability of fuel recycling as a means of extending these fuel resources. A substantial, government-sponsored effort is in progress to examine alternate fuel cycles and advanced reactor concepts which can lead to improved resource utilization while minimizing proliferation potential. This paper evaluates several improved fuel cycles for use in current design PWRs and develops selected scenarios for their use within the framework of the safeguarded Nuclear Energy Center (NEC) concept

  17. ESR studies of heat denaturation in Cu myoglobin complexes

    International Nuclear Information System (INIS)

    Louro, S.R.W.; Ribeiro, S.C.; Bemski, G.

    1975-01-01

    An investigation is made on the behaviour of Cu2 + and Fe3 + in copper doped myoglobin, subjected to heat treatment. ESR is observed at X-band. The amplitude of the g = 5.9 line of the high spin F 3 + in met-myoglobin is studied as a function of the temperature of the heat treatment, the pH and the length of time of exposure to the heat treatment. These experiments are performed for both the pure protein and for the copper-myoglobin complex, at pH between 5 and 8.5. Results concerning the decrease of the amplitude of Fe 3+ high spin signal are discussed in comparison with Hollocher's results in hemoglobin. The Cu spectra obtained are interpreted, with the aid of a computer program, Kivelson and Neiman's results being used to analyse the authors data

  18. Denaturalizing the technique: feminists contributions to think the cinematography

    Directory of Open Access Journals (Sweden)

    Marina Cavalcanti Tedesco

    2014-07-01

    Full Text Available Cinematography is generally understood as a technological area, which means neutral for many people. This may account for not deserving the due attention into the cinema and the audiovisual studies. However, as any technique, it was developed starting from some hypotheses to reach specific objectives. In this study, we aim to analyze it with a feministic perspective to identify the gender views that contributed to its expression, which were influenced by it. We ́ll do that by the sudy of some cinematographic manuals edited since the years 1930, comparing their guidelines with some important classic/industrial Mexican films.

  19. The anionic biosurfactant rhamnolipid does not denature industrial enzymes

    Directory of Open Access Journals (Sweden)

    Jens Kvist Madsen

    2015-04-01

    Full Text Available Biosurfactants (BS are surface-active molecules produced by microorganisms. Their combination of useful properties and sustainable production make them promising industrial alternatives to petrochemical and oleochemical surfactants. Here we compare the impact of the anionic BS rhamnolipid (RL and the conventional/synthetic anionic surfactant sodium dodecyl sulfate (SDS on the structure and stability of three different commercially used enzymes, namely the cellulase Carezyme® (CZ, the phospholipase Lecitase Ultra® (LT and the α-amylase Stainzyme® (SZ. Our data reveal a fundamental difference in their mode of interaction. SDS shows great diversity of interaction towards the different enzymes. It efficiently unfolds both LT and CZ, but LT is unfolded by SDS through formation of SDS clusters on the protein well below the cmc, while CZ is only unfolded by bulk micelles and on average binds significantly less SDS than LT. SDS binds with even lower stoichiometry to SZ and leads to an increase in thermal stability. In contrast, RL does not affect the tertiary or secondary structure of any enzyme at room temperature, has little impact on thermal stability and only binds detectably (but at low stoichiometries to SZ. Furthermore all enzymes maintain activity at both monomeric and micellar concentrations of RL. We conclude that RL, despite its anionic charge, is a surfactant that does not compromise the structural integrity of industrially relevant proteins. This makes RL a promising alternative to current synthetic anionic surfactants in a wide range of commercial applications.

  20. Ultraviolet light-denatured DNA/anti-ultraviolet light-denatured DNA immune-complex nephritis in rabbits

    International Nuclear Information System (INIS)

    Sweny, P.

    1980-01-01

    Two groups of preimmunized rabbits were studied during a 3-month course of daily intravenous injections of uv DNA in amounts sufficient to neuralize circulating antibody. One group was given high-molecular-weight uv DNA, and the other group, US uv DNA. Rabbits receiving US uv DNA formed potentially more damaging immune complexes, since this group of animals developed greater rises in blood urea and greater falls in C3. Both groups of animals developed evidence of immune complex-mediated glomerular nephritis as evidenced by heavy granular deposits of IgG and C3 in the glomeruli. The results suggest that immune complexes formed with US uv DNA may be more nephrotoxic

  1. O estatuto da diversidade sexual nas políticas de educação no Brasil e na França: a comparação como ferramenta de desnaturalização do cotidiano de pesquisa Sexual diversity status in Brazil's and France's educational policies: comparison as a denaturalizing tool for the quotidian of research

    Directory of Open Access Journals (Sweden)

    Henrique Caetano Nardi

    2008-01-01

    Full Text Available O objetivo deste artigo é descrever problematizando o cotidiano da construção de uma pesquisa que, ela mesma, busca entender como se produz, a partir da implantação de políticas públicas, ações sobre um cotidiano naturalizado de discriminações de caráter homofóbico e sexista nas escolas do Brasil e da França. Além de apresentar informações e hipóteses que emergem do trabalho de campo, a intenção do texto é apresentar os desafios de uma pesquisa que mergulha no cotidiano de culturas e contextos socio-econômicos distintos. A análise da utilização da ferramenta da comparação, pensada como efeito de distorção da imagem no espelho e colocação em suspenso de formas pré-construídas de compreensão do vivido, indica que uma intervenção analítica desta ordem permite, de forma distinta da pretensão de generalização pópria a um exercício de comparação de base positivista, a desnaturalização de formas de pesquisar e de compreender as problemáticas sobre as quais nos debruçamos.The goal of this article is to describe and problematize the quotidian of producing a research, which, in itself, seeks to understand how naturalized homophobic and sexist discriminatory actions are produced in the quotidian of schools in Brazil and France, based on the implementation of public policies. Besides presenting information and hypotheses derived from fieldwork, the intention of this text is to present the challenges of a research that immerses into the quotidian of different socioeconomic and cultural contexts. The analysis of using the comparative tool, considered as having a distortion effect of an image in the mirror and suspension effect of pre-constructed forms of comprehension of what is experienced, indicates that this order of analytic intervention allows to denaturalize research practices and the understanding of problems, distinctively from positivist comparison exercises that intend to generalize.

  2. Superficial artery aneurysms underlying nummular headache--2 cases and proposed diagnostic work-up.

    Science.gov (United States)

    López-Ruiz, Pedro; Cuadrado, María-Luz; Aledo-Serrano, Angel; Alonso-Oviés, Almudena; Porta-Etessam, Jesús; Ganado, Tomás

    2014-01-01

    Nummular headache, considered a primary headache in the 3rd edition of the International Classification of Headache Disorders, has nonetheless been attributed in several occasions to underlying epicranial anomalies. Vascular imaging of the head in 2 patients with nummular headache revealed fusiform aneurysms of the scalp vessel in close relation to the painful area. One of the patients underwent surgical resection of the aneurysm with excellent response. A response to triptans was noted in both. In selected patients with nummular headache, vascular imaging of the scalp may reveal anomalies amenable to surgical treatment or triptan administration, sometimes resulting in disappearance of the pain. © 2014 American Headache Society.

  3. Approach to pediatric epilepsy surgery: State of the art, Part I: General principles and presurgical workup.

    Science.gov (United States)

    Obeid, Makram; Wyllie, Elaine; Rahi, Amal C; Mikati, Mohamad A

    2009-03-01

    In 1990, the National Institute of Health adopted epilepsy surgery in children as an option when medications fail. In the past few years several concepts have become increasingly recognized as key to a successful approach to epilepsy surgery in children. These include the concepts of neuronal plasticity, the epileptogenic lesion, the ictal onset, symptomatogenic, irritative, and epileptogenic zones. In addition, several techniques have increasingly been utilized to delineate the above areas in an attempt to determine, in each patient, the epileptogenic zone, defined as the zone the resection of which leads to seizure freedom. When seizure semiology (which defines the symptomatogenic zone), ictal EEG (which identifies the ictal onset zone), and structural imaging (which identifies the epileptogenic lesion) can be reconciled to infer the location of the epileptogenic zone, surgery is usually, subsequently, undertaken. When these diagnostic modalities are discordant, not definitive, or when the epileptogenic zone is close to eloquent cortex, invasive EEG, complemented by other imaging techniques may be needed. These include magnetoencephalography, single photon emission tomography, various types of positron emission tomography, various magnetic resonance imaging modalities (functional, diffusion weighted, other) and other emerging and experimental techniques. While MRI, video-EEG, and neuropsychological assessments are well established components of the presurgical evaluation, the use of the new emerging imaging technologies is dictated by the degree of anatomo-electro-clinical correlations, and, awaiting multicentric studies and more detailed guidelines, remains center-dependent.

  4. Should the post-coital test (PCT) be part of the routine fertility work-up?

    NARCIS (Netherlands)

    van der Steeg, Jan W.; Steures, Pieternel; Eijkemans, Marinus J. C.; Habbema, J. Dik; van der Veen, Fulco; Bossuyt, Patrick M. M.; Hompes, Peter G. A.; Mol, Ben W. J.

    2004-01-01

    BACKGROUND: This study aimed to determine whether medical history and semen analysis can predict the result of the post-coital test (PCT). METHODS: A previously reported data set of Dutch patients collected between 1985 and 1993 was used. Our study was limited to just patients with an ovulatory

  5. Anemia in children: prevalence, causes, diagnostic work-up, and long-term consequences.

    Science.gov (United States)

    Allali, Slimane; Brousse, Valentine; Sacri, Anne-Sylvia; Chalumeau, Martin; de Montalembert, Mariane

    2017-11-01

    Anemia in children is a major public health problem throughout the world. It is often multifactorial, iron deficiency being the most frequent etiology. Consequences are diverse and largely under evaluated. Areas covered: This paper briefly reviews the main causes and focus on the potential consequences of acute and chronic anemia in children. Expert commentary: Anemia in children should never be trivialized. Even if iron deficiency is frequently involved, other potentially life-threatening causes are possible and should be looked for. The exact contribution of anemia to child mortality and morbidity is difficult to assess because of overlapping comorbidities. Chronic anemia may impair growth, cardiac function and cognitive development in infants but other consequences are rather poorly described and should be explored more thoroughly.

  6. Molecular biomarkers have the potential to improve the diagnostic work-up of pancreatic cystic lesions

    DEFF Research Database (Denmark)

    Plougmann, Julie Isabelle; Klausen, Pia; Karstensen, John Gásdal

    2017-01-01

    of diagnostic tools are used to predict the malignant potential of these cysts, but specificity and sensitivity are limited. Thus, many patients undergo unnecessary operations for benign cysts. Balancing the risks of watchful waiting with those of operative management is key in managing these lesions. During...... the last decade, genetic changes of pancreatic cysts have been examined extensively to estimate their malignant potential. In this review, we provide an overview of the latest molecular and genetic aspects of pancreatic cysts and how they may contribute to the differential diagnosis in patients...

  7. Diagnostic work-up and loss of tuberculosis suspects in Jogjakarta, Indonesia

    NARCIS (Netherlands)

    R.A. Ahmad (Riris); F. Matthys (Francine); B. Dwihardiani (Bintari); N. Rintiswati (Ning); S.J. de Vlas (Sake); Y. Mahendradhata (Yodi); P. van der Stuyft (Patrick)

    2012-01-01

    textabstractBackground: Early and accurate diagnosis of pulmonary tuberculosis (TB) is critical for successful TB control. To assist in the diagnosis of smear-negative pulmonary TB, the World Health Organisation (WHO) recommends the use of a diagnostic algorithm. Our study evaluated the

  8. The Appropriate Use of Neuroimaging in the Diagnostic Work-Up of Dementia

    Science.gov (United States)

    Bermingham, SL

    2014-01-01

    Background Structural brain imaging is often performed to establish the underlying causes of dementia. However, recommendations differ as to who should receive neuroimaging and whether computed tomography (CT) or magnetic resonance imaging (MRI) should be used. Objectives This study aimed to determine the cost-effectiveness in Ontario of offering structural imaging to all patients with mild to moderate dementia compared with offering it selectively according to guidelines from the Canadian Consensus Conference on the Diagnosis and Treatment of Dementia (CCC). We compared the cost-effectiveness of CT and MRI as first-line strategies. Methods We performed a systematic literature search (2000 to 2013) to identify cost-effectiveness studies of clinical prediction rules and structural imaging modalities. Studies were assessed for quality and applicability to Ontario. We also developed a model to evaluate the cost-effectiveness of clinical guidelines (image all versus according to CCC) and modalities (CT versus MRI). Transition probabilities, utilities, and costs were obtained from published literature or expert opinion. Results were expressed in terms of costs and quality adjusted life years (QALYs). Results No relevant cost-effectiveness analyses were identified in the published literature. According to the base-case results of our model, the most effective and cost-effective strategy is to image patients who meet CCC criteria with CT and to follow-up with MRI for suspected cases of space-occupying lesions (SOL). However, the results were sensitive to the specificity of MRI for detecting vascular causes of dementia. At a specificity of 64%, the most cost-effective strategy is CCC followed by MRI. Limitations Studies used to estimate diagnostic accuracy were limited by a lack of a gold standard test for establishing the cause of dementia. The model does not include costs to patients and their families, nor does it account for patient preferences about diagnostic information. Conclusions Given the relative prevalence of vascular dementia and SOLs, and the improvement in QALYs associated with treatment, the strategy with the greatest combined sensitivity (CCC with CT followed by MRI for patients with SOLs) results in the greatest number of QALYs and is the least costly. Due to limitations in the clinical data and challenges in the interpretation of this evidence, the model should be considered a framework for assessing uncertainty in the evidence base rather than providing definitive answers to the research questions. Plain Language Summary There is wide debate about whether or not brain scans should routinely be used to assess patients with mild to moderate dementia. Proponents say that imaging is important to detect or rule out possible underlying causes of dementia, such as silent strokes and tumours. Opponents call for a more selective approach, considering the need for clinical judgement and the cost of the technology. Using data from published research, a model was developed to study the cost-effectiveness of different approaches to brain imaging for a hypothetical group of patients with dementia. The model compared 2 strategies: imaging all patients and imaging selectively based on clinical practice guidelines from the Canadian Consensus Conference on the Diagnosis and Treatment of Dementia (CCC). It also compared 2 types of technology: computed tomography (CT) and magnetic resonance imaging (MRI). The results of the model depended on the accuracy of CT and MRI in diagnosing dementia caused by vascular disease. Unfortunately, because there is no “gold standard” approach to diagnosing dementia, interpreting the published research is challenging. Based on current evidence, in which diagnostic strategies are assessed using a mix of methods, the model showed that the most effective and least costly strategy is to image selectively according to the CCC guidelines, using CT first and then MRI as a follow-up for patients suspected of having space-occupying lesions such as tumours. However, if we assumed that MRI plus clinical assessment is the gold standard, then imaging all patients with MRI is the most cost-effective strategy, despite the higher cost of this technology. The model did not take into account the value that physicians, patients, and families place on having diagnostic information, even if effective treatment does not yet exist. The model was not able to answer the specific research questions with confidence, but it provides a framework for identifying areas where more research is needed to support decision-making in the diagnosis of dementia. PMID:24592297

  9. Work-up of thyroid incidentalomas identified by 18F-fluorodeoxyglucose PET/CT

    DEFF Research Database (Denmark)

    Asmar, Ali; Simonsen, Lene; Bülow, Jens

    2017-01-01

    Several reports have described dramatic increase over recent decades in the incidence of thyroid cancer, even as thyroid cancer-related mortality rates have not changed substantially. Nevertheless, in several retrospective studies the incidence of malignancy in focal18F-fluorodeoxyglucose (FDG......) thyroid uptake discovered on whole body18F-FDG PET/CT, carried out for non-thyroid cancers, is 13-64%. Our aim was to design a practical algorithm for management of an increasing number of thyroid incidentalomas, identified by18F-FDG PET/CT....

  10. Diagnostic Work-up and Treatment of Severe and/or Refractory Atopic Dermatitis

    NARCIS (Netherlands)

    A.C.A. Devillers (Arjan)

    2009-01-01

    textabstractAtopic dermatitis (AD) or atopic eczema , is a chronic inflammatory skin disease characterized by dry skin, itching and recurrent red and scaly skin lesions. It is a relatively common skin disease with an estimated prevalence of 10-20%. The majority of patients show their first clinical

  11. The basic fertility workup in women with polycystic ovary syndrome: a systematic review

    NARCIS (Netherlands)

    Nahuis, Marleen J.; Oosterhuis, G. Jurjen E.; Hompes, Peter G. A.; van Wely, Madelon; Mol, Ben Willem J.; van der Veen, Fulco

    2013-01-01

    To summarize the evidence for the use of commonly accepted fertility tests in subfertile women with ovulation problems. Systematic review. Not applicable. The study population included women starting with clomiphene citrate (CC) as first-line treatment, women starting with second-line treatment if

  12. Overdiagnosing Vascular Dementia using Structural Brain Imaging for Dementia Work-Up

    NARCIS (Netherlands)

    Niemantsverdriet, Ellis; Feyen, Bart F. E.; Le Bastard, Nathalie; Martin, Jean-Jacques; Goeman, Johan; De Deyn, Peter Paul; Engelborghs, Sebastiaan

    2015-01-01

    Hypothesizing that non-significant cerebrovascular lesions on structural brain imaging lead to overdiagnosis of a vascular etiology of dementia as compared to autopsy-confirmed diagnosis, we set up a study including 71 patients with autopsy-confirmed diagnoses. Forty-two patients in the population

  13. Assessment of short stature in children : auxological screening and diagnostic work-up

    NARCIS (Netherlands)

    Grote, Floor Katelijn

    2007-01-01

    Growth impairment is considered a relatively early sign of poor health in children. Depending on its setting and the age of the child the impairment is expressed in several ways. In industrialized countries, where malnutrition is rare, the major purpose of growth monitoring, implying regular

  14. Preoperative identification of synchronous ovarian and endometrial cancers: the importance of appropriate workup

    NARCIS (Netherlands)

    Broeders, F.M.; Wurff, A.A. van der; Pijnenborg, J.M.A.; Vos, M.C.

    2012-01-01

    OBJECTIVE: For treatment of patients with both endometrial and ovarian cancer, it is important to discriminate between 2 primary tumors and metastatic disease. Currently, criteria are based on postoperative findings. The aim of this study was to determine whether clinical parameters can discriminate

  15. Peptide release, side-chain deprotection, work-up, and isolation

    DEFF Research Database (Denmark)

    Pedersen, Søren Ljungberg; Jensen, Knud Jørgen

    2013-01-01

    After having successfully synthesized a peptide, it has to be released from the solid support, unless it is being used for on-resin display. The linker and, in some cases, the cleavage mixture determine the C-terminal functionality of the released peptide. In most cases, the peptide is released w...

  16. Neonatal seizures : Aetiology by means of a standardized work-up

    NARCIS (Netherlands)

    Loman, Annemiek M. W.; ter Horst, Henk J.; Lambrechtsen, Florise A. C. P.; Lunsing, Roelineke J.

    Neonatal seizures are an alarming symptom and are frequent in neonates. It is important to find the cause of neonatal seizures to start a specific treatment and to give a meaningful prognosis. The aim of this study is to investigate the incidence of different aetiologies of neonatal seizures in our

  17. Use of Bone Scan During Initial Prostate Cancer Workup, Downstream Procedures, and Associated Medicare Costs

    Energy Technology Data Exchange (ETDEWEB)

    Falchook, Aaron D. [Department of Radiation Oncology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina (United States); Salloum, Ramzi G. [Department of Health Services Policy and Management, University of South Carolina, Columbia, South Carolina (United States); Hendrix, Laura H. [Department of Radiation Oncology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina (United States); Chen, Ronald C., E-mail: ronald_chen@med.unc.edu [Department of Radiation Oncology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina (United States); Cecil G. Sheps Center for Health Services Research, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina (United States); Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina (United States)

    2014-06-01

    Purpose: For patients with a high likelihood of having metastatic disease (high-risk prostate cancer), bone scan is the standard, guideline-recommended test to look for bony metastasis. We quantified the use of bone scans and downstream procedures, along with associated costs, in patients with high-risk prostate cancer, and their use in low- and intermediate-risk patients for whom these tests are not recommended. Methods and Materials: Patients in the Surveillance, Epidemiology, and End Results (SEER)-Medicare database diagnosed with prostate cancer from 2004 to 2007 were included. Prostate specific antigen (PSA), Gleason score, and clinical T stage were used to define D'Amico risk categories. We report use of bone scans from the date of diagnosis to the earlier of treatment or 6 months. In patients who underwent bone scans, we report use of bone-specific x-ray, computed tomography (CT), and magnetic resonance imaging (MRI) scans, and bone biopsy within 3 months after bone scan. Costs were estimated using 2012 Medicare reimbursement rates. Results: In all, 31% and 48% of patients with apparent low- and intermediate-risk prostate cancer underwent a bone scan; of these patients, 21% underwent subsequent x-rays, 7% CT, and 3% MRI scans. Bone biopsies were uncommon. Overall, <1% of low- and intermediate-risk patients were found to have metastatic disease. The annual estimated Medicare cost for bone scans and downstream procedures was $11,300,000 for low- and intermediate-risk patients. For patients with apparent high-risk disease, only 62% received a bone scan, of whom 14% were found to have metastasis. Conclusions: There is overuse of bone scans in patients with low- and intermediate-risk prostate cancers, which is unlikely to yield clinically actionable information and results in a potential Medicare waste. However, there is underuse of bone scans in high-risk patients for whom metastasis is likely.

  18. Abdominal ultrasonography in the diagnostic work-up in children with recurrent abdominal pain

    DEFF Research Database (Denmark)

    Wewer, Anne Vibeke; Strandberg, C; Pærregaard, Anders

    1997-01-01

    We report on our experience with routine abdominal ultrasonography in 120 children (aged 3-15 years) with recurrent abdominal pain, in order to determine the diagnostic value of this investigation. Eight children (7%) revealed sonographic abnormalities: gallbladder stone (n = 2), splenomegaly (n...... = 1) and urogenital abnormalities (n = 5). The recurrent abdominal pain could be explained by these findings in only two (may be three) cases. CONCLUSION: The diagnostic value of abdominal ultrasonography in unselected children with recurrent abdominal pain is low. However, the direct visualization...... of the abdominal structures as being normal may be helpful to the parents and the child in their understanding and acceptance of the benign nature of recurrent abdominal pain....

  19. Low-dose helical computed tomography (CT) in the perioperative workup of adolescent idiopathic scoliosis

    Energy Technology Data Exchange (ETDEWEB)

    Abul-Kasim, Kasim; Overgaard, Angelica; Maly, Pavel [Malmoe University Hospital, Department of Radiology, Section of Neuroradiology, University of Lund, Malmoe (Sweden); Ohlin, Acke [Malmoe University Hospital, Department of Orthopaedic Surgery, University of Lund, Malmoe (Sweden); Gunnarsson, Mikael [Malmoe University Hospital, Department of Radiation Physics, University of Lund, Malmoe (Sweden); Sundgren, Pia C. [University of Michigan Health Systems, Department of Radiology, Division of Neuroradiology, Ann Arbor (United States)

    2009-03-15

    The study aims were to estimate the radiation dose in patients examined with low dose spine CT and to compare it with that received by patients undergoing standard CT for trauma of the same region, as well as to evaluate the impact of dose reduction on image quality. Radiation doses in 113 consecutive low dose spine CTs were compared with those in 127 CTs for trauma. The inter- and intraobserver agreement in measurements of pedicular width, and vertebral rotation, measurements of signal-to-noise ratio and assessment of hardware status were the indicators in the evaluation of image quality. The effective dose of the low dose spine CT (0.37 mSv) was 20 times lower than that of a standard CT for trauma (13.09 mSv). This dose reduction conveyed no impact on image quality. This low dose spine CT protocol allows detailed evaluation that is necessary for preoperative planning and postoperative evaluation. (orig.)

  20. Is the routine use of intravenous contrast justified in the workup of ...

    African Journals Online (AJOL)

    The indications for the administration of intravenous contrast media to adult patients presenting with firsttime seizures and a normal pre-contrast study are not clear. A retrospective audit of such patients was done and compared with the post-contrast study. In our study sample of 205 patients all the post-contrast scans were ...

  1. Cetrorelix suppression test in the diagnostic work-up of severe hyperandrogenism in adolescence.

    NARCIS (Netherlands)

    Man, M. de; Derksen, E.; Pieters, G.F.F.M.; Boer, H. de

    2008-01-01

    Conventional diagnostic procedures failed to provide a definitive diagnosis in a 15 year-old girl presenting with severe hirsutism, oligomenorrhea, and markedly elevated serum testosterone levels. To examine whether androgen overproduction was luteinizing hormone (LH) dependent and thus likely of

  2. Genetic predisposition syndromes: when should they be considered in the work-up of MDS?

    Science.gov (United States)

    Babushok, Daria V; Bessler, Monica

    2015-03-01

    Myelodysplastic syndromes (MDS) are clonal hematopoietic disorders characterized by cytopenias, ineffective hematopoiesis, myelodysplasia, and an increased risk of acute myeloid leukemia (AML). While sporadic MDS is primarily a disease of the elderly, MDS in children and young and middle-aged adults is frequently associated with underlying genetic predisposition syndromes. In addition to the classic hereditary bone marrow failure syndromes (BMFS) such as Fanconi Anemia and Dyskeratosis Congenita, in recent years there has been an increased awareness of non-syndromic familial MDS/AML predisposition syndromes such as those caused by mutations in GATA2, RUNX1, CEBPA, and SRP72 genes. Here, we will discuss the importance of recognizing an underlying genetic predisposition syndrome a patient with MDS, will review clinical scenarios when genetic predisposition should be considered, and will provide a practical overview of the common BMFS and familial MDS/AML syndromes which may be encountered in adult patients with MDS. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. [Ischemic stroke. Prevalence of cardiovascular causes documented by an extensive cardiovascular workup in 110 patients].

    Science.gov (United States)

    Bendriss, L; Khatouri, A

    2012-08-01

    The ischemic cerebrovascular accidents (I CVA) correspond to a pathology widely dominated by atherosclerosis and embolic cardiopathies. Our work aimed to determine the frequency of the cardiovascular diseases among the patients who were previously victims of an I CVA and the interest of the cardiovascular assessment in the etiologic inquest. We led a retrospective study in the cardiology service of the Avicenne military hospital of Marrakech about 110 cases of I CVA between January 2005 and August 2008. The electrocardiogram (ECG), Holter ECG, transthoracic echocardiography and Doppler echography of the cervical vessels were systematically made for all the patients. The transesophageal echocardiography was practice in a few patients. The average age of the patients was 60.8 years old (±12.14) with a male predominance (72%). Ninety-one percent of the patients presented at least one cardiovascular risk factor: hypertension (66.45%), diabetes (41.8%), smoking (35.45%). Cardiovascular antecedents were noted among 18.2% of the patients, the continuous atrial fibrillation comes first (9%). A carotid atheromatous excess was noted in 74 cases of which 24 with a significant plaque. The transesophageal echocardiography made to 13 patients showed a spontaneous echo contrast with a left atrial thrombus in four cases and a left atrial myxoma in one patient. Penetrating artery disease occupies 39%, large artery atherosclerosis 28% and cardiogenic stroke 18%. The cardiovascular assessment is indispensable, and the echocardiography is more interesting in presence of cardiopathy. Its therapeutic repercussion is modest. Copyright © 2012. Published by Elsevier SAS.

  4. Cytogenetic analysis after evaluation of 750 fetal deaths : proposal for diagnostic workup

    NARCIS (Netherlands)

    Korteweg, Fleurisca J.; Bouman, Katelijne; Erwich, Jan Jaap H. M.; Timmer, Albertus; Veeger, Nic J. G. M.; Ravise, Joke M.; Nijman, Thomas H.; Holm, Andjozien P.

    OBJECTIVE: To estimate success rates for cytogenetic analysis in different tissues after intrauterine fetal death, and study selection criteria and value of cytogenetic testing in determining cause of death. METHODS: Cytogenetic analyses and the value of this test in determining cause by a

  5. Cushing Syndrome: Diagnostic Workup and Imaging Features, With Clinical and Pathologic Correlation.

    Science.gov (United States)

    Wagner-Bartak, Nicolaus A; Baiomy, Ali; Habra, Mouhammed Amir; Mukhi, Shalini V; Morani, Ajaykumar C; Korivi, Brinda R; Waguespack, Steven G; Elsayes, Khaled M

    2017-07-01

    Cushing syndrome (CS) is a constellation of clinical signs and symptoms resulting from chronic exposure to excess cortisol, either exogenous or endogenous. Exogenous CS is most commonly caused by administration of glucocorticoids. Endogenous CS is subdivided into two types: adrenocorticotropic hormone (ACTH) dependent and ACTH independent. Cushing disease, which is caused by a pituitary adenoma, is the most common cause of ACTH-dependent CS for which pituitary MRI can be diagnostic, with bilateral inferior petrosal sinus sampling useful in equivocal cases. In ectopic ACTH production, which is usually caused by a tumor in the thorax (e.g., small cell lung carcinoma, bronchial and thymic carcinoids, or medullary thyroid carcinoma) or abdomen (e.g., gastroenteropancreatic neuroendocrine tumors or pheochromocytoma), CT, MRI, and nuclear medicine tests are used for localizing the source of ACTH. In ACTH-independent CS, which is caused by various adrenal abnormalities, adrenal protocol CT or MRI is usually diagnostic.

  6. The importance of social history in the work-up of painful ophthalmoplegia

    Directory of Open Access Journals (Sweden)

    Jun Mu, MD, PhD

    2014-12-01

    Full Text Available Painful ophthalmoplegia is not one uncommon clinic phenomenon in both ophthalmological and neurological clinics, which can be caused by various pathological conditions and easily be misdiagnosed by some clinical practitioners. Extensive clinical, laboratory and imaging assessment is required to exclude or confirm the diagnosis. Herein, we report an unusual case of painful ophthalmoplegia caused by orbital wall fracture in one young lady. Our case indicates that orbit wall fracture can be a rare cause of painful ophthalmoplegia, which may enrich our etiological diagnosis and avoid some unnecessary mistakes in certain cases of ophthalmoplegia.

  7. Anaerobiospirillum succiniciproducens sepsis in an autopsy patient: A troublesome diagnostic workup

    Directory of Open Access Journals (Sweden)

    Jason Koshy

    2014-01-01

    Full Text Available Anaerobiospirillum succiniciproducens is an uncommon yet potentially lethal gram-negative bacterium typically affecting patients with comorbidities. We report a case of A. succiniciproducens infection in an autopsy patient who had hepatitis C and type 2 diabetes and describe the difficulties in the laboratory identification of this pathogen.

  8. Confirmation of translatability and functionality certifies the dual endothelin1/VEGFsp receptor (DEspR) protein.

    Science.gov (United States)

    Herrera, Victoria L M; Steffen, Martin; Moran, Ann Marie; Tan, Glaiza A; Pasion, Khristine A; Rivera, Keith; Pappin, Darryl J; Ruiz-Opazo, Nelson

    2016-06-14

    In contrast to rat and mouse databases, the NCBI gene database lists the human dual-endothelin1/VEGFsp receptor (DEspR, formerly Dear) as a unitary transcribed pseudogene due to a stop [TGA]-codon at codon#14 in automated DNA and RNA sequences. However, re-analysis is needed given prior single gene studies detected a tryptophan [TGG]-codon#14 by manual Sanger sequencing, demonstrated DEspR translatability and functionality, and since the demonstration of actual non-translatability through expression studies, the standard-of-excellence for pseudogene designation, has not been performed. Re-analysis must meet UNIPROT criteria for demonstration of a protein's existence at the highest (protein) level, which a priori, would override DNA- or RNA-based deductions. To dissect the nucleotide sequence discrepancy, we performed Maxam-Gilbert sequencing and reviewed 727 RNA-seq entries. To comply with the highest level multiple UNIPROT criteria for determining DEspR's existence, we performed various experiments using multiple anti-DEspR monoclonal antibodies (mAbs) targeting distinct DEspR epitopes with one spanning the contested tryptophan [TGG]-codon#14, assessing: (a) DEspR protein expression, (b) predicted full-length protein size, (c) sequence-predicted protein-specific properties beyond codon#14: receptor glycosylation and internalization, (d) protein-partner interactions, and (e) DEspR functionality via DEspR-inhibition effects. Maxam-Gilbert sequencing and some RNA-seq entries demonstrate two guanines, hence a tryptophan [TGG]-codon#14 within a compression site spanning an error-prone compression sequence motif. Western blot analysis using anti-DEspR mAbs targeting distinct DEspR epitopes detect the identical glycosylated 17.5 kDa pull-down protein. Decrease in DEspR-protein size after PNGase-F digest demonstrates post-translational glycosylation, concordant with the consensus-glycosylation site beyond codon#14. Like other small single-transmembrane proteins, mass

  9. Simple plasma work-up for a fast chromatographic analysis of homocysteine, cysteine, methionine and aromatic amino acids

    Czech Academy of Sciences Publication Activity Database

    Hušek, Petr; Matucha, P.; Vránková, A.; Šimek, Petr

    2003-01-01

    Roč. 789, - (2003), s. 311-322 ISSN 1570-0232 R&D Projects: GA AV ČR IPP1050128; GA MZd NB6708 Institutional research plan: CEZ:AV0Z5007907 Keywords : Homocysteine * cysteine * methionine Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 2.085, year: 2003

  10. Conventional 3T brain MRI and diffusion tensor imaging in the diagnostic workup of early stage parkinsonism

    International Nuclear Information System (INIS)

    Meijer, Frederick J.A.; Rumund, Anouke van; Tuladhar, Anil M.; Aerts, Marjolein B.; Titulaer, Imke; Esselink, Rianne A.J.; Bloem, Bastiaan R.; Verbeek, Marcel M.; Goraj, Bozena

    2015-01-01

    The aim of this study is to evaluate whether the diagnostic accuracy of 3 T brain MRI is improved by region of interest (ROI) measures of diffusion tensor imaging (DTI), to differentiate between neurodegenerative atypical parkinsonism (AP) and Parkinson's disease (PD) in early stage parkinsonism. We performed a prospective observational cohort study of 60 patients presenting with early stage parkinsonism and initial uncertain diagnosis. At baseline, patients underwent a 3 T brain MRI including DTI. After clinical follow-up (mean 28.3 months), diagnoses could be made in 49 patients (30 PD and 19 AP). Conventional brain MRI was evaluated for regions of atrophy and signal intensity changes. Tract-based spatial statistics and ROI analyses of DTI were performed to analyze group differences in mean diffusivity (MD) and fractional anisotropy (FA), and diagnostic thresholds were determined. Diagnostic accuracy of conventional brain MRI and DTI was assessed with the receiver operating characteristic (ROC). Significantly higher MD of the centrum semiovale, body corpus callosum, putamen, external capsule, midbrain, superior cerebellum, and superior cerebellar peduncles was found in AP. Significantly increased MD of the putamen was found in multiple system atrophy-parkinsonian form (MSA-P) and increased MD in the midbrain and superior cerebellar peduncles in progressive supranuclear palsy (PSP). The diagnostic accuracy of brain MRI to identify AP as a group was not improved by ROI measures of MD, though the diagnostic accuracy to identify MSA-P was slightly increased (AUC 0.82 to 0.85). The diagnostic accuracy of brain MRI to identify AP as a group was not improved by the current analysis approach to DTI, though DTI measures could be of added value to identify AP subgroups. (orig.)

  11. Problems of radiologic diagnosis in the work-up and treatment of urologic diseases in the pelvis

    International Nuclear Information System (INIS)

    Marberger, M.

    1979-01-01

    From a clinical standpoint problems are set forth the answers which the urologist awaits from radiologic diagnosis in obstruction of the distal ureter, space-occupying lesions of the true pelvis, infravesical obstruction, recurrent urinary tract infections in childhood, incontinence, and injuries of the lower urinary tract. Presented at the same time are suggestions as to how the desired information can be most simply and reliably obtained, with regard to the rationale and risk for the patient. From this there follows an appraisal of the different examination methods, whereby emphasis is placed upon the socalled urologic basic study. (orig.) 891 MG/orig. 892 MKO [de

  12. Non-invasive diagnostic work-up of patients with clinically suspected pulmonary embolism; results of a management study

    NARCIS (Netherlands)

    ten Wolde, M.; Hagen, P. J.; Macgillavry, M. R.; Pollen, I. J.; Mairuhu, A. T. A.; Koopman, M. M. W.; Prins, M. H.; Hoekstra, O. S.; Brandjes, D. P. M.; Postmus, P. E.; Büller, H. R.

    2004-01-01

    Background: Clinicians often deviate from the recommended algorithm for the diagnosis of pulmonary embolism consisting of ventilation-perfusion scintigraphy and pulmonary angiography. Objectives: To assess the safety and feasibility of a diagnostic algorithm which reduces the need for lung

  13. Conventional 3T brain MRI and diffusion tensor imaging in the diagnostic workup of early stage parkinsonism

    Energy Technology Data Exchange (ETDEWEB)

    Meijer, Frederick J.A. [Radboud University Nijmegen Medical Center, Department of Radiology and Nuclear Medicine, Nijmegen (Netherlands); Rumund, Anouke van; Tuladhar, Anil M.; Aerts, Marjolein B.; Titulaer, Imke; Esselink, Rianne A.J.; Bloem, Bastiaan R. [Radboud University Nijmegen Medical Center, Department of Neurology, Donders Institute for Brain, Cognition and Behaviour, Nijmegen (Netherlands); Verbeek, Marcel M. [Radboud University Nijmegen Medical Center, Department of Neurology, Donders Institute for Brain, Cognition and Behaviour, Nijmegen (Netherlands); Radboud University Nijmegen Medical Center, Department of Laboratory Medicine, Nijmegen (Netherlands); Goraj, Bozena [Radboud University Nijmegen Medical Center, Department of Radiology and Nuclear Medicine, Nijmegen (Netherlands); Medical Center of Postgraduate Education, Department of Diagnostic Imaging, Warsaw (Poland)

    2015-07-15

    The aim of this study is to evaluate whether the diagnostic accuracy of 3 T brain MRI is improved by region of interest (ROI) measures of diffusion tensor imaging (DTI), to differentiate between neurodegenerative atypical parkinsonism (AP) and Parkinson's disease (PD) in early stage parkinsonism. We performed a prospective observational cohort study of 60 patients presenting with early stage parkinsonism and initial uncertain diagnosis. At baseline, patients underwent a 3 T brain MRI including DTI. After clinical follow-up (mean 28.3 months), diagnoses could be made in 49 patients (30 PD and 19 AP). Conventional brain MRI was evaluated for regions of atrophy and signal intensity changes. Tract-based spatial statistics and ROI analyses of DTI were performed to analyze group differences in mean diffusivity (MD) and fractional anisotropy (FA), and diagnostic thresholds were determined. Diagnostic accuracy of conventional brain MRI and DTI was assessed with the receiver operating characteristic (ROC). Significantly higher MD of the centrum semiovale, body corpus callosum, putamen, external capsule, midbrain, superior cerebellum, and superior cerebellar peduncles was found in AP. Significantly increased MD of the putamen was found in multiple system atrophy-parkinsonian form (MSA-P) and increased MD in the midbrain and superior cerebellar peduncles in progressive supranuclear palsy (PSP). The diagnostic accuracy of brain MRI to identify AP as a group was not improved by ROI measures of MD, though the diagnostic accuracy to identify MSA-P was slightly increased (AUC 0.82 to 0.85). The diagnostic accuracy of brain MRI to identify AP as a group was not improved by the current analysis approach to DTI, though DTI measures could be of added value to identify AP subgroups. (orig.)

  14. Screening for lung cancer with digital chest radiography: sensitivity and number of secondary work-up CT examinations

    NARCIS (Netherlands)

    de Hoop, Bartjan; Schaefer-Prokop, Cornelia; Gietema, Hester A.; de Jong, Pim A.; van Ginneken, Bram; van Klaveren, Rob J.; Prokop, Mathias

    2010-01-01

    To estimate the performance of digital chest radiography for detection of lung cancer. The study had ethics committee approval, and a nested case-control design was used and included 55 patients with lung cancer detected at computed tomography (CT) and confirmed with histologic examination and a

  15. NON-INVASIVE METHODS OF THE WORK-UP FOR ASSESSMENT OF MORPHOLOGIC AND FUNCTIONAL STATE OF THE SIGMOID WALL

    Directory of Open Access Journals (Sweden)

    A. E. Mashkov

    2015-01-01

    Full Text Available Background: Prolonged colonic congestion in children with chronic constipation and dolichosigma are characterized  by a permanent imbalance of gut microflora, secondary inflammation and degeneration of the sigmoid wall. There is plenty of research papers on the optic non-invasive diagnostics in medicine, based on spectrophotometry and laser spectral analysis. Aim: To study morphologic and functional state  of the sigmoid wall for detection of inflammation  and  degeneration in the  sigmoid  wall and  optimization  of treatment of children with dolichosigma  and long-standing constipation. Materials and methods: From 2009 to 2014, 30 children with dolichosigma  were seen in the Department of Pediatric surgery of MONIKI. All patients  were  hospitalized  after unsuccessful conservative  treatment in in-patient  clinics of the Moscow Region. The children underwent a set of investigations  for objective assessment of degree of the secondary  inflammatory and degenerative abnormalities  in the sigmoid wall, such as microbiological assessment, cytological assessment and fluorescent  diagnostics.  Results:  There  was  no   caused  by dolichosigma. It maintains  chronic inflammation  and  may play an indirect  role in abnormalities  of gut  motor  function. Inflammatory and  degenerative abnormalities  were  confirmed by a cytological investigation  of wall-adjacent biopsy of the  sigma. The results of the  complex assessment showed  moderate inflammation  and degeneration in the  sigmoid wall in 20 children; subsequent conservative treatment of chronic colostasis was effective. Ten children had advanced secondary inflammatory and degenerative abnormalities of the  sigmoid  wall, with high  levels of elastin and collagen in the colon wall. Surgery was performed in 6 children with the highest degree of fibrous transformation of the sigma. Conclusion: Complex assessment of the sigmoid wall, including  fluorescent  diagnostics  and  microbiological and  cytological  investigations, allows for detection  and  description  of secondary  inflammatory and  degenerative abnormalities  and  subsequent choice of individualized treatment.

  16. Diagnostic work-up of 449 consecutive girls who were referred to be evaluated for precocious puberty

    DEFF Research Database (Denmark)

    Mogensen, Signe Sloth; Aksglaede, Lise; Mouritsen, Annette

    2011-01-01

    A decrease in age at pubertal onset has been observed internationally. The aim of this study was to describe a large cohort of Caucasian girls referred with signs of early puberty according to etiology and compare biochemical characteristics.......A decrease in age at pubertal onset has been observed internationally. The aim of this study was to describe a large cohort of Caucasian girls referred with signs of early puberty according to etiology and compare biochemical characteristics....

  17. Additive value of amyloid-PET in routine cases of clinical dementia work-up after FDG-PET

    International Nuclear Information System (INIS)

    Brendel, Matthias; Schnabel, Jonas; Wagner, Leonie; Brendel, Eva; Meyer-Wilmes, Johanna; Unterrainer, Marcus; Schoenecker, Sonja; Prix, Catharina; Ackl, Nibal; Schildan, Andreas; Patt, Marianne; Barthel, Henryk; Sabri, Osama; Catak, Cihan; Pogarell, Oliver; Levin, Johannes; Danek, Adrian; Buerger, Katharina; Bartenstein, Peter; Rominger, Axel

    2017-01-01

    In recent years, several [ 18 F]-labeled amyloid-PET tracers have been developed and have obtained clinical approval. Despite their widespread scientific use, studies in routine clinical settings are limited. We therefore investigated the impact of [ 18 F]-florbetaben (FBB)-PET on the diagnostic management of patients with suspected dementia that was still unclarified after [ 18 F]-fluordeoxyglucose (FDG)-PET. All subjects were referred in-house with a suspected dementia syndrome due to neurodegenerative disease. After undergoing an FDG-PET exam, the cases were discussed by the interdisciplinary dementia board, where the most likely diagnosis as well as potential differential diagnoses were documented. Because of persistent diagnostic uncertainty, the patients received an additional FBB-PET exam. Results were interpreted visually and classified as amyloid-positive or amyloid-negative, and we then compared the individual clinical diagnoses before and after additional FBB-PET. A total of 107 patients (mean age 69.4 ± 9.7y) were included in the study. The FBB-PET was rated as amyloid-positive in 65/107. In 83% of the formerly unclear cases, a final diagnosis was reached through FBB-PET, and the most likely prior diagnosis was changed in 28% of cases. The highest impact was observed for distinguishing Alzheimer's dementia (AD) from fronto-temporal dementia (FTLD), where FBB-PET altered the most likely diagnosis in 41% of cases. FBB-PET has a high additive value in establishing a final diagnosis in suspected dementia cases when prior investigations such as FDG-PET are inconclusive. The differentiation between AD and FTLD was particularly facilitated by amyloid-PET, predicting a considerable impact on patient management, especially in the light of upcoming disease-modifying therapies. (orig.)

  18. Preanalytical blood sample workup for cell-free DNA analysis using Droplet Digital PCR for future molecular cancer diagnostics

    NARCIS (Netherlands)

    van Ginkel, Joost H.; van den Broek, Daan A.; van Kuik, Joyce; Linders, Dorothé; de Weger, Roel; Willems, Stefan M.; Huibers, Manon M.H.

    2017-01-01

    In current molecular cancer diagnostics, using blood samples of cancer patients for the detection of genetic alterations in plasma (cell-free) circulating tumor DNA (ctDNA) is an emerging practice. Since ctDNA levels in blood are low, highly sensitive Droplet Digital PCR (ddPCR) can be used for

  19. Safety analysis of holmium-166 microsphere scout dose imaging during radioembolisation work-up. A cohort study

    Energy Technology Data Exchange (ETDEWEB)

    Braat, Arthur J.A.T.; Prince, Jip F.; Rooij, Rob van; Bruijnen, Rutger C.G.; Bosch, Maurice A.A.J. van den; Lam, Marnix G.E.H. [University Medical Center Utrecht, Department of Radiology and Nuclear Medicine, Utrecht (Netherlands)

    2018-03-15

    Radioembolisation is generally preceded by a scout dose of technetium-99m-macroaggregated albumin to estimate extrahepatic shunting of activity. Holmium-166 microspheres can be used as a scout dose (±250 MBq) and as a therapeutic dose. The general toxicity of a holmium-166 scout dose ({sup 166}Ho-SD) and safety concerns of an accidental extrahepatic deposition of {sup 166}Ho-SD were investigated. All patients who received a {sup 166}Ho-SD in our institute were reviewed for general toxicity and extrahepatic depositions. The absorbed dose in extrahepatic tissue was calculated on SPECT/CT and correlated to clinical toxicities. In total, 82 patients were included. No relevant clinical toxicity occurred. Six patients had an extrahepatic deposition of {sup 166}Ho-SD (median administered activity 270 MBq). The extrahepatic depositions (median activity 3.7 MBq) were located in the duodenum (3x), gastric fundus, falciform ligament and the lesser curvature of the stomach, and were deposited in a median volume of 15.3 ml, which resulted in an estimated median absorbed dose of 3.6 Gy (range 0.3-13.8 Gy). No adverse events related to the extrahepatic deposition of the {sup 166}Ho-SD occurred after a median follow-up of 4 months (range 1-12 months). These results support the safety of 250 MBq {sup 166}Ho-SD in a clinical setting. (orig.)

  20. Tailoring Stress Incontinence Surgery : “How to install proper diagnostic work-up and individualised treatment”

    NARCIS (Netherlands)

    Houwert, R.M.

    2009-01-01

    In this thesis a different principle to determine the value of diagnostic parameters and to install the proper, individualized, surgical treatment for stress urinary incontinence is presented. The aim of the first part of this thesis was to evaluate the value of urodynamic investigation (UDI) in the

  1. Accuracy of FDG-PET-CT in the Diagnostic Work-up of Vascular Prosthetic Graft Infection

    NARCIS (Netherlands)

    Bruggink, J. L. M.; Glaudemans, A. W. J. M.; Saleem, B. R.; Meerwaldt, R.; Alkefaji, H.; Prins, T. R.; Start, R. H. J. A.; Zeebregts, C. J.

    Objectives: To investigate the diagnostic accuracy of fluoro-2-deoxy-D-glucose positron emission tomography (FDG-PET) compared with computed tomography (CT) scanning and added value of fused FDG-PET CT in diagnosing vascular prosthetic graft infection. Design: Prospective cohort study with

  2. Idiopathic interstitial pneumonias. From classification to diagnostic work-up; Idiopathische interstitielle Pneumonien. Von der Klassifikation zur Diagnostik

    Energy Technology Data Exchange (ETDEWEB)

    Mueller-Mang, C.; Bankier, A. [Medizinische Universitaet, Universitaetsklinik fuer Radiodiagnostik, Wien (Austria); Stiebellehner, L. [Medizinische Universitaet, Klinische Abteilung fuer Pulmologie, Universitaetsklinik fuer Innere Medizin, Wien (Austria); Schmid, K. [Medizinische Universitaet, Klinisches Institut fuer Pathologie, Wien (Austria)

    2007-05-15

    Idiopathic interstitial pneumonias (IIP) comprise seven entities with distinct histologic patterns. In their idiopathic form IIP are rare diseases. They are, nevertheless, considered prototypes of the much more common secondary interstitial pneumonias. The advent of high-resolution computed tomography (HRCT) has had a profound impact on the imaging of IIP, because the detailed delineation of the lung anatomy allows a close correlation between the histologic patterns of IIP and the CT features. On the basis of CT morphology and in the correct clinical context, the radiologist can achieve an accurate diagnosis in many cases. However, due to overlap between the various entities, complementary lung biopsy is recommended in virtually all cases. This article reviews the CT pattern of IIP and offers relevant clinical and histological information for the purpose of enabling the radiologist to understand and participate in the multidisciplinary concept of IIP. (orig.) [German] Die Bezeichnung idiopathische interstitielle Pneumonien (IIP) umfasst 7 Entitaeten mit definierten histologischen Mustern. In ihrer idiopathischen Form sind die interstitiellen Pneumonien selten, allerdings werden die einzelnen Muster als Prototypen fuer die haeufigeren sekundaeren interstitiellen Pneumonien betrachtet. Mit der Einfuehrung der hochaufloesenden Computertomographie (HRCT) hat die Bildgebung der IIP einen deutlichen Aufschwung erfahren, da durch die detaillierte Darstellung der Lungenanatomie eine sehr gute Korrelation zwischen den histologischen Mustern der IIP und den CT-Veraenderungen gegeben ist. Der Radiologe kann anhand der CT-Morphologie und unter Beruecksichtigung der Klinik des Patienten in vielen Faellen eine korrekte Diagnose erstellen. Dennoch wird aufgrund von Ueberlappungen zwischen den verschiedenen Entitaeten bei nahezu allen Formen der IIP eine ergaenzende Lungenbiopsie empfohlen. Dieser Uebersichtsartikel veranschaulicht die verschiedenen CT-Merkmale der IIP und beinhaltet relevante klinische und histologische Informationen, mit dem Ziel, dem Radiologen das interdisziplinaere Konzept der IIP naeher zu bringen und ihm dadurch ein Instrument zur verbesserten Kommunikation mit den zuweisenden Klinikern in die Hand zu geben. (orig.)

  3. Additive value of amyloid-PET in routine cases of clinical dementia work-up after FDG-PET

    Energy Technology Data Exchange (ETDEWEB)

    Brendel, Matthias; Schnabel, Jonas; Wagner, Leonie; Brendel, Eva; Meyer-Wilmes, Johanna; Unterrainer, Marcus [University Hospital, LMU Munich, Department of Nuclear Medicine, Munich (Germany); Schoenecker, Sonja; Prix, Catharina; Ackl, Nibal [University Hospital, LMU Munich, Department of Neurology, Munich (Germany); Schildan, Andreas; Patt, Marianne; Barthel, Henryk; Sabri, Osama [University of Leipzig, Department of Nuclear Medicine, Leipzig (Germany); Catak, Cihan [Klinikum der Universitaet Muenchen, Institute for Stroke and Dementia Research, Munich (Germany); Pogarell, Oliver [University Hospital, LMU Munich, Department of Psychiatry, Munich (Germany); Levin, Johannes; Danek, Adrian [University Hospital, LMU Munich, Department of Neurology, Munich (Germany); DZNE - German Center for Neurodegenerative Diseases, Munich (Germany); Buerger, Katharina [Klinikum der Universitaet Muenchen, Institute for Stroke and Dementia Research, Munich (Germany); DZNE - German Center for Neurodegenerative Diseases, Munich (Germany); Bartenstein, Peter; Rominger, Axel [University Hospital, LMU Munich, Department of Nuclear Medicine, Munich (Germany); Munich Cluster for Systems Neurology (SyNergy), Munich (Germany)

    2017-12-15

    In recent years, several [{sup 18}F]-labeled amyloid-PET tracers have been developed and have obtained clinical approval. Despite their widespread scientific use, studies in routine clinical settings are limited. We therefore investigated the impact of [{sup 18}F]-florbetaben (FBB)-PET on the diagnostic management of patients with suspected dementia that was still unclarified after [{sup 18}F]-fluordeoxyglucose (FDG)-PET. All subjects were referred in-house with a suspected dementia syndrome due to neurodegenerative disease. After undergoing an FDG-PET exam, the cases were discussed by the interdisciplinary dementia board, where the most likely diagnosis as well as potential differential diagnoses were documented. Because of persistent diagnostic uncertainty, the patients received an additional FBB-PET exam. Results were interpreted visually and classified as amyloid-positive or amyloid-negative, and we then compared the individual clinical diagnoses before and after additional FBB-PET. A total of 107 patients (mean age 69.4 ± 9.7y) were included in the study. The FBB-PET was rated as amyloid-positive in 65/107. In 83% of the formerly unclear cases, a final diagnosis was reached through FBB-PET, and the most likely prior diagnosis was changed in 28% of cases. The highest impact was observed for distinguishing Alzheimer's dementia (AD) from fronto-temporal dementia (FTLD), where FBB-PET altered the most likely diagnosis in 41% of cases. FBB-PET has a high additive value in establishing a final diagnosis in suspected dementia cases when prior investigations such as FDG-PET are inconclusive. The differentiation between AD and FTLD was particularly facilitated by amyloid-PET, predicting a considerable impact on patient management, especially in the light of upcoming disease-modifying therapies. (orig.)

  4. Molecular diagnostics of catecholaminergic polymorphic ventricular tachycardia using denaturing high-performance liquid chromatography and sequencing

    NARCIS (Netherlands)

    Postma, Alex V.; Bhuiyan, Zahurul A.; Bikker, Hennie

    2006-01-01

    Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an arrhythmogenic disease characterized by adrenergic-induced arrhythmias in the form of bidirectional and PVT. CPVT is a distinct clinical entity associated with a high mortality rate of up to 50% by the age of 30 yr. Recently, the

  5. Induced refolding of a temperature denatured llama heavy-chain antibody fragment by its antigen

    NARCIS (Netherlands)

    Dolk, E.; Vliet, C. van; Perez, J.M.J.; Vriend, G.; Darbon, H.; Ferrat, G.; Cambillau, C.; Frenken, L.G.J.; Verrips, T.

    2005-01-01

    In a previous study we have shown that llama VHH antibody fragments are able to bind their antigen after a heat shock of 90°C, in contrast to the murine monoclonal antibodies. However, the molecular mechanism by which antibody:antigen interaction occurs under these extreme conditions remains

  6. Quantifying why urea is a protein denaturant, whereas glycine betaine is a protein stabilizer

    Science.gov (United States)

    Guinn, Emily J.; Pegram, Laurel M.; Capp, Michael W.; Pollock, Michelle N.; Record, M. Thomas

    2011-01-01

    To explain the large, opposite effects of urea and glycine betaine (GB) on stability of folded proteins and protein complexes, we quantify and interpret preferential interactions of urea with 45 model compounds displaying protein functional groups and compare with a previous analysis of GB. This information is needed to use urea as a probe of coupled folding in protein processes and to tune molecular dynamics force fields. Preferential interactions between urea and model compounds relative to their interactions with water are determined by osmometry or solubility and dissected using a unique coarse-grained analysis to obtain interaction potentials quantifying the interaction of urea with each significant type of protein surface (aliphatic, aromatic hydrocarbon (C); polar and charged N and O). Microscopic local-bulk partition coefficients Kp for the accumulation or exclusion of urea in the water of hydration of these surfaces relative to bulk water are obtained. Kp values reveal that urea accumulates moderately at amide O and weakly at aliphatic C, whereas GB is excluded from both. These results provide both thermodynamic and molecular explanations for the opposite effects of urea and glycine betaine on protein stability, as well as deductions about strengths of amide NH—amide O and amide NH—amide N hydrogen bonds relative to hydrogen bonds to water. Interestingly, urea, like GB, is moderately accumulated at aromatic C surface. Urea m-values for protein folding and other protein processes are quantitatively interpreted and predicted using these urea interaction potentials or Kp values. PMID:21930943

  7. Fast-scan cyclic voltammetry with thiol-modified mercury electrodes distinguishes native from denatured BSA

    Czech Academy of Sciences Publication Activity Database

    Černocká, Hana; Ostatná, Veronika; Paleček, Emil

    2015-01-01

    Roč. 61, DEC2015 (2015), s. 114-116 ISSN 1388-2481 R&D Projects: GA ČR(CZ) GA15-15479S Institutional support: RVO:68081707 Keywords : catalyzed hydrogen evolution * proteins * electrocatalysis Subject RIV: BO - Biophysics Impact factor: 4.569, year: 2015

  8. Thermodynamic stability and denaturation kinetics of a benign natural transthyretin mutant identified in a Danish kindred

    DEFF Research Database (Denmark)

    Jensen, Minna Grønning; Campos, Raul I.; Fagerberg, Christina

    2011-01-01

    The disease phenotype of transthyretin (TTR) is dramatically influenced by single point mutations in the TTR gene. Herein, we report on a novel mutation D99N (Asp99Asn) in TTR found in a Danish kindred. None of the family members carrying this mutation have so far shown any clinical signs...... of amyloidosis. One carrier found compound heterozygous for TTR D99N and L111M (Leu111Met) associated with cardiac amyloid is asymptomatic (42 years). Disease severity can often be linked to both the kinetics of fibril formation and the degree of destabilisation of the native state. In this study, we show...

  9. Legumin allergens from peanuts and soybeans: Effects of denaturation and aggregation on allergenicity

    NARCIS (Netherlands)

    Boxtel, E.L. van; Broek, L.A.M. van den; Koppelman, S.J.; Gruppen, H.

    2008-01-01

    Legumin proteins Ara h 3 from peanuts and glycinin from soybeans are increasingly described as important allergens. The stability of an allergen's IgE binding capacity towards heating and digestion is considered an important characteristic for food allergens. We investigated the effects of heating

  10. Legumin allergens from peanuts and soybeans : Effects of denaturation and aggregation on allergenicity

    NARCIS (Netherlands)

    Boxtel, van E.L.; Broek, van den L.A.M.; Koppelman, S.J.; Gruppen, H.

    2008-01-01

    Legumin proteins Ara h 3 from peanuts and glycinin from soybeans are increasingly described as important allergens. The stability of an allergen's IgE binding capacity towards heating and digestion is considered an important characteristic for food allergens. We investigated the effects of heating

  11. Na,K-ATPase structure/function relationships probed by the denaturant urea.

    Science.gov (United States)

    Esmann, Mikael; Fedosova, Natalya U; Olesen, Claus

    2015-05-01

    Urea interacts with the Na,K-ATPase, leading to reversible as well as irreversible inhibition of the hydrolytic activity. The enzyme purified from shark rectal glands is more sensitive to urea than Na,K-ATPase purified from pig kidney. An immediate and reversible inhibition under steady-state conditions of hydrolytic activity at 37°C is demonstrated for the three reactions studied: the overall Na,K-ATPase activity, the Na-ATPase activity observed in the absence of K+ as well as the K+-dependent phosphatase reaction (K-pNPPase) seen in the absence of Na+. Half-maximal inhibition is seen with about 1M urea for shark enzyme and about 2M urea for pig enzyme. In the presence of substrates there is also an irreversible inhibition in addition to the reversible process, and we show that ATP protects against the irreversible inhibition for both the Na,K-ATPase and Na-ATPase reaction, whereas the substrate paranitrophenylphosphate leads to a slight increase in the rate of irreversible inhibition of the K-pNPPase. The rate of the irreversible inactivation in the absence of substrates is much more rapid for shark enzyme than for pig enzyme. The larger number of potentially urea-sensitive hydrogen bonds in shark enzyme compared to pig enzyme suggests that interference with the extensive hydrogen bonding network might account for the higher urea sensitivity of shark enzyme. The reversible inactivation is interpreted in terms of domain interactions and domain accessibilities using as templates the available crystal structures of Na,K-ATPase. It is suggested that a few interdomain hydrogen bonds are those mainly affected by urea during reversible inactivation. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Detection and Identification of Ciprofloxacin-Resistant Yersinia pestis Denaturing High-Performance Liquid Chromatography

    Science.gov (United States)

    2003-07-01

    analysis in hereditary breast and ovarian cancers . Hum. Mutat. 14:333– 339. 2. Bauer, A. W., W. M. Kirby, J. C. Sherris, and M. Turck. 1966. Antibiotic...Listeria monocytogenes lineage group classification by MAMA -PCR of the listeriolysin gene. Curr. Microbiol. 43:129–133. 17. Klein, B., G. Weirich...Germline and somatic mutation anal- yses in the DNA mismatch repair gene MLH3: evidence for somatic muta- tion in colorectal cancers . Hum. Mutat. 17:389–396

  13. Denatured state is critical in determining the properties of model proteins designed on different folds

    DEFF Research Database (Denmark)

    Amatori, Andrea; Ferkinghoff-Borg, Jesper; Tiana, Guido

    2008-01-01

    The thermodynamics of proteins designed on three common folds (SH3, chymotrypsin inhibitor 2 [CI2], and protein G) is studied with a simplified C alpha, model and compared with the thermodynamics of proteins designed on random-generated folds. The model allows to design sequences to fold within a...

  14. Unrecorded alcohol consumption in Russia: toxic denaturants and disinfectants pose additional risks.

    Science.gov (United States)

    Solodun, Yuriy V; Monakhova, Yulia B; Kuballa, Thomas; Samokhvalov, Andriy V; Rehm, Jürgen; Lachenmeier, Dirk W

    2011-12-01

    In 2005, 30% of all alcohol consumption in Russia was unrecorded. This paper describes the chemical composition of unrecorded and low cost alcohol, including a toxicological evaluation. Alcohol products (n=22) from both recorded and unrecorded sources were obtained from three Russian cities (Saratov, Lipetsk and Irkutsk) and were chemically analyzed. Unrecorded alcohols included homemade samogons, medicinal alcohols and surrogate alcohols. Analysis included alcoholic strength, levels of volatile compounds (methanol, acetaldehyde, higher alcohols), ethyl carbamate, diethyl phthalate (DEP) and polyhexamethyleneguanidine hydrochloride (PHMG). Single samples showed contamination with DEP (275-1269 mg/l) and PHMG (515 mg/l) above levels of toxicological concern. Our detailed chemical analysis of Russian alcohols showed that the composition of vodka, samogon and medicinal alcohols generally did not raise major public health concerns other than for ethanol. It was shown, however, that concentration levels of DEP and PHMG in some surrogate alcohols make these samples unfit for human consumption as even moderate drinking would exceed acceptable daily intakes.

  15. Denaturing gradient gel electrophoresis as a fingerprinting method for the analysis of soil microbial communities

    Czech Academy of Sciences Publication Activity Database

    Valášková, Vendula; Baldrian, Petr

    2009-01-01

    Roč. 55, č. 10 (2009), s. 413-423 ISSN 1214-1178 R&D Projects: GA ČR GA526/08/0751; GA MŠk LC06066 Institutional research plan: CEZ:AV0Z50200510 Keywords : DGGE * bacteria * fungi Subject RIV: EE - Microbiology, Virology Impact factor: 0.697, year: 2009

  16. Detection of AGEs as markers for carbohydrate metabolism and protein denaturation

    OpenAIRE

    Nagai, Ryoji; Shirakawa, Jun-ichi; Fujiwara, Yukio; Ohno, Rei-ichi; Moroishi, Narumi; Sakata, Noriyuki; Nagai, Mime

    2014-01-01

    Approximately 100 years have passed since the Maillard reaction was first reported in the field of food chemistry as a condensation reaction between reducing sugars and amino acids. This reaction is thought to progress slowly primarily from glucose with proteins in vivo. An early-stage product, called the ”Amadori product”, is converted into advanced glycation end products. Those accumulate in the body in accordance with age, with such accumulation being enhanced by lifestyle-related diseases...

  17. Sorbitol counteracts high hydrostatic pressure-induced denaturation of inulin fructotransferase.

    Science.gov (United States)

    Li, Yungao; Miao, Ming; Liu, Miao; Jiang, Bo; Zhang, Tao; Chen, Xiangyin

    2014-09-01

    Inulin fructotransferase (IFTase), a novel hydrolase for inulin, was exposed to high hydrostatic pressure (HHP) at 400 and 600 MPa for 15 min in the presence or absence of sorbitol. Sorbitol protected the enzyme against HHP-induced activity loss. The relative residual activity increased nearly 3.1- and 3.8-fold in the presence of 3 mol/L sorbitol under 400 MPa and 600 MPa for 15 min, respectively. Circular dichroism results indicated that the original chaotic unfolding conformation of the enzyme under HHP shifted toward more ordered and impact with 3 mol/L sorbitol. Fluorescence and UV spectra results suggested that sorbitol prevented partially the unfolding of the enzyme and stabilized the conformation under high pressure. These results might be attributed to the binding of sorbitol on the surface of IFTase to rearrange and strengthen intra- and intermolecular hydrogen bonds. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. C5-Alkynyl-Functionalized ?-L-LNA: Synthesis, Thermal Denaturation Experiments and Enzymatic Stability

    OpenAIRE

    Kumar, Pawan; Baral, Bharat; Anderson, Brooke A.; Guenther, Dale C.; ?stergaard, Michael E.; Sharma, Pawan K.; Hrdlicka, Patrick J.

    2014-01-01

    Major efforts are currently being devoted to improving the binding affinity, target specificity, and enzymatic stability of oligonucleotides used for nucleic acid targeting applications in molecular biology, biotechnology, and medicinal chemistry. One of the most popular strategies toward this end has been to introduce additional modifications to the sugar ring of affinity-inducing conformationally restricted nucleotide building blocks such as locked nucleic acid (LNA). In the preceding artic...

  19. Determination of pulse energy dependence for skin denaturation from 585nm fibre laser

    Science.gov (United States)

    Mujica-Ascencio, S.; Velazquez-Gonzalez, J. S.; Mujica-Ascencio, C.; Alvarez-Chavez, J. A.

    2014-05-01

    In this paper, simulation and mathematical analysis for the determination of pulse energy from a Q-switched Yb3+-doped fibre laser is required in Port Wine Stain (PWS) treatment. The pulse energy depends on average power, gain, volume, repetition rate and pulse duration. In some treatments such as Selective Photothermolysis (SP), the peak power at the end of the optical fibre and pulse duration can be obtained and modified via a cavity design. For that purpose, a 585nm optical fibre laser full design which considers all of the above besides the average losses through the optical devices proposed for the design and the Ytterbium optical fibre overall gain will be presented.

  20. ASP53, a thermostable protein from Acacia erioloba seeds that protects target proteins against thermal denaturation

    CSIR Research Space (South Africa)

    Mtwisha, L

    2007-02-01

    Full Text Available ) and the Typha pollen D7 protein was found to stabilise sugar glasses in an in vitro system (Wolkers et al. 2001). The cupin family of proteins comprises a wide variety of proteins from both prokaryotes and eukaryotes and includes the seed storage proteins...–268. Garay-Arroyo A, Colmenero-Flores JM, Garciarrubio A, Covarrubias AA (2000) Highly hydrophilic proteins in prokaryotes and eukaryotes are common during conditions of water deficit. Journal of Biological Chemistry 275, 5668–5674. doi: 10.1074/jbc.275...