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Sample records for biochemical assays showed

  1. Automated optical sensing system for biochemical assays

    Science.gov (United States)

    Oroszlan, Peter; Duveneck, Gert L.; Ehrat, Markus; Widmer, H. M.

    1994-03-01

    In this paper, we present a new system called FOBIA that was developed and optimized with respect to automated operation of repetitive assay cycles with regenerable bioaffinity sensors. The reliability and precision of the new system is demonstrated by an application in a competitive assay for the detection of the triazine herbicide Atrazine. Using one sensor in more than 300 repetitive cycles, a signal precision better than 5% was achieved.

  2. Use of laminar flow patterning for miniaturised biochemical assays

    DEFF Research Database (Denmark)

    Regenberg, Birgitte; Krühne, Ulrich; Beyer, M.

    2004-01-01

    Laminar flow in microfluidic chambers was used to construct low (one dimensional) density arrays suitable for miniaturized biochemical assays. By varying the ratio of flows of two guiding streams flanking a sample stream, precise focusing and positioning of the latter was achieved, and reactive s...... species carried in the sample stream were deposited on functionalized chip surfaces as discrete 50 mm wide lanes. Using different model systems we have confirmed the method's suitability for qualitative screening and quantification tasks in receptor-ligand assays, recording biotin...

  3. Analytical interference by contrast agents in biochemical assays

    DEFF Research Database (Denmark)

    Otnes, Sigrid; Fogh-Andersen, Niels; Rømsing, Janne

    2017-01-01

    /4 of these CA concentrations. The results were assessed statistically by a paired Student’s t-test. Results. Iodixanol produced a negative interference on the bicarbonate (p = 0.011), lactate dehydrogenase (p ....0062), calcium (p bicarbonate (p = 0.0057) and magnesium (p = 0.0001) assays and a positive interference on the calcium (p .... Gadoxetate disodium produced a negative interference on the iron (p sodium (p = 0.032) assay. Conclusion. CAs cause analytical interference. Attention should be given to the above-mentioned analyte-CA combinations when assessing...

  4. CLSI-based transference of the CALIPER database of pediatric reference intervals to Beckman Coulter DxC biochemical assays.

    Science.gov (United States)

    Araújo, Petra A T; Thomas, Dylan; Sadeghieh, Tara; Bevilacqua, Victoria; Chan, Man Khun; Chen, Yunqi; Randell, Edward; Adeli, Khosrow

    2015-09-01

    The CALIPER program has established a comprehensive database of age- and sex-stratified pediatric reference intervals for over 85 common biochemical markers, largely using the Abbott ARCHITECT assays. To allow a broader application of the CALIPER database, we examined transference to 36 Beckman Coulter Synchron Unicel DxC800 assays, based on the CLSI C28-A3/EP9-A3 guidelines. Patient sample comparisons were performed for 36 biochemical assays using 200 serum specimens obtained from pediatric patients on the Abbott ARCHITECT ci8200 and the Beckman Coulter DxC800. For each analyte, R(2) values were calculated to assess the quality of correlation between the platforms. Statistical criteria used to assess transferability included a) regression analysis to create the equation of the line of best fit, b) standardized residual, c) Bland-Altman, and d) quantile-quantile plots. Transferred reference intervals were further verified by analyzing serum samples from 100 healthy children from the CALIPER cohort on the Beckman Coulter system. The reference intervals for most of the assessed analytes were transferable to Beckman Coulter assays (31 out of 36 studied) and the newly calculated reference intervals were verified through analysis of CALIPER reference samples (28 out of 31). Eighteen assays demonstrated excellent correlation (R(2)≥0.95), and 13 assays showed strong correlation (0.77≤R2≤0.94). The current study allowed successful transference of a large number of biochemical markers from the CALIPER database to assays on the Beckman Coulter DxC800 platform. Transference should facilitate broader application of CALIPER reference intervals at pediatric centers using DxC biochemical assays. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  5. Transference of CALIPER pediatric reference intervals to biochemical assays on the Roche cobas 6000 and the Roche Modular P.

    Science.gov (United States)

    Higgins, Victoria; Chan, Man Khun; Nieuwesteeg, Michelle; Hoffman, Barry R; Bromberg, Irvin L; Gornall, Doug; Randell, Edward; Adeli, Khosrow

    2016-01-01

    The Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) has recently established pediatric age- and sex-specific reference intervals for over 85 biochemical markers on the Abbott Architect system. Previously, CALIPER reference intervals for several biochemical markers were successfully transferred from Abbott assays to Roche, Beckman, Ortho, and Siemens assays. This study further broadens the CALIPER database by performing transference and verification for 52 biochemical assays on the Roche cobas 6000 and the Roche Modular P. Using CLSI C28-A3 and EP9-A2 guidelines, transference of the CALIPER reference intervals was attempted for 16 assays on the Roche cobas 6000 and 36 on the Modular P. Calculated reference intervals were further verified using 100 healthy CALIPER samples. Most assays showed strong correlation between assay systems and were transferable from Abbott to the Roche cobas 6000 (81%) and the Modular P (86%). Bicarbonate and magnesium were not transferable on either system and calcium and prealbumin were not transferable to the Modular P. Of the transferable analytes, 62% and 61% were verified on the cobas 6000 and the Modular P, respectively. This study extends the utility of the CALIPER database to two additional analytical systems, which facilitates the broad application of CALIPER reference intervals at pediatric centers utilizing Roche biochemical assays. Transference studies across different analytical platforms can later be collectively analyzed in an attempt to develop common reference intervals across all clinical chemistry instruments to harmonize laboratory test interpretation in diagnosis and monitoring of pediatric disease. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  6. A new trend to determine biochemical parameters by quantitative FRET assays.

    Science.gov (United States)

    Liao, Jia-yu; Song, Yang; Liu, Yan

    2015-12-01

    Förster resonance energy transfer (FRET) has been widely used in biological and biomedical research because it can determine molecule or particle interactions within a range of 1-10 nm. The sensitivity and efficiency of FRET strongly depend on the distance between the FRET donor and acceptor. Historically, FRET assays have been used to quantitatively deduce molecular distances. However, another major potential application of the FRET assay has not been fully exploited, that is, the use of FRET signals to quantitatively describe molecular interactive events. In this review, we discuss the use of quantitative FRET assays for the determination of biochemical parameters, such as the protein interaction dissociation constant (K(d)), enzymatic velocity (k(cat)) and K(m). We also describe fluorescent microscopy-based quantitative FRET assays for protein interaction affinity determination in cells as well as fluorimeter-based quantitative FRET assays for protein interaction and enzymatic parameter determination in solution.

  7. CLSI-based transference of CALIPER pediatric reference intervals to Beckman Coulter AU biochemical assays.

    Science.gov (United States)

    Abou El Hassan, Mohamed; Stoianov, Alexandra; Araújo, Petra A T; Sadeghieh, Tara; Chan, Man Khun; Chen, Yunqi; Randell, Edward; Nieuwesteeg, Michelle; Adeli, Khosrow

    2015-11-01

    The CALIPER program has established a comprehensive database of pediatric reference intervals using largely the Abbott ARCHITECT biochemical assays. To expand clinical application of CALIPER reference standards, the present study is aimed at transferring CALIPER reference intervals from the Abbott ARCHITECT to Beckman Coulter AU assays. Transference of CALIPER reference intervals was performed based on the CLSI guidelines C28-A3 and EP9-A2. The new reference intervals were directly verified using up to 100 reference samples from the healthy CALIPER cohort. We found a strong correlation between Abbott ARCHITECT and Beckman Coulter AU biochemical assays, allowing the transference of the vast majority (94%; 30 out of 32 assays) of CALIPER reference intervals previously established using Abbott assays. Transferred reference intervals were, in general, similar to previously published CALIPER reference intervals, with some exceptions. Most of the transferred reference intervals were sex-specific and were verified using healthy reference samples from the CALIPER biobank based on CLSI criteria. It is important to note that the comparisons performed between the Abbott and Beckman Coulter assays make no assumptions as to assay accuracy or which system is more correct/accurate. The majority of CALIPER reference intervals were transferrable to Beckman Coulter AU assays, allowing the establishment of a new database of pediatric reference intervals. This further expands the utility of the CALIPER database to clinical laboratories using the AU assays; however, each laboratory should validate these intervals for their analytical platform and local population as recommended by the CLSI. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  8. Development of a rapid ferricyanide mediated assay for biochemical oxygen demand using a mixed microbial consortium.

    Science.gov (United States)

    Catterall, Kylie; Zhao, Huijun; Pasco, Neil; John, Richard

    2003-06-01

    Ferricyanide-mediated (FM) microbial reactions were used for the rapid determination of the biochemical oxygen demand (BOD) of a range of synthetic and real wastewater samples. Four single-species microbial seeds and a synthetically prepared microbial consortium were compared. In all cases, the microbial consortium exhibited a greater extent and rate of biodegradation compared to the individual microbial seeds. Markedly improved correlation to the standard BOD5 method was also noted for the microbial consortium (compared to the single-species seeds). A linear dynamic range up to 200 mg BOD5 L(-1) was observed, which is considerably greater than the linear range of the standard BOD5 assay and most other rapid BOD assays reported. In addition, biodegradation efficiencies comparable to the 5-day BOD5 assay (and much greater than other rapid BOD assays) were observed in 3 h. A highly significant correlation (R = 0.935, p = 0.000, n = 30) between the FM-BOD method and the standard BOD5 method was found for a wide diversity of real wastewater samples. The results indicate that the FM-BOD assay is a promising, rapid, alternative to the standard 5-day BOD5 assay.

  9. Evaluation of intramyocellular lipid breakdown during exercise by biochemical assay, NMR spectroscopy, and Oil Red O staining

    OpenAIRE

    De Bock, Katrien; Dresselaers, Tom; Kiens, B; Richter, E. A; Van Hecke, Paul; Hespel, Peter

    2007-01-01

    The study compared the net decline of intramyocellular lipids (IMCL) during exercise (n = 18) measured by biochemical assay (BIO) and Oil Red O (ORO) staining on biopsy samples from vastus lateralis muscle and by (1)H-MR spectroscopy (MRS) sampled in an 11 x 11 x 18-mm(3) voxel in the same muscle. IMCL was measured before and after a 2-h cycling bout ( approximately 75% Vo(2 peak)). ORO and MRS measurements showed substantial IMCL use during exercise of 31 +/- 12 and 47 +/- 6% of preexercise ...

  10. Amoxicillin-associated interference in an HPLC-EC assay for urinary fractionated metanephrines: potential pitfall in pheochromocytoma biochemical diagnosis.

    Science.gov (United States)

    Barco, Sebastiano; Alpigiani, Maria Giannina; Ghiggeri, Gian Marco; Talio, Marina; Maffia, Angelo; Tripodi, Gino; Cangemi, Giuliana

    2014-10-01

    Measurement of urinary fractionated metanephrines represents a first-line test for the biochemical diagnosis of pheochromocytoma. The high performance liquid chromatography coupled to electrochemical detection (HPLC-EC) assays used in the routine clinical laboratory can be subjected to analytical interferences by the presence of drugs or their metabolites. In this paper we describe the interference on urinary normetanephrine (uNMN) caused by amoxicillin. Two pediatric patients suspected of pheochromocytoma had very high uNMN levels (2543 and 4227μg/g Cr respectively; upper reference value: 339μg/g Cr). Amoxicillin interference was assessed by comparison for co-elution with uNMN and by LC-MS/MS analysis. After amoxicillin interference was suspected and the therapy was stopped uNMN levels returned to normal (149 and 214μg/g Cr respectively). Chromatograms obtained by HPLC-EC clearly showed that amoxicillin co-elutes with uNMN. Patients' uNMN levels measured by LC-MS/MS were in the normal range. Amoxicillin is responsible for analytical interference on HPLC-EC assay for uNMN. This finding can be of help in distinguishing true-positive from false-positive results in the course of a biochemical diagnosis for pheochromocytoma. Copyright © 2014 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  11. Chronic exposure to quinalphos shows biochemical changes and genotoxicty in erythrocytes of silver barb, Barbonymus gonionotus

    Directory of Open Access Journals (Sweden)

    Sadiqul Islam M.

    2017-11-01

    Full Text Available An in vivo study was carried out on the freshwater fish Barbonymus gonionotus to evaluate the genotoxic effects of the organophosphate quinalphos. The fish were exposed to sub-lethal doses of quinalphos (0%, 10%, 25%, and 50% of LC50 for a period of 30 days. Analysis of biochemical characteristics (protein and lipid contents of different organs, nuclear abnormalities of erythrocytes (NAE and morphological abnormalities of erythrocytes (MAE were performed on peripheral erythrocytes sampled at post-treatment intervals of 0 and 30 days. The biochemical results revealed a significant dose-dependent decline in protein and lipid contents and increase in the frequencies of NAE as well as MAE. Our findings also confirmed that the morphological deformations of erythrocytes in addition to NAE on fish erythrocytes in vivo are effective tools in determining the potential genotoxicity of organophosphates.

  12. Interaction of ZnO nanoparticles with microbes--a physio and biochemical assay.

    Science.gov (United States)

    Padmavathy, Nagarajan; Vijayaraghavan, Rajagopalan

    2011-12-01

    The antibacterial activity of ZnO nanoparticles (nano ZnO) on various microorganisms has been investigated in this study and acting mechanism is proposed for E. coli a model organism by analyzing the growth, permeability and morphology of the bacterial cells. The experimental results indicated 1 mmoL (81.4 microg) nano ZnO could completely inhibit the growth of 10(7) cfu/mL bacterial cells in liquid Luria-Betrani medium. In the biochemical study, it has been observed that, nano ZnO resulted in the leakage of proteins. Chemiluminescence assay proved the generation of the reactive oxygen species into active state. When the cells of E. coli were exposed to nano ZnO, many pits were observed in bacterial cells by TEM and Fluorescence microscopy and the cells were found to be fragmentary. Released zinc ion from has been monitored by AAS. It suggested that nano ZnO is able to destroy the permeability of the bacterial membranes. Our study demonstrated that nano ZnO damages the structure of bacterial cell membrane and depressed the activity of some membranous enzymes by ROS production which caused E. coli bacteria to die eventually.

  13. Enhancing anaerobic digestion of food waste through biochemical methane potential assays at different substrate: inoculum ratios.

    Science.gov (United States)

    Hobbs, Shakira R; Landis, Amy E; Rittmann, Bruce E; Young, Michelle N; Parameswaran, Prathap

    2018-01-01

    Food waste has a high energy potential that can be converted into useful energy in the form of methane via anaerobic digestion. Biochemical Methane Potential assays (BMPs) were conducted to quantify the impacts on methane production of different ratios of food waste. Anaerobic digester sludge (ADS) was used as the inoculum, and BMPs were performed at food waste:inoculum ratios of 0.42, 1.42, and 3.0g chemical oxygen demand/g volatile solids (VS). The 1.42 ratio had the highest CH 4 -COD recovery: 90% of the initial total chemical oxygen demand (TCOD) was from food waste, followed by ratios 0.42 and 3.0 at 69% and 57%, respectively. Addition of food waste above 0.42 caused a lag time for CH 4 production that increased with higher ratios, which highlighted the negative impacts of overloading with food waste. The Gompertz equation was able to represent the results well, and it gave lag times of 0, 3.6 and 30days and maximum methane productions of 370, 910, and 1950mL for ratios 0.42, 1.42 and 3.0, respectively. While ratio 3.0 endured a long lag phase and low VSS destruction, ratio 1.42 achieved satisfactory results for all performance criteria. These results provide practical guidance on food-waste-to-inoculum ratios that can lead to optimizing methanogenic yield. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Alternative Methods for the Detection of Emerging Marine Toxins: Biosensors, Biochemical Assays and Cell-Based Assays

    Directory of Open Access Journals (Sweden)

    Laia Reverté

    2014-11-01

    Full Text Available The emergence of marine toxins in water and seafood may have a considerable impact on public health. Although the tendency in Europe is to consolidate, when possible, official reference methods based on instrumental analysis, the development of alternative or complementary methods providing functional or toxicological information may provide advantages in terms of risk identification, but also low cost, simplicity, ease of use and high-throughput analysis. This article gives an overview of the immunoassays, cell-based assays, receptor-binding assays and biosensors that have been developed for the screening and quantification of emerging marine toxins: palytoxins, ciguatoxins, cyclic imines and tetrodotoxins. Their advantages and limitations are discussed, as well as their possible integration in research and monitoring programs.

  15. Alternative Methods for the Detection of Emerging Marine Toxins: Biosensors, Biochemical Assays and Cell-Based Assays

    Science.gov (United States)

    Reverté, Laia; Soliño, Lucía; Carnicer, Olga; Diogène, Jorge; Campàs, Mònica

    2014-01-01

    The emergence of marine toxins in water and seafood may have a considerable impact on public health. Although the tendency in Europe is to consolidate, when possible, official reference methods based on instrumental analysis, the development of alternative or complementary methods providing functional or toxicological information may provide advantages in terms of risk identification, but also low cost, simplicity, ease of use and high-throughput analysis. This article gives an overview of the immunoassays, cell-based assays, receptor-binding assays and biosensors that have been developed for the screening and quantification of emerging marine toxins: palytoxins, ciguatoxins, cyclic imines and tetrodotoxins. Their advantages and limitations are discussed, as well as their possible integration in research and monitoring programs. PMID:25431968

  16. Micromachined filter-chamber array with passive valves for biochemical assays on beads.

    Science.gov (United States)

    Andersson, H; van der Wijngaart, W; Stemme, G

    2001-01-01

    The filter-chamber array presented here enables a real-time parallel analysis of three different samples on beads in a volume of 3 nL, on a 1 cm2 chip. The filter-chamber array is a system containing three filter-chambers, three passive valves at the inlet channels and a common outlet. The design enables parallel sample handling and time-controlled analysis. The device is microfabricated in silicon and sealed with a Pyrex lid to enable real-time analysis. Single nucleotide polymorphism analysis by using pyrosequencing has successfully been performed in single filter-chamber devices. The passive valves consist of plasma-deposited octafluorocyclobutane and show a much higher resistance towards water and surface-active solutions than previous hydrophobic patches. The device is not sensitive to gas bubbles, clogging is rare and reversible, and the filter-chamber array is reusable. More complex (bio)chemical reactions on beads can be performed in the devices with passive valves than in the devices without valves.

  17. Biochemical techniques for the characterization of G-quadruplex structures: EMSA, DMS footprinting, and DNA polymerase stop assay.

    Science.gov (United States)

    Sun, Daekyu; Hurley, Laurence H

    2010-01-01

    The proximal promoter region of many human growth-related genes contains a polypurine/polypyrimidine tract that serves as multiple binding sites for Sp1 or other transcription factors. These tracts often contain a guanine-rich sequence consisting of four runs of three or more contiguous guanines separated by one or more bases, corresponding to a general motif known for the formation of an intramolecular G-quadruplex. Recent results provide strong evidence that specific G-quadruplex structures form naturally within these polypurine/polypyrimidine tracts in many human promoter regions, raising the possibility that the transcriptional control of these genes can be modulated by G-quadruplex-interactive agents. In this chapter, we describe three general biochemical methodologies, electrophoretic mobility shift assay (EMSA), dimethylsulfate (DMS) footprinting, and the DNA polymerase stop assay, which can be useful for initial characterization of G-quadruplex structures formed by G-rich sequences.

  18. Towards developing a representative biochemical methane potential (BMP) assay for landfilled municipal solid waste - A review.

    Science.gov (United States)

    Pearse, Lauretta Feyisetan; Hettiaratchi, Joseph Patrick; Kumar, Sunil

    2018-04-01

    The applicability of slurry-based (semi-liquids) BMP assay in determining biodegradation kinetic parameters of landfilled waste is critically reviewed. Factors affecting the amount and rate of methane (CH 4 ) production during anaerobic degradation of municipal solid waste (MSW) and optimal values of these factors specific to landfill conditions are presented. The history of conventional BMP, and some existing procedures are reviewed. A landfill BMP (LBMP) assay is proposed that manipulates some of the key factors, such as moisture content, particle and sample size, that affects the rate of CH 4 production and the CH 4 generation potential of landfilled MSW (LMSW). By selecting proper conditions for these factors, a representative BMP assay could be conducted to ensure accurate determinations of CH 4 potential and the kinetic parameters k; first order rate coefficient and L o ; methane generation potential. Crown Copyright © 2018. Published by Elsevier Ltd. All rights reserved.

  19. A sensitive ferricyanide-mediated biochemical oxygen demand assay for analysis of wastewater treatment plant influents and treated effluents.

    Science.gov (United States)

    Jordan, Mark A; Welsh, David T; John, Richard; Catterall, Kylie; Teasdale, Peter R

    2013-02-01

    Representative and fast monitoring of wastewater influent and effluent biochemical oxygen demand (BOD) is an elusive goal for the wastewater industry and regulatory bodies alike. The present study describes a suitable assay, which incorporates activated sludge as the biocatalyst and ferricyanide as the terminal electron acceptor for respiration. A number of different sludges and sludge treatments were investigated, primarily to improve the sensitivity of the assay. A limit of detection (LOD) (2.1 mg BOD₅ L⁻¹) very similar to that of the standard 5-day BOD₅ method was achieved in 4 h using raw influent sludge that had been cultured overnight as the biocatalyst. Reducing the microbial concentration was the most effective means to improve sensitivity and reduce the contribution of the sludge's endogenous respiration to total ferricyanide-mediated (FM) respiration. A strong and highly significant relationship was found (n = 33; R = 0.96; p < 0.001; slope = 0.94) between BOD₅ and FM-BOD equivalent values for a diverse range of samples including wastewater treatment plant (WWTP) influent and treated effluent, as well as several grey water samples. The activated sludge FM-BOD assay presented here is an exceptional surrogate method to the standard BOD₅ assay, providing representative, same-day BOD analysis of WWTP samples with a comparable detection limit, a 4-fold greater analytical range and much faster analysis time. The industry appeal of such an assay is tremendous given that ~90% of all BOD₅ analysis is dedicated to measurement of WWTP samples, for which this assay is specifically designed. Copyright © 2012 Elsevier Ltd. All rights reserved.

  20. Assays for identification of Hsp90 inhibitors and biochemical methods for discriminating their mechanism of action.

    Science.gov (United States)

    Matts, Robert L; Manjarrez, Jacob R

    2009-01-01

    The Hsp90-dependence of many oncogenic proteins has precipitated a great deal of interest in Hsp90 as a drug target, as evidence mounts that Hsp90 inhibitors may be effective chemotherapeutic agents for the treatment of cancer. In addition, Hsp90-inhibitors have shown promise for the treatment of neurodegenerative diseases characterized by the accumulation of toxic denatured protein aggregates. The development of assays for the identification of novel Hsp90 inhibitors began in earnest when it became apparent that the Hsp90 inhibitors available at the time had less than ideal pharmacological properties. This review summarizes what is known about Hsp90's structure and function, its ATPase cycle, its interactions with co-chaperones and clients, and the effect of Hsp90-inhibitors on these processes. It further summarizes various high throughput assays (and secondary confirmatory assays) developed to identify new Hsp90 inhibitors from chemical libraries based on the inhibitors ability to: inhibit Hsp90's ATPase activity; compete for ligand binding to Hsp90 and its N-terminal ATP-binding domain; inhibit Hsp90-dependent refolding of denatured luciferase; and deplete culture cells of Hsp90-dependnet client protein or induce Hsp70 expression. In addition, in vitro assays are described that help determine the site of inhibitor binding to Hsp90 (N- or C-terminal domain) and there mechanism of action based on effects of inhibitors on Hsp90/ co-chaperone and client interactions, and Hsp90 conformation characterized by proteolytic fingerprinting.

  1. Determining the functional significance of mismatch repair gene missense variants using biochemical and cellular assays

    DEFF Research Database (Denmark)

    Heinen, Christopher D; Juel Rasmussen, Lene

    2012-01-01

    ABSTRACT: With the discovery that the hereditary cancer susceptibility disease Lynch syndrome (LS) is caused by deleterious germline mutations in the DNA mismatch repair (MMR) genes nearly 20 years ago, genetic testing can now be used to diagnose this disorder in patients. A definitive diagnosis...... of LS can direct how clinicians manage the disease as well as prevent future cancers for the patient and their families. A challenge emerges, however, when a germline missense variant is identified in a MMR gene in a suspected LS patient. The significance of a single amino acid change in these large...... some of these assays along with the challenges of using such assays to determine the functional consequences of MMR VUS which, in turn, can provide valuable insight into their clinical significance. With increased gene sequencing in patients, the number of identified VUS has expanded dramatically...

  2. Centrifugal multiplexing fixed-volume dispenser on a plastic lab-on-a-disk for parallel biochemical single-end-point assays.

    Science.gov (United States)

    La, Moonwoo; Park, Sang Min; Kim, Dong Sung

    2015-01-01

    In this study, a multiple sample dispenser for precisely metered fixed volumes was successfully designed, fabricated, and fully characterized on a plastic centrifugal lab-on-a-disk (LOD) for parallel biochemical single-end-point assays. The dispenser, namely, a centrifugal multiplexing fixed-volume dispenser (C-MUFID) was designed with microfluidic structures based on the theoretical modeling about a centrifugal circumferential filling flow. The designed LODs were fabricated with a polystyrene substrate through micromachining and they were thermally bonded with a flat substrate. Furthermore, six parallel metering and dispensing assays were conducted at the same fixed-volume (1.27 μl) with a relative variation of ±0.02 μl. Moreover, the samples were metered and dispensed at different sub-volumes. To visualize the metering and dispensing performances, the C-MUFID was integrated with a serpentine micromixer during parallel centrifugal mixing tests. Parallel biochemical single-end-point assays were successfully conducted on the developed LOD using a standard serum with albumin, glucose, and total protein reagents. The developed LOD could be widely applied to various biochemical single-end-point assays which require different volume ratios of the sample and reagent by controlling the design of the C-MUFID. The proposed LOD is feasible for point-of-care diagnostics because of its mass-producible structures, reliable metering/dispensing performance, and parallel biochemical single-end-point assays, which can identify numerous biochemical.

  3. Structural, Biochemical, and Evolutionary Characterizations of Glyoxylate/Hydroxypyruvate Reductases Show Their Division into Two Distinct Subfamilies.

    Science.gov (United States)

    Kutner, Jan; Shabalin, Ivan G; Matelska, Dorota; Handing, Katarzyna B; Gasiorowska, Olga; Sroka, Piotr; Gorna, Maria W; Ginalski, Krzysztof; Wozniak, Krzysztof; Minor, Wladek

    2018-02-13

    The d-2-hydroxyacid dehydrogenase (2HADH) family illustrates a complex evolutionary history with multiple lateral gene transfers and gene duplications and losses. As a result, the exact functional annotation of individual members can be extrapolated to a very limited extent. Here, we revise the previous simplified view on the classification of the 2HADH family; specifically, we show that the previously delineated glyoxylate/hydroxypyruvate reductase (GHPR) subfamily consists of two evolutionary separated GHRA and GHRB subfamilies. We compare two representatives of these subfamilies from Sinorhizobium meliloti (SmGhrA and SmGhrB), employing a combination of biochemical, structural, and bioinformatics approaches. Our kinetic results show that both enzymes reduce several 2-ketocarboxylic acids with overlapping, but not equivalent, substrate preferences. SmGhrA and SmGhrB show highest activity with glyoxylate and hydroxypyruvate, respectively; in addition, only SmGhrB reduces 2-keto-d-gluconate, and only SmGhrA reduces pyruvate (with low efficiency). We present nine crystal structures of both enzymes in apo forms and in complexes with cofactors and substrates/substrate analogues. In particular, we determined a crystal structure of SmGhrB with 2-keto-d-gluconate, which is the biggest substrate cocrystallized with a 2HADH member. The structures reveal significant differences between SmGhrA and SmGhrB, both in the overall structure and within the substrate-binding pocket, offering insight into the molecular basis for the observed substrate preferences and subfamily differences. In addition, we provide an overview of all GHRA and GHRB structures complexed with a ligand in the active site.

  4. Development and validation of a quick easily used biochemical assay for evaluating the viability of small immobile arthropods.

    Science.gov (United States)

    Phillips, Craig B; Iline, Ilia I; Richards, Nicola K; Novoselov, Max; McNeill, Mark R

    2013-10-01

    Quickly, accurately, and easily assessing the efficacy of treatments to control sessile arthropods (e.g., scale insects) and stationary immature life stages (e.g., eggs and pupae) is problematic because it is difficult to tell whether treated organisms are alive or dead. Current approaches usually involve either maintaining organisms in the laboratory to observe them for development, gauging their response to physical stimulation, or assessing morphological characters such as turgidity and color. These can be slow, technically difficult, or subjective, and the validity of methods other than laboratory rearing has seldom been tested. Here, we describe development and validation of a quick easily used biochemical colorimetric assay for measuring the viability of arthropods that is sufficiently sensitive to test even very small organisms such as white fly eggs. The assay was adapted from a technique for staining the enzyme hexokinase to signal the presence of adenosine triphosphate in viable specimens by reducing a tetrazolium salt to formazan. Basic laboratory facilities and skills are required for production of the stain, but no specialist equipment, expertise, or facilities are needed for its use.

  5. A novel high throughput biochemical assay to evaluate the HuR protein-RNA complex formation.

    Directory of Open Access Journals (Sweden)

    Vito G D'Agostino

    Full Text Available The RNA binding protein HuR/ELAVL1 binds to AU-rich elements (AREs promoting the stabilization and translation of a number of mRNAs into the cytoplasm, dictating their fate. We applied the AlphaScreen technology using purified human HuR protein, expressed in a mammalian cell-based system, to characterize in vitro its binding performance towards a ssRNA probe whose sequence corresponds to the are present in TNFα 3' untranslated region. We optimized the method to titrate ligands and analyzed the kinetic in saturation binding and time course experiments, including competition assays. The method revealed to be a successful tool for determination of HuR binding kinetic parameters in the nanomolar range, with calculated Kd of 2.5±0.60 nM, k on of 2.76±0.56*10(6 M(-1 min(-1, and k off of 0.007±0.005 min(-1. We also tested the HuR-RNA complex formation by fluorescent probe-based RNA-EMSA. Moreover, in a 384-well plate format we obtained a Z-factor of 0.84 and an averaged coefficient of variation between controls of 8%, indicating that this biochemical assay fulfills criteria of robustness for a targeted screening approach. After a screening with 2000 small molecules and secondary verification with RNA-EMSA we identified mitoxantrone as an interfering compound with rHuR and TNFα probe complex formation. Notably, this tool has a large versatility and could be applied to other RNA Binding Proteins recognizing different RNA, DNA, or protein species. In addition, it opens new perspectives in the identification of small-molecule modulators of RNA binding proteins activity.

  6. Unsaturated fatty acids show clear elicitation responses in a modified local lymph node assay with an elicitation phase, and test positive in the direct peptide reactivity assay.

    Science.gov (United States)

    Yamashita, Kunihiko; Shinoda, Shinsuke; Hagiwara, Saori; Miyazaki, Hiroshi; Itagaki, Hiroshi

    2015-12-01

    The Organisation for Economic Co-operation and Development (OECD) Test Guidelines (TG) adopted the murine local lymph node assay (LLNA) and guinea pig maximization test (GPMT) as stand-alone skin sensitization test methods. However, unsaturated carbon-carbon double-bond and/or lipid acids afforded false-positive results more frequently in the LLNA compared to those in the GPMT and/or in human subjects. In the current study, oleic, linoleic, linolenic, undecylenic, fumaric, maleic, and succinic acid and squalene were tested in a modified LLNA with an elicitation phase (LLNA:DAE), and in a direct peptide reactivity assay (DPRA) to evaluate their skin-sensitizing potential. Oleic, linoleic, linolenic, undecylenic and maleic acid were positive in the LLNA:DAE, of which three, linoleic, linolenic, and maleic acid were positive in the DPRA. Furthermore, the results of the cross-sensitizing tests using four LLNA:DAE-positive chemicals were negative, indicating a chemical-specific elicitation response. In a previous report, the estimated concentration needed to produce a stimulation index of 3 (EC3) of linolenic acid, squalene, and maleic acid in the LLNA was undecylenic, and maleic acid had skin-sensitizing potential, and that the LLNA overestimated the skin-sensitizing potential compared to that estimated by the LLNA:DAE.

  7. MMP Mediated Degradation of Type VI Collagen Is Highly Associated with Liver Fibrosis - Identification and Validation of a Novel Biochemical Marker Assay

    DEFF Research Database (Denmark)

    Veidal, Sanne Skovgard; Karsdal, Morten Asser; Vassiliadis, Efstathios

    2011-01-01

    Background and Aims: During fibrogenesis, in which excessive remodeling of the extracellular matrix occurs, both the quantity of type VI collagen and levels of matrix metalloproteinases, including MMP-2 and MMP-9, increase significantly. Proteolytic degradation of type VI collagen into small...... fragments, so-called neo-epitopes, may be specific biochemical marker of liver fibrosis. The aim of this study was to develop an ELISA detecting a fragment of type VI collagen generated by MMP-2 and MMP-9, and evaluate this assay in two preclinical models of liver fibrosis. Methods: Mass spectrometric...... analysis of cleaved type VI collagen revealed a large number of protease-generated neo-epitopes. A fragment unique to type VI collagen generated by MMP-2 and MMP-9 was selected for ELISA development. The CO6-MMP assay was evaluated in two rat models of liver fibrosis: bile duct ligation (BDL) and carbon...

  8. Biochemical Assay Detects Feeding Damage to Loblolly Pine Seeds Caused by the Leaffooted Pine Seed Bug (Hemiptera: Coreidae)

    Science.gov (United States)

    Cameron G. Lait; Daniel R. Miller; Sarah L. Bates; John H. Borden; Allison R. Kermode

    2003-01-01

    A large number of proteins in salivary gland extracts of the leaffooted pine seed bug, Leptoglossus corculus Say, were strongly recognized by a polyclonal antibody-based assay developed for detecting saliva of the western conifer seed bug, Lepfoglossus occidentalis Heidemann, in lodgepole pine, Pinus contorta var...

  9. Secukinumab, a novel anti–IL-17A antibody, shows low immunogenicity potential in human in vitro assays comparable to other marketed biotherapeutics with low clinical immunogenicity

    Science.gov (United States)

    Karle, Anette; Spindeldreher, Sebastian; Kolbinger, Frank

    2016-01-01

    ABSTRACT Secukinumab is a human monoclonal antibody that selectively targets interleukin-17A and has been demonstrated to be highly efficacious in the treatment of moderate to severe plaque psoriasis, starting at early time points, with a sustained effect and a favorable safety profile. Biotherapeutics—including monoclonal antibodies (mAbs)—can be immunogenic, leading to formation of anti-drug antibodies (ADAs) that can result in unwanted effects, including hypersensitivity reactions or compromised therapeutic efficacy. To gain insight into possible explanations for the clinically observed low immunogenicity of secukinumab, we evaluated its immunogenicity potential by applying 2 different in vitro assays: T-cell activation and major histocompatibility complex–associated peptide proteomics (MAPPs). For both assays, monocyte-derived dendritic cells (DCs) from healthy donors were exposed in vitro to biotherapeutic proteins. DCs naturally process proteins and present the derived peptides in the context of human leukocyte antigen (HLA)-class II. HLA-DR–associated biotherapeutic-derived peptides, representing potential T–cell epitopes, were identified in the MAPPs assay. In the T-cell assay, autologous CD4+ T cells were co-cultured with secukinumab-exposed DCs and T-cell activation was measured by proliferation and interleukin-2 secretion. In the MAPPs analysis and T-cell activation assays, secukinumab consistently showed relatively low numbers of potential T-cell epitopes and low T-cell response rates, respectively, comparable to other biotherapeutics with known low clinical immunogenicity. In contrast, biotherapeutics with elevated clinical immunogenicity rates showed increased numbers of potential T-cell epitopes and increased T-cell response rates in T-cell activation assays, indicating an approximate correlation between in vitro assay results and clinical immunogenicity incidence. PMID:26817498

  10. Proteomic and biochemical assays of glutathione-related proteins in susceptible and multiple herbicide resistant Avena fatua L.

    Science.gov (United States)

    Burns, Erin E; Keith, Barbara K; Refai, Mohammed Y; Bothner, Brian; Dyer, William E

    2017-08-01

    Extensive herbicide usage has led to the evolution of resistant weed populations that cause substantial crop yield losses and increase production costs. The multiple herbicide resistant (MHR) Avena fatua L. populations utilized in this study are resistant to members of all selective herbicide families, across five modes of action, available for A. fatua control in U.S. small grain production, and thus pose significant agronomic and economic threats. Resistance to ALS and ACCase inhibitors is not conferred by target site mutations, indicating that non-target site resistance mechanisms are involved. To investigate the potential involvement of glutathione-related enzymes in the MHR phenotype, we used a combination of proteomic, biochemical, and immunological approaches to compare their constitutive activities in herbicide susceptible (HS1 and HS2) and MHR (MHR3 and MHR4) A. fatua plants. Proteomic analysis identified three tau and one phi glutathione S-transferases (GSTs) present at higher levels in MHR compared to HS plants, while immunoassays revealed elevated levels of lambda, phi, and tau GSTs. GST specific activity towards 1-chloro-2,4-dinitrobenzene was 1.2-fold higher in MHR4 than in HS1 plants and 1.3- and 1.2-fold higher in MHR3 than in HS1 and HS2 plants, respectively. However, GST specific activities towards fenoxaprop-P-ethyl and imazamethabenz-methyl were not different between untreated MHR and HS plants. Dehydroascorbate reductase specific activity was 1.4-fold higher in MHR than HS plants. Pretreatment with the GST inhibitor NBD-Cl did not affect MHR sensitivity to fenoxaprop-P-ethyl application, while the herbicide safener and GST inducer mefenpyr reduced the efficacy of low doses of fenoxaprop-P-ethyl on MHR4 but not MHR3 plants. Mefenpyr treatment also partially reduced the efficacy of thiencarbazone-methyl or mesosulfuron-methyl on MHR3 or MHR4 plants, respectively. Overall, the GSTs described here are not directly involved in enhanced rates of

  11. Purification of Yeast Native Reagents for the Analysis of Chromatin Function-II: Multiprotein Complexes and Biochemical Assays.

    Science.gov (United States)

    Lacoste, Nicolas; Bhat, Wajid; Côté, Jacques

    2017-01-01

    Post-translational modifications of histones play essential roles in regulating chromatin structure and function. These are tightly regulated in vivo and there is an intricate cross-talk between different marks as they are recognized by specific reader modules present in a large number of nuclear factors. In order to precisely dissect these processes in vitro native reagents like purified chromatin and histone modifying/remodeling enzymes are required to more accurately reproduce physiological conditions. The vast majority of these enzymes need to be part of stable multiprotein complexes with cofactors enabling them to act on chromatin substrates and/or read specific histone marks. In the accompanying chapter, we have described the protocol for purification of native chromatin from yeast cells (Chapter 3 ). Here, we present the methods to obtain highly purified native chromatin modifying complexes from Saccharomyces cerevisiae, based on Tandem Affinity Purification (TAP). We also present possible applications and useful functional assays that can be performed using these yeast native reagents.

  12. Clinical characteristics of Graves' orbitopathy in patients showing discrepancy between levels from TBII assays and TSI bioassay.

    Science.gov (United States)

    Jang, Sun Young; Shin, Dong Yeob; Lee, Eun Jig; Yoon, Jin Sook

    2014-04-01

    To investigate clinical characteristics of patients with Graves' orbitopathy (GO) who showed discrepancies between levels of thyroid-stimulating immunoglobulin (TSI) and thyrotropin-binding inhibitory immunoglobulin (TBII). Comparative case series. A total of 317 patients with GO in whom Mc4-TSI and M22-TRAb (third-generation TBII) were measured simultaneously. Patients were divided into four groups according to TRAb levels as followings: Group 1, TBII and TSI TSI TSI ≥ median; Group 4, both TBII and TSI ≥ median. Endocrine and ophthalmic clinical manifestations in each group. The median value of M22-TRAb was 6·11 IU/l and that of Mc4-TSI was 415·1 (SRR%). One hundred seventeen patients were classified as Group 1, 41 patients as Group 2, 41 patients as group 3 and 118 patients as group 4. Mean CAS was significantly higher in Groups 3 (2·2) and 4 (2·2) than in Groups 1 (1·6) and 2 (1·4; P = 0·001, ANOVA). Mean modified NOSPECS scores were significantly higher (P TSI than in patients with predominant M22-TRAb. Patients with hyperthyroidism were more likely to be included with patients with predominant M22-TRAb than with predominant Mc4-TSI. © 2013 John Wiley & Sons Ltd.

  13. Larval development assays reveal the presence of sub-populations showing high- and low-level resistance in a monepantel (Zolvix®)-resistant isolate of Haemonchus contortus.

    Science.gov (United States)

    Raza, Ali; Lamb, Jane; Chambers, Michael; Hunt, Peter W; Kotze, Andrew C

    2016-04-15

    Resistance to the amino-acetonitrile derivative monepantel has been reported in several species of gastrointestinal nematodes over recent years. We were interested in the use of in vitro assays with free-living worm life-stages to detect resistance to this drug. We therefore used larval development and larval migration assays to examine dose response relationships for the drug against two susceptible and one resistant isolate of Haemonchus contortus. The resistant isolate was established by laboratory propagation of the survivors of a field treatment with Zolvix(®) that had originally resulted in a drug efficacy of over 99%. Drug efficacy against this field-derived laboratory-propagated resistant isolate in vivo was approximately 15%. The larval development assay proved able to discriminate between the susceptible and resistant isolates, with larvae of the resistant isolate showing an ability to develop at higher drug concentrations than the two susceptible isolates. The resistant isolate showed the presence of two distinct subpopulations, separated by a plateau in the dose-response curve. Sub-population 1 (approximately 40% of the total population) showed a low level of resistance with an IC50 increased approximately 7-fold compared to the baseline susceptible isolate, while sub-population 2 (the remaining 60% of the total population) showed an IC50 increased over 1000-fold compared to the baseline susceptible isolate. This level of resistance is unusually high for any gastrointestinal nematode species in drug dose-response in vitro assays. In contrast, the migration assay could not discriminate between the three isolates, with migration not reduced to zero at any of the drug concentrations tested. This study demonstrates that a larval development assay is able to detect resistance to monepantel in H. contortus, and that resistance can exist in two distinct forms. This suggests that at least two separate monepantel resistance mechanisms are acting within the worm

  14. New sub-family of lysozyme-like proteins shows no catalytic activity: crystallographic and biochemical study of STM3605 protein from Salmonella Typhimurium

    Energy Technology Data Exchange (ETDEWEB)

    Michalska, Karolina; Brown, Roslyn N.; Li, Hui; Jedrzejczak, Robert; Niemann, George; Heffron, Fred; Cort, John R.; Adkins, Joshua N.; Babnigg, Gyorgy; Joachimiak, Andrzej

    2013-03-01

    Phage viruses that infect prokaryotes integrate their genome into the host chromosome; thus, microbial genomes typically contain genetic remnants of both recent and ancient phage infections. Often phage genes occur in clusters of atypical G+C content that reflect integration of the foreign DNA. However, some phage genes occur in isolation without other phage gene neighbors, probably resulting from horizontal gene transfer. In these cases, the phage gene product is unlikely to function as a component of a mature phage particle, and instead may have been co-opted by the host for its own benefit. The product of one such gene from Salmonella enterica serovar Typhimurium, STM3605, encodes a protein with modest sequence similarity to phage-like lysozyme (N-acetylmuramidase) but appears to lack essential catalytic residues that are strictly conserved in all lysozymes. Close homologs in other bacteria share this characteristic. The structure of the STM3605 protein was characterized by X-ray crystallography, and functional assays showed that it is a stable, folded protein whose structure closely resembles lysozyme. However, this protein is unlikely to hydrolyze peptidoglycan. Instead, STM3605 is presumed to have evolved an alternative function because it shows some lytic activity and partitions to micelles.

  15. New sub-family of lysozyme-like proteins shows no catalytic activity: crystallographic and biochemical study of STM3605 protein from Salmonella Typhimurium.

    Science.gov (United States)

    Michalska, Karolina; Brown, Roslyn N; Li, Hui; Jedrzejczak, Robert; Niemann, George S; Heffron, Fred; Cort, John R; Adkins, Joshua N; Babnigg, Gyorgy; Joachimiak, Andrzej

    2013-03-01

    Phage viruses that infect prokaryotes integrate their genome into the host chromosome; thus, microbial genomes typically contain genetic remnants of both recent and ancient phage infections. Often phage genes occur in clusters of atypical G+C content that reflect integration of the foreign DNA. However, some phage genes occur in isolation without other phage gene neighbors, probably resulting from horizontal gene transfer. In these cases, the phage gene product is unlikely to function as a component of a mature phage particle, and instead may have been co-opted by the host for its own benefit. The product of one such gene from Salmonella enterica serovar Typhimurium, STM3605, encodes a protein with modest sequence similarity to phage-like lysozyme (N-acetylmuramidase) but appears to lack essential catalytic residues that are strictly conserved in all lysozymes. Close homologs in other bacteria share this characteristic. The structure of the STM3605 protein was characterized by X-ray crystallography, and functional assays showed that it is a stable, folded protein whose structure closely resembles lysozyme. However, this protein is unlikely to hydrolyze peptidoglycan. Instead, STM3605 is presumed to have evolved an alternative function because it shows some lytic activity and partitions to micelles.

  16. {sup 11}C-Choline PET/CT in patients with hormone-resistant prostate cancer showing biochemical relapse after radical prostatectomy

    Energy Technology Data Exchange (ETDEWEB)

    Ceci, Francesco; Ambrosini, Valentina; Boschi, Stefano; Fanti, Stefano [University of Bologna, Nuclear Medicine Unit, Department of Haematology Oncology and Laboratory Medicine, Azienda Ospedaliero-Universitaria di Bologna, Policlinico Sant' Orsola-Malpighi, Bologna (Italy); Castellucci, Paolo [University of Bologna, Nuclear Medicine Unit, Department of Haematology Oncology and Laboratory Medicine, Azienda Ospedaliero-Universitaria di Bologna, Policlinico Sant' Orsola-Malpighi, Bologna (Italy); Azienda Ospedaliero-Unversitaria di Bologna Policlinico Sant' Orsola-Malpighi, UO di Medicina Nucleare, PAD. 30, Bologna (Italy); Mamede, Marcelo [Universidade Federal de Minas Gerais, Molecular Imaging Center, Belo Horizonte (Brazil); Schiavina, Riccardo; Martorana, Giuseppe [University of Bologna, Department of Urology, Azienda Ospedaliero-Universitaria di Bologna, Policlinico Sant' Orsola-Malpighi, Bologna (Italy); Rubello, Domenico [' Santa Maria della Misericordia' Hospital, Department of Nuclear Medicine and PET/CT Centre, Rovigo (Italy); Fuccio, Chiara [Fondazione Salvatore Maugeri, Service of Nuclear Medicine, Pavia (Italy)

    2013-02-15

    To determine the diagnostic efficacy of {sup 11}C-choline PET/CT in patients with prostate cancer (PC) after radical prostatectomy who presented with increasing PSA levels during follow-up in spite of being on hormone treatment (HT), and therefore showing HT resistance. We evaluated a large series of 157 consecutive PC patients previously treated by radical prostatectomy who presented with biochemical recurrence with increasing PSA levels in spite of ongoing HT (HT-resistant patients). At the time of {sup 11}C-choline PET/CT, the mean value of trigger PSA level was 8.3 (range 0.2 - 60.6 ng/mL), the mean PSA doubling time (PSAdt) was 5.3 (range 0.4 - 35 months), and the mean PSA velocity (PSAvel) was 22.1 ng/mL/year (range 0.12 - 82 ng/mL/year). {sup 11}C-Choline PET/CT was performed following a standard procedure at our centre to investigate increasing PSA levels, either as the first imaging procedure or in patients with negative conventional imaging. At the time of {sup 11}C-choline PET/CT all patients were receiving HT (61 were receiving monotherapy and 96 multidrug therapy). PET-positive findings were validated by: (a) transrectal US-guided biopsy in patients with recurrence in the prostatic bed, (b) surgical pelvic lymphadenectomy, (c) other imaging modalities, including repeated {sup 11}C-choline PET/CT, performed during a minimum follow-up of 12-months. {sup 11}C-Choline PET/CT showed positive findings in 104 of the 157 patients (66 %). {sup 11}C-choline PET/CT detected: a single lesion in 40 patients (7 in the prostate bed, 10 in lymph nodes, 22 in bone, 1 at another site); two lesions in 18 patients (7 in lymph nodes, 7 in bone, 4 in both lymph nodes and bone); three or four lesions in 7 patients (4 in lymph nodes, 2 in bone, 1 at another site); and more than four lesions in the remaining 39 patients (2 in the prostate bed, 12 in lymph nodes, 12 in bone, 11 in both lymph nodes and bone, 2 at other sites). In {sup 11}C-choline PET-negative patients, the mean

  17. A rapid quantitative activity assay shows that the Vibrio cholerae colonization factor GbpA is an active lytic polysaccharide monooxygenase

    NARCIS (Netherlands)

    Loose, Jennifer S. M.; Forsberg, Zarah; Fraaije, Marco W.; Eijsink, Vincent G. H.; Vaaje-Kolstad, Gustav

    2014-01-01

    The discovery of the copper-dependent lytic polysaccharide monooxygenases (LPMOs) has revealed new territory for chemical and biochemical analysis. These unique mononuclear copper enzymes are abundant, suggesting functional diversity beyond their established roles in the depolymerization of biomass

  18. Development of a microfluidic confocal fluorescence detection system for the hyphenation of nano-LC to on-line biochemical assays

    NARCIS (Netherlands)

    Heus, F.; Giera, M.A.; de Kloe, G.E.; van Iperen, D.; Buijs, J.B.; Nahar, T.T.; Smit, A.B.; Lingeman, H.; de Esch, I.J.P.; Niessen, W.M.A.; Irth, H.; Kool, J.

    2010-01-01

    One way to profile complex mixtures for receptor affinity is to couple liquid chromatography (LC) on-line to biochemical detection (BCD). A drawback of this hyphenated screening approach is the relatively high consumption of sample, receptor protein and (fluorescently labeled) tracer ligand. Here,

  19. Comprehensive genotyping for 18 blood group systems using a multiplex ligation-dependent probe amplification assay shows a high degree of accuracy

    NARCIS (Netherlands)

    Haer-Wigman, Lonneke; Ji, Yanli; Lodén, Martin; de Haas, Masja; van der Schoot, C. Ellen; Veldhuisen, Barbera

    2013-01-01

    In recent years genotyping methods have been implemented in blood banks as alternative to comprehensive serologic typing. We evaluated a newly developed assay for convenient and comprehensive genotyping of blood group alleles based on multiplex ligation-dependent probe amplification (MLPA)

  20. Biochemical studies of mouse brain tubulin: colchicine binding (DEAE-cellulose filter) assay and subunits (α and β) biosynthesis and degradation (in newborn brain)

    International Nuclear Information System (INIS)

    Tse, C.F.

    1978-01-01

    A DEAE-cellulose filter assay, measuring [ 3 H]colchicine bound to colchicine binding protein (CBP) absorbed on filter discs, has been modified to include lM sucrose in the incubation medium for complexing colchicine to CBP in samples before applying the samples to filter discs (single point assay). Due to the much greater stability of colchicine binding capacity in the presence of lM sucrose, multiple time-point assays and least squares linear regression analysis were not necessary for accurate determination of CBP in hybrid mouse brain at different stages of development. The highest concentrations of CBP were observed in the 160,000g supernatant and pellet of newborn brain homogenate. Further studies of the modified filter assay documented that the assay has an overall counting efficiency of 27.3%, that DEAE-cellulose filters bind and retain all tubulin in the assay samples, and that one molecule of colchicine binds approximately one molecule of tubulin dimer. Therefore, millimoles of colchicine bound per milligram total protein can be used to calculate tubulin content. With this technique tubulin content of brain supernatant was found to be 11.9% for newborn, and 7.15% for 11 month old mice. Quantitative densitometry was also used to measure mouse brain supernatant actin content for these two stages. In vivo synthesis and degradation rates of tubulin α and β subunits of two day mouse brain 100,000g supernatant were studied after intracerebral injection of [ 3 H]leucine. Quantitative changes of the ratio of tritium specific activities of tubulin α and β subunits with time were determined. The pattern of change was biphasic. During the first phase the ratio decreased; during the second phase the ratio increased continuously. An interpretation consistent with all the data in this study is that the α subunit is synthesized at a more rapid rate than the β subunit

  1. Biochemical studies of mouse brain tubulin: colchicine binding (DEAE-cellulose filter) assay and subunits ( α and β) biosynthesis and degradation (in newborn brain)

    Energy Technology Data Exchange (ETDEWEB)

    Tse, Cek-Fyne [Univ. of Rochester, NY (United States)

    1978-01-01

    A DEAE-cellulose filter assay, measuring (3H)colchicine bound to colchicine binding protein (CBP) absorbed on filter discs, has been modified to include lM sucrose in the incubation medium for complexing colchicine to CBP in samples before applying the samples to filter discs (single point assay). Due to the much greater stability of colchicine binding capacity in the presence of lM sucrose, multiple time-point assays and least squares linear regression analysis were not necessary for accurate determination of CBP in hybrid mouse brain at different stages of development. The highest concentrations of CBP were observed in the 160,000g supernatant and pellet of newborn brain homogenate. Further studies of the modified filter assay documented that the assay has an overall counting efficiency of 27.3%, that DEAE-cellulose filters bind and retain all tubulin in the assay samples, and that one molecule of colchicine binds approximately one molecule of tubulin dimer. Therefore, millimoles of colchicine bound per milligram total protein can be used to calculate tubulin content. With this technique tubulin content of brain supernatant was found to be 11.9% for newborn, and 7.15% for 11 month old mice. Quantitative densitometry was also used to measure mouse brain supernatant actin content for these two stages. In vivo synthesis and degradation rates of tubulin ..cap alpha.. and ..beta.. subunits of two day mouse brain 100,000g supernatant were studied after intracerebral injection of (3H)leucine. Quantitative changes of the ratio of tritium specific activities of tubulin ..cap alpha.. and ..beta.. subunits with time were determined. The pattern of change was biphasic. During the first phase the ratio decreased; during the second phase the ratio increased continuously. An interpretation consistent with all the data in this study is that the ..cap alpha.. subunit is synthesized at a more rapid rate than the ..beta.. subunit. (ERB)

  2. Analysis of Flavonoids in Lotus (Nelumbo nucifera Leaves and Their Antioxidant Activity Using Macroporous Resin Chromatography Coupled with LC-MS/MS and Antioxidant Biochemical Assays

    Directory of Open Access Journals (Sweden)

    Ming-Zhi Zhu

    2015-06-01

    Full Text Available Lotus (Nelumbo nucifera leaves, a traditional Chinese medicinal herb, are rich in flavonoids. In an effort to thoroughly analyze their flavonoid components, macroporous resin chromatography coupled with HPLC-MS/MS was employed to simultaneously enrich and identify flavonoids from lotus leaves. Flavonoids extracted from lotus leaves were selectively enriched in the macroporous resin column, eluted subsequently as fraction II, and successively subjected to analysis with the HPLC-MS/MS and bioactivity assays. Altogether, fourteen flavonoids were identified, four of which were identified from lotus leaves for the first time, including quercetin 3-O-rhamnopyranosyl-(1→2-glucopyranoside, quercetin 3-O-arabinoside, diosmetin 7-O-hexose, and isorhamnetin 3-O-arabino- pyranosyl-(1→2-glucopyranoside. Further bioactivity assays revealed that these flavonoids from lotus leaves possess strong antioxidant activity, and demonstrate very good potential to be explored as food supplements or even pharmaceutical products to improve human health.

  3. A Biochemical/Biophysical Assay Dyad for HTS-Compatible Triaging of Inhibitors of the HIV-1 Nef/Hck SH3 Interaction

    KAUST Repository

    Breuer, Sebastian

    2013-07-26

    The current treatment regimens for HIV include over 20 anti-retrovirals. However, adverse drug effects and the emergence of drug resistance necessitates the continued improvement of the existing drug classes as well as the development of novel drugs that target as yet therapeutically unexploited viral and cellular pathways. Here we demonstrate a strategy for the discovery of protein-protein interaction inhibitors of the viral pathogenicity factor HIV-1 Nef and its interaction with the host factor SH3. A combination of a time-resolved fluorescence resonance energy resonance energy transfer-based assay and a label-free resonant waveguide grating-based assay was optimized for high-throughput screening formats.

  4. Biochemical Abnormalities in Batten's Syndrome

    DEFF Research Database (Denmark)

    Clausen, Jytte Lene; Nielsen, Gunnar Gissel; Jensen, Gunde Egeskov

    1978-01-01

    The present data indicate that a group of ten patients with Batten's syndrome showed reduced activity of erythrocyte glutathione (GSH) peroxidase (Px) (glutathione: H2O2 oxidoreductase, EC 1.1.1.9.) using H2O2 as peroxide donor. Assay of erythrocyte GSHPx using H2O2, cumene hydroperoxide and t......-butyl hydroperoxide as donors also makes it possible biochemically to divide Batten's syndrome into two types: (1) one type with decreased values when H2O2 and cumene hydroperoxide are used, and (2) one type with increased values when t-butyl hydroperoxide is used. Furthermore an increased content of palmitic, oleic...

  5. Jasmonate response locus JAR1 and several related Arabidopsis genes encode enzymes of the firefly luciferase superfamily that show activity on jasmonic, salicylic, and indole-3-acetic acids in an assay for adenylation.

    Science.gov (United States)

    Staswick, Paul E; Tiryaki, Iskender; Rowe, Martha L

    2002-06-01

    Jasmonic acid (JA) and related cyclopentanones are critical plant signaling molecules, but their mode of action at the molecular level is unclear. A map-based approach was used to identify the defective gene in the Arabidopsis JA response mutant jar1-1. JAR1 is 1 of 19 closely related Arabidopsis genes that are similar to the auxin-induced soybean GH3 gene. Analysis of fold predictions for this protein family suggested that JAR1 might belong to the acyl adenylate-forming firefly luciferase superfamily. These enzymes activate the carboxyl groups of a variety of substrates for their subsequent biochemical modification. An ATP-PPi isotope exchange assay was used to demonstrate adenylation activity in a glutathione S-transferase-JAR1 fusion protein. Activity was specific for JA, suggesting that covalent modification of JA is important for its function. Six other Arabidopsis genes were specifically active on indole-3-acetic acid (IAA), and one was active on both IAA and salicylic acid. These findings suggest that the JAR1 gene family is involved in multiple important plant signaling pathways.

  6. Complete Comparison Display (CCD) evaluation of ethanol extracts of Centella asiatica and Withania somnifera shows that they can non-synergistically ameliorate biochemical and behavioural damages in MPTP induced Parkinson's model of mice

    Science.gov (United States)

    Bhatnagar, Maheep; Goel, Ishan; Roy, Tathagato; Khurana, Sukant

    2017-01-01

    Parkinson’s disease remains as one of the most common debilitating neurodegenerative disorders. With the hopes of finding agents that can cure or reduce the pace of progression of the disease, we studied two traditional medicinal plants: Centella asiatica and Withania somnifera that have been explored in some recent studies. In agreement with the previous work on ethanol extracts of these two plants in mice model, we saw an improvement in oxidative stress profile as well as behavioral performance in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced Parkinson-like symptoms in Balb/c mice. Given the known potential of both the herbal extracts in improving Parkinson-like symptoms, we expected the combination of the two to show better results than either of the two but surprisingly there was no additivity in either oxidative stress or behavioural recovery. In fact, in some assays, the combination performed worse than either of the two individual constituents. This effect of mixtures highlights the need of testing mixtures in supplements market using enthomedicine. The necessity of comparing multiple groups in this study to get most information from the experiments motivated us to design a ladder-like visualization to show comparison with different groups that we call complete comparison display (CCD). In summary, we show the potential of Centella asiatica and Withania somnifera to ameliorate Parkinson’s disorder. PMID:28510600

  7. Complete Comparison Display (CCD evaluation of ethanol extracts of Centella asiatica and Withania somnifera shows that they can non-synergistically ameliorate biochemical and behavioural damages in MPTP induced Parkinson's model of mice.

    Directory of Open Access Journals (Sweden)

    Maheep Bhatnagar

    Full Text Available Parkinson's disease remains as one of the most common debilitating neurodegenerative disorders. With the hopes of finding agents that can cure or reduce the pace of progression of the disease, we studied two traditional medicinal plants: Centella asiatica and Withania somnifera that have been explored in some recent studies. In agreement with the previous work on ethanol extracts of these two plants in mice model, we saw an improvement in oxidative stress profile as well as behavioral performance in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP induced Parkinson-like symptoms in Balb/c mice. Given the known potential of both the herbal extracts in improving Parkinson-like symptoms, we expected the combination of the two to show better results than either of the two but surprisingly there was no additivity in either oxidative stress or behavioural recovery. In fact, in some assays, the combination performed worse than either of the two individual constituents. This effect of mixtures highlights the need of testing mixtures in supplements market using enthomedicine. The necessity of comparing multiple groups in this study to get most information from the experiments motivated us to design a ladder-like visualization to show comparison with different groups that we call complete comparison display (CCD. In summary, we show the potential of Centella asiatica and Withania somnifera to ameliorate Parkinson's disorder.

  8. Complete Comparison Display (CCD) evaluation of ethanol extracts of Centella asiatica and Withania somnifera shows that they can non-synergistically ameliorate biochemical and behavioural damages in MPTP induced Parkinson's model of mice.

    Science.gov (United States)

    Bhatnagar, Maheep; Goel, Ishan; Roy, Tathagato; Shukla, Sunil Dutt; Khurana, Sukant

    2017-01-01

    Parkinson's disease remains as one of the most common debilitating neurodegenerative disorders. With the hopes of finding agents that can cure or reduce the pace of progression of the disease, we studied two traditional medicinal plants: Centella asiatica and Withania somnifera that have been explored in some recent studies. In agreement with the previous work on ethanol extracts of these two plants in mice model, we saw an improvement in oxidative stress profile as well as behavioral performance in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced Parkinson-like symptoms in Balb/c mice. Given the known potential of both the herbal extracts in improving Parkinson-like symptoms, we expected the combination of the two to show better results than either of the two but surprisingly there was no additivity in either oxidative stress or behavioural recovery. In fact, in some assays, the combination performed worse than either of the two individual constituents. This effect of mixtures highlights the need of testing mixtures in supplements market using enthomedicine. The necessity of comparing multiple groups in this study to get most information from the experiments motivated us to design a ladder-like visualization to show comparison with different groups that we call complete comparison display (CCD). In summary, we show the potential of Centella asiatica and Withania somnifera to ameliorate Parkinson's disorder.

  9. The use of a modified [3H]4-DAMP radioligand binding assay with increased selectivity for muscarinic M3 receptor shows that cortical CHRM3 levels are not altered in mood disorders.

    Science.gov (United States)

    Jeon, Won Je; Gibbons, Andrew S; Dean, Brian

    2013-12-02

    [(3)H]4-DAMP is a radioligand that has been used to quantify levels of the muscarinic receptor CHRM3 protein in situ. However, in addition to high affinity binding to CHRM3, [(3)H]4-DAMP binds with low affinity to CHRM1 confounding the potential to discriminate between changes in these two muscarinic receptors. We have developed a [(3)H]4-DAMP binding assay, optimised for measuring CHRM3 protein levels in the cortex, with minimal selectivity towards CHRM1. The selectivity of our assay towards CHRM3 was confirmed using recombinant receptor-expressing, cell lysate preparations. [(3)H]4-DAMP binding levels were similar between wildtype and CHRM1 knockout mice, confirming that the amount of [(3)H]4-DAMP binding to CHRM1 was negligible. We used this assay to measure CHRM3 protein levels in the frontal pole, obtained post-mortem from subjects with bipolar disorder (n = 15), major depressive disorder (n = 15) and matched controls (n = 20) and showed that [(3)H]4-DAMP binding was not altered in either bipolar disorder or major depressive disorder. Western blotting confirmed that CHRM3 protein levels were unchanged in these subjects. © 2013.

  10. Cloning and Biochemical Characterization of HIV Integrase

    Science.gov (United States)

    1989-12-15

    biochemical assays for the interaction of the HIV Intebration protein, IN, with a specific DNA target, the viral LTRs. The gene encoding IN has been subcloned ...FILE COPY CO CONTRACT NO.: DAMD7-88-C-8126 r TITLE: CLONING AND BIOCHEMICAL CHARACTERIZATION OF HIV INTEGRASE I- SPRINCIPAL INVESTIGATOR: ELLEN...ELEMENT No. NO. 392- NO. IACCESSION NO. 1 . TITLE (indude Security Classification)630A I315D49C08 (U) Cloning and Biochemical Characterization of the HIV

  11. Lateral flow assays.

    Science.gov (United States)

    Koczula, Katarzyna M; Gallotta, Andrea

    2016-06-30

    Lateral flow assays (LFAs) are the technology behind low-cost, simple, rapid and portable detection devices popular in biomedicine, agriculture, food and environmental sciences. This review presents an overview of the principle of the method and the critical components of the assay, focusing on lateral flow immunoassays. This type of assay has recently attracted considerable interest because of its potential to provide instantaneous diagnosis directly to patients. The range and interpretation of results and parameters used for evaluation of the assay will also be discussed. The main advantages and disadvantages of LFAs will be summarized and relevant future improvements to testing devices and strategies will be proposed. Finally, the major recent advances and future diagnostic applications in the LFA field will be explored. © 2016 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  12. Isolation of Cronobacter spp. (formerly Enterobacter sakazakii) from infant food, herbs and environmental samples and the subsequent identification and confirmation of the isolates using biochemical, chromogenic assays, PCR and 16S rRNA sequencing.

    Science.gov (United States)

    Jaradat, Ziad W; Ababneh, Qotaiba O; Saadoun, Ismail M; Samara, Nawal A; Rashdan, Abrar M

    2009-10-27

    Cronobacter spp. (formerly Enterobacter sakazakii), are a group of Gram-negative pathogens that have been implicated as causative agents of meningitis and necrotizing enterocolitis in infants. The pathogens are linked to infant formula; however, they have also been isolated from a wide range of foods and environmental samples. In this study, 233 samples of food, infant formula and environment were screened for the presence of Cronobacter spp. in an attempt to find its source. Twenty nine strains were isolated from samples of spices, herbs, infant foods, and dust obtained from household vacuum cleaners. Among the 76 samples of infant food, infant formula, milk powder and non-milk dairy products tested, only one sample of infant food contained Cronobacter spp. (1.4%). The other Cronobacter spp. isolates recovered include two from household vacuum dust, and 26 from 67 samples of herbs and spices. Among the food categories analyzed, herbs and spices harbored the highest number of isolates, indicating plants as a possible reservoir of this pathogen. Initial screening with API 20E test strips yielded 42 presumptive isolates. Further characterization using 3 chromogenic media (alpha-MUG, DFI and EsPM) and 8 sets of PCR primers detecting ITS (internal transcribed spacer sequences), 16S rRNA, zpx, gluA, gluB, OmpA genes followed by nucleotide sequencing of some PCR amplicons did not confirm the identity of all the isolates as none of the methods proved to be free of both false positives or false negatives. The final confirmation step was done by 16S rRNA sequence analysis identifying only 29 of the 42 isolates as Cronobacter spp. Our studies showed that Cronobacter spp. are highly diverse and share many phenotypic traits with other Enterobacteriaceae members highlighting the need to use several methods to confirm the identity of this pathogen. None of the biochemical, chromogenic or PCR primers proved to be a reliable method for confirmation of the identity of the isolates

  13. Isolation of Cronobacter spp. (formerly Enterobacter sakazakii from infant food, herbs and environmental samples and the subsequent identification and confirmation of the isolates using biochemical, chromogenic assays, PCR and 16S rRNA sequencing

    Directory of Open Access Journals (Sweden)

    Samara Nawal A

    2009-10-01

    Full Text Available Abstract Background Cronobacter spp. (formerly Enterobacter sakazakii, are a group of Gram-negative pathogens that have been implicated as causative agents of meningitis and necrotizing enterocolitis in infants. The pathogens are linked to infant formula; however, they have also been isolated from a wide range of foods and environmental samples. Results In this study, 233 samples of food, infant formula and environment were screened for the presence of Cronobacter spp. in an attempt to find its source. Twenty nine strains were isolated from samples of spices, herbs, infant foods, and dust obtained from household vacuum cleaners. Among the 76 samples of infant food, infant formula, milk powder and non-milk dairy products tested, only one sample of infant food contained Cronobacter spp. (1.4%. The other Cronobacter spp. isolates recovered include two from household vacuum dust, and 26 from 67 samples of herbs and spices. Among the food categories analyzed, herbs and spices harbored the highest number of isolates, indicating plants as a possible reservoir of this pathogen. Initial screening with API 20E test strips yielded 42 presumptive isolates. Further characterization using 3 chromogenic media (α-MUG, DFI and EsPM and 8 sets of PCR primers detecting ITS (internal transcribed spacer sequences, 16S rRNA, zpx, gluA, gluB, OmpA genes followed by nucleotide sequencing of some PCR amplicons did not confirm the identity of all the isolates as none of the methods proved to be free of both false positives or false negatives. The final confirmation step was done by 16S rRNA sequence analysis identifying only 29 of the 42 isolates as Cronobacter spp. Conclusion Our studies showed that Cronobacter spp. are highly diverse and share many phenotypic traits with other Enterobacteriaceae members highlighting the need to use several methods to confirm the identity of this pathogen. None of the biochemical, chromogenic or PCR primers proved to be a reliable

  14. Binding of peptides from the N-terminal region of alpha-gliadin to the celiac disease-associated HLA-DQ2 molecule assessed in biochemical and T cell assays

    DEFF Research Database (Denmark)

    Johansen, B H; Gjertsen, H A; Vartdal, F

    1996-01-01

    -purified DQ2 and DR3 (i.e., DR(alpha, beta1*0301)) molecules. The peptides were also tested for binding to DQ2 in a functional binding assay, where binding was measured as the capacity to inhibit the stimulation of a gliadin-specific, DQ2-restricted T lymphocyte clone RNnTalpha33. In both assay systems...... here provide new methods for the screening of potentially toxic peptides....

  15. Enzyme assays.

    Science.gov (United States)

    Brodelius, P E

    1991-02-01

    The past year or so has seen the development of new enzyme assays, as well as the improvement of existing ones. Assays are becoming more rapid and sensitive as a result of modifications such as amplification of the enzyme product(s). Recombinant DNA technology is now being recognized as a particularly useful tool in the search for improved assay systems.

  16. Clinical impact of prostate specific antigen (PSA) inter-assay variability on management of prostate cancer.

    Science.gov (United States)

    Murthy, Vedang; Rishi, Anupam; Gupta, Sanjeev; Kannan, Sadhana; Mahantshetty, Umesh; Tongaonkar, Hemant; Bakshi, Ganesh; Prabhash, Kumar; Bhanushali, Paresh; Shinde, Bhoopal; Inamdar, Nitin; Shrivastava, Shyamkishore

    2016-01-01

    To evaluate the inter-assay variability of six commercially available prostate specific antigen (PSA) assays, its clinical impact in prostate cancer (PCa) and comparison of automated versus manual assays. Sera from 495 patients (425 with PCa and 70 men with Benign Prostatic Hyperplasia (BPH), were measured with six different assays [three automated assays (a-PSA) and three manual ELISA based assay (m-PSA)]. Variability, agreement and bias were measured and compared among assays using Bland Altman plots and Passing and Bablok regression analysis. The possible impact of inter-assay variability on important clinical scenarios was also studied. All the assays were well correlated (r: 0.88-0.98); however there was significant disagreement and bias between the systems, which were more pronounced among the a-PSA assays. The Bland Altman plot showed that the variability was high between the m-PSA assays and the standard Abbott system with mean difference of 3.8-5.8ng/ml. In contrast, the a-PSA had better agreement with mean difference of 0.8-2.3ng/ml. Beckman Coulter showed the best agreement to the institutional reference (slope-1.097; 95% CI: 1.06-1.14; p<0.05, and intercept-0.20; 95% CI-0.38-0.58; p<0.05, Passing Bablok). It led to significant variability in PCa risk stratification and failure to detect biochemical failure in more than 50% cases. The discrepancies between the assays lead to significant clinical misinterpretation with risk group migration and detection of biochemical failure post radiotherapy. There are significant discordances between automated and ELISA based assays. Copyright © 2015 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  17. Biochemical Characterization of Prion Strains in Bank Voles

    Directory of Open Access Journals (Sweden)

    Romolo Nonno

    2013-07-01

    Full Text Available Prions exist as different strains exhibiting distinct disease phenotypes. Currently, the identification of prion strains is still based on biological strain typing in rodents. However, it has been shown that prion strains may be associated with distinct PrPSc biochemical types. Taking advantage of the availability of several prion strains adapted to a novel rodent model, the bank vole, we investigated if any prion strain was actually associated with distinctive PrPSc biochemical characteristics and if it was possible to univocally identify strains through PrPSc biochemical phenotypes. We selected six different vole-adapted strains (three human-derived and three animal-derived and analyzed PrPSc from individual voles by epitope mapping of protease resistant core of PrPSc (PrPres and by conformational stability and solubility assay. Overall, we discriminated five out of six prion strains, while two different scrapie strains showed identical PrPSc types. Our results suggest that the biochemical strain typing approach here proposed was highly discriminative, although by itself it did not allow us to identify all prion strains analyzed.

  18. Comparison of the effect of chemical composition of anthocyanin-rich plant extracts on colon cancer cell proliferation and their potential mechanism of action using in vitro, in silico, and biochemical assays.

    Science.gov (United States)

    Mazewski, Candice; Liang, Katie; Gonzalez de Mejia, Elvira

    2018-03-01

    The objective was to compare the anti-proliferative effect of anthocyanin-rich plant extracts on human colon cancer cells and determine their mechanism of action. Eleven extracts were tested: red (RG) and purple grape, purple sweet potato, purple carrot, black and purple bean, black lentil (BL), black peanut, sorghum (SH), black rice, and blue wheat. HCT-116 and HT-29 inhibition correlated with total phenolics (r=0.87 and 0.77, respectively), delphinidin-3-O-glucoside concentration with HT-29 inhibition (r=0.69). The concentration inhibition fifty (IC 50 ) for BL, SH, RG on HT-29 and HCT-116 cell proliferation ranged 0.9-2.0mg/mL. Extracts decreased expression of anti-apoptotic proteins (survivin, cIAP-2, XIAP), induced apoptosis, and arrested cells in G1. Anthocyanins exhibited tyrosine kinase inhibitory potential in silico and biochemically; cyanidin-3-O-glucoside had one of the highest binding affinities with all kinases, especially ABL1 (-8.5kcal/mol). Cyanidin-3-O-glucoside and delphinidin-3-O-glucoside inhibited EGFR (IC 50 =0.10 and 2.37µM, respectively). Cyanidin-3-O-glucoside was the most potent anthocyanin on kinase inhibition. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Morpho-Physiological and Biochemical Criteria of Acanthamoeba spp. Isolated from the Egyptian Aquatic Environment

    Science.gov (United States)

    Al-Herrawy, A; Bahgat, M; Mohammed, A; Ashour, A; Hikal, W

    2013-01-01

    Background The free-living amoebae Acanthamoeba spp., have been recognized as etiologic agents of amoebic encephalitis, keratitis, otitis, lung lesions and other skin infections mainly in immuno-compromised individuals. In this study, morpho-physiological and biochemical characterization of Acanthamoeba strains isolated from the Egyptian aquatic environment were surveyed. Methods Some Acanthamoeba species were cultivated on non-nutrient agar. Isolated strains of Acanthamoeba were identification based on the morphology of trophic and cyst forms in addition to temperature and osmo-tolerance assays. Biochemical characterization of the isolated amoeba strains was performed using quantitative assay as well as qualitative determination of proteolytic activity in zymograph analysis. Results Potentially pathogenic Acanthamoeba species were isolated from all of the examined water sources. Colorimetric assays showed protease activity in the heat-tolerant isolates of Acanthamoeba. All pathogenic isolates of Acanthamoeba exhibited higher protease activity than did the non-pathogenic ones. The zymographic protease assays showed various banding patterns for different strains of Acanthamoeba. Conclusion The incidence and prevalence of the pathogenic Acanthamoeba species in the aquatic environment using parasitological and biochemical diagnostic tools will provide baseline data against which the risk factors associated with waterborne transmission can be identified. PMID:23914245

  20. Preliminary biochemical and haematological effects of aqueous ...

    African Journals Online (AJOL)

    Some biochemical and haematological parameters were assayed in rats fed aqueous pulp suspension of hyphaene thebaica (L) mart. Sixteen white albino rats of the wistar strain weighing between 90-110g were grouped into four groups of four rats each. Group 1 served as control while groups 2, 3 and 4 were given ...

  1. Discordant results between biochemical and molecular transthyretin

    Indian Academy of Sciences (India)

    Discordant results between biochemical and molecular transthyretin assays: lessons learned from a unique testing algorithm at the Mayo Clinic. Honey V. Reddi Brittany C. Thomas Kurt S. Willkomm Matthew J. Ferber Kandelaria M. Rumilla Kimiyo M. Raymond John F. O'Brien W. Edward Highsmith. Research Note Volume ...

  2. Possible Biochemical Markers in Protein-Energy Malnutrition and ...

    African Journals Online (AJOL)

    This study was carried out to determine possible biochemical markers in children suffering from Plasmodium falciparum malaria and Protein-Energy Malnutrition in a Hospital setting in Western Kenya. Spectrophotometric assays of selected biochemical parameters namely, albumin, total proteins, glucose, glutamate ...

  3. Biochemical Hypermedia: Galactose Metabolism.

    Directory of Open Access Journals (Sweden)

    J.K. Sugai

    2013-05-01

    Full Text Available Introduction: Animations of biochemical processes and virtual laboratory environments lead to true molecular simulations. The use of interactive software’s in education can improve cognitive capacity, better learning and, mainly, it makes information acquisition easier. Material and Methods: This work presents the development of a biochemical hypermedia to understanding of the galactose metabolism. It was developed with the help of concept maps, ISIS Draw, ADOBE Photoshop and FLASH MX Program. Results and Discussion: A step by step animation process shows the enzymatic reactions of galactose conversion to glucose-1-phosphate (to glycogen synthesis, glucose-6-phosphate (glycolysis intermediary, UDP-galactose (substrate to mucopolysaccharides synthesis and collagen’s glycosylation. There are navigation guide that allow scrolling the mouse over the names of the components of enzymatic reactions of via the metabolism of galactose. Thus, explanatory text box, chemical structures and animation of the actions of enzymes appear to navigator. Upon completion of the module, the user’s response to the proposed exercise can be checked immediately through text box with interactive content of the answer. Conclusion: This hypermedia was presented for undergraduate students (UFSC who revealed that it was extremely effective in promoting the understanding of the theme.

  4. Biochemical markers of bone turnover

    International Nuclear Information System (INIS)

    Kim, Deog Yoon

    1999-01-01

    Biochemical markers of bone turnover has received increasing attention over the past few years, because of the need for sensitivity and specific tool in the clinical investigation of osteoporosis. Bone markers should be unique to bone, reflect changes of bone less, and should be correlated with radiocalcium kinetics, histomorphometry, or changes in bone mass. The markers also should be useful in monitoring treatment efficacy. Although no bone marker has been established to meet all these criteria, currently osteocalcin and pyridinium crosslinks are the most efficient markers to assess the level of bone turnover in the menopausal and senile osteoporosis. Recently, N-terminal telopeptide (NTX), C-terminal telopeptide (CTX) and bone specific alkaline phosphatase are considered as new valid markers of bone turnover. Recent data suggest that CTX and free deoxypyridinoline could predict the subsequent risk of hip fracture of elderly women. Treatment of postmenopausal women with estrogen, calcitonin and bisphosphonates demonstrated rapid decrease of the levels of bone markers that correlated with the long-term increase of bone mass. Factors such as circadian rhythms, diet, age, sex, bone mass and renal function affect the results of biochemical markers and should be appropriately adjusted whenever possible. Each biochemical markers of bone turnover may have its own specific advantages and limitations. Recent advances in research will provide more sensitive and specific assays

  5. Biochemical markers of bone turnover

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Deog Yoon [College of Medicine, Kyunghee Univ., Seoul (Korea, Republic of)

    1999-08-01

    Biochemical markers of bone turnover has received increasing attention over the past few years, because of the need for sensitivity and specific tool in the clinical investigation of osteoporosis. Bone markers should be unique to bone, reflect changes of bone less, and should be correlated with radiocalcium kinetics, histomorphometry, or changes in bone mass. The markers also should be useful in monitoring treatment efficacy. Although no bone marker has been established to meet all these criteria, currently osteocalcin and pyridinium crosslinks are the most efficient markers to assess the level of bone turnover in the menopausal and senile osteoporosis. Recently, N-terminal telopeptide (NTX), C-terminal telopeptide (CTX) and bone specific alkaline phosphatase are considered as new valid markers of bone turnover. Recent data suggest that CTX and free deoxypyridinoline could predict the subsequent risk of hip fracture of elderly women. Treatment of postmenopausal women with estrogen, calcitonin and bisphosphonates demonstrated rapid decrease of the levels of bone markers that correlated with the long-term increase of bone mass. Factors such as circadian rhythms, diet, age, sex, bone mass and renal function affect the results of biochemical markers and should be appropriately adjusted whenever possible. Each biochemical markers of bone turnover may have its own specific advantages and limitations. Recent advances in research will provide more sensitive and specific assays.

  6. Disagreement between Human Papillomavirus Assays

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Preisler, Sarah; Ejegod, Ditte Møller

    2014-01-01

    -65 years (n = 2,881), 23% tested positive on at least one assay, and 42 to 58% of these showed positive agreement on any compared pair of the assays. While 4% of primary screening samples showed abnormal cytology, 6 to 10% were discordant on any pair of assays. A literature review corroborated our findings...

  7. Histological and Biochemical Evaluation of the Kidney following Chronic Consumption of Hibiscus sabdariffa

    Directory of Open Access Journals (Sweden)

    U. U. Ukoha

    2015-01-01

    Full Text Available Hibiscus sabdariffa L. has been used traditionally as herbal medicine and has been documented to have a broad range of therapeutic effects. The present study aimed to investigate the effects of chronic administration of aqueous extract of flowers of Hibiscus sabdariffa on the histology of the kidney and some biochemical indices of renal function in male Wistar rats. Twenty (20 Wistar rats were randomly divided into four (4 groups of five rats each. The extract was administered orally in doses 200, 500, and 800 mg/kg body weight for 21 days. The kidney was harvested and processed histologically and blood samples were taken for biochemical assays. The histological results showed dose dependent pathological states and the biochemical analysis revealed a dose dependant variation in renal indices. These results suggest that chronic administration of aqueous extract of Hibiscus sabdariffa may be toxic to the kidney.

  8. Hormone assay

    International Nuclear Information System (INIS)

    Eisentraut, A.M.

    1977-01-01

    An improved radioimmunoassay is described for measuring total triiodothyronine or total thyroxine levels in a sample of serum containing free endogenous thyroid hormone and endogenous thyroid hormone bound to thyroid hormone binding protein. The thyroid hormone is released from the protein by adding hydrochloric acid to the serum. The pH of the separated thyroid hormone and thyroid hormone binding protein is raised in the absence of a blocking agent without interference from the endogenous protein. 125 I-labelled thyroid hormone and thyroid hormone antibodies are added to the mixture, allowing the labelled and unlabelled thyroid hormone and the thyroid hormone antibody to bind competitively. This results in free thyroid hormone being separated from antibody bound thyroid hormone and thus the unknown quantity of thyroid hormone may be determined. A thyroid hormone test assay kit is described for this radioimmunoassay. It provides a 'single tube' assay which does not require blocking agents for endogenous protein interference nor an external solid phase sorption step for the separation of bound and free hormone after the competitive binding step; it also requires a minimum number of manipulative steps. Examples of the assay are given to illustrate the reproducibility, linearity and specificity of the assay. (UK)

  9. Binding of peptides from the N-terminal region of alpha-gliadin to the celiac disease-associated HLA-DQ2 molecule assessed in biochemical and T cell assays

    DEFF Research Database (Denmark)

    Johansen, B H; Gjertsen, H A; Vartdal, F

    1996-01-01

    Celiac disease (CD) is most probably an immunological disease, precipitated in susceptible individuals by ingestion of wheat gliadin and related proteins from other cereals. The disease shows a strong HLA association predominantly to the cis- or trans-encoded HLA-DQ(alpha1*0501, beta1*02) (i.e., DQ...

  10. Thyroglobulin measurement by highly sensitive assays

    DEFF Research Database (Denmark)

    Giovanella, Luca; Feldt-Rasmussen, Ulla; Verburg, Frederik A

    2015-01-01

    of thyroglobulin (Tg), circulating Tg serves as a biochemical marker of persistent or recurrent disease in the follow-up of DTC. Due to the suboptimal clinical detection rate of older Tg assays endogenous or exogenous thyrotropin (TSH) stimulations are recommended for unmasking occult disease. However...... in current clinical guidelines, such assays are now increasingly used in clinical practice. As serum Tg measurement is a technically challenging assay and criteria to define a 'highly sensitive' assay may be different, a good knowledge of the technical difficulties and interpretation criteria is of paramount...

  11. Radioreceptor assays

    International Nuclear Information System (INIS)

    Lapka, R.

    1985-01-01

    Radioreceptor assay (RRA) is an analytical method using the specific interaction of some pharmaceuticals and endogenic substances (ligands) with specific receptors present in certin tissues of living organisms. RRA uses the principle of isotope dilution. The method is described in detail of the preparation of receptors, samples and radioligands, conditions of incubation, the separation of free and bound radioligand, and the mathematical evaluation of RRA. The sensitivity of RRA is measured in units to tens of pg. The specificity of RRA relates to a group of substances with similar pharmacological effect. RRA may be used for identifying neuroleptics, antidepressants, anxiolytics, ergot alkaloids, beta blockers, anticholinergic drugs, certain hormones and neuropeptides. (M.D.)

  12. Serum Biochemical Phenotypes in the Domestic Dog.

    Directory of Open Access Journals (Sweden)

    Yu-Mei Chang

    Full Text Available The serum or plasma biochemical profile is essential in the diagnosis and monitoring of systemic disease in veterinary medicine, but current reference intervals typically take no account of breed-specific differences. Breed-specific hematological phenotypes have been documented in the domestic dog, but little has been published on serum biochemical phenotypes in this species. Serum biochemical profiles of dogs in which all measurements fell within the existing reference intervals were retrieved from a large veterinary database. Serum biochemical profiles from 3045 dogs were retrieved, of which 1495 had an accompanying normal glucose concentration. Sixty pure breeds plus a mixed breed control group were represented by at least 10 individuals. All analytes, except for sodium, chloride and glucose, showed variation with age. Total protein, globulin, potassium, chloride, creatinine, cholesterol, total bilirubin, ALT, CK, amylase, and lipase varied between sexes. Neutering status significantly impacted all analytes except albumin, sodium, calcium, urea, and glucose. Principal component analysis of serum biochemical data revealed 36 pure breeds with distinctive phenotypes. Furthermore, comparative analysis identified 23 breeds with significant differences from the mixed breed group in all biochemical analytes except urea and glucose. Eighteen breeds were identified by both principal component and comparative analysis. Tentative reference intervals were generated for breeds with a distinctive phenotype identified by comparative analysis and represented by at least 120 individuals. This is the first large-scale analysis of breed-specific serum biochemical phenotypes in the domestic dog and highlights potential genetic components of biochemical traits in this species.

  13. Biochemical genetics of some Indian fishes

    Digital Repository Service at National Institute of Oceanography (India)

    Menezes, M.R.; Qasim, S.Z.

    Studies on biochemical genetics of fishes, using electrophoretic methods, are relatively of recent origin. Earlier serum and eye lens protein were used to identify marine populations. This technique showed that closely related species have...

  14. Dactylifera L) on the biochemical indicators of lead poisoning in ...

    African Journals Online (AJOL)

    ... in the biochemical parameters. Thus, the treatment by the pectin of dates reduced the high concentration of these parameters. Our results show that the pectins of dates may have a corrective effect on the biochemical disturbances induced by the lead. Keywords: Phoenix Dactylifera, Pectin, Lead, Biochemical parameters ...

  15. Comparative biochemical analysis of recombinant reverse transcriptase enzymes of HIV-1 subtype B and subtype C

    Directory of Open Access Journals (Sweden)

    Moisi Daniella

    2010-10-01

    Full Text Available Abstract Background HIV-1 subtype C infections account for over half of global HIV infections, yet the vast focus of HIV-1 research has been on subtype B viruses which represent less than 12% of the global pandemic. Since HIV-1 reverse transcriptase (RT is a major target of antiviral therapy, and since differential drug resistance pathways have been observed among different HIV subtypes, it is important to study and compare the enzymatic activities of HIV-1 RT derived from each of subtypes B and C as well as to determine the susceptibilities of these enzymes to various RT inhibitors in biochemical assays. Methods Recombinant subtype B and C HIV-1 RTs in heterodimeric form were purified from Escherichia coli and enzyme activities were compared in cell-free assays. The efficiency of (- ssDNA synthesis was measured using gel-based assays with HIV-1 PBS RNA template and tRNA3Lys as primer. Processivity was assayed under single-cycle conditions using both homopolymeric and heteropolymeric RNA templates. Intrinsic RNase H activity was compared using 5'-end labeled RNA template annealed to 3'-end recessed DNA primer in a time course study in the presence and absence of a heparin trap. A mis-incorporation assay was used to assess the fidelity of the two RT enzymes. Drug susceptibility assays were performed both in cell-free assays using recombinant enzymes and in cell culture phenotyping assays. Results The comparative biochemical analyses of recombinant subtype B and subtype C HIV-1 reverse transcriptase indicate that the two enzymes are very similar biochemically in efficiency of tRNA-primed (- ssDNA synthesis, processivity, fidelity and RNase H activity, and that both enzymes show similar susceptibilities to commonly used NRTIs and NNRTIs. Cell culture phenotyping assays confirmed these results. Conclusions Overall enzyme activity and drug susceptibility of HIV-1 subtype C RT are comparable to those of subtype B RT. The use of RT inhibitors (RTIs

  16. Biochemical Neuroscience Laboratory

    Data.gov (United States)

    Federal Laboratory Consortium — This biochemistry lab is set up for protein analysis using Western blot, enzyme linked immunosorbent assays, immunohistochemistry, and bead-based immunoassays. The...

  17. Molecular and biochemical characterisation of Trypanosoma cruzi phosphofructokinase.

    Science.gov (United States)

    Rodríguez, Evelyn; Lander, Noelia; Ramirez, Jose Luis

    2009-08-01

    The characterisation of the gene encoding Trypanosoma cruzi CL Brener phosphofructokinase (PFK) and the biochemical properties of the expressed enzyme are reported here. In contradiction with previous reports, the PFK genes of CL Brener and YBM strain T. cruzi were found to be similar to their Leishmania mexicana and Trypanosoma brucei homologs in terms of both kinetic properties and size, with open reading frames encoding polypeptides with a deduced molecular mass of 53,483. The predicted amino acid sequence contains the C-terminal glycosome-targeting tripeptide SKL; this localisation was confirmed by immunofluorescence assays. In sequence comparisons with the genes of other eukaryotes, it was found that, despite being an adenosine triphosphate-dependent enzyme, T. cruzi PFK shows significant sequence similarity with inorganic pyrophosphate-dependent PFKs.

  18. Molecular and biochemical characterisation of Trypanosoma cruzi phosphofructokinase

    Directory of Open Access Journals (Sweden)

    Evelyn Rodríguez

    2009-08-01

    Full Text Available The characterisation of the gene encoding Trypanosoma cruzi CL Brener phosphofructokinase (PFK and the biochemical properties of the expressed enzyme are reported here. In contradiction with previous reports, the PFK genes of CL Brener and YBM strain T. cruzi were found to be similar to their Leishmania mexicana and Trypanosoma brucei homologs in terms of both kinetic properties and size, with open reading frames encoding polypeptides with a deduced molecular mass of 53,483. The predicted amino acid sequence contains the C-terminal glycosome-targeting tripeptide SKL; this localisation was confirmed by immunofluorescence assays. In sequence comparisons with the genes of other eukaryotes, it was found that, despite being an adenosine triphosphate-dependent enzyme, T. cruzi PFK shows significant sequence similarity with inorganic pyrophosphate-dependent PFKs.

  19. Cytotoxicity assay automation

    Science.gov (United States)

    Levinthal, E. C.; Payne, R. O.

    1971-01-01

    The design and construction of a system to automatically test HLP antigens are described. Major efforts were made to test and evaluate the performance of such a system, and compare its performance with nonautomatic tissue typing techniques. The system is based on the fluorochromatic cytotoxicity assay. Results show the system will work but is subject to malfunctions after a few samplings, and poses problems in showing correctly the necessary readings.

  20. Measures of Biochemical Sociology

    Science.gov (United States)

    Snell, Joel; Marsh, Mitchell

    2008-01-01

    In a previous article, the authors introduced a new sub field in sociology that we labeled "biochemical sociology." We introduced the definition of a sociology that encompasses sociological measures, psychological measures, and biological indicators Snell & Marsh (2003). In this article, we want to demonstrate a research strategy that would assess…

  1. Biochemical Education in Brazil.

    Science.gov (United States)

    Vella, F.

    1988-01-01

    Described are discussions held concerning the problems of biochemical education in Brazil at a meeting of the Sociedade Brazileira de Bioquimica in April 1988. Also discussed are other visits that were made to universities in Brazil. Three major recommendations to improve the state of biochemistry education in Brazil are presented. (CW)

  2. 78 FR 19499 - Request for Information: The National Toxicology Program Requests Information On Assays and...

    Science.gov (United States)

    2013-04-01

    ... (e.g., 25-50) in biochemical- or cell- based assays or alternative (non-rodent) animal models, that might be used to prioritize compounds for in vivo neurotoxicity testing. DATES: The deadline for receipt... biochemical- or cell-based assays or alternative animal models that assess the potential ability of compounds...

  3. Metabolic profiles show specific mitochondrial toxicities in vitro in myotube cells

    International Nuclear Information System (INIS)

    Xu Qiuwei; Vu, Heather; Liu Liping; Wang, Ting-Chuan; Schaefer, William H.

    2011-01-01

    Mitochondrial toxicity has been a serious concern, not only in preclinical drug development but also in clinical trials. In mitochondria, there are several distinct metabolic processes including fatty acid β-oxidation, the tricarboxylic acid (TCA) cycle, and oxidative phosphorylation (OXPHOS), and each process contains discrete but often intimately linked steps. Interruption in any one of those steps can cause mitochondrial dysfunction. Detection of inhibition to OXPHOS can be complicated in vivo because intermediate endogenous metabolites can be recycled in situ or circulated systemically for metabolism in other organs or tissues. Commonly used assays for evaluating mitochondrial function are often applied to ex vivo or in vitro samples; they include various enzymatic or protein assays, as well as functional assays such as measurement of oxygen consumption rate, membrane potential, or acidification rates. Metabolomics provides quantitative profiles of overall metabolic changes that can aid in the unraveling of explicit biochemical details of mitochondrial inhibition while providing a holistic view and heuristic understanding of cellular bioenergetics. In this paper, we showed the application of quantitative NMR metabolomics to in vitro myotube cells treated with mitochondrial toxicants, rotenone and antimycin A. The close coupling of the TCA cycle to the electron transfer chain (ETC) in OXPHOS enables specific diagnoses of inhibition to ETC complexes by discrete biochemical changes in the TCA cycle.

  4. Ouroboros - Playing A Biochemical

    Directory of Open Access Journals (Sweden)

    D. T. Rodrigues

    2014-08-01

    Full Text Available Ouroboros: Playing A Biochemical RODRIGUES,D.T.1,2;GAYER, M.C.1,2; ESCOTO, D.F.1; DENARDIN, E.L.G.2, ROEHRS, R.1,2 1Interdisciplinary Research Group on Teaching Practice, Graduate Program in Biochemistry, Unipampa, RS, Brazil 2Laboratory of Physicochemical Studies and Natural Products, Post Graduate Program in Biochemistry, Unipampa, RS, Brazil Introduction: Currently, teachers seek different alternatives to enhance the teaching-learning process. Innovative teaching methodologies are increasingly common tools in educational routine. The use of games, electronic or conventional, is an effective tool to assist in learning and also to raise the social interaction between students. Objective: In this sense our work aims to evaluate the card game and "Ouroboros" board as a teaching and learning tool in biochemistry for a graduating class in Natural Sciences. Materials and methods: The class gathered 22 students of BSc in Natural Sciences. Each letter contained a question across the board that was drawn to a group to answer within the allotted time. The questions related concepts of metabolism, organic and inorganic chemical reactions, bioenergetics, etc.. Before the game application, students underwent a pre-test with four issues involving the content that was being developed. Soon after, the game was applied. Then again questions were asked. Data analysis was performed from the ratio of the number of correct pre-test and post-test answers. Results and discussion: In the pre-test 18.1% of the students knew all issues, 18.1% got 3 correct answers, 40.9% answered only 2 questions correctly and 22.7% did not hit any. In post-test 45.4% answered all the questions right, 31.8% got 3 questions and 22.7% got 2 correct answers. The results show a significant improvement of the students about the field of content taught through the game. Conclusion: Generally, traditional approaches of chemistry and biochemistry are abstract and complex. Thus, through games

  5. A novel real-time PCR assay for quantitative detection of Campylobacter fetus based on ribosomal sequences.

    Science.gov (United States)

    Iraola, Gregorio; Pérez, Ruben; Betancor, Laura; Marandino, Ana; Morsella, Claudia; Méndez, Alejandra; Paolicchi, Fernando; Piccirillo, Alessandra; Tomás, Gonzalo; Velilla, Alejandra; Calleros, Lucía

    2016-12-15

    Campylobacter fetus is a pathogen of major concern for animal and human health. The species shows a great intraspecific variation, with three subspecies: C. fetus subsp. fetus, C. fetus subsp. venerealis, and C. fetus subsp. testudinum. Campylobacter fetus fetus affects a broad range of hosts and induces abortion in sheep and cows. Campylobacter fetus venerealis is restricted to cattle and causes the endemic disease bovine genital campylobacteriosis, which triggers reproductive problems and is responsible for major economic losses. Campylobacter fetus testudinum has been proposed recently based on genetically divergent strains isolated from reptiles and humans. Both C. fetus fetus and C. fetus testudinum are opportunistic pathogens for immune-compromised humans. Biochemical tests remain as the gold standard for identifying C. fetus but the fastidious growing requirements and the lack of reliability and reproducibility of some biochemical tests motivated the development of molecular diagnostic tools. These methods have been successfully tested on bovine isolates but fail to detect some genetically divergent strains isolated from other hosts. The aim of the present study was to develop a highly specific molecular assay to identify and quantify C. fetus strains. We developed a highly sensitive real-time PCR assay that targets a unique region of the 16S rRNA gene. This assay successfully detected all C. fetus strains, including those that were negative for the cstA gene-based assay used as a standard for molecular C. fetus identification. The assay showed high specificity and absence of cross-reactivity with other bacterial species. The analytical testing of the assay was determined using a standard curve. The assay demonstrated a wide dynamic range between 10 2 and 107 genome copies per reaction, and a good reproducibility with small intra- and inter-assay variability. The possibility to characterize samples in a rapid, sensitive and reproducible way makes this assay

  6. Biochemical blood parameters in semi-adult rabbits experimentally ...

    African Journals Online (AJOL)

    Biochemical parameters in blood specimens obtained from semi-adult rabbits of both sexes fed crude oil contaminated diets were examined. The diets had crude oil inclusions of 0, 0.05, 0.10, 0.15 and 0.20%. Blood samples were obtained from the marginal ear vein of representatives in each treatment group and assayed ...

  7. Molecular and biochemical diagnosis of Salmonella in wastewater ...

    African Journals Online (AJOL)

    This study aimed to employ biochemical and molecular assays to detect and diagnose Salmonella in wastewater. For this reason, two water samples were collected from Alexandria wastewater treatment plant (S1) and septic tank of a hospital at Alexandria governorate (S2). Selective culture media specific for Salmonella ...

  8. Dynamic changes in biochemical markers of renal function with ...

    African Journals Online (AJOL)

    Thyroid dysfunction is known to cause significant changes in glomerular filtration rate. The present cross-sectional study was performed to evaluate the changes in biochemical markers of renal function in hypothyroid subjects before and after treatment. Thyroid function tests (T3, T4 and TSH levels) were assayed in 385 ...

  9. Biochemical quality indices of sorghum genotypes from east Africa ...

    African Journals Online (AJOL)

    Biochemical characteristics assayed for 131 sorghum [Sorghum bicolor (L) Moench] accessions included total starch, amylopectin, amylose, proteins, tannins contents, germination energy and germination capacity. Results indicate that starch contents ranged from 22.8 - 81.2%, amylose from 11.5 - 30.2% while the ...

  10. Estrogen and progesterone receptors in endometrial carcinoma: comparison of immunohistochemical and biochemical analysis

    DEFF Research Database (Denmark)

    Nyholm, H C; Nielsen, Anette Lynge; Lyndrup, J

    1993-01-01

    In 159 endometrial carcinomas, estrogen (ER) and progesterone receptors (PR) were determined biochemically by dextran-coated charcoal (DCC) assay and immunohistochemically (ICA) on frozen sections. ICA receptor content was estimated by a total histologic score (HSCORE), including all tissue...

  11. Early Biochemical Screening for Fetal Aneuploidy in the First Trimester

    DEFF Research Database (Denmark)

    Tørring, Niels

    2013-01-01

    8+0 to 14+0 weeks, and clinical test performance of risk assesment was conducted. Results The in-between day imprecision of the Elecsys® free βhCG and PAPP-A assays were between 1.0 and 2.8%. Comparison (Passing/Bablok regression) of free βhCG and PAPP-A from Roche Elecsys® and the Brahms Kryptor...... with the standards for biochemical assays for prenatal screening set by the Fetal Medicine Foundation, with low assay imprecision, and a high clinical performance of prenatal screening for fetal trisomy in the first trimester....

  12. Mammalian cells exposed to ionizing radiation: structural and biochemical aspects

    International Nuclear Information System (INIS)

    Sabanero, M.; Flores V, L. L.; Azorin V, J. C.; Vallejo, M. A.; Cordova F, T.; Sosa A, M.; Castruita D, J. P.; Barbosa S, G.

    2015-10-01

    Acute or chronic exposure to ionizing radiation is a factor that may be hazardous to health. It has been reported that exposure to low doses of radiation (less than 50 mSv / year) and subsequently exposure to high doses have greater effects in people. However, it is unknown molecular and biochemical level alteration. This study, analyzes the susceptibility of a biological system (HeLa Atcc CCL-2 human cervix cancer cell line) to ionizing radiation (6 and 60 mSv/ 90). Our evaluate multiple variables such as: total protein profile, mitochondrial metabolic activity (XTT assay), cell viability (Trypan blue exclusion assay), cytoskeleton (actin micro filaments), nuclei (D API), genomic DNA. The results indicate, that cells exposed to ionizing radiation structurally show alterations in nuclear phenotype and aneuploidy, further disruption in the tight junctions and consequently on the distribution of actin micro filaments. Similar alterations were observed in cells treated with a genotoxic agent (200μM H 2 O 2 /1 h). In conclusion, this multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems. However, it is necessary to determine stress markers for integration of the effects of ionizing radiation. (Author)

  13. Mammalian cells exposed to ionizing radiation: structural and biochemical aspects

    Energy Technology Data Exchange (ETDEWEB)

    Sabanero, M.; Flores V, L. L. [Universidad de Guanajuato, Departamento de Biologia, DCNE, Noria Alta s/n, 36250 Guanajuato, Gto. (Mexico); Azorin V, J. C.; Vallejo, M. A.; Cordova F, T.; Sosa A, M. [Universidad de Guanajuato, Departamento de Ingenieria Fisica, DCI, Loma del Bosque 103, Col. Lomas del Campestre, 37150 Leon, Guanajuato (Mexico); Castruita D, J. P. [Universidad de Guadalajara, Departamento de Ecologia, CUCBA, Las Agujas, 45100 Zapopan, Jalisco (Mexico); Barbosa S, G., E-mail: myrna.sabanero@gmail.com [Universidad de Guanajuato, Departamento de Ciencias Medicas, DCS, 20 de Enero No. 929, Col. Obregon, 37000 Leon, Guanajuato (Mexico)

    2015-10-15

    Acute or chronic exposure to ionizing radiation is a factor that may be hazardous to health. It has been reported that exposure to low doses of radiation (less than 50 mSv / year) and subsequently exposure to high doses have greater effects in people. However, it is unknown molecular and biochemical level alteration. This study, analyzes the susceptibility of a biological system (HeLa Atcc CCL-2 human cervix cancer cell line) to ionizing radiation (6 and 60 mSv/ 90). Our evaluate multiple variables such as: total protein profile, mitochondrial metabolic activity (XTT assay), cell viability (Trypan blue exclusion assay), cytoskeleton (actin micro filaments), nuclei (D API), genomic DNA. The results indicate, that cells exposed to ionizing radiation structurally show alterations in nuclear phenotype and aneuploidy, further disruption in the tight junctions and consequently on the distribution of actin micro filaments. Similar alterations were observed in cells treated with a genotoxic agent (200μM H{sub 2}O{sub 2}/1 h). In conclusion, this multi-criteria assessment enables precise comparisons of the effects of radiation between any biological systems. However, it is necessary to determine stress markers for integration of the effects of ionizing radiation. (Author)

  14. Molecular and biochemical toxicology

    National Research Council Canada - National Science Library

    Smart, Robert C; Hodgson, Ernest

    2008-01-01

    ... Expression and Regulation 2.6.1 Northern Analysis 2.6.2 Nuclear Run-On 2.6.3 Promoter Deletion Analysis/Reporter Gene Assays 2.6.4 Microarrays 2.6.5 Reverse Transcriptase-PCR (RT-PCR) and Real-Time P...

  15. Biochemical activity of fullerenes and related derivatives

    International Nuclear Information System (INIS)

    Huczko, A.; Lange, H.; Calko, E.

    1999-01-01

    An astonishing scientific interest, embodied in over 15000 research articles so far, has been encountered since 1985 when fullerenes were discovered. From new superconductors to a rich electrochemistry and reaction chemistry, fullerene nanostructures continue to excite the scientific world, and new findings continue at record pace. This review presents many examples of the biochemical activities of fullerenes and derivatives, e. g. cytotoxic activity, selective DNA cleavage and antiviral activity against HIV. We also present some results of our testing which show that, despite its chemical and biochemical activity, fullerene matter does not present any health hazard directly related to skin irritation and allergic risks. (author)

  16. Structural and biochemical characterization of autotaxin

    NARCIS (Netherlands)

    Hausmann, Jens

    2013-01-01

    Autotaxin (ATX) was originally discovered as an “autocrine motility factor” from melanoma cells, more than two decades ago, but its biochemical function remained elusive. It took another decade to show that ATX functions as a lysophospholipase D that generates the lipid growth factor

  17. Biochemical and secondary metabolites changes under moisture ...

    African Journals Online (AJOL)

    The study showed the importance of carbohydrate and nitrogen cycle related metabolites in mediating tolerance in cassava by affecting their phenotypic expression in the plant. Keywords: Hydrothermal stress, bio-chemicals, pigments, secondary metabolites, cassava. African Journal of Biotechnology, Vol 13(31) 3173-3186 ...

  18. Radiorespirometic assay device

    International Nuclear Information System (INIS)

    Levin, G.V.; Straat, P.A.

    1981-01-01

    A radiorespirometic assay device is described in which the presence of microorganisms in a sample is determined by placing the sample in contact with a metabolisable radioactive labelled substrate, collecting any gas evolved, exposing a photosensitive material to the gas and determining if a spot is produced on the material. A spot indicates the presence of radioactivity showing that the substrate has been metabolized by a microorganism. Bacteria may be detected in body fluids, hospital operating rooms, water, food, cosmetics and drugs. (U.K.)

  19. Biochemical investigation of cypermethrin toxicity in rabbits.

    Science.gov (United States)

    Dahamna, S; Harzallah, D; Guemache, A; Sekfali, N

    2009-01-01

    Pyrethroids are the most frequently used pesticides in agriculture, forestry, horticulture, hospitals public health, homes and textile industry. Cypermethrin, a composite pyrethroid is moderately toxic to mammals. Exposure to the pyrethroids occurs by inhalation, dermal and oral routes both accidentally as well as from the environment. Cypermethrin and DDT have been detected in human breast milk from malaria endemic area in South Africa. The WHO has recommended that the level of permethrin in drinking water not exceed 20 micrograms per liter (microg/L). The effects of exposure to any hazardous substance depend on the dose, the duration, how you are exposed, personal traits, habits and whether other chemicals are present. Pyrethroids are often combined commercially with other chemicals called synergists, which enhance the insecticidal activity of the pyrethrins and pyrethroids. The synergists prevent some enzymes from breaking down the pyrethrins and pyrethroids, thus increasing their toxicity. Because these compounds are broken down in the body quickly, there are several ways to measure the metabolites of these chemicals in human blood and urine. In this study the pyrethroid cypermethrin Sherpa 25% (active substance 250 g/l cypermethrin) was used, rabbits (1 kg weight), were gavaged by 1/20 LD50 for 3 weeks (one dose every week). Blood was collected before dosing and after 24, 72, 144 hours after the treatment. Enzyme activities were assayed in the plasma samples obtained. GOT, GPT, ALPH, CREA, GGT, Glucose and Total Pro were measured. Rabbits showed depression, decrease in feed intake, body weight and loose faeces. Livers exhibited fatty change, necrosis, lesions in kidney included tubular necrosis and pink homogeneous tubular casts. Serum ALT and creatinine concentrations increased while those of total proteins, albumin, serum cholesterol and triglycerides decreased.The results showed a decrease in RBC; WBC and Hb. This probably explained by the effect of

  20. 3-Hydroxypyrimidine-2,4-diones as Selective Active Site Inhibitors of HIV Reverse Transcriptase-Associated RNase H: Design, Synthesis, and Biochemical Evaluations.

    Science.gov (United States)

    Tang, Jing; Liu, Feng; Nagy, Eva; Miller, Lena; Kirby, Karen A; Wilson, Daniel J; Wu, Bulan; Sarafianos, Stefan G; Parniak, Michael A; Wang, Zhengqiang

    2016-03-24

    Human immunodeficiency virus (HIV) reverse transcriptase (RT) associated ribonuclease H (RNase H) remains an unvalidated antiviral target. A major challenge of specifically targeting HIV RNase H arises from the general lack of selectivity over RT polymerase (pol) and integrase (IN) strand transfer (ST) inhibitions. We report herein the synthesis and biochemical evaluations of three novel 3-hydroxypyrimidine-2,4-dione (HPD) subtypes carefully designed to achieve selective RNase H inhibition. Biochemical studies showed the two subtypes with an N-1 methyl group (9 and 10) inhibited RNase H in low micromolar range without significantly inhibiting RT polymerase, whereas the N-1 unsubstituted subtype 11 inhibited RNase H in submicromolar range and RT polymerase in low micromolar range. Subtype 11 also exhibited substantially reduced inhibition in the HIV-1 INST assay and no significant cytotoxicity in the cell viability assay, suggesting that it may be amenable to further structure-activity relationship (SAR) for identifying RNase H inhibitors with antiviral activity.

  1. Glucuronoyl Esterase Screening and Characterization Assays Utilizing Commercially Available Benzyl Glucuronic Acid Ester

    Directory of Open Access Journals (Sweden)

    Hampus Sunner

    2015-09-01

    Full Text Available Research on glucuronoyl esterases (GEs has been hampered by the lack of enzyme assays based on easily obtainable substrates. While benzyl d-glucuronic acid ester (BnGlcA is a commercially available substrate that can be used for GE assays, several considerations regarding substrate instability, limited solubility and low apparent affinities should be made. In this work we discuss the factors that are important when using BnGlcA for assaying GE activity and show how these can be applied when designing BnGlcA-based GE assays for different applications: a thin-layer chromatography assay for qualitative activity detection, a coupled-enzyme spectrophotometric assay that can be used for high-throughput screening or general activity determinations and a HPLC-based detection method allowing kinetic determinations. The three-level experimental procedure not merely facilitates routine, fast and simple biochemical characterizations but it can also give rise to the discovery of different GEs through an extensive screening of heterologous Genomic and Metagenomic expression libraries.

  2. Diagnosis Of Inherited Neurometabolic Disorders : A Biochemical Approach

    Directory of Open Access Journals (Sweden)

    Christopher R

    1999-01-01

    Full Text Available The past two decades have witnessed a rapid increase in the knowledge of the inherited neurometabolic disorders. The precise diagnosis of these disorders which is a challenge to the physician can be best accomplished by biochemical methods. Screening of clinically selected patients with simple chemical urine tests and routine blood chemistry investigations followed by measurement of specific metabolites and assay of the relevant enzymes confirms the diagnosis in most cases. Biochemical diagnosis of inherited neurometabolic disorders although expensive is rapid and confirmatory and therefore aids in treatment and further prevention of these rare disorders.

  3. Show-Bix &

    DEFF Research Database (Denmark)

    2014-01-01

    The anti-reenactment 'Show-Bix &' consists of 5 dias projectors, a dial phone, quintophonic sound, and interactive elements. A responsive interface will enable the Dias projectors to show copies of original dias slides from the Show-Bix piece ”March på Stedet”, 265 images in total. The copies are...

  4. A fluorimetric assay for cortisol.

    Science.gov (United States)

    Appel, Daniel; Schmid, Rolf D; Dragan, Calin-Aurel; Bureik, Matthias; Urlacher, Vlada B

    2005-09-01

    A simple, rapid and sensitive fluorimetric assay for the quantitative determination of cortisol is reported. The assay is based on the formation of a fluorescent dye when cortisol is incubated with a mixture of sulfuric acid and acetic acid. The fluorescence spectrum recorded for the resulting dye shows a maximum extinction at 475 nm and a maximum emission at 525 nm. The solvent 2-methyl-4-pentanone was used for extraction and was found to act as a fluorescence amplifier. A limit of detection of 2.7 muM was achieved, making it possible to forego solvent evaporation. The assay suffers minor interference from 11-deoxycortisol which exhibits low fluorescence at lambda (ex): 460 nm; lambda (em): 505 nm. Typical standard deviations were below 4%. We validated the assay using a biotransformation with recombinant Schizosaccharomyces pombe which regioselectively hydroxylates 11-deoxycortisol to cortisol. The method described herein is suitable for preliminary screening of microorganisms capable of steroid hydroxylation.

  5. Cross-resistance of bisultap resistant strain of Nilaparvata lugens and its biochemical mechanism.

    Science.gov (United States)

    Ling, Shanfeng; Zhang, Runjie

    2011-02-01

    The resistant (R) strain of the planthopper Nilaparvata lugens (Stål) selected for bisultap resistance displayed 7.7-fold resistance to bisultap and also had cross-resistance to nereistoxin (monosultap, thiocyclam, and cartap), chlorpyrifos, dimethoate, and malathion but no cross-resistance to buprofezin, imidacloprid, and fipronil. To find out the biochemical mechanism of resistance to bisultap, biochemical assay was done. The results showed that cytochrome P450 monooxygenases (P450) activity in R strain was 2.71-fold that in susceptible strain (S strain), in which the changed activity for general esterase (EST) was 1.91 and for glutathione S-transferases only 1.32. Piperonyl butoxide (PBO) could significantly inhibit P450 activity (percentage of inhibition [PI]: 37.31%) in the R strain, with ESTs PI = 16.04% by triphenyl phosphate (TPP). The results also demonstrated that diethyl maleate had no synergism with bisultap. However, PBO displayed significant synergism in three different strains, and the synergism increased with resistance (S strain 1.42, Lab strain, 2.24 and R strain, 3.23). TPP also showed synergism for three strains, especially in R strain (synergistic ratio = 2.47). An in vitro biochemical study and in vivo synergistic study indicated that P450 might be play important role in the biochemical mechanism of bisultap resistance and that esterase might be the important factor of bisultap resistance. Acetylcholinesterase (AChE) insensitivity play important role in bisultap resistance. We suggest that buprofezin, imidacloprid, and fipronil could be used in resistance management programs for N. lugens via alternation and rotation with bisultap.

  6. A peptide-binding assay for the disease-associated HLA-DQ8 molecule

    DEFF Research Database (Denmark)

    Straumfors, A; Johansen, B H; Vartdal, F

    1998-01-01

    The study of peptide binding to HLA class II molecules has mostly concentrated on DR molecules. Since many autoimmune diseases show a primary association to particular DQ molecules rather than DR molecules, it is also important to study the peptide-binding properties of DQ molecules. Here we report...... a biochemical peptide-binding assay for the type I diabetes-associated DQ8, i.e. DQ (alpha1*0301, beta1*0302), molecule. Affinity-purified DQ8 molecules were tested in peptide-binding assays using a radiolabelled influenza haemagglutinin (Ha) peptide encompassing positions 255-271(Y) as an indicator peptide...... of 43 peptides of different lengths and sequences. The DQ8 molecules showed a different pattern of peptide binding compared to a previously studied DQ2 molecule. Peptides derived from thyroid peroxidase, HLA-DQ(alpha1*0301), HLA-DQ(alpha1*0302), retinol receptor and p21ras were among the high...

  7. Biochemical constituents of seaweeds along the Maharashtra coast

    Digital Repository Service at National Institute of Oceanography (India)

    Dhargalkar, V.K.; Jagtap, T.G.; Untawale, A.G.

    Protein, carbohydrate and organic carbon were estimated in 43 marine algal species from different stations along the Maharashtra Coast in India These species showed variation in their biochemical contents Protein varied from 10 to 33% Chlorophyceae...

  8. Talk Show Science.

    Science.gov (United States)

    Moore, Mitzi Ruth

    1992-01-01

    Proposes having students perform skits in which they play the roles of the science concepts they are trying to understand. Provides the dialog for a skit in which hot and cold gas molecules are interviewed on a talk show to study how these properties affect wind, rain, and other weather phenomena. (MDH)

  9. Biochemical principle of Limulus test for detecting bacterial endotoxins

    OpenAIRE

    Iwanaga, Sadaaki

    2007-01-01

    A hemocyte lysate from horseshoe crab (Limulus) produced a gel, when exposed to Gram-negative bacterial endotoxins, lipopolysaccharides (LPS). This gelation reaction of the lysate, so-called Limulus test, has been widely employed as a simple and very sensitive assay method for endotoxins. Recent biochemical studies on the principle of Limulus test indicate that the hemocytes contain several serine protease zymogens, which constitute a coagulation cascade triggered by endotoxins, and that ther...

  10. 40 CFR 158.2083 - Experimental use permit biochemical pesticides human health assessment data requirements table.

    Science.gov (United States)

    2010-07-01

    ... Biochemical Pesticides § 158.2083 Experimental use permit biochemical pesticides human health assessment data... following table: (d) Table. The following table shows the data requirements for experimental use permit... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Experimental use permit biochemical...

  11. Physics Reality Show

    Science.gov (United States)

    Erukhimova, Tatiana

    The attention span of K-12 students is very short; they are used to digesting information in short snippets through social media and TV. To get the students interested in physics, we created the Physics Reality Show: a series of staged short videos with duration no longer than a few minutes. Each video explains and illustrates one physics concept or law through a fast-paced sequence of physics demonstrations and experiments. The cast consists entirely of physics undergraduate students with artistic abilities and substantial experience in showing physics demonstrations at current outreach events run by the department: Physics Shows and Physics & Engineering Festival. Undergraduate students are of almost the same age as their high-school audience. They are in the best position to connect with kids and convey their fascination with physics. The PI and other faculty members who are involved in the outreach advise and coach the cast. They help students in staging the episodes and choosing the most exciting and relevant demonstrations. Supported by the APS mini-outreach Grant.

  12. Kombucha tea fermentation: Microbial and biochemical dynamics.

    Science.gov (United States)

    Chakravorty, Somnath; Bhattacharya, Semantee; Chatzinotas, Antonis; Chakraborty, Writachit; Bhattacharya, Debanjana; Gachhui, Ratan

    2016-03-02

    Kombucha tea, a non-alcoholic beverage, is acquiring significant interest due to its claimed beneficial properties. The microbial community of Kombucha tea consists of bacteria and yeast which thrive in two mutually non-exclusive compartments: the soup or the beverage and the biofilm floating on it. The microbial community and the biochemical properties of the beverage have so far mostly been described in separate studies. This, however, may prevent understanding the causal links between the microbial communities and the beneficial properties of Kombucha tea. Moreover, an extensive study into the microbial and biochemical dynamics has also been missing. In this study, we thus explored the structure and dynamics of the microbial community along with the biochemical properties of Kombucha tea at different time points up to 21 days of fermentation. We hypothesized that several biochemical properties will change during the course of fermentation along with the shifts in the yeast and bacterial communities. The yeast community of the biofilm did not show much variation over time and was dominated by Candida sp. (73.5-83%). The soup however, showed a significant shift in dominance from Candida sp. to Lachancea sp. on the 7th day of fermentation. This is the first report showing Candida as the most dominating yeast genus during Kombucha fermentation. Komagateibacter was identified as the single largest bacterial genus present in both the biofilm and the soup (~50%). The bacterial diversity was higher in the soup than in the biofilm with a peak on the seventh day of fermentation. The biochemical properties changed with the progression of the fermentation, i.e., beneficial properties of the beverage such as the radical scavenging ability increased significantly with a maximum increase at day 7. We further observed a significantly higher D-saccharic acid-1,4-lactone content and caffeine degradation property compared to previously described Kombucha tea fermentations. Our

  13. Development of multiplex-PCR assay for rapid detection of Candida spp.

    Directory of Open Access Journals (Sweden)

    Ni Made A. Tarini

    2010-05-01

    Full Text Available Aim Candida spp. infection commonly occur in immunocompromised patients. Biochemical assay for identification of Candida spp. is time-consuming and shows many undetermined results. Specific detection for antibody, antigen and metabolites of Candida spp. had low sensitivity and specificity. In this study, we developed a rapid diagnostic method, Multiplex-PCR, to identify Candida spp.Methods Five Candida spp. isolates were cultured, identifi ed with germ tube and API® 20 C AUX (BioMerieux® SA kit. Furthermore, DNA was purified by QIAamp DNA mini (Qiagen® kit for Multiplex-PCR assay.Results DNA detection limit by Multiplex-PCR assays for C. albicans, C. tropicalis, C. parapsilosis, C. krusei and C. glabrata were 4 pg, 0.98 pg, 0.98 pg, 0.5 pg and 16 pg respectively. This assay was also more sensitive than culture in that Multiplex-PCR could detect 2.6-2.9 x 100 CFU/ml, whereas culture 2.6-2.9 x 102 CFU/ml.Conclusion Multiplex-PCR is much more sensitive than culture and thus, can be recommended as a sensitive and specific assay for identification of Candida spp. (Med J Indones 2010; 19:83-7Keywords: Candida spp., multiplex-PCR

  14. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas

    2013-01-21

    We report a simple and rapid room temperature assay for point-of-care (POC) testing that is based on specific agglutination. Agglutination tests are based on aggregation of microbeads in the presence of a specific analyte thus enabling the macroscopic observation. Such tests are most often used to explore antibody-antigen reactions. Agglutination has been used for protein assays using a biotin/streptavidin system as well as a hybridization based assay. The agglutination systems are prone to selftermination of the linking analyte, prone to active site saturation and loss of agglomeration at high analyte concentrations. We investigated the molecular target/ligand interaction, explaining the common agglutination problems related to analyte self-termination, linkage of the analyte to the same bead instead of different microbeads. We classified the agglutination process into three kinds of assays: a two- component assay, a three-component assay and a stepped three- component assay. Although we compared these three kinds of assays for recognizing DNA and protein molecules, the assay can be used for virtually any molecule, including ions and metabolites. In total, the optimized assay permits detecting analytes with high sensitivity in a short time, 5 min, at room temperature. Such a system is appropriate for POC testing.

  15. New enzymatic assay, parasite lactate dehydrogenase in diagnosis ...

    African Journals Online (AJOL)

    Background: The unique ability of plasmodial lactate dehydrogenase p(LDH) to utilise 3-acetyl pyridine dinucleotide (APAD) in lieu of NAD as a coenzyme in the conversion of pyruvate to lactate, led to the development of a biochemical assay for the detection of plasmodial parasitaemia. Researchers have reported that ...

  16. Not a "reality" show.

    Science.gov (United States)

    Wrong, Terence; Baumgart, Erica

    2013-01-01

    The authors of the preceding articles raise legitimate questions about patient and staff rights and the unintended consequences of allowing ABC News to film inside teaching hospitals. We explain why we regard their fears as baseless and not supported by what we heard from individuals portrayed in the filming, our decade-long experience making medical documentaries, and the full un-aired context of the scenes shown in the broadcast. The authors don't and can't know what conversations we had, what documents we reviewed, and what protections we put in place in each televised scene. Finally, we hope to correct several misleading examples cited by the authors as well as their offhand mischaracterization of our program as a "reality" show.

  17. Radioreceptor opioid assay

    International Nuclear Information System (INIS)

    Miller, R.J.; Chang, K.-J.

    1981-01-01

    A radioreceptor assay is described for assaying opioid drugs in biological fluids. The method enables the assay of total opioid activity, being specific for opioids as a class but lacking specificity within the class. A radio-iodinated opioid and the liquid test sample are incubated with an opiate receptor material. The percentage inhibition of the binding of the radio-iodinated compound to the opiate receptor is calculated and the opioid activity of the test liquid determined from a standard curve. Examples of preparing radio-iodinated opioids and assaying opioid activity are given. A test kit for the assay is described. Compared to other methods, this assay is cheap, easy and rapid. (U.K.)

  18. Showing Value (Editorial

    Directory of Open Access Journals (Sweden)

    Denise Koufogiannakis

    2009-06-01

    Full Text Available When Su Cleyle and I first decided to start Evidence Based Library and Information Practice, one of the things we agreed upon immediately was that the journal be open access. We knew that a major obstacle to librarians using the research literature was that they did not have access to the research literature. Although Su and I are both academic librarians who can access a wide variety of library and information literature from our institutions, we belong to a profession where not everyone has equal access to the research in our field. Without such access to our own body of literature, how can we ever hope for practitioners to use research evidence in their decision making? It would have been contradictory to the principles of evidence based library and information practice to do otherwise.One of the specific groups we thought could use such an open access venue for discovering research literature was school librarians. School librarians are often isolated and lacking access to the research literature that may help them prove to stakeholders the importance of their libraries and their role within schools. Certainly, school libraries have been in decline and the use of evidence to show value is needed. As Ken Haycock noted in his 2003 report, The Crisis in Canada’s School Libraries: The Case for Reform and Reinvestment, “Across the country, teacher-librarians are losing their jobs or being reassigned. Collections are becoming depleted owing to budget cuts. Some principals believe that in the age of the Internet and the classroom workstation, the school library is an artifact” (9. Within this context, school librarians are looking to our research literature for evidence of the impact that school library programs have on learning outcomes and student success. They are integrating that evidence into their practice, and reflecting upon what can be improved locally. They are focusing on students and showing the impact of school libraries and

  19. Absolute nuclear material assay

    Science.gov (United States)

    Prasad, Manoj K [Pleasanton, CA; Snyderman, Neal J [Berkeley, CA; Rowland, Mark S [Alamo, CA

    2010-07-13

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  20. Absolute nuclear material assay

    Science.gov (United States)

    Prasad, Manoj K [Pleasanton, CA; Snyderman, Neal J [Berkeley, CA; Rowland, Mark S [Alamo, CA

    2012-05-15

    A method of absolute nuclear material assay of an unknown source comprising counting neutrons from the unknown source and providing an absolute nuclear material assay utilizing a model to optimally compare to the measured count distributions. In one embodiment, the step of providing an absolute nuclear material assay comprises utilizing a random sampling of analytically computed fission chain distributions to generate a continuous time-evolving sequence of event-counts by spreading the fission chain distribution in time.

  1. Tricyclic antidepressant radioreceptor assay

    International Nuclear Information System (INIS)

    Innis, R.B.; Tune, L.; Rock, R.; Depaulo, R.; U'Prichard, D.C.; Snyder, S.M.

    1979-01-01

    A receptor assay for tricyclic antidepressants described here is based on the ability of these drugs to compete with [ 3 H]-3-guinuclidnyl benzilate ( 3 H-QNB) for binding to muscarinic cholinergic receptors in rat brain membranes. The assay is sensitive, in that it can detect, for example, 2ng/ml nortriptyline in plasma. Seven plasma samples from depressed patients treated with nortriptyline were assayed with the radioreceptor and gas liquid chromatographic methods, and the results from these two methods were almost identical. This assay should be used cautiously, if at all, in patients treated with other drugs that have potent anticholinergic effects. (Auth.)

  2. Fitness effects of fluctuations in biochemical networks

    Science.gov (United States)

    Tanase-Nicola, Sorin

    2009-03-01

    The concentration of many cellular components fluctuates not only as a response to external and internal inputs but also due to random birth and death events of individual molecules. This biochemical noise affects the capacity of every individual cell in a population to respond and adapt to the environment. While the sources and effects of biochemical fluctuations on individual cells have been intensively studied, the effects of noise on the growth rate of a population of cells are much less understood. We present a model of the cell cycle in which the growth and division of individual cells are coupled with the noisy dynamics of their internal components. The model allows us to compute the contribution of the biochemical noise to the average growth rate of a population of cells as a function of the noise strength and the correlation time of the fluctuations. We show that, due to fluctuations, the growth rate of a population of cells is always larger than the average growth rate of a individual cell and can be larger even than a corresponding deterministic model. In most relevant cases it is assumed that the average concentration of a cellular component is close to a value that maximizes the population growth as given by the external, environmental, conditions and the internal cellular regulation. In such cases we show that contribution of fluctuations to the growth rate is negative and increases with the sensitivity of the biochemical networks to the noise sources and the noise correlation time. We also discuss how the selection pressure due to fluctuations affects the structure and parameters of genetic regulatory networks.

  3. Endogenous Locus Reporter Assays.

    Science.gov (United States)

    Liu, Yaping; Hermes, Jeffrey; Li, Jing; Tudor, Matthew

    2018-01-01

    Reporter gene assays are widely used in high-throughput screening (HTS) to identify compounds that modulate gene expression. Traditionally a reporter gene assay is built by cloning an endogenous promoter sequence or synthetic response elements in the regulatory region of a reporter gene to monitor transcriptional activity of a specific biological process (exogenous reporter assay). In contrast, an endogenous locus reporter has a reporter gene inserted in the endogenous gene locus that allows the reporter gene to be expressed under the control of the same regulatory elements as the endogenous gene, thus more accurately reflecting the changes seen in the regulation of the actual gene. In this chapter, we introduce some of the considerations behind building a reporter gene assay for high-throughput compound screening and describe the methods we have utilized to establish 1536-well format endogenous locus reporter and exogenous reporter assays for the screening of compounds that modulate Myc pathway activity.

  4. Explorations into Chemical Reactions and Biochemical Pathways.

    Science.gov (United States)

    Gasteiger, Johann

    2016-12-01

    A brief overview of the work in the research group of the present author on extracting knowledge from chemical reaction data is presented. Methods have been developed to calculate physicochemical effects at the reaction site. It is shown that these physicochemical effects can quite favourably be used to derive equations for the calculation of data on gas phase reactions and on reactions in solution such as aqueous acidity of alcohols or carboxylic acids or the hydrolysis of amides. Furthermore, it is shown that these physicochemical effects are quite effective for assigning reactions into reaction classes that correspond to chemical knowledge. Biochemical reactions constitute a particularly interesting and challenging task for increasing our understanding of living species. The BioPath.Database is a rich source of information on biochemical reactions and has been used for a variety of applications of chemical, biological, or medicinal interests. Thus, it was shown that biochemical reactions can be assigned by the physicochemical effects into classes that correspond to the classification of enzymes by the EC numbers. Furthermore, 3D models of reaction intermediates can be used for searching for novel enzyme inhibitors. It was shown in a combined application of chemoinformatics and bioinformatics that essential pathways of diseases can be uncovered. Furthermore, a study showed that bacterial flavor-forming pathways can be discovered. © 2016 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Reelin exerts structural, biochemical and transcriptional regulation over presynaptic and postsynaptic elements in the adult hippocampus

    Directory of Open Access Journals (Sweden)

    Carles eBosch

    2016-05-01

    Full Text Available Reelin regulates neuronal positioning and synaptogenesis in the developing brain, and adult brain plasticity. Here we used transgenic mice overexpressing Reelin (Reelin-OE mice to perform a comprehensive dissection of the effects of this protein on the structural and biochemical features of dendritic spines and axon terminals in the adult hippocampus. Electron microscopy (EM revealed both higher density of synapses and structural complexity of both pre- and postsynaptic elements in transgenic mice than in WT mice. Dendritic spines had larger spine apparatuses, which correlated with a redistribution of Synaptopodin. Most of the changes observed in Reelin-OE mice were reversible after blockade of transgene expression, thus supporting the specificity of the observed phenotypes. Western blot and transcriptional analyses did not show major changes in the expression of pre- or postsynaptic proteins, including SNARE proteins, glutamate receptors, and scaffolding and signaling proteins. However, EM immunogold assays revealed that the NMDA receptor subunits NR2a and NR2b, and p-Cofilin showed a redistribution from synaptic to extrasynaptic pools. Taken together with previous studies, the present results suggest that Reelin regulates the structural and biochemical properties of adult hippocampal synapses by increasing their density and morphological complexity and by modifying the distribution and trafficking of major glutamatergic components.

  6. Solid phase assays

    International Nuclear Information System (INIS)

    Reese, M.G.; Johnson, L.R.; Ransom, D.K.

    1980-01-01

    In a solid phase assay for quantitative determination of biological and other analytes, a sample such as serum is contacted with a receptor for the analyte being assayed, the receptor being supported on a solid support. No tracer for the analyte is added to the sample before contacting with the receptor; instead the tracer is contacted with the receptor after unbound analyte has been removed from the receptor. The assay can be otherwise performed in a conventional manner but can give greater sensitivity. (author)

  7. [Pre-analytical stability before centrifugation of 7 biochemical analytes in whole blood].

    Science.gov (United States)

    Perrier-Cornet, Andreas; Moineau, Marie-Pierre; Narbonne, Valérie; Plee-Gautier, Emmanuelle; Le Saos, Fabienne; Carre, Jean-Luc

    2015-01-01

    The pre-analytical stability of 7 biochemical parameters (parathyroid hormone -PTH-, vitamins A, C E and D, 1,25-dihydroxyvitamin D and insulin) at +4 °C, was studied on whole blood samples before centrifugation. The impact of freezing at -20°C was also analyzed/performed for PTH and vitamin D. The differences in the results of assays for whole blood samples, being kept for different times between sampling time and analysis, from 9 healthy adults, were compaired by using a Student t test. The 7 analytes investigated remained stable up to 4 hours at +4°C in whole blood. This study showed that it is possible to accept uncentrifuged whole blood specimens kept at +4°C before analysis. PTH is affected by freezing whereas vitamin D is not.

  8. Biochemical Alterations Of Amino Acids, Neurotransmitters And ...

    African Journals Online (AJOL)

    Egyptian Journal of Biochemistry and Molecular Biology ... At the sametime, brain neurotransmitters, serum ALT, AST, ALP and γ-GT levels were also assayed. ... There was increasing tendency in brain concentration of norepinephrine and dopamine, while the level of brain serotonin showed a pronounced decrease after ...

  9. Autotaxin : biochemical and functional studies

    NARCIS (Netherlands)

    Houben, Anna Jacoba Sara

    2012-01-01

    This thesis focuses on autotaxin (ATX), the main enzyme responsible for the production of lysophosphatidic acid (LPA). The ATX-LPA receptor axis has a wide implication in health and disease. The studies described in this thesis aim at characterizing the biochemical and functional properties of ATX,

  10. Serum biochemical changes accompanying prolonged ...

    African Journals Online (AJOL)

    Jane

    2010-10-18

    Oct 18, 2010 ... Toxicological evaluation of the whole fruit of Lagenaria breviflora was carried out using the serum biochemical changes accompanying prolonged administration of the ethanolic extract of the fruit in ... glucose in rats administered with the extract of 8000 mg/kg body weight increased two and half-fold.

  11. Comparison of Batch Assay and Random Assay Using Automatic Dispenser in Radioimmunoassay

    Energy Technology Data Exchange (ETDEWEB)

    Moon, Seung Hwan; Jang, Su Jin; Kang, Ji Yeon; Lee, Dong Soo; Chung, June Key; Lee, Myung Chul [Seoul Metropolitan Government Seoul National University Boramae Medical Center, Seoul (Korea, Republic of); Lee, Ho Young; Shin, Sun Young; Min, Gyeong Sun; Lee, Hyun Joo [Seoul National University college of Medicine, Seoul (Korea, Republic of)

    2009-08-15

    Radioimmunoassay (RIA) was usually performed by the batch assay. To improve the efficiency of RIA without increase of the cost and time, random assay could be a choice. We investigated the possibility of the random assay using automatic dispenser by assessing the agreement between batch assay and random assay. The experiments were performed with four items; Triiodothyronine (T3), free thyroxine (fT4), Prostate specific antigen (PSA), Carcinoembryonic antigen (CEA). In each item, the sera of twenty patients, the standard, and the control samples were used. The measurements were done 4 times with 3 hour time intervals by random assay and batch assay. The coefficient of variation (CV) of the standard samples and patients' data in T3, fT4, PSA, and CEA were assessed. ICC (Intraclass correlation coefficient) and coefficient of correlation were measured to assessing the agreement between two methods. The CVs (%) of T3, fT4, PSA, and CEA measured by batch assay were 3.2+-1.7%, 3.9+-2.1%, 7.1+-6.2%, 11.2+-7.2%. The CVs by random assay were 2.1+-1.7%, 4.8+-3.1%, 3.6+-4.8%, and 7.4+-6.2%. The ICC between the batch assay and random assay were 0.9968 (T3), 0.9973 (fT4), 0.9996 (PSA), and 0.9901 (CEA). The coefficient of correlation between the batch assay and random assay were 0.9924(T3), 0.9974 (fT4), 0.9994 (PSA), and 0.9989 (CEA) (p<0.05). The results of random assay showed strong agreement with the batch assay in a day. These results suggest that random assay using automatic dispenser could be used in radioimmunoassay

  12. A Multichannel Calorimetric Simultaneous Assay Platform Using a Microampere Constant-Current Looped Enthalpy Sensor Array.

    Science.gov (United States)

    Wei, Hsien-Chin; Huang, Su-Hua; Jiang, Joe-Air; Lee, Yeun-Chung

    2017-02-04

    Calorimetric biochemical measurements offer various advantages such as low waste, low cost, low sample consumption, short operating time, and labor-savings. Multichannel calorimeters can enhance the possibility of performing higher-throughput biochemical measurements. An enthalpy sensor (ES) array is a key device in multichannel calorimeters. Most ES arrays use Wheatstone bridge amplifiers to condition the sensor signals, but such an approach is only suitable for null detection and low resistance sensors. To overcome these limitations, we have developed a multichannel calorimetric simultaneous assay (MCSA) platform. An adjustable microampere constant-current (AMCC) source was designed for exciting the ES array using a microampere current loop measurement circuit topology. The MCSA platform comprises a measurement unit, which contains a multichannel calorimeter and an automatic simultaneous injector, and a signal processing unit, which contains multiple ES signal conditioners and a data processor. This study focused on the construction of the MCSA platform; in particular, construction of the measurement circuit and calorimeter array in a single block. The performance of the platform, including current stability, temperature sensitivity and heat sensitivity, was evaluated. The sensor response time and calorimeter constants were given. The capability of the platform to detect relative enzyme activity was also demonstrated. The experimental results show that the proposed MCSA is a flexible and powerful biochemical measurement device with higher throughput than existing alternatives.

  13. A Multichannel Calorimetric Simultaneous Assay Platform Using a Microampere Constant-Current Looped Enthalpy Sensor Array

    Directory of Open Access Journals (Sweden)

    Hsien-Chin Wei

    2017-02-01

    Full Text Available Calorimetric biochemical measurements offer various advantages such as low waste, low cost, low sample consumption, short operating time, and labor-savings. Multichannel calorimeters can enhance the possibility of performing higher-throughput biochemical measurements. An enthalpy sensor (ES array is a key device in multichannel calorimeters. Most ES arrays use Wheatstone bridge amplifiers to condition the sensor signals, but such an approach is only suitable for null detection and low resistance sensors. To overcome these limitations, we have developed a multichannel calorimetric simultaneous assay (MCSA platform. An adjustable microampere constant-current (AMCC source was designed for exciting the ES array using a microampere current loop measurement circuit topology. The MCSA platform comprises a measurement unit, which contains a multichannel calorimeter and an automatic simultaneous injector, and a signal processing unit, which contains multiple ES signal conditioners and a data processor. This study focused on the construction of the MCSA platform; in particular, construction of the measurement circuit and calorimeter array in a single block. The performance of the platform, including current stability, temperature sensitivity and heat sensitivity, was evaluated. The sensor response time and calorimeter constants were given. The capability of the platform to detect relative enzyme activity was also demonstrated. The experimental results show that the proposed MCSA is a flexible and powerful biochemical measurement device with higher throughput than existing alternatives.

  14. Lateral flow assays

    Science.gov (United States)

    Koczula, Katarzyna M.

    2016-01-01

    Lateral flow assays (LFAs) are the technology behind low-cost, simple, rapid and portable detection devices popular in biomedicine, agriculture, food and environmental sciences. This review presents an overview of the principle of the method and the critical components of the assay, focusing on lateral flow immunoassays. This type of assay has recently attracted considerable interest because of its potential to provide instantaneous diagnosis directly to patients. The range and interpretation of results and parameters used for evaluation of the assay will also be discussed. The main advantages and disadvantages of LFAs will be summarized and relevant future improvements to testing devices and strategies will be proposed. Finally, the major recent advances and future diagnostic applications in the LFA field will be explored. PMID:27365041

  15. The Antioxidant Activity and the Effects of Convolvulus Aucheri (Convolvulaceae Extract on Biochemical Indices in Rats

    Directory of Open Access Journals (Sweden)

    R. MAMMADOV

    2014-06-01

    Full Text Available Convolvulus L., the second largest genus of the family Convolvulaceae, has about 250 species distributed mainly in the temperate and tropical regions of the world, with a cosmopolitan distribution. According to recent studies, this genus is represented in Turkey by 33 species, 9 of which are endemic. Convolvulus species are extensively used in traditional medicine for various purposes as in ulcer treatment, diabetes, and tension. The aim of this study was to investigate the antioxidant activity and the effects of Convolvulus aucheri extract on biochemical indices in rats.The antioxidant activities of various solvent extracts (methanol, ethanol, acetone and benzene obtained from C. aucheri were evaluated by using 2,2-diphenyl-1-picrylhydrazyl (DPPH and β-carotene-linoleic acid assays. In addition, total phenolic contents in all the extracts of C. aucheri were determined as gallic acid equivalents. As for the biochemical assay, the extracts of the plant at the concentrations of 0.5 and 1 ml/100 g body weight/day were administered orally to the experimental groups for 36 days. Blood samples were taken by cardiac venipuncture on the 2nd and 4th weeks after the initial treatment. Aspartate aminotransferase (AST, alanine aminotransferase (ALT, gamma-glutamyltransferase (GGT and blood urea nitrogen (BUN were measured for the determination of liver function.Among all the extracts, the ethanolic extracts of C. aucheri showed the highest antioxidant activity (66.88 ± 0.8%. The highest free radical scavenging activity (59.50 ± 1.2% was recorded on the ethanolic extracts. The phenolic contents of the ethanolic extracts are higher than the other types of extracts (23.03 mg/g GAE. In biochemical assay, it was found a significant increase in the levels of serum ALT, AST and decrease the serum GGT levels in the experimental groups when compared to the controls (p<0.05. On the other hand, we found significant increase in the level of BUN.

  16. [Chronic fatigue syndrome: biochemical examination of blood].

    Science.gov (United States)

    Hakariya, Yukiko; Kuratsune, Hirohiko

    2007-06-01

    Though patients with chronic fatigue syndrome (CFS) have lots of complaints, abnormal findings cannot be detected by biochemical screening tests. However, some specialized blood tests have revealed neuroendocrine immune axis abnormalities, which is closely associated with each other. Recent studies indicate that CFS can be understood as a special condition based on abnormality of the psycho-neuro-endocrino-immunological system, with the distinguishing feature of CFS seeming to be the secondary brain dysfunction caused by several cytokines and/or autoantibodies. In this paper, we summarize these abnormalities found in CFS and show the neuro-molecular mechanism leading to chronic fatigue.

  17. Biochemical characterization and antioxidant and antiproliferative activities of different Ganoderma collections.

    Science.gov (United States)

    Saltarelli, Roberta; Ceccaroli, Paola; Buffalini, Michele; Vallorani, Luciana; Casadei, Lucia; Zambonelli, Alessandra; Iotti, Mirco; Badalyan, Susanna; Stocchi, Vilberto

    2015-01-01

    The aim of this study was to conduct a molecular and biochemical characterization and to compare the antioxidant and antiproliferative activities of four Ganoderma isolates belonging to Ganoderma lucidum (Gl-4, Gl-5) and Ganoderma resinaceum (F-1, F-2) species. The molecular identification was performed by ITS and IGS sequence analyses and the biochemical characterization by enzymatic and proteomic approaches. The antioxidant activity of the ethanolic extracts was compared by three different methods and their flavonoid contents were also analyzed by high-performance liquid chromatography. The antiproliferative effect on U937 cells was determined by MTT assay. The studied mycelia differ both in the enzymatic activities and protein content. The highest content in total phenol and the highest antioxidant activity for DPPH free radical scavenging and chelating activity on Fe(2+) were observed with the Gl-4 isolate of G. lucidum. The presence of quercetin, rutin, myricetin, and morin as major flavonoids with effective antioxidant activity was detected. The ethanolic extracts from mycelia of G. lucidum isolates possess a substantial antiproliferative activity against U937 cells in contrast to G. resinaceum in which the antiproliferative effects were insignificant. This study provides a comparison between G. lucidum and G. resinaceum mycelial strains, and shows that G. resinaceum could be utilized to obtain several bioactive compounds. © 2015 S. Karger AG, Basel.

  18. THE SIDE-EFFECT OF ORGANIC INSECTICIDE SPINOSAD ON BIOCHEMICAL AND MICROBIOLOGICAL PROPERTIES OF CLAY SOIL

    Directory of Open Access Journals (Sweden)

    Arkadiusz Telesiński

    2015-09-01

    Full Text Available The aim of the study was to determine the effect of spinosad on soil biochemical and microbiological properties. The experiment was carried out on sandy loam with Corg content 10.91 g·kg-l. Spinosad, as Spintor 240 SC was added into soil in dosages: a recommended field dosage, and fivefold, tenfold, and twenty-fivefold higher dosages. The amount of spinosad introduced into soil was between 12.55 and 313.75 g·kg-l. Moreover, soil samples without spinosad supplement were prepared as a reference. Respective Spintor 240 SC doses were converted into 1 kg soil, taking into account 10 cm depth. After application of insecticide water emulsions, soil moisture was brought to 60% maximum holding water capacity. The soil was thoroughly mixed and stored in tightly-closed polyethylene bags at 20 °C for a period 4 weeks. During the experiment dissipation of spinosad, soil enzymes (dehydrogenase, alkaline phosphatase, acid phosphatase, urease and number of bacteria, fungi, actinomycetes were assayed. Obtained results showed, that dissipation of spinosad in soil was relatively fast – the DT50 of this insecticide was ranged between 1.11 and 2.21 days. Spinosad residues had different effects on soil microbiological and biochemical properties. However, over time the impact of this insecticide definitely decreased. This indicated that the use of spinosad in organic farming, particularly in the field dosage, does not pose a long-term threat to the soil environment.

  19. Structural and biochemical analysis of atypically low dephosphorylating activity of human dual-specificity phosphatase 28.

    Directory of Open Access Journals (Sweden)

    Bonsu Ku

    Full Text Available Dual-specificity phosphatases (DUSPs constitute a subfamily of protein tyrosine phosphatases, and are intimately involved in the regulation of diverse parameters of cellular signaling and essential biological processes. DUSP28 is one of the DUSP subfamily members that is known to be implicated in the progression of hepatocellular and pancreatic cancers, and its biological functions and enzymatic characteristics are mostly unknown. Herein, we present the crystal structure of human DUSP28 determined to 2.1 Å resolution. DUSP28 adopts a typical DUSP fold, which is composed of a central β-sheet covered by α-helices on both sides and contains a well-ordered activation loop, as do other enzymatically active DUSP proteins. The catalytic pocket of DUSP28, however, appears hardly accessible to a substrate because of the presence of nonconserved bulky residues in the protein tyrosine phosphatase signature motif. Accordingly, DUSP28 showed an atypically low phosphatase activity in the biochemical assay, which was remarkably improved by mutations of two nonconserved residues in the activation loop. Overall, this work reports the structural and biochemical basis for understanding a putative oncological therapeutic target, DUSP28, and also provides a unique mechanism for the regulation of enzymatic activity in the DUSP subfamily proteins.

  20. Assessing sediment contamination using six toxicity assays

    Directory of Open Access Journals (Sweden)

    Allen G. BURTON Jr.

    2001-08-01

    Full Text Available An evaluation of sediment toxicity at Lake Orta, Italy was conducted to compare a toxicity test battery of 6 assays and to evaluate the extent of sediment contamination at various sediment depths. Lake Orta received excessive loadings of copper and ammonia during the 1900’s until a large remediation effort was conducted in 1989-90 using lime addition. Since that time, the lake has shown signs of a steady recovery of biological communities. The study results showed acute toxicity still exists in sediments at a depth of 5 cm and greater. Assays that detected the highest levels of toxicity were two whole sediment exposures (7 d using Hyalella azteca and Ceriodaphnia dubia. The MicrotoxR assay using pore water was the third most sensitive assay. The Thamnotox, Rototox, Microtox solid phase, and Seed Germination-Root Elongation (pore and solid phase assays showed occasional to no toxicity. Based on similarity of responses and assay sensitivity, the two most useful assays were the C. dubia (or H. azteca and Microtox pore water. These assays were effective at describing sediment toxicity in a weight-of-evidence approach.

  1. Assay method and compositions

    International Nuclear Information System (INIS)

    1977-01-01

    Methods are described for measuring catecholamine levels in human and animal body fluids and tissues using the catechol-O-methyl-transferase (COMT) radioassay. The assay involves incubating the biological sample with COMT and the tritiated methyl donor, S-adenosyl-L-methionine( 3 H)-methyl. The O-methylated ( 3 H) epinephrine and/or norepinephrine are extracted and oxidised to vanillin- 3 H which in turn is extracted and its radioactivity counted. When analysing dopamine levels the assay is extended by vanillin- 3 H and raising the pH of the aqueous periodate phase from which O-methylated ( 3 H) dopamine is extracted and counted. The assay may be modified depending on whether measurements of undifferentiated total endogenous catecholamine levels or differential analyses of the catecholamine levels are being performed. The sensitivity of the assay can be as low as 5 picograms for norepinephrine and epinephrine and 12 picograms for dopamine. The assemblance of the essential components of the assay into a kit for use in laboratories is also described. (U.K.)

  2. Extracellular α-Galactosidase from Trichoderma sp. (WF-3: Optimization of Enzyme Production and Biochemical Characterization

    Directory of Open Access Journals (Sweden)

    Aishwarya Singh Chauhan

    2015-01-01

    Full Text Available Trichoderma spp. have been reported earlier for their excellent capacity of secreting extracellular α-galactosidase. This communication focuses on the optimization of culture conditions for optimal production of enzyme and its characterization. The evaluation of the effects of different enzyme assay parameters such as stability, pH, temperature, substrate concentrations, and incubation time on enzyme activity has been made. The most suitable buffer for enzyme assay was found to be citrate phosphate buffer (50 mM, pH 6.0 for optimal enzyme activity. This enzyme was fairly stable at higher temperature as it exhibited 72% activity at 60°C. The enzyme when incubated at room temperature up to two hours did not show any significant loss in activity. It followed Michaelis-Menten curve and showed direct relationship with varying substrate concentrations. Higher substrate concentration was not inhibitory to enzyme activity. The apparent Michaelis-Menten constant (Km, maximum rate of reaction (Vmax, Kcat, and catalytic efficiency values for this enzyme were calculated from the Lineweaver-Burk double reciprocal plot and were found to be 0.5 mM, 10 mM/s, 1.30 U mg−1, and 2.33 U mg−1 mM−1, respectively. This information would be helpful in understanding the biophysical and biochemical characteristics of extracellular α-galactosidase from other microbial sources.

  3. Biochemical Characterisation of Phage Pseudomurein Endoisopeptidases PeiW and PeiP Using Synthetic Peptides

    Science.gov (United States)

    Schofield, Linley R.; Beattie, Amy K.; Tootill, Catherine M.; Dey, Debjit; Ronimus, Ron S.

    2015-01-01

    Pseudomurein endoisopeptidases cause lysis of the cell walls of methanogens by cleaving the isopeptide bond Ala-ε-Lys in the peptide chain of pseudomurein. PeiW and PeiP are two thermostable pseudomurein endoisopeptidases encoded by phage ΨM100 of Methanothermobacter wolfei and phages ΨM1 and ΨM2 of Methanothermobacter marburgensis, respectively. A continuous assay using synthetic peptide substrates was developed and used in the biochemical characterisation of recombinant PeiW and PeiP. The advantages of these synthetic peptide substrates over natural substrates are sensitivity, high purity, and characterisation and the fact that they are more easily obtained than natural substrates. In the presence of a reducing agent, purified PeiW and PeiP each showed similar activity under aerobic and anaerobic conditions. Both enzymes required a divalent metal for activity and showed greater thermostability in the presence of Ca2+. PeiW and PeiP involve a cysteine residue in catalysis and have a monomeric native conformation. The kinetic parameters, K M and k cat, were determined, and the ε-isopeptide bond between alanine and lysine was confirmed as the bond lysed by these enzymes in pseudomurein. The new assay may have wider applications for the general study of peptidases and the identification of specific methanogens susceptible to lysis by specific pseudomurein endoisopeptidases. PMID:26483615

  4. Radioreceptor assay for insulin

    International Nuclear Information System (INIS)

    Suzuki, Kazuo

    1975-01-01

    Radioreceptor assay of insulin was discussed from the aspects of the measuring method, its merits and problems to be solved, and its clinical application. Rat liver 10 x g pellet was used as receptor site, and enzymatic degradation of insulin by the system contained in this fraction was inhibited by adding 1 mM p-CMB. 125 I-labelled porcine insulin was made by lactoperoxidase method under overnight incubation at 4 0 C and later purification by Sephadex G-25 column and Whatman CF-11 cellulose powder. Dog pancreatic vein serum insulin during and after the glucose load was determined by radioreceptor assay and radioimmunoassay resulting that both measurements accorded considerably. Radioreceptor assay would clarify the pathology of disorders of glucose metabolism including diabetes. (Tsukamoto, Y.)

  5. Rover waste assay system

    International Nuclear Information System (INIS)

    Akers, D.W.; Stoots, C.M.; Kraft, N.C.; Marts, D.J.

    1997-01-01

    The Rover Waste Assay System (RWAS) is a nondestructive assay system designed for the rapid assay of highly-enriched 235 U contaminated piping, tank sections, and debris from the Rover nuclear rocket fuel processing facility at the Idaho Chemical Processing Plant. A scanning system translates a NaI(Tl) detector/collimator system over the structural components where both relative and calibrated measurements for 137 Cs are made. Uranium-235 concentrations are in operation and is sufficiently automated that most functions are performed by the computer system. These functions include system calibration, problem identification, collimator control, data analysis, and reporting. Calibration of the system was done through a combination of measurements on calibration standards and benchmarked modeling. A description of the system is presented along with the methods and uncertainties associated with the calibration and analysis of the system for components from the Rover facility. 4 refs., 2 figs., 4 tabs

  6. Biological and biochemical soil indicators: monitoring tools of different agricultural managements

    Science.gov (United States)

    Scotti, Riccardo; Sultana, Salma; Scelza, Rosalia; Marzaioli, Rossana; D'Ascoli, Rosaria; Rao, Maria A.

    2010-05-01

    on fertility of both soils under intensive farming. In general, all enzymatic activities and organic carbon content increased after 1 month, and they were still higher after 4 months from amendment application. Microbial biomass and soil potential activity (respiration) showed significantly higher values in soils added with organic amendments, for both farms and samplings, with more marked effects on respiration in the first sampling. In conclusion results showed, in general, a quick response as indicators of the assayed biological and biochemical soil properties and a good recovery in fertility of the studied agricultural soils. The project was founded by CCIIAA of Salerno

  7. Serum biochemical and liver enzymes changes in dogs with single ...

    African Journals Online (AJOL)

    The liver enzymes: aspartate aminotransferase (AST) and alanine aminotransferase (ALT) showed significant increase in the infected groups, while alkaline phosphatase (ALP) showed a significant decrease (P < 0.05). These biochemical changes were in all cases more profound in the conjunct infection, and could thus be ...

  8. Physiological and biochemical responses of halophyte Kalidium ...

    African Journals Online (AJOL)

    In this study, the physiological and biochemical responses of a halophyte Kalidium foliatum to salinity were studied. In order to reflect salt-tolerance in K. foliatum and to analyze the physiological and biochemical mechanism for its salt tolerance, salinity threshold and biochemical parameters were studied. A halophyte ...

  9. Radioreceptor assay for GH

    International Nuclear Information System (INIS)

    Tsushima, Toshio; Matsuzaki, Fukashi

    1975-01-01

    Radioreceptor assay (RRA) of growth hormone (GH) was studied using the protein which specifically bound to GH presenting in the liver of rabbits. 100,000g pellet of the liver homogenate was used as receptor source. The factors which affected the results of RRA such as salt, temperature and incubation time, were discussed. As same as in other RRA methods, serum protein inhibited non-specifically 125 I-GH binding in this method. In this assay, serum GH less than 5ng/ml could not be detected. The difference between the value obtained by RRA and that by radioimmunoassay was compared with reference to the patients with acromegalia. (Tsukamoto, Y.)

  10. Lateral flow strip assay

    Science.gov (United States)

    Miles, Robin R [Danville, CA; Benett, William J [Livermore, CA; Coleman, Matthew A [Oakland, CA; Pearson, Francesca S [Livermore, CA; Nasarabadi, Shanavaz L [Livermore, CA

    2011-03-08

    A lateral flow strip assay apparatus comprising a housing; a lateral flow strip in the housing, the lateral flow strip having a receiving portion; a sample collection unit; and a reagent reservoir. Saliva and/or buccal cells are collected from an individual using the sample collection unit. The sample collection unit is immersed in the reagent reservoir. The tip of the lateral flow strip is immersed in the reservoir and the reagent/sample mixture wicks up into the lateral flow strip to perform the assay.

  11. Biochemical adaptation to ocean acidification.

    Science.gov (United States)

    Stillman, Jonathon H; Paganini, Adam W

    2015-06-01

    The change in oceanic carbonate chemistry due to increased atmospheric PCO2  has caused pH to decline in marine surface waters, a phenomenon known as ocean acidification (OA). The effects of OA on organisms have been shown to be widespread among diverse taxa from a wide range of habitats. The majority of studies of organismal response to OA are in short-term exposures to future levels of PCO2 . From such studies, much information has been gathered on plastic responses organisms may make in the future that are beneficial or harmful to fitness. Relatively few studies have examined whether organisms can adapt to negative-fitness consequences of plastic responses to OA. We outline major approaches that have been used to study the adaptive potential for organisms to OA, which include comparative studies and experimental evolution. Organisms that inhabit a range of pH environments (e.g. pH gradients at volcanic CO2 seeps or in upwelling zones) have great potential for studies that identify adaptive shifts that have occurred through evolution. Comparative studies have advanced our understanding of adaptation to OA by linking whole-organism responses with cellular mechanisms. Such optimization of function provides a link between genetic variation and adaptive evolution in tuning optimal function of rate-limiting cellular processes in different pH conditions. For example, in experimental evolution studies of organisms with short generation times (e.g. phytoplankton), hundreds of generations of growth under future conditions has resulted in fixed differences in gene expression related to acid-base regulation. However, biochemical mechanisms for adaptive responses to OA have yet to be fully characterized, and are likely to be more complex than simply changes in gene expression or protein modification. Finally, we present a hypothesis regarding an unexplored area for biochemical adaptation to ocean acidification. In this hypothesis, proteins and membranes exposed to the

  12. Biochemical Markers for Assessing Aquatic Contamination

    Directory of Open Access Journals (Sweden)

    Zdeňka Svobodová

    2007-11-01

    Full Text Available Biochemical markers, specifically enzymes of the first phase of xenobiotic transformation - cytochrome P450 and ethoxyresorufin-O-deethylase (EROD - were used to determine the quantities of persistent organic pollutants (POPs in fish muscle (PCB, HCB, HCH, OCS, DDT. Eight rivers were monitored (Orlice, Chrudimka, Cidlina, Jizera, Vltava, Ohře and Bílina; and the River Blanice was used as a control. The indicator species selected was the chub (Leuciscus cephalus L.. There were no significant differences in cytochrome P450 content between the locations monitored. The highest concentration of cytochrome P450 in fish liver was in the Vltava (0.241 nmol mg-1 protein, and the lowest was in the Orlice (0.120 nmol mg-1 protein. Analysis of EROD activity showed a significant difference between the Blanice and the Vltava (P< 0.05, and also between the Orlice and the Vltava (P< 0.01, the Orlice and the Bílina (P< 0.01, and the Orlice and the Ohře (P< 0.05. The highest EROD activity in fish liver was in the Vltava (576.4 pmol min-1 mg-1 protein, and the lowest was in the Orlice (63.05 pmol min-1 mg-1 protein. In individual locations, results of chemical monitoring and values of biochemical markers were compared. A significant correlation (P< 0.05 was found between biochemical markers and OCS, and PCB. Among the tributaries studied those that contaminated the Elbe most were the Vltava and the Bílina. These tributaries should not be considered the main sources of industrial contamination of the River Elbe, because the most important contamination sources were along the river Elbe itself.

  13. Biomonitoring of genotoxic risk in radar facility workers: comparison of the comet assay with micronucleus assay and chromatid breakage assay

    International Nuclear Information System (INIS)

    Garaj-Vrhovac, V.; Kopjar, N.

    2003-01-01

    Genotoxic risks of occupational exposure in a radar facility were evaluated by using alkaline comet assay, micronucleus assay and chromatid breakage assay on peripheral blood leukocytes in exposed subjects and corresponding controls. Results show that occupational exposure to microwave radiation correlates with an increase of genome damage in somatic cells. The levels of DNA damage in exposed subjects determined by using alkaline comet assay were increased compared to control and showed interindividual variations. Incidence of micronuclei was also significantly increased compared to baseline control values. After short exposure of cultured lymphocytes to bleomycin, cells of occupationally exposed subjects responded with high numbers of chromatid breaks. Although the level of chromosome damage generated by bleomycin varied greatly between individuals, in exposed subjects a significantly elevated number of chromatid breaks was observed. Our results support data reported in literature indicating that microwave radiation represents a potential DNA-damaging hazard. Alkaline comet assay is confirmed as a sensitive and highly reproducible technique for detection of primary DNA damage inflicted in somatic cells. Micronucleus assay was confirmed as reliable bio-markers of effect and chromatid breakage assay as sensitive bio-marker of individual cancer susceptibility. The results obtained also confirm the necessity to improve measures and to perform accurate health surveillance of individuals occupationally exposed to microwave radiation

  14. Hyaluronic Acid Assays

    DEFF Research Database (Denmark)

    Itenov, Theis Skovsgaard; Kirkby, Nikolai S; Bestle, Morten H

    2016-01-01

    BACKGROUD: Hyaluronic acid (HA) is proposed as a marker of functional liver capacity. The aim of the present study was to compare a new turbidimetric assay for measuring HA with the current standard method. METHODS: HA was measured by a particle-enhanced turbidimetric immunoassay (PETIA) and enzyme...

  15. Lateral flow assays

    NARCIS (Netherlands)

    Posthuma-Trumpie, G.A.; Amerongen, van A.

    2012-01-01

    A simple version of immunochemical-based methods is the Lateral Flow Assay (LFA). It is a dry chemistry technique (reagents are included); the fluid from the sample runs through a porous membrane (often nitrocellulose) by capillary force. Typically the membrane is cut as a strip of 0.5*5 cm. In most

  16. FLUIDICS DEVICE FOR ASSAY

    DEFF Research Database (Denmark)

    2007-01-01

    The present invention relates to a device for use in performing assays on standard laboratory solid supports whereon chemical entities are attached. The invention furthermore relates to the use of such a device and a kit comprising such a device. The device according to the present invention is a...

  17. Biochemical and nutritional characterization of coconut (Cocos nucifera L.) haustorium.

    Science.gov (United States)

    Manivannan, Arivalagan; Bhardwaj, Rakesh; Padmanabhan, Sugatha; Suneja, Poonam; Hebbar, K B; Kanade, Santosh R

    2018-01-01

    Study was conducted to determine the biochemical constituents in coconut (Cocos nucifera L.) haustorium, a spongy tissue formed during coconut germination. Results indicated that 100g of dried coconut haustorium contained 1.05±0.2% ash, 44.2±4.6% soluble sugar, 24.5±3.2% starch, 5.50±0.3% protein, 1.99±0.9% fat, 5.72±0.4% soluble dietary fibre, 20.3±1.9% insoluble dietary fibre, and 146±14.3mg phenolics. Mineral profiling showed that it contained 145±8.6, 104±9.6, 33.9±8.2, 30.9±1.9, 9.45±2.1, 0.292±0.1, 2.53±0.2 and 1.20±0.1mg of K, Mg, Ca, P, Mn, Cu, Fe and Zn, respectively. Antioxidant activity assay indicated that 100g haustorium was equivalent to 1918±173, 170±20.4, 72.8±14.7 and 860±116mg of Trolox as measured by CUPRAC, FRAP, DPPH and ABTS, respectively. Amino acid score indicated that methionine+cysteine (57.6%), phenylalanine+tyrosine (32.6%), leucine (45.7%) and isoleucine (68%) are found less in haustorium. Further studies needed in developing nutritionally balanced formulations using coconut haustorium, which will be useful for lactose intolerant children. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Molecular and biochemical characterization of calmodulin from Echinococcus granulosus.

    Science.gov (United States)

    Wang, Ning; Zhong, Xiuqin; Song, Xingju; Gu, Xiaobin; Lai, Weiming; Xie, Yue; Peng, Xuerong; Yang, Guangyou

    2017-12-04

    Echinococcus granulosus is a harmful cestode parasite that causes cystic echinococcosis in humans as well as various livestock species and wild animals. Calmodulin (CaM), a Ca 2+ sensor protein, is widely expressed in eukaryotes and mediates a variety of cellular signaling activities. In the present study, the cDNA encoding CaM in Echinococcus granulosus (rEgCaM) was successfully cloned and the molecular and biochemical characterizations carried out. The antigenicity and immunoreactivity of rEgCaM was detected and the preliminary enzyme-linked immunosorbent assay (ELISA)-based serodiagnostic potential of EgCaM was assessed. The locations of this protein in the adult worm and larval stage, and the mRNA expression in different states of E. granulosus protoscoleces (PSCs) were defined clearly. Moreover, the Ca 2+ -binding properties of EgCaM were measured. rEgCaM is a highly conserved calcium-binding protein, consisting of 149 amino acids. Immunoblotting analysis revealed that rEgCaM could be identified using E. granulosus infected sheep serum. The use of rEgCaM as an antigen was evaluated by indirect ELISA which exhibited a high sensitivity (90.3%), but low specificity (47.1%). rEgCaM was ubiquitously expressed in protoscoleces and adults of E. granulosus, as well as in the germinal layer of the cyst wall. The mRNA expression level of rEgCaM was increased from the start of H 2 O 2 exposure and then gradually decreased because of the increased apoptosis of PSCs. In electrophoretic mobility tests and 1-anilinonaphthalene-8-sulfonic acid assays, rEgCaM showed a typical characteristic of a calcium-binding protein. To our knowledge, this is the first report on CaM from E. granulosus and rEgCaM is likely to be involved in some important biological function of E. granulosus as a calcium-binding protein.

  19. Rebinding in biochemical reactions on membranes

    Science.gov (United States)

    Lawley, Sean D.; Keener, James P.

    2017-10-01

    The behavior of many biochemical processes depends crucially on molecules rapidly rebinding after dissociating. In the case of multisite protein modification, the importance of rebinding has been demonstrated both experimentally and through several recent computational studies involving stochastic spatial simulations. As rebinding stems from spatio-temporal correlations, theorists have resorted to models that explicitly include space to properly account for the effects of rebinding. However, for reactions in three space dimensions it was recently shown that well-mixed ordinary differential equation (ODE) models can incorporate rebinding by adding connections to the reaction network. The rate constants for these new connections involve the probability that a pair of molecules rapidly rebinds after dissociation. In order to study biochemical reactions on membranes, in this paper we derive an explicit formula for this rebinding probability for reactions in two space dimensions. We show that ODE models can use the formula to replicate detailed stochastic spatial simulations, and that the formula can predict ultrasensitivity for reactions involving multisite modification of membrane-bound proteins. Further, we compute a new concentration-dependent rebinding probability for reactions in three space dimensions. Our analysis predicts that rebinding plays a much larger role in reactions on membranes compared to reactions in cytoplasm.

  20. Thermodynamic analysis of biochemical systems

    International Nuclear Information System (INIS)

    Yuan, Y.; Fan, L.T.; Shieh, J.H.

    1989-01-01

    Introduction of the concepts of the availability (or exergy), datum level materials, and the dead state has been regarded as some of the most significant recent developments in classical thermodynamics. Not only the available energy balance but also the material and energy balances of a biological system may be established in reference to the datum level materials in the dead state or environment. In this paper these concepts are illustrated with two examples of fermentation and are shown to be useful in identifying sources of thermodynamic inefficiency, thereby leading naturally to the rational definition of thermodynamic efficiency of a biochemical process

  1. Biochemical structure of Calendula officinalis.

    Science.gov (United States)

    Korakhashvili, A; Kacharava, T; Kiknavelidze, N

    2007-01-01

    Calendula officinalis is a well known medicinal herb. It is common knowledge that its medicinal properties are conditioned on biologically active complex substances of Carotin (Provitamin A), Stearin, Triterpiniod, Plavonoid, Kumarin, macro and micro compound elements. Because of constant need in raw material of Calendula officinalis, features of its ontogenetic development agro-biological qualities in various eco regions of Georgia were investigated. The data of biologically active compounds, biochemical structure and the maintenance both in flowers and in others parts of plant is presented; the pharmacological activity and importance in medicine was reviewed.

  2. Enzyme and biochemical producing fungi

    DEFF Research Database (Denmark)

    Lübeck, Peter Stephensen; Lübeck, Mette; Nilsson, Lena

    2010-01-01

    We are developing a biorefinery concept for biological production of chemicals, drugs, feed and fuels using plant biomass as raw material in well-defined cell-factories. Among the important goals is the discovery of new biocatalysts for production of enzymes, biochemicals and fuels and already our...... screening of a large collection of fungal strains isolated from natural habitats have resulted in identification of strains with high production of hydrolytic enzymes and excretion of organic acids. Our research focuses on creating a fungal platform based on synthetic biology for developing new cell...

  3. Reporter gene assays for investigating GPCR signaling.

    Science.gov (United States)

    Azimzadeh, Pedram; Olson, John A; Balenga, Nariman

    2017-01-01

    Luciferase-based assays are applied to evaluate various cellular processes due to their sensitivity and feasibility. The field of GPCR research has also benefited from this enzymatic reaction both in deorphanization campaigns and in delineation of the signaling pathways. Here, we describe the details of this assay in GPCR studies in 96-well format and will provide examples where the assay can show constitutive activity of an orphan GPCR and demonstrate the impact of cell type on the efficacy and potency of ligands. © 2017 Elsevier Inc. All rights reserved.

  4. Dual isotope assays

    International Nuclear Information System (INIS)

    Smith, G.F.W.; Stevens, R.A.J.; Jacoby, B.

    1980-01-01

    Dual isotope assays for thyroid function are performed by carrying out a radio-immunoassay for two of thyroxine (T4), tri-iodothyronine (T3), thyroid stimulating hormone (TSH), and thyroxine binding globulin (TBG), by a method wherein a version of one of the thyroid components, preferably T4 or T3 is labelled with Selenium-75 and the version of the other thyroid component is labelled with a different radionuclide, preferably Iodine-125. (author)

  5. Biochemical principle of Limulus test for detecting bacterial endotoxins.

    Science.gov (United States)

    Iwanaga, Sadaaki

    2007-05-01

    A hemocyte lysate from horseshoe crab (Limulus) produced a gel, when exposed to Gram-negative bacterial endotoxins, lipopolysaccharides (LPS). This gelation reaction of the lysate, so-called Limulus test, has been widely employed as a simple and very sensitive assay method for endotoxins. Recent biochemical studies on the principle of Limulus test indicate that the hemocytes contain several serine protease zymogens, which constitute a coagulation cascade triggered by endotoxins, and that there is a (1,3)-β-D-glucan-mediated coagulation pathway which also results in the formation of gel. Up to now, six protein components, designated coagulogen, proclotting enzyme, factor B, factor C, and factor G, all of which are closely associated with the endotoxin-mediated coagulation pathway, have been purified and biochemically characterized. The molecular structures of these proteins have also been elucidated. Moreover, the reconstitution experiments using the isolated clotting factors, factor C, factor B, proclotting enzyme and coagulogen in the presence of endotoxin, leads to the formation of coagulin gel. Here, I will focus on the biochemical principle of Limulus test for detecting bacterial endotoxins, and its activation and regulation mechanism on the LPS-mediated coagulation cascade.

  6. Bistability in biochemical signaling models.

    Science.gov (United States)

    Sobie, Eric A

    2011-09-20

    This Teaching Resource provides lecture notes, slides, and a student assignment for a two-part lecture on the principles underlying bistability in biochemical signaling networks, which are illustrated with examples from the literature. The lectures cover analog, or graded, versus digital, all-or-none, responses in cells, with examples from different types of biological processes requiring each. Rate-balance plots are introduced as a method for determining whether generic one-variable systems exhibit one or several stable steady states. Bifurcation diagrams are presented as a more general method for detecting the presence of bistability in biochemical signaling networks. The examples include an artificial toggle switch, the lac operon in bacteria, and the mitogen-activated protein kinase cascade in both Xenopus oocytes and mammalian cells. The second part of the lecture links the concepts of bistability more closely to the mathematical tools provided by dynamical systems analysis. The examples from the first part of the lecture are analyzed with phase-plane techniques and bifurcation analysis, using the scientific programming language MATLAB. Using these programs as a template, the assignment requires the students to implement a model from the literature and analyze the stability of this model's steady states.

  7. Biochemical abnormalities in Pearson syndrome.

    Science.gov (United States)

    Crippa, Beatrice Letizia; Leon, Eyby; Calhoun, Amy; Lowichik, Amy; Pasquali, Marzia; Longo, Nicola

    2015-03-01

    Pearson marrow-pancreas syndrome is a multisystem mitochondrial disorder characterized by bone marrow failure and pancreatic insufficiency. Children who survive the severe bone marrow dysfunction in childhood develop Kearns-Sayre syndrome later in life. Here we report on four new cases with this condition and define their biochemical abnormalities. Three out of four patients presented with failure to thrive, with most of them having normal development and head size. All patients had evidence of bone marrow involvement that spontaneously improved in three out of four patients. Unique findings in our patients were acute pancreatitis (one out of four), renal Fanconi syndrome (present in all patients, but symptomatic only in one), and an unusual organic aciduria with 3-hydroxyisobutyric aciduria in one patient. Biochemical analysis indicated low levels of plasma citrulline and arginine, despite low-normal ammonia levels. Regression analysis indicated a significant correlation between each intermediate of the urea cycle and the next, except between ornithine and citrulline. This suggested that the reaction catalyzed by ornithine transcarbamylase (that converts ornithine to citrulline) might not be very efficient in patients with Pearson syndrome. In view of low-normal ammonia levels, we hypothesize that ammonia and carbamylphosphate could be diverted from the urea cycle to the synthesis of nucleotides in patients with Pearson syndrome and possibly other mitochondrial disorders. © 2015 Wiley Periodicals, Inc.

  8. Diurnal changes of biochemical metabolic markers in healthy young males

    DEFF Research Database (Denmark)

    Sennels, Henriette P; Jørgensen, Henrik L; Fahrenkrug, Jan

    2015-01-01

    .06 mmol/L) did not show significant oscillations. CONCLUSIONS: When diagnosing and monitoring metabolic disorders compensation for the 24-h variation of the biochemical metabolic markers is needed especially C-peptide, triglyceride and glucose. Furthermore, the stable HbA1c level through 24 h makes...... it an accurate diagnostic test for diabetes mellitus....

  9. Biochemical and pathological studies in rats following dietary ...

    African Journals Online (AJOL)

    Biochemical and pathological studies in rats following dietary supplementation with high levels of polyunsaturated fatty acids and vitamin E. ... Furthermore, high dietary supplementation of vitamin E showed no deleterious effects on rats and no pathological changes in the liver, kidney and heart tissues were observed in the ...

  10. Biochemical response of ouda sheep to water contaminated with ...

    African Journals Online (AJOL)

    Also, biochemical examinations of fasting blood glucose(FBGL), total serum protein(TSP), serum albumin(S.ALB), blood urea nitrogen(BUN), serum creatinine(S.CREAT.), serum phosphate(S.PO4), aspartate ... hours of the experiment following standard procedures. The result showed that all the parameters measured were ...

  11. Haematological and Serum Bio-Chemical Parameters of West ...

    African Journals Online (AJOL)

    Haematological and Serum Bio-Chemical Parameters of West African Dwarf and Kalahari Red Goats in the Humid Tropics. ... Haematological results showed that white blood cell count, haemoglobin concentration, haematocrit, mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin ...

  12. Some biochemical properties of guaiacol peroxidases as modified ...

    African Journals Online (AJOL)

    Some biochemical properties of guaiacol peroxidases as modified by salt stress in leaves of salt-tolerant and salt-sensitive safflower ( Carthamus tinctorius L.cv.) ... The pH profile of GP activity in leaves extract of two cultivars in control and salt stressed plants showed different pattern of pH dependency with three maxima ...

  13. Germination potential index of Sindh rice cultivars on biochemical ...

    African Journals Online (AJOL)

    use

    2011-12-14

    Dec 14, 2011 ... Full Length Research Paper. Germination potential index of Sindh ... α-amylase activity is a biochemical indicator showing different germination abilities of rice varieties, leading to different seed vigor. Among ... production of this foreign commodity may be due to many constraints such as poor seed quality, ...

  14. Biochemical and functional characterization of Bothropoidin: the first haemorrhagic metalloproteinase from Bothrops pauloensis snake venom.

    Science.gov (United States)

    Gomes, Mário Sérgio R; Naves de Souza, Dayane L; Guimarães, Denise O; Lopes, Daiana S; Mamede, Carla C N; Gimenes, Sarah Natalie C; Achê, David C; Rodrigues, Renata S; Yoneyama, Kelly A G; Borges, Márcia H; de Oliveira, Fábio; Rodrigues, Veridiana M

    2015-03-01

    We present the biochemical and functional characterization of Bothropoidin, the first haemorrhagic metalloproteinase isolated from Bothrops pauloensis snake venom. This protein was purified after three chromatographic steps on cation exchange CM-Sepharose fast flow, size-exclusion column Sephacryl S-300 and anion exchange Capto Q. Bothropoidin was homogeneous by SDS-PAGE under reducing and non-reducing conditions, and comprised a single chain of 49,558 Da according to MALDI TOF analysis. The protein presented an isoelectric point of 3.76, and the sequence of six fragments obtained by MS (MALDI TOF\\TOF) showed a significant score when compared with other PIII Snake venom metalloproteinases (SVMPs). Bothropoidin showed proteolytic activity on azocasein, Aα-chain of fibrinogen, fibrin, collagen and fibronectin. The enzyme was stable at pH 6-9 and at lower temperatures when assayed on azocasein. Moreover, its activity was inhibited by EDTA, 1.10-phenanthroline and β-mercaptoethanol. Bothropoidin induced haemorrhage [minimum haemorrhagic dose (MHD) = 0.75 µg], inhibited platelet aggregation induced by collagen and ADP, and interfered with viability and cell adhesion when incubated with endothelial cells in a dose and time-dependent manner. Our results showed that Bothropoidin is a haemorrhagic metalloproteinase that can play an important role in the toxicity of B. pauloensis envenomation and might be used as a tool for studying the effects of SVMPs on haemostatic disorders and tumour metastasis. © The Authors 2014. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  15. Development of a Surface Plasmon Resonance Assay for the Characterization of Small-Molecule Binding Kinetics and Mechanism of Binding to Kynurenine 3-Monooxygenase.

    Science.gov (United States)

    Poda, Suresh B; Kobayashi, Masakazu; Nachane, Ruta; Menon, Veena; Gandhi, Adarsh S; Budac, David P; Li, Guiying; Campbell, Brian M; Tagmose, Lena

    2015-10-01

    Kynurenine 3-monooxygenase (KMO), a pivotal enzyme in the kynurenine pathway, was identified as a potential therapeutic target for treating neurodegenerative and psychiatric disorders. In this article, we describe a surface plasmon resonance (SPR) assay that delivers both kinetics and the mechanism of binding (MoB) data, enabling a detailed characterization of KMO inhibitors for the enzyme in real time. SPR assay development included optimization of the protein construct and the buffer conditions. The stability and inhibitor binding activity of the immobilized KMO were significantly improved when the experiments were performed at 10°C using a buffer containing 0.05% n-dodecyl-β-d-maltoside (DDM) as the detergent. The KD values of the known KMO inhibitors (UPF648 and RO61-8048) from the SPR assay were in good accordance with the biochemical LC/MS/MS assay. Also, the SPR assay was able to differentiate the binding kinetics (k(a) and k(d)) of the selected unknown KMO inhibitors. For example, the inhibitors that showed comparable IC50 values in the LC/MS/MS assay displayed differences in their residence time (τ = 1/k(d)) in the SPR assay. To better define the MoB of the inhibitors to KMO, an SPR-based competition assay was developed, which demonstrated that both UPF648 and RO61-8048 bound to the substrate-binding site. These results demonstrate the potential of the SPR assay for characterizing the affinity, the kinetics, and the MoB profiles of the KMO inhibitors.

  16. Development of biochemical heterogeneity of articular cartilage: Influences of age and exercise

    NARCIS (Netherlands)

    Brama, P.A.J.; Tekoppele, J.M.; Bank, R.A.; Barneveld, A.; Weeren, P.R. van

    2002-01-01

    The objective of this study was to document the development of biochemical heterogeneity from birth to maturity in equine articular cartilage, and to test the hypothesis that the amount of exercise during early life may influence this process. Neonatal foals showed no biochemical heterogeneity

  17. A Modified Hydroxyproline Assay Based on Hydrochloric Acid in Ehrlich's Solution Accurately Measures Tissue Collagen Content.

    Science.gov (United States)

    Cissell, Derek D; Link, Jarrett M; Hu, Jerry C; Athanasiou, Kyriacos A

    2017-04-01

    Collagen quantification has long been relevant to biomedical research and clinical practice to characterize tissues and determine disease states. The hydroxyproline assay, while a broadly employed method of quantifying collagen, uses perchloric acid to dissolve Ehrlich's reagent. Since perchloric acid poses occupational safety hazards and high costs, in this study, a new hydroxyproline assay was developed that replaces perchloric acid with a relatively safer and cheaper alternative, hydrochloric acid (HCl). To validate this biochemical technique, first, using either acid to dissolve Ehrlich's reagent, the assays were completed for native and engineered collagenous tissues. No statistical differences were identified between the assays (p = 0.32). Subsequently, both biochemical techniques were compared to amino acid analysis, considered a proteomics gold standard. Interestingly, utilizing HCl in lieu of perchloric acid yielded greater concordance with amino acid analysis (ρ c  = 0.980) than did the traditional assay (ρ c  = 0.947); that is, the HCl-based assay more closely estimates hydroxyproline content, and, consequently, true collagen content. Thus, using Ehrlich's reagent containing HCl in the hydroxyproline assay represents an advance in both mitigating laboratory safety hazards and improving biochemical collagen quantification.

  18. Biochemical adaptation of phytopathogenic fungi, Sclerotium rolfsii ...

    African Journals Online (AJOL)

    Biochemical adaptation of phytopathogenic fungi, Sclerotium rolfsii, in response to temperature stress. Natthiya Buensanteai, Kanjana Thumanu, Khanistha Kooboran, Dusit Athinuwat, Sutruedee Prathuangwong ...

  19. Radon assay for SNO+

    Energy Technology Data Exchange (ETDEWEB)

    Rumleskie, Janet [Laurentian University, Greater Sudbury, Ontario (Canada)

    2015-12-31

    The SNO+ experiment will study neutrinos while located 6,800 feet below the surface of the earth at SNOLAB. Though shielded from surface backgrounds, emanation of radon radioisotopes from the surrounding rock leads to back-grounds. The characteristic decay of radon and its daughters allows for an alpha detection technique to count the amount of Rn-222 atoms collected. Traps can collect Rn-222 from various positions and materials, including an assay skid that will collect Rn-222 from the organic liquid scintillator used to detect interactions within SNO+.

  20. Biochemical Markers in Neurocritical Care

    Directory of Open Access Journals (Sweden)

    Omidvar Rezae

    2016-07-01

    Full Text Available During the past two decades, a variety of serum or cerebrospinal fluid (CSF biochemical markers in daily clinical practice have been recommended to diagnose and monitor diverse diseases or pathologic situations. It will be essential to develop a panel of biomarkers, to be suitable for evaluation of treatment efficacy, representing distinct phases of injury and recovery and consider the temporal profile of those. Among the possible and different biochemical markers, S100b appeared to fulfill many of optimized criteria of an ideal marker. S100b, a cytosolic low molecular weight dimeric calciumbinding protein from chromosome 21, synthesized in glial cells throughout the CNS, an homodimeric diffusible, belongs to a family of closely related protein, predominantly expressed by astrocytes and Schwann cells and a classic immunohistochemical marker for these cells, is implicated in brain development and neurophysiology. Of the 3 isoforms of S-100, the BB subunit (S100B is present in high concentrations in central and peripheral glial and Schwann cells, Langerhans and anterior pituitary cells, fat, muscle, and bone marrow tissues. The biomarker has shown to be a sensitive marker of clinical and subclinical cerebral damage, such as stroke, traumatic brain injury, and spinal cord injury. Increasing evidence suggests that the biomarker plays a double function as an intracellular regulator and an extracellular signal of the CNS. S100b is found in the cytoplasm in a soluble form and also is associated with intracellular membranes, centrosomes, microtubules, and type III intermediate filaments. Their genomic organization now is known, and many of their target proteins have been identified, although the mechanisms of regulating S100b secretion are not completely understood and appear to be related to many factors, such as the proinflammatory cytokines, tumor necrosis factor alpha (TNF-a, interleukin (IL-1b, and metabolic stress. 

  1. Biochemical evidence for deficient DNA repair leading to enhanced G2 chromatid radiosensitivity and susceptibility to cancer

    International Nuclear Information System (INIS)

    Gantt, R.; Parshad, R.; Price, F.M.; Sanford, K.K.

    1986-01-01

    Human tumor cells and cells from cancer-prone individuals, compared with those from normal individuals, show a significantly higher incidence of chromatid breaks and gaps seen in metaphase cells immediately after G2 X irradiation. Previous studies with DNA repair-deficient mutants and DNA repair inhibitors strongly indicate that the enhancement results from a G2 deficiency(ies) in DNA repair. We report here biochemical evidence for a DNA repair deficiency that correlates with the cytogenetic studies. In the alkaline elution technique, after a pulse label with radioactive thymidine in the presence of 3-acetylaminobenzamide (a G2-phase blocker) and X irradiation, DNA from tumor or cancer-prone cells elutes more rapidly during the postirradiation period than that from normal cells. These results indicate that the DNA of tumor and cancer-prone cells either repairs more slowly or acquires more breaks than that of normal cells; breaks can accumulate during incomplete or deficient repair processes. The kinetic difference between normal and tumor or cancer-prone cells in DNA strand-break repair reaches a maximum within 2 h, and this maximum corresponds to the kinetic difference in chromatid aberration incidence following X irradiation reported previously. These findings support the concept that cells showing enhanced G2 chromatid radiosensitivity are deficient in DNA repair. The findings could also lead to a biochemical assay for cancer susceptibility

  2. Random assay in radioimmunoassay: Feasibility and application compared with batch assay

    International Nuclear Information System (INIS)

    Lee, Jung Min; Lee, Hwan Hee; Park, Sohyun; Kim, Tae Sung; Kim, Seok Ki

    2016-01-01

    The batch assay has been conventionally used for radioimmunoassay (RIA) because of its technical robustness and practical convenience. However, it has limitations in terms of the relative lag of report time due to the necessity of multiple assays in a small number of samples compared with the random assay technique. In this study, we aimed to verify whether the random assay technique can be applied in RIA and is feasible in daily practice. The coefficients of variation (CVs) of eight standard curves within a single kit were calculated in a CA-125 immunoradiometric assay (IRMA) for the reference of the practically ideal CV of the CA-125 kit. Ten standard curves of 10 kits from 2 prospectively collected lots (pLot) and 85 standard curves of 85 kits from 3 retrospectively collected lots (Lot) were obtained. Additionally, the raw measurement data of both 170 control references and 1123 patients' sera were collected retrospectively between December 2015 and January 2016. A standard curve of the first kit of each lot was used as a master standard curve for a random assay. The CVs of inter-kits were analyzed in each lot, respectively. All raw measurements were normalized by decay and radioactivity. The CA-125 values from control samples and patients' sera were compared using the original batch assay and random assay. In standard curve analysis, the CVs of inter-kits in pLots and Lots were comparable to those within a single kit. The CVs from the random assay with normalization were similar to those from the batch assay in the control samples (CVs % of low/high concentration; Lot1 2.71/1.91, Lot2 2.35/1.83, Lot3 2.83/2.08 vs. Lot1 2.05/1.21, Lot2 1.66/1.48, Lot3 2.41/2.14). The ICCs between the batch assay and random assay using patients' sera were satisfactory (Lot1 1.00, Lot2 0.999, Lot3 1.00). The random assay technique could be successfully applied to the conventional CA-125 IRMA kits. The random assay showed strong agreement with the batch assay. The

  3. Random assay in radioimmunoassay: Feasibility and application compared with batch assay

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jung Min; Lee, Hwan Hee; Park, Sohyun; Kim, Tae Sung; Kim, Seok Ki [Dept. of Nuclear MedicineNational Cancer Center, Goyang (Korea, Republic of)

    2016-12-15

    The batch assay has been conventionally used for radioimmunoassay (RIA) because of its technical robustness and practical convenience. However, it has limitations in terms of the relative lag of report time due to the necessity of multiple assays in a small number of samples compared with the random assay technique. In this study, we aimed to verify whether the random assay technique can be applied in RIA and is feasible in daily practice. The coefficients of variation (CVs) of eight standard curves within a single kit were calculated in a CA-125 immunoradiometric assay (IRMA) for the reference of the practically ideal CV of the CA-125 kit. Ten standard curves of 10 kits from 2 prospectively collected lots (pLot) and 85 standard curves of 85 kits from 3 retrospectively collected lots (Lot) were obtained. Additionally, the raw measurement data of both 170 control references and 1123 patients' sera were collected retrospectively between December 2015 and January 2016. A standard curve of the first kit of each lot was used as a master standard curve for a random assay. The CVs of inter-kits were analyzed in each lot, respectively. All raw measurements were normalized by decay and radioactivity. The CA-125 values from control samples and patients' sera were compared using the original batch assay and random assay. In standard curve analysis, the CVs of inter-kits in pLots and Lots were comparable to those within a single kit. The CVs from the random assay with normalization were similar to those from the batch assay in the control samples (CVs % of low/high concentration; Lot1 2.71/1.91, Lot2 2.35/1.83, Lot3 2.83/2.08 vs. Lot1 2.05/1.21, Lot2 1.66/1.48, Lot3 2.41/2.14). The ICCs between the batch assay and random assay using patients' sera were satisfactory (Lot1 1.00, Lot2 0.999, Lot3 1.00). The random assay technique could be successfully applied to the conventional CA-125 IRMA kits. The random assay showed strong agreement with the batch assay. The

  4. Study on color difference estimation method of medicine biochemical analysis

    Science.gov (United States)

    Wang, Chunhong; Zhou, Yue; Zhao, Hongxia; Sun, Jiashi; Zhou, Fengkun

    2006-01-01

    The biochemical analysis in medicine is an important inspection and diagnosis method in hospital clinic. The biochemical analysis of urine is one important item. The Urine test paper shows corresponding color with different detection project or different illness degree. The color difference between the standard threshold and the test paper color of urine can be used to judge the illness degree, so that further analysis and diagnosis to urine is gotten. The color is a three-dimensional physical variable concerning psychology, while reflectance is one-dimensional variable; therefore, the estimation method of color difference in urine test can have better precision and facility than the conventional test method with one-dimensional reflectance, it can make an accurate diagnose. The digital camera is easy to take an image of urine test paper and is used to carry out the urine biochemical analysis conveniently. On the experiment, the color image of urine test paper is taken by popular color digital camera and saved in the computer which installs a simple color space conversion (RGB -> XYZ -> L *a *b *)and the calculation software. Test sample is graded according to intelligent detection of quantitative color. The images taken every time were saved in computer, and the whole illness process will be monitored. This method can also use in other medicine biochemical analyses that have relation with color. Experiment result shows that this test method is quick and accurate; it can be used in hospital, calibrating organization and family, so its application prospect is extensive.

  5. Opium and heroin alter biochemical parameters of human's serum.

    Science.gov (United States)

    Kouros, Divsalar; Tahereh, Haghpanah; Mohammadreza, Afarinesh; Minoo, Mahmoudi Zarandi

    2010-05-01

    Iran is a significant consumer of opium, and, generally, of opioids, in the world. Addiction is one of the important issues of the 21st century and is an imperative issue in Iran. Long-term consumption of opioids affects homeostasis. To determine the effects of opium and heroin consumption on serum biochemical parameters. In a cross-sectional study, subjects who had consumed heroin (n = 35) or opium (n = 42) for more than two years and 35 nonaddict volunteers as the control group were compared in regard to various biochemical parameters such as fasting blood sugar (FBS), Na(+), K(+), Ca(2+), blood urea nitrogen (BUN), uric acid (UA), triglyceride (TG), cholesterol, creatinine, and total protein. Chromatography was used to confirm opioid consumption, and the concentration of biochemical parameters was determined by laboratory diagnostic tests on serum. No significant differences were found in Na(+), Ca(2+), BUN, UA, TG, creatinine, and total protein concentrations among the three groups. FBS, K(+), and UA levels were significantly lower in opium addicts compared to the control group. Serum Ca(2+) concentration of heroin addicts showed a significant decrease compared to that of the control group. Both addict groups showed a significant decrease in serum cholesterol levels. Chronic use of opium and heroin can change serum FBS, K(+), Ca(2+), UA, and cholesterol. This study, one of few on the effects of opium on serum biochemical parameters in human subjects, has the potential to contribute to the investigation of new approaches for further basic studies.

  6. immunological arthritis Prevalence of biochemical and abnormalities ...

    African Journals Online (AJOL)

    1991-02-02

    Feb 2, 1991 ... Tile prevalence of biochemical and immunological abnormali- ties was studied in a group of 256 patients with rheumatoid arthritis (104 coloureds, 100 whites and 52 blacks). The most common biochemical abnormalities detected were a reduction in the serum creatinine value (43,4%), raised globulins (39 ...

  7. Prevalence of biochemical and immunological abnormalities in ...

    African Journals Online (AJOL)

    Tile prevalence of biochemical and immunological abnormalities was studied in a group of 256 patients with rheumatoid arthritis (104 coloureds, 100 whites and 52 blacks). The most common biochemical abnormalities detected were a reduction in the serum creatinine value (43,4%), raised globulins (39,7%), raised serum ...

  8. Evaluation of Haematological and Biochemical Parameters of ...

    African Journals Online (AJOL)

    After 84 days of exposure, blood was collected and used in conducting haematological and biochemical analyses. Exposure of water to crude oil caused increased levels in chloride, conductivity, salinity, magnesium, biochemical oxygen demand, chemical oxygen demand, turbidity and Nitrate. The crude oil contaminated ...

  9. Diversity in Biochemical Characteristics and Antibiotics Resistant ...

    African Journals Online (AJOL)

    In this study, biochemical and antibiotic susceptibility tests were carried out on one hundred and fifty poultry isolates of Escherichia coli using Microscan® Dried Gram-negative Breakpoint Combo Pannels. The microscan panel analysed bacterial isolates for 24 biochemical tests and 23 to 25 antimicrobial agents following ...

  10. An acoustic prion assay

    Directory of Open Access Journals (Sweden)

    Gordon Hayward

    2016-12-01

    Full Text Available An acoustic prion assay has been demonstrated for sheep brain samples. Only five false positives and no false negatives were observed in a test of 45 positive and 45 negative samples. The acoustic prion sensor was constructed using a thickness shear mode quartz resonator coated with a covalently bound recombinant prion protein. The characteristic indicator of a scrapie infected sheep brain sample was an observed shoulder in the frequency decrease in response to a sample.The response of the sensor aligns with a conformational shift in the surface protein and with the propagation mechanism of the disease. This alignment is evident in the response timing and shape, dependence on concentration, cross species behaviour and impact of blood plasma. This alignment is far from sufficient to prove the mechanism of the sensor but it does offer the possibility of a rapid and inexpensive additional tool to explore prion disease. Keywords: Prions, Thickness shear mode quartz sensor

  11. Assay of oestrogen

    International Nuclear Information System (INIS)

    Edwards, J.C.

    1981-01-01

    A particular problem with the direct radioimmunoassay of unconjugated oestriol in pregnancy is caused by the increased amount of steroid-binding proteins present in pregnancy serum and plasma. The steroid-binding proteins react with oestriol and 125 I-labelled oestriol during the assay procedure and the steroid-protein bound 125 I-labelled oestriol is precipitated along with the antibody-bound 125 I-labelled oestriol by the ammonium sulphate solution separation system. A novel method is described whereby progesterone (1-20 μg/ml) is used to block the action of steroid-binding proteins in pregnancy serum and plasma samples, thus minimizing interference in a direct radioimmunoassay for unconjugated oestriol using a specific anti-oestriol serum. (U.K.)

  12. Coagulation assays and anticoagulant monitoring.

    Science.gov (United States)

    Funk, Dorothy M Adcock

    2012-01-01

    Anticoagulant therapy, including conventional agents and a variety of new oral, fast-acting drugs, is prescribed for millions of patients annually. Each anticoagulant varies in its effect on routine and specialty coagulation assays and each drug may require distinct laboratory assay(s) to measure drug concentration or activity. This review provides an overview of the assorted assays that can measure anticoagulant drug concentration or activity and includes key assay interferences. The effect of these conventional and new anticoagulant agents on specialty coagulation assays used to evaluate for bleeding or clotting disorders, and whether this impact is physiological or factitious, is included. Also provided is a short review of superwarfarin poisoning and features distinguishing this from warfarin overdose. Knowledge of clinically significant pearls and pitfalls pertinent to coagulation assays in relation to anticoagulant therapy are important to optimize patient care.

  13. Organic and biochemical synthesis group

    International Nuclear Information System (INIS)

    Anon.

    1976-01-01

    Stable isotopes, because of their unique properties and non-radioactive nature, have great potential for many fields of science and technology. In particular, isotopes of carbon, nitrogen, oxygen, and sulfur (the basic building blocks of all biological molecules) would be widely used in biomedical and environmental research if they were economically available in sufficient quantities and in the required chemical forms. The major objective of our program continues to be stimulation of the widespread utilization of stable isotopes and commercial involvement through development and demonstration of applications which have potential requirements for large quantities of isotopes. Thus, demand will be created which is necessary for large-scale production of stable isotopes and labeled compounds and concomitant low unit costs. The program continues to produce a variety of labeled materials needed for clinical, biomedical, chemical, and environmental applications which serve as effective demonstrations of unique and advantageous utilization of stable isotopes. Future commercial involvement should benefit, and is a consideration in our research and development, from the technology transfer that can readily be made as a result of our organic and biochemical syntheses and also of various techniques involved in applications

  14. Complex reference value distributions and partitioned reference intervals across the pediatric age range for 14 specialized biochemical markers in the CALIPER cohort of healthy community children and adolescents.

    Science.gov (United States)

    Kelly, Jacalyn; Raizman, Joshua E; Bevilacqua, Victoria; Chan, Man Khun; Chen, Yunqi; Quinn, Frank; Shodin, Beth; Armbruster, David; Adeli, Khosrow

    2015-10-23

    The CALIPER program has previously reported a comprehensive database of pediatric reference intervals for 63 biochemical and immunochemical markers. Here, covariate-stratified reference intervals were determined for a number of special assays not previously reported. A total of 1917 healthy children and adolescents were recruited and serum concentrations of 14 biochemical markers were measured using the Abbott Architect ci4100 system. Age and gender partitions were statistically determined, outliers removed and reference intervals calculated using CSLI C28-A3 guidelines. Many analytes showed dynamic changes in concentration requiring at least 3 age partitions. Unique intervals were required within the first year of life for: pancreatic amylase, C-peptide, ceruloplasmin, insulin, β-2-microglobulin, cystatin C, dehydroepiandrosterone sulfate (DHEA-S), and α-1-glycoprotein. Cholinesterase, cholinesterase-dibucaine number, and immunoglobulin E required only 2 age partitions and α-1-antitrypsin required only one. Anti-CCP and anti-TPO levels were below the detection limit of the assay. Some analytes including insulin and DHEA-S required additional gender partitions for specific age groups. Complex profiles were observed for endocrine and special chemistry markers, requiring establishment of age- and gender-specific reference intervals. These updated reference intervals will allow improved laboratory assessment of pediatric patients but should be validated for each analytical platform and local population as recommended by CLSI. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Heart-type Fatty Acid-Binding Protein (H-FABP as a Biochemical Marker to Differentiate Ischemic and Hemorrhagic Stroke

    Directory of Open Access Journals (Sweden)

    Eli Halimah

    2017-03-01

    Full Text Available Stroke is an acute neurological syndrome that occurs due to a decrease in blood flow caused by blocked or rupture of blood vessels of the brain (cerebrovascular that causes damage to brain tissue. Based on the pathogenesis, there are two types of stroke, the ischemic and hemorrhagic stroke, in which the handling of treatment in both types of stroke are very different, so the differential diagnosis is required to distinguish the two types of stroke. The purpose of this study is to determine whether Heart-type Fatty Acid-Binding Protein (H-FABP can be used as a parameter of biochemical marker to distinguish between ischemic and hemorrhagic stroke. H-FABP assay is performed using blood serum and analyzed by Enzyme Linked Immunosorbent Assay (ELISA sandwich method, each using H-FABP test kit. Blood serum taken from 20 patients with ischemic strokes and 18 hemorrhagic stroke patients from one of a hospital in Jakarta. The results showed that the average H-FABP concentration in ischemic stroke‑patients is 9,07 ng/mL and hemorrhagic stroke‑patients is 18,54 ng/mL; statistically there are significant difference between H-FABP concentration in ischemic stroke‑patients and hemorrhagic stroke-patients (α=0.05. Thus Heart-type Fatty Acid-Binding Protein (H-FABP can be used as one of the parameters of biochemical markers to distinguish between ischemic and hemorrhagic stroke.

  16. The DNA comet assay and the germination test in detection of food treated by ionizing radiation

    International Nuclear Information System (INIS)

    Huachaca, Nelida Simona Marin

    2002-01-01

    Two methods of irradiated food detection, one biochemical, the comet assay and, other biological, the germination test, were applied in bovine meat and fruit samples. The comet assay detects the damage on DNA caused by ionizing radiation. The germination test evaluates the sensitivity to radiation of seeds as for germination ability, shooting and, rooting. The samples were irradiated in gamma font and electron accelerator. For bovine meat samples, the doses were 0.0; 2.5; 4.5 e 7.0 kGy at chilled condition and, 0.0; 2.5; 4.5; 7.0 e 8.5 kGy at frozen conditions. For fruit samples such as melon, watermelon, apple, orange, papaya and, tomato, the doses were: 0.0; 0.5; 0.75; 1.0; 2.0 e 4.0 kGy. The differences between the gamma rays and the electron beam effects on extent of DNA migration and, on shooting and rooting, showed to be similar. The comet assay, under neutral conditions, permitted to discriminate between irradiated and unirradiated bovine meat samples, until one month of storage. Also, it was possible to distinguish, by the comet assay, the control sample with regard to irradiated fruit, at doses as low as 0,5 kGy. In the germination test, the root length was the best parameter to discriminate irradiated and unirradiated samples of melon, watermelon and tomato, while the germination percent was the best parameter for apple and orange. (author)

  17. Solar radiation (PAR and UVA) and water temperature in relation to biochemical performance of Gelidium corneum (Gelidiales, Rhodophyta) in subtidal bottoms off the Basque coast

    Science.gov (United States)

    Quintano, Endika; Ganzedo, Unai; Díez, Isabel; Figueroa, Félix L.; Gorostiaga, José M.

    2013-10-01

    Gelidium corneum (Hudson) J.V. Lamouroux is a very important primary producer in the Cantabrian coastal ecosystem. Some local declines in their populations have been recently detected in the Basque coast. Occurrences of yellowing and an unusual branch breakdown pattern have also been reported for some G. corneum populations. In order to gain further insight into those environmental stressors operating at a local scale, here we investigate if shallow subtidal populations of G. corneum living under potentially different conditions of irradiance (PAR and UVA) and water temperature exhibit differences in some biochemical indicators of stress, namely C:N, antioxidant activity (radical cation of 2,2‧-azino-bis (3-ethylbenzothiazoline-6-sulfonate); ABTS+ assay) and mycosporine-like amino acids (MAAs) (Asterine 330 and Palythine). We hypothesised that G. corneum subjected to higher ambient levels of irradiance and water temperature would show higher C:N ratios, lower antioxidant activity and higher MAA concentrations. Our results partially support this hypothesis. We found that G. corneum exposed to increased levels of irradiance (PAR, UVA) exhibited greater C:N ratios and lower antioxidant activity (higher IC50), whereas no relationship was found regarding MAAs. No differences in biochemical performance in relation to temperature were detected among G. corneum exposed to comparable high light. Similarly, G. corneum growing under lower UVA radiation levels showed no differences in any of the measured biochemical variables with regard to PAR and water temperature. These findings suggest that, among the environmental factors examined, UVA radiation may be an important driver in regulating the along-shore variation in G. corneum biochemical performance. Therefore, the role of irradiance, especially UV radiation, in potential future alterations in Cantabrian G. corneum populations cannot be ruled out as a potential underlying factor.

  18. Micromachined filter-chamber array with passive valves for biochemical assays on beads

    NARCIS (Netherlands)

    Lichtenberg, Jan; Verpoorte, Elisabeth; De Rooij, Nico F.

    2001-01-01

    The filter-chamber array presented here enables a real-time parallel analysis of three different samples on beads in a volume of 3 nL, on a 1 cm2chip. The filter-chamber array is a system containing three filter-chambers, three passive valves at the inlet channels and a common outlet. The design

  19. Effect of Aqueous Stem Bark Extract of Khaya senegalensis on Some Biochemical, Haematological, and Histopathological Parameters of Rats.

    Science.gov (United States)

    Onu, A; Saidu, Y; Ladan, M J; Bilbis, L S; Aliero, A A; Sahabi, S M

    2013-01-01

    The subchronic effect of aqueous stem bark extract of Khaya senegalensis on some biochemical, haematological, and histopathological parameters of rats was investigated. The rats were divided into six groups of five rats per group. Groups I to VI were administered graded doses of 0, 400, 800, 1200, 1600, and 2000 mg/kg bw, respectively. The result of study revealed that administration of the Khaya senegalensis for twenty-eight days at the experimental dose resulted in significant (P < 0.05) increase in urea, electrolytes (Na(+), K(+)), and creatinine levels. The extract also significantly (P < 0.05) increased serum activity of ALT, AST, and ALP. The levels of protein, albumin, and bilirubin were significantly changed when compared to their control values, but they were not dose dependent. The hematological indices assayed in this study were not significantly affected at the experimental dose when compared to the control values. Histological studies of the liver showed cellular degeneration and necrosis and bile duct hyperplasia and fibrosis with lymphocytic infiltration of the hepatocyte, providing supportive evidence for discussing the biochemical findings, indicative of functional derangement. The histological architecture of the kidney and that of the heart were however preserved. The result of this study indicates that the aqueous stem bark extract of K. senegalensis may affect the cellular integrity of vital organs of the body.

  20. Biochemical Analysis Reveals the Multifactorial Mechanism of Histone H3 Clipping by Chicken Liver Histone H3 Protease

    KAUST Repository

    Chauhan, Sakshi

    2016-09-02

    Proteolytic clipping of histone H3 has been identified in many organisms. Despite several studies, the mechanism of clipping, the substrate specificity, and the significance of this poorly understood epigenetic mechanism are not clear. We have previously reported histone H3 specific proteolytic clipping and a protein inhibitor in chicken liver. However, the sites of clipping are still not known very well. In this study, we attempt to identify clipping sites in histone H3 and to determine the mechanism of inhibition by stefin B protein, a cysteine protease inhibitor. By employing site-directed mutagenesis and in vitro biochemical assays, we have identified three distinct clipping sites in recombinant human histone H3 and its variants (H3.1, H3.3, and H3t). However, post-translationally modified histones isolated from chicken liver and Saccharomyces cerevisiae wild-type cells showed different clipping patterns. Clipping of histone H3 N-terminal tail at three sites occurs in a sequential manner. We have further observed that clipping sites are regulated by the structure of the N-terminal tail as well as the globular domain of histone H3. We also have identified the QVVAG region of stefin B protein to be very crucial for inhibition of the protease activity. Altogether, our comprehensive biochemical studies have revealed three distinct clipping sites in histone H3 and their regulation by the structure of histone H3, histone modifications marks, and stefin B.

  1. Risk Aversion in Game Shows

    DEFF Research Database (Denmark)

    Andersen, Steffen; Harrison, Glenn W.; Lau, Morten I.

    2008-01-01

    We review the use of behavior from television game shows to infer risk attitudes. These shows provide evidence when contestants are making decisions over very large stakes, and in a replicated, structured way. Inferences are generally confounded by the subjective assessment of skill in some games...

  2. Biochemical correlates in an animal model of depression

    International Nuclear Information System (INIS)

    Johnson, J.O.

    1986-01-01

    A valid animal model of depression was used to explore specific adrenergic receptor differences between rats exhibiting aberrant behavior and control groups. Preliminary experiments revealed a distinct upregulation of hippocampal beta-receptors (as compared to other brain regions) in those animals acquiring a response deficit as a result of exposure to inescapable footshock. Concurrent studies using standard receptor binding techniques showed no large changes in the density of alpha-adrenergic, serotonergic, or dopaminergic receptor densities. This led to the hypothesis that the hippocampal beta-receptor in responses deficient animals could be correlated with the behavioral changes seen after exposure to the aversive stimulus. Normalization of the behavior through the administration of antidepressants could be expected to reverse the biochemical changes if these are related to the mechanism of action of antidepressant drugs. This study makes three important points: (1) there is a relevant biochemical change in the hippocampus of response deficient rats which occurs in parallel to a well-defined behavior, (2) the biochemical and behavioral changes are normalized by antidepressant treatments exhibiting both serotonergic and adrenergic mechanisms of action, and (3) the mode of action of antidepressants in this model is probably a combination of serotonergic and adrenergic influences modulating the hippocampal beta-receptor. These results are discussed in relation to anatomical and biochemical aspects of antidepressant action

  3. Biosensors and bioelectronics on smartphone for portable biochemical detection.

    Science.gov (United States)

    Zhang, Diming; Liu, Qingjun

    2016-01-15

    Smartphone has been widely integrated with sensors, such as test strips, sensor chips, and hand-held detectors, for biochemical detections due to its portability and ubiquitous availability. Utilizing built-in function modules, smartphone is often employed as controller, analyzer, and displayer for rapid, real-time, and point-of-care monitoring, which can significantly simplify design and reduce cost of the detecting systems. This paper presents a review of biosensors and bioelectronics on smartphone for portable biochemical detections. The biosensors and bioelectronics based on smartphone can mainly be classified into biosensors using optics, surface plasmon resonance, electrochemistry, and near-field communication. The developments of these biosensors and bioelectronics on smartphone are reviewed along with typical biochemical detecting cases. Sensor strategies, detector attachments, and coupling methods are highlighted to show designs of the compact, lightweight, and low-cost sensor systems. The performances and advantages of these designs are introduced with their applications in healthcare diagnosis, environment monitoring, and food evaluation. With advances in micro-manufacture, sensor technology, and miniaturized electronics, biosensor and bioelectronic devices on smartphone can be used to perform biochemical detections as common and convenient as electronic tag readout in foreseeable future. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Biochemical and immunological characteristics of Peruvian Loxosceles laeta spider venom: neutralization of its toxic effects by anti-loxoscelic antivenoms.

    Science.gov (United States)

    Guimarães, G; Dias-Lopes, C; Duarte, C G; Felicori, L; Machado de Avila, R A; Figueiredo, L F M; de Moura, J; Faleiro, B T; Barro, J; Flores, K; Silva, W; Tintaya, B; Yarleque, A; Bonilla, C; Kalapothakis, E; Salas, C E; Chávez-Olortegui, C

    2013-08-01

    This manuscript describes the general biochemical properties and immunological characteristics of Peruvian spider Loxosceles laeta venom (PLlv), which is responsible for the largest number of accidents involving venomous animals in Peru. In this work, we observed that the venom of this spider is more lethal to mice when compared with L. laeta venom from Brazil (BLlv). The LD₅₀ of PLlv was 1.213 mg/kg when the venom was intradermally injected. The venom displayed sphingomyelinase activity and produced dermonecrotic, hemorrhagic and edema effects in rabbits. 2-D SDS-PAGE separation of the soluble venoms resulted in a protein profile ranging from 20 to 205 kDa. Anti-PLlv and anti-BLlv sera produced in rabbits and assayed by ELISA showed that rabbit antibodies cross-reacted with PLlv and BLlv and also with other Brazilian Loxosceles venoms. Western blotting analysis showed that bands corresponding to 25-35 kDa are the proteins best recognized in every Loxosceles spp venoms analyzed. The immunized rabbits displayed protective effect after challenge with PLlv and BLlv. In vitro assays with horse anti-loxoscelic antivenoms produced in Brazil and Peru demonstrated that these commercial antivenoms were efficient to inhibit the sphingomyelinase activity of PLlv and BLlv. Copyright © 2013 Elsevier Ltd. All rights reserved.

  5. Measuring performance at trade shows

    DEFF Research Database (Denmark)

    Hansen, Kåre

    2004-01-01

    Trade shows is an increasingly important marketing activity to many companies, but current measures of trade show performance do not adequately capture dimensions important to exhibitors. Based on the marketing literature's outcome and behavior-based control system taxonomy, a model is built...... that captures a outcome-based sales dimension and four behavior-based dimensions (i.e. information-gathering, relationship building, image building, and motivation activities). A 16-item instrument is developed for assessing exhibitors perceptions of their trade show performance. The paper presents evidence...... of the scale's reliability, factor structure, and validity on the basis of analyzing data from independent samples of exhibitors at the international trade shows SIAL (Paris) and ANUGA (Cologne); and it concludes with a discussion of potential managerial applications and implications for future research. New...

  6. Biochemical analysis of Phytolacca DOPA dioxygenase.

    Science.gov (United States)

    Takahashi, Kana; Yoshida, Kazuko; Yura, Kei; Ashihara, Hiroshi; Sakuta, Masaaki

    2015-05-01

    The biochemical analysis of Phytolacca americana DOPA dioxygenases (PaDOD1 and PaDOD2) was carried out. The recombinant protein of PaDOD1 catalyzed the conversion of DOPA to betalamic acid, whereas DOD activity was not detected in PaDOD2 in vitro. While the reported motif conserved in DODs from betalain-producing plants was found in PaDOD1, a single amino acid residue alteration was detected in PaDOD2. A mutated PaDOD1 protein with a change of 177 Asn to Gly showed reduced specific activity compared with PaDOD1, while DOPA dioxygenase activity was not observed for a mutated PaDOD2 protein which had its conserved motif replaced with that of PaDOD. A three-dimensional (3D) structural model of PaDOD1 and PaDOD2 showed that the conserved motif in DODs was located in the N-terminal side of a loop, which was found close to the putative active site. The difference in stability of the loop may affect the enzymatic activity of PaDOD2.

  7. Skin biochemical composition analysis by Raman spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Oliveira, Patricia Karen; Tosato, Maira Gaspar; Alves, Rani de Souza; Martin, Airton Abrahao; Favero, Priscila Pereira; Raniero, Leandro, E-mail: amartin@univap.br [Laboratorio de Espectroscopia Vibracional Biomedica, Instituto de Pesquisa e Desenvolvimento - IP e D, Universidade do Vale do Paraiba - UniVap, Sao Jose dos Campos, SP (Brazil)

    2012-09-15

    Skin aging is characterized by cellular and molecular alterations. In this context, Confocal Raman spectroscopy was used in vivo to measure these biochemical changes as function of the skin depth. In this study we have tried to correlate spectra from pure amino acids to in vivo spectra from volunteers with different ages. This study was performed on 32 volunteers: 11 from Group A (20-23 years), 11 from Group B (39-42 years) and 10 from Group C (59-62 years). For each group, the Raman spectra were measured on the surface (0 mm), 30 +- 3 mm and 60 +- 3 {mu}m below the surface. The results from intergroup comparisons showed that the oldest group had a prevalence of the tyrosine band, but it also presented a decrease in the band centered at 875 cm{sup -1} of pyrrolidone acid. The amide I band centered at 1637 cm{sup -1} that is attributed to collagen, as well as other proteins and lipid, showed a smaller amount of these biomolecules for Group C, which can be explained by the decrease in collagen concentration as a function of age. (author)

  8. Measurement of peptide-MHC interactions in solution using the spin column filtration assay

    DEFF Research Database (Denmark)

    Buus, Soren; Lise Lauemøller, S; Stryhn, A

    2001-01-01

    This unit describes how peptide-MHC complexes can be generated in vitro using affinity-purified MHC and synthetic peptide. The unit first describes how the interaction between peptide and MHC interaction can be measured in an accurate, quantitative biochemical assay. This procedure has been...

  9. Biochemical methane potential of kraft bleaching effluent and codigestion with other in-mill streams

    DEFF Research Database (Denmark)

    Fitamo, Temesgen Mathewos; Dahl, Olli; Master, Emma

    2016-01-01

    A biochemical methane potential assay was conducted to investigate the anaerobic digestibility of bleaching effluent from hardwood kraft pulping and the potential of codigestion with other effluents from an integrated pulp and paper mill. Four in-mill streams were tested individually and in combi......A biochemical methane potential assay was conducted to investigate the anaerobic digestibility of bleaching effluent from hardwood kraft pulping and the potential of codigestion with other effluents from an integrated pulp and paper mill. Four in-mill streams were tested individually...... of anaerobic inhibitors such as adsorbable organic halogens (36 mg/L), total sulfur (170 mg/L), and resin and fatty acids (3.2 mg/L). Therefore, the total bleaching effluent from hardwood kraft pulping may be considered for full-scale anaerobic wastewater treatment, either as a singular stream or as part...... of a composite stream including other in-mill effluents....

  10. Biochemical and genetic functional dissection of the P38 viral suppressor of RNA silencing.

    Science.gov (United States)

    Iki, Taichiro; Tschopp, Marie-Aude; Voinnet, Olivier

    2017-05-01

    Phytoviruses encode viral suppressors of RNA silencing (VSRs) to counteract the plant antiviral silencing response, which relies on virus-derived small interfering (si)RNAs processed by Dicer RNaseIII enzymes and subsequently loaded into ARGONAUTE (AGO) effector proteins. Here, a tobacco cell-free system was engineered to recapitulate the key steps of antiviral RNA silencing and, in particular, the most upstream double-stranded (ds)RNA processing reaction, not kinetically investigated thus far in the context of plant VSR studies. Comparative biochemical analyses of distinct VSRs in the reconstituted assay showed that in all cases tested, VSR interactions with siRNA duplexes inhibited the loading, but not the activity, of antiviral AGO1 and AGO2. Turnip crinkle virus P38 displayed the additional and unique property to bind both synthetic and RNA-dependent-RNA-polymerase-generated long dsRNAs, and inhibited the processing into siRNAs. Single amino acid substitutions in P38 could dissociate dsRNA-processing from AGO-loading inhibition in vitro and in vivo, illustrating dual-inhibitory strategies discriminatively deployed within a single viral protein, which, we further show, are bona fide suppressor functions that evolved independently of the conserved coat protein function of P38. © 2017 Iki et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  11. Activity profiles of 309 ToxCastTM chemicals evaluated across 292 biochemical targets

    International Nuclear Information System (INIS)

    Knudsen, Thomas B.; Houck, Keith A.; Sipes, Nisha S.; Singh, Amar V.; Judson, Richard S.; Martin, Matthew T.; Weissman, Arthur; Kleinstreuer, Nicole C.; Mortensen, Holly M.; Reif, David M.; Rabinowitz, James R.; Setzer, R. Woodrow; Richard, Ann M.; Dix, David J.; Kavlock, Robert J.

    2011-01-01

    Understanding the potential health risks posed by environmental chemicals is a significant challenge elevated by the large number of diverse chemicals with generally uncharacterized exposures, mechanisms, and toxicities. The present study is a performance evaluation and critical analysis of assay results for an array of 292 high-throughput cell-free assays aimed at preliminary toxicity evaluation of 320 environmental chemicals in EPA's ToxCast TM project (Phase I). The chemicals (309 unique, 11 replicates) were mainly precursors or the active agent of commercial pesticides, for which a wealth of in vivo toxicity data is available. Biochemical HTS (high-throughput screening) profiled cell and tissue extracts using semi-automated biochemical and pharmacological methodologies to evaluate a subset of G-protein coupled receptors (GPCRs), CYP450 enzymes (CYPs), kinases, phosphatases, proteases, HDACs, nuclear receptors, ion channels, and transporters. The primary screen tested all chemicals at a relatively high concentration 25 μM concentration (or 10 μM for CYP assays), and a secondary screen re-tested 9132 chemical-assay pairs in 8-point concentration series from 0.023 to 50 μM (or 0.009-20 μM for CYPs). Mapping relationships across 93,440 chemical-assay pairs based on half-maximal activity concentration (AC50) revealed both known and novel targets in signaling and metabolic pathways. The primary dataset, summary data and details on quality control checks are available for download at (http://www.epa.gov/ncct/toxcast/).

  12. Genotyping of hepatitis C virus-comparison of three assays.

    Science.gov (United States)

    Haushofer, Alexander C; Berg, Jörg; Hauer, René; Trubert-Exinger, Doris; Stekel, Herbert G; Kessler, Harald H

    2003-08-01

    Genotyping of hepatitis C virus (HCV) is clinically relevant to epidemiology, prognosis, and therapeutical management of HCV infection. Accuracy and specificity of three assays for HCV genotyping/subtyping were determined. The TruGene HCV 5'NC Genotyping Kit (TruGene), which is a direct sequencing test and two assays based on reversed hybridization, Inno-LiPA HCV II assay and ViennaLab HCV Strip Assay, were compared. Amplification products generated by the Cobas Amplicor HCV Test were used. A total of 100 consecutive HCV RNA positive samples derived from patients with chronic hepatitis C were examined for their genotypes/subtypes by the three assays. Identification of genotypes and subtypes by the TruGene assay as reference test for the Inno-LiPA HCV II assay and the ViennaLab HCV Strip Assay or Inno-LiPA HCV II assay as reference test for the TruGene and the ViennaLab HCV Strip Assay showed similar results for overall accuracies (TruGene as reference test for Inno-LiPA HCV II and ViennaLab HCV Strip Assay, genotypes/subtypes: 100%/95.5% and 97%/92%; Inno-LiPA HCV II as reference test for TruGene and ViennaLab HCV Strip Assay, genotypes/subtypes: 99%/85.9% and 97%/87.9%) and specificities (TruGene as reference test for Inno-LiPA HCV II and ViennaLab HCV Strip Assay, genotypes/subtypes: 100%/97.8% and 99%/97.7%; Inno-LiPA HCV II as reference test for TruGene and ViennaLab HCV Strip Assay, genotypes/subtypes: 100%/99.4% and 99.7%/98%). The three assays were found to be reliable for the detection and discrimination of all HCV genotypes common in Europe and in North America and to be suitable for the routine diagnostic laboratory.

  13. Evaluation of total PSA assay on vitros ECi and correlation with Kryptor-PSA assay.

    Science.gov (United States)

    Cassinat, B; Wacquet, M; Toubert, M E; Rain, J D; Schlageter, M H

    2001-01-01

    An increasing number of multiparametric immuno-analysers for PSA assays are available. As different immuno-assays may vary in their analytical quality and their accuracy for the follow-up of patients, expertise is necessary for each new assay. The PSA assay on the Vitros-ECi analyser has been evaluated and compared with the PSA assay from the Kryptor analyser. Variation coefficients were 0.91 to 1.98% for within-run assays, and 4.2% to 5.4% for interassay (PSA levels = 0.8 microgram/L to 33.6 micrograms/L). Dilution tests showed 93 to 136% recovery until 70 micrograms/L PSA. Functional sensitivity was estimated at 0.03 microgram/L. Equimolarity of the test was confirmed. Correlation of PSA levels measured with Vitros-ECi and Kryptor analysers displayed a correlation coefficient r2 of 0.9716. The half-lives and doubling times of PSA were similar using both methods. Vitros-ECi PSA assay meets the major criteria for the management of prostate cancer patients.

  14. Effect of Substrate-Inoculum Ratio on the Biochemical Methane Potential of Municipal Biowastes

    OpenAIRE

    Parra-Orobio Brayan Alexis; Torres-Lozada Patricia; Marmolejo-Rebellón Luis Fernando; Cárdenas-Cleves Lina Marcela; Vásquez-Franco Carlos; Torres-López Wilmar Alexander; Ordoñez-Andrade José Abdón

    2015-01-01

    Biowastes are the predominant fraction of municipal solid waste (MSW) of developing countries. Biowastes are characterized by a high content of organic matter; characteristics which facilitates its transformation through biological processes such as anaerobic digestion (AD). Using biochemical methane potential (PBM) assays, in this study we evaluated the influence of substrate-inoculum (S/I) ratio on the AD of biowaste from MSW. The S/I ranged between 0.25 to 9 gSVsubstrate * ...

  15. Accurate atom-mapping computation for biochemical reactions.

    Science.gov (United States)

    Latendresse, Mario; Malerich, Jeremiah P; Travers, Mike; Karp, Peter D

    2012-11-26

    The complete atom mapping of a chemical reaction is a bijection of the reactant atoms to the product atoms that specifies the terminus of each reactant atom. Atom mapping of biochemical reactions is useful for many applications of systems biology, in particular for metabolic engineering where synthesizing new biochemical pathways has to take into account for the number of carbon atoms from a source compound that are conserved in the synthesis of a target compound. Rapid, accurate computation of the atom mapping(s) of a biochemical reaction remains elusive despite significant work on this topic. In particular, past researchers did not validate the accuracy of mapping algorithms. We introduce a new method for computing atom mappings called the minimum weighted edit-distance (MWED) metric. The metric is based on bond propensity to react and computes biochemically valid atom mappings for a large percentage of biochemical reactions. MWED models can be formulated efficiently as Mixed-Integer Linear Programs (MILPs). We have demonstrated this approach on 7501 reactions of the MetaCyc database for which 87% of the models could be solved in less than 10 s. For 2.1% of the reactions, we found multiple optimal atom mappings. We show that the error rate is 0.9% (22 reactions) by comparing these atom mappings to 2446 atom mappings of the manually curated Kyoto Encyclopedia of Genes and Genomes (KEGG) RPAIR database. To our knowledge, our computational atom-mapping approach is the most accurate and among the fastest published to date. The atom-mapping data will be available in the MetaCyc database later in 2012; the atom-mapping software will be available within the Pathway Tools software later in 2012.

  16. DMPD: Innate immunity minireview series: making biochemical sense of nucleic acidsensors that trigger antiviral innate immunity. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 17395579 Innate immunity minireview series: making biochemical sense of nucleic aci...007 Mar 29. (.png) (.svg) (.html) (.csml) Show Innate immunity minireview series: making biochemical sense o...itle Innate immunity minireview series: making biochemical sense of nucleic acidsensors that trigger antivir

  17. Tokyo Motor Show 2003; Tokyo Motor Show 2003

    Energy Technology Data Exchange (ETDEWEB)

    Joly, E.

    2004-01-01

    The text which follows present the different techniques exposed during the 37. Tokyo Motor Show. The report points out the great tendencies of developments of the Japanese automobile industry. The hybrid electric-powered vehicles or those equipped with fuel cells have been highlighted by the Japanese manufacturers which allow considerable budgets in the research of less polluting vehicles. The exposed models, although being all different according to the manufacturer, use always a hybrid system: fuel cell/battery. The manufacturers have stressed too on the intelligent systems for navigation and safety as well as on the design and comfort. (O.M.)

  18. A critical appraisal of the phene-plate biochemical fingerprinting system for epidemiological subtyping of Salmonella typhimurium

    DEFF Research Database (Denmark)

    On, S.L.W.; Baggesen, Dorte Lau

    1996-01-01

    The efficacy and reproducibility of the Phene-Plate (PhP) system (Biosys Inova, Stockholm, Sweden) for biochemical fingerprinting of Salmonella typhimurium was investigated. Duplicate and replicate assays on 40 epidemiologically related and unrelated strains were performed in two batches of PhP-48...

  19. Biochemical and genetical analysis reveal a new clade of biovar 3 Dickeya spp. strains isolated from potato in Europe

    NARCIS (Netherlands)

    Slawiak, M.; Beckhoven, van J.R.C.M.; Speksnijder, A.G.C.L.; Czajkowski, R.L.; Grabe, G.; Wolf, van der J.M.

    2009-01-01

    Sixty-five potato strains of the soft rot-causing plant pathogenic bacterium Dickeya spp., and two strains from hyacinth, were characterised using biochemical assays, REP-PCR genomic finger printing, 16S rDNA and dnaX sequence analysis. These methods were compared with nineteen strains representing

  20. A miniaturized cell-based fluorescence resonance energy transfer assay for insulin-receptor activation.

    Science.gov (United States)

    Marine, Shane; Zamiara, Elize; Smith, S Todd; Stec, Erica M; McGarvey, Jeremy; Kornienko, Oleg; Jiang, Guoqiang; Wong, Kenny K; Stack, Jeffrey H; Zhang, Bei B; Ferrer, Marc; Strulovici, Berta

    2006-08-15

    This report describes the development, optimization, and implementation of a miniaturized cell-based assay for the identification of small-molecule insulin mimetics and potentiators. Cell-based assays are attractive formats for compound screening because they present the molecular targets in their cellular environment. A fluorescence resonance energy transfer (FRET) cell-based assay that measures the insulin-dependent colocalization of Akt2 fused with either cyan fluorescent protein or yellow fluorescent protein to the cellular membrane was developed. This ratiometric FRET assay was miniaturized into a robust, yet sensitive 3456-well nanoplate assay with Z' factors of approximately 0.6 despite a very small assay window (less than twofold full activation with insulin). The FRET assay was used for primary screening of a large compound collection for insulin-receptor agonists and potentiators. To prioritize compounds for further development, primary hits were tested in two additional assays, a biochemical time-resolved fluorescence resonance energy transfer assay to measure insulin-receptor phosphorylation and a translocation-based imaging assay. Results from the three assays were combined to yield 11 compounds as potential leads for the development of insulin mimetics or potentiators.

  1. Create a Polarized Light Show.

    Science.gov (United States)

    Conrad, William H.

    1992-01-01

    Presents a lesson that introduces students to polarized light using a problem-solving approach. After illustrating the concept using a slinky and poster board with a vertical slot, students solve the problem of creating a polarized light show using Polya's problem-solving methods. (MDH)

  2. Producing Talent and Variety Shows.

    Science.gov (United States)

    Szabo, Chuck

    1995-01-01

    Identifies key aspects of producing talent shows and outlines helpful hints for avoiding pitfalls and ensuring a smooth production. Presents suggestions concerning publicity, scheduling, and support personnel. Describes types of acts along with special needs and problems specific to each act. Includes a list of resources. (MJP)

  3. A Review : Dietary Fiber Biochemical Inductions and Its Hypocholesterolemic Effects

    OpenAIRE

    Asj'ari, Sri Rahajoe

    2014-01-01

    Most people already know that dietary_ fiber can have detrimental or beneficial effects on health, primarily with regard to nutritional status. In this writing one of the beneficial effects of the consumption ofdietary fiber especially its hypocholesterolemic effect will be discussed. Many research papers have already showed regarding the connection between the biochemical induction of soluble fiber and the hypocholesterolemic effect of soluble fiber. In these investigations treatments were g...

  4. Widespread nanoparticle-assay interference: implications for nanotoxicity testing.

    Science.gov (United States)

    Ong, Kimberly J; MacCormack, Tyson J; Clark, Rhett J; Ede, James D; Ortega, Van A; Felix, Lindsey C; Dang, Michael K M; Ma, Guibin; Fenniri, Hicham; Veinot, Jonathan G C; Goss, Greg G

    2014-01-01

    The evaluation of engineered nanomaterial safety has been hindered by conflicting reports demonstrating differential degrees of toxicity with the same nanoparticles. The unique properties of these materials increase the likelihood that they will interfere with analytical techniques, which may contribute to this phenomenon. We tested the potential for: 1) nanoparticle intrinsic fluorescence/absorbance, 2) interactions between nanoparticles and assay components, and 3) the effects of adding both nanoparticles and analytes to an assay, to interfere with the accurate assessment of toxicity. Silicon, cadmium selenide, titanium dioxide, and helical rosette nanotubes each affected at least one of the six assays tested, resulting in either substantial over- or under-estimations of toxicity. Simulation of realistic assay conditions revealed that interference could not be predicted solely by interactions between nanoparticles and assay components. Moreover, the nature and degree of interference cannot be predicted solely based on our current understanding of nanomaterial behaviour. A literature survey indicated that ca. 95% of papers from 2010 using biochemical techniques to assess nanotoxicity did not account for potential interference of nanoparticles, and this number had not substantially improved in 2012. We provide guidance on avoiding and/or controlling for such interference to improve the accuracy of nanotoxicity assessments.

  5. Widespread nanoparticle-assay interference: implications for nanotoxicity testing.

    Directory of Open Access Journals (Sweden)

    Kimberly J Ong

    Full Text Available The evaluation of engineered nanomaterial safety has been hindered by conflicting reports demonstrating differential degrees of toxicity with the same nanoparticles. The unique properties of these materials increase the likelihood that they will interfere with analytical techniques, which may contribute to this phenomenon. We tested the potential for: 1 nanoparticle intrinsic fluorescence/absorbance, 2 interactions between nanoparticles and assay components, and 3 the effects of adding both nanoparticles and analytes to an assay, to interfere with the accurate assessment of toxicity. Silicon, cadmium selenide, titanium dioxide, and helical rosette nanotubes each affected at least one of the six assays tested, resulting in either substantial over- or under-estimations of toxicity. Simulation of realistic assay conditions revealed that interference could not be predicted solely by interactions between nanoparticles and assay components. Moreover, the nature and degree of interference cannot be predicted solely based on our current understanding of nanomaterial behaviour. A literature survey indicated that ca. 95% of papers from 2010 using biochemical techniques to assess nanotoxicity did not account for potential interference of nanoparticles, and this number had not substantially improved in 2012. We provide guidance on avoiding and/or controlling for such interference to improve the accuracy of nanotoxicity assessments.

  6. haematological, lipid profile and other biochemical parameters

    African Journals Online (AJOL)

    hi-tech

    2006-01-01

    Jan 1, 2006 ... HAEMATOLOGICAL, LIPID PROFILE AND OTHER BIOCHEMICAL PARAMETERS IN NORMAL AND HYPERTENSIVE SUBJECTS AMONG THE ... Hypertensive individuals had significantly higher serum sodium, chloride and calcium levels but a .... exposed to urbanisation and associated changes in diet,.

  7. Biochemical and immunohistochemical characterisation of mucins in ...

    African Journals Online (AJOL)

    Biochemical and immunohistochemical characterisation of mucins in 8 cases of colonic disease - a pilot study. N Chirwa, A Mall, M Tyler, B Kavin, P Goldberg, JEJ Krige, Z Lotz, D Khan, D Govender, A Hunter ...

  8. Biochemical, haematological and morphological variations in ...

    African Journals Online (AJOL)

    Biochemical, haematological and morphological variations in juvenile Clarias gariepinus exposed to Carbendazim® fungicide. ... protein levels and neutrophil numbers. Consequently, the use and environmental concentrations of CBZ should be closely monitored to safeguard fish health condition in aquatic ecosystems.

  9. Morphological, physiological and biochemical studies on Pyricularia ...

    African Journals Online (AJOL)

    SARAH

    2014-02-28

    Magnaporthe grisea) causes significant yield loss in Ethiopia. This study was conducted to isolate, identify and characterize the pathogen (using morphological, physiological and biochemical methods). Methodology and ...

  10. Raman spectroscopic biochemical mapping of tissues

    Science.gov (United States)

    Stone, Nicholas; Hart Prieto, Maria C.; Kendall, Catherine A.; Shetty, Geeta; Barr, Hugh

    2006-02-01

    Advances in technologies have brought us closer to routine spectroscopic diagnosis of early malignant disease. However, there is still a poor understanding of the carcinogenesis process. For example it is not known whether many cancers follow a logical sequence from dysplasia, to carcinoma in situ, to invasion. Biochemical tissue changes, triggered by genetic mutations, precede morphological and structural changes. These can be probed using Raman or FTIR microspectroscopy and the spectra analysed for biochemical constituents. Local microscopic distribution of various constituents can then be visualised. Raman mapping has been performed on a number of tissues including oesophagus, breast, bladder and prostate. The biochemical constituents have been calculated at each point using basis spectra and least squares analysis. The residual of the least squares fit indicates any unfit spectral components. The biochemical distribution will be compared with the defined histopathological boundaries. The distribution of nucleic acids, glycogen, actin, collagen I, III, IV, lipids and others appear to follow expected patterns.

  11. A Program on Biochemical and Biomedical Engineering.

    Science.gov (United States)

    San, Ka-Yiu; McIntire, Larry V.

    1989-01-01

    Presents an introduction to the Biochemical and Biomedical Engineering program at Rice University. Describes the development of the academic and enhancement programs, including organizational structure and research project titles. (YP)

  12. Short Report Biochemical derangements prior to emergency ...

    African Journals Online (AJOL)

    diagnoses frequently associated with biochemical derangement. A substantial proportion displayed clinically significant derangements in their blood tests. These derangements can result in anorexia, nausea, muscle weakness, paralytic ileus, and ...

  13. Contaminations Occurring in Fungal PCR Assays

    Science.gov (United States)

    Loeffler, Juergen; Hebart, Holger; Bialek, Ralf; Hagmeyer, Lars; Schmidt, Diethard; Serey, Francois-Prâseth; Hartmann, Matthias; Eucker, Jan; Einsele, Hermann

    1999-01-01

    Successful in vitro amplification of fungal DNA in clinical specimens has been reported recently. In a collaboration among five European centers, the frequency and risk of contamination due to airborne spore inoculation or carryover contamination in fungal PCR were analyzed. The identities of all contaminants were specified by cycle sequencing and GenBank analysis. Twelve of 150 PCR assays that together included over 2,800 samples were found to be contaminated (3.3% of the negative controls were contaminated during the DNA extraction, and 4.7% of the PCR mixtures were contaminated during the amplification process). Contaminants were specified as Aspergillus fumigatus, Saccharomyces cerevisiae, and Acremonium spp. Further analysis showed that commercially available products like zymolyase powder or 10× PCR buffer may contain fungal DNA. In conclusion, the risk of contamination is not higher in fungal PCR assays than in other diagnostic PCR-based assays if general precautions are taken. PMID:10074553

  14. Human skeletal muscle biochemical diversity.

    Science.gov (United States)

    Tirrell, Timothy F; Cook, Mark S; Carr, J Austin; Lin, Evie; Ward, Samuel R; Lieber, Richard L

    2012-08-01

    The molecular components largely responsible for muscle attributes such as passive tension development (titin and collagen), active tension development (myosin heavy chain, MHC) and mechanosensitive signaling (titin) have been well studied in animals but less is known about their roles in humans. The purpose of this study was to perform a comprehensive analysis of titin, collagen and MHC isoform distributions in a large number of human muscles, to search for common themes and trends in the muscular organization of the human body. In this study, 599 biopsies were obtained from six human cadaveric donors (mean age 83 years). Three assays were performed on each biopsy - titin molecular mass determination, hydroxyproline content (a surrogate for collagen content) and MHC isoform distribution. Titin molecular mass was increased in more distal muscles of the upper and lower limbs. This trend was also observed for collagen. Percentage MHC-1 data followed a pattern similar to collagen in muscles of the upper extremity but this trend was reversed in the lower extremity. Titin molecular mass was the best predictor of anatomical region and muscle functional group. On average, human muscles had more slow myosin than other mammals. Also, larger titins were generally associated with faster muscles. These trends suggest that distal muscles should have higher passive tension than proximal ones, and that titin size variability may potentially act to 'tune' the protein's mechanotransduction capability.

  15. [Biochemical diagnostics of fatal opium intoxication].

    Science.gov (United States)

    Papyshev, I P; Astashkina, O G; Tuchik, E S; Nikolaev, B S; Cherniaev, A L

    2013-01-01

    Biochemical diagnostics of fatal opium intoxication remains a topical problem in forensic medical science and practice. We investigated materials obtained in the course of forensic medical expertise of the cases of fatal opium intoxication. The study revealed significant differences between myoglobin levels in blood, urine, myocardium, and skeletal muscles. The proposed approach to biochemical diagnostics of fatal opium intoxication enhances the accuracy and the level of evidence of expert conclusions.

  16. Improving Marine Ecosystem Models with Biochemical Tracers

    Science.gov (United States)

    Pethybridge, Heidi R.; Choy, C. Anela; Polovina, Jeffrey J.; Fulton, Elizabeth A.

    2018-01-01

    Empirical data on food web dynamics and predator-prey interactions underpin ecosystem models, which are increasingly used to support strategic management of marine resources. These data have traditionally derived from stomach content analysis, but new and complementary forms of ecological data are increasingly available from biochemical tracer techniques. Extensive opportunities exist to improve the empirical robustness of ecosystem models through the incorporation of biochemical tracer data and derived indices, an area that is rapidly expanding because of advances in analytical developments and sophisticated statistical techniques. Here, we explore the trophic information required by ecosystem model frameworks (species, individual, and size based) and match them to the most commonly used biochemical tracers (bulk tissue and compound-specific stable isotopes, fatty acids, and trace elements). Key quantitative parameters derived from biochemical tracers include estimates of diet composition, niche width, and trophic position. Biochemical tracers also provide powerful insight into the spatial and temporal variability of food web structure and the characterization of dominant basal and microbial food web groups. A major challenge in incorporating biochemical tracer data into ecosystem models is scale and data type mismatches, which can be overcome with greater knowledge exchange and numerical approaches that transform, integrate, and visualize data.

  17. Data quality in drug discovery: the role of analytical performance in ligand binding assays.

    Science.gov (United States)

    Wätzig, Hermann; Oltmann-Norden, Imke; Steinicke, Franziska; Alhazmi, Hassan A; Nachbar, Markus; El-Hady, Deia Abd; Albishri, Hassan M; Baumann, Knut; Exner, Thomas; Böckler, Frank M; El Deeb, Sami

    2015-09-01

    Despite its importance and all the considerable efforts made, the progress in drug discovery is limited. One main reason for this is the partly questionable data quality. Models relating biological activity and structures and in silico predictions rely on precisely and accurately measured binding data. However, these data vary so strongly, such that only variations by orders of magnitude are considered as unreliable. This can certainly be improved considering the high analytical performance in pharmaceutical quality control. Thus the principles, properties and performances of biochemical and cell-based assays are revisited and evaluated. In the part of biochemical assays immunoassays, fluorescence assays, surface plasmon resonance, isothermal calorimetry, nuclear magnetic resonance and affinity capillary electrophoresis are discussed in details, in addition radiation-based ligand binding assays, mass spectrometry, atomic force microscopy and microscale thermophoresis are briefly evaluated. In addition, general sources of error, such as solvent, dilution, sample pretreatment and the quality of reagents and reference materials are discussed. Biochemical assays can be optimized to provide good accuracy and precision (e.g. percental relative standard deviation data quality are still advancing and will further advance the progress in drug development.

  18. "Medicine show." Alice in Doctorland.

    Science.gov (United States)

    1987-01-01

    This is an excerpt from the script of a 1939 play provided to the Institute of Social Medicine and Community Health by the Library of Congress Federal Theater Project Collection at George Mason University Library, Fairfax, Virginia, pages 2-1-8 thru 2-1-14. The Federal Theatre Project (FTP) was part of the New Deal program for the arts 1935-1939. Funded by the Works Progress Administration (WPA) its goal was to employ theater professionals from the relief rolls. A number of FTP plays deal with aspects of medicine and public health. Pageants, puppet shows and documentary plays celebrated progress in medical science while examining social controversies in medical services and the public health movement. "Medicine Show" sharply contrasts technological wonders with social backwardness. The play was rehearsed by the FTP but never opened because funding ended. A revised version ran on Broadway in 1940. The preceding comments are adapted from an excellent, well-illustrated review of five of these plays by Barabara Melosh: "The New Deal's Federal Theatre Project," Medical Heritage, Vol. 2, No. 1 (Jan/Feb 1986), pp. 36-47.

  19. Screening of xanthine oxidase inhibitors in complex mixtures using online HPLC coupled with postcolumn fluorescence-based biochemical detection.

    Science.gov (United States)

    Li, Deqiang; Li, Shaoping; Zhao, Jing

    2014-02-01

    Xanthine oxidase (XO) catalyzes the metabolism of hypoxanthine and xanthine to uric acid, the overproduction and/or underexcretion of which could cause the incidence of hyperuricemia such as gout. Herein, the inhibition of XO is recognized as one of the therapeutic approaches to treat gout. In the present study, an off-line fluorescence-based microplate method was first developed for an XO assay in which the enzyme converted pterin to its fluorescent metabolite isoxanthopterin. Then, a postcolumn continuous XO assay as a means of bioactivity assessment was coupled to HPLC separation to establish the online HPLC with diode array detection, biochemical detection, and MS/MS system for the screening of XO inhibitors. The availability of the online system was first tested with a positive drug, allopurinol, a well-known XO inhibitor, and subsequent analysis of Scutellaria baicalensis extract showed that two main bioactive compounds with XO inhibitory activities were observed, indicating that the developed online system was applicable to complex mixtures. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Varicose veins show enhanced chemokine expression.

    Science.gov (United States)

    Solá, L del Rio; Aceves, M; Dueñas, A I; González-Fajardo, J A; Vaquero, C; Crespo, M Sanchez; García-Rodríguez, C

    2009-11-01

    Leucocyte infiltration in the wall of varicose veins has been reported previously. This study was designed to investigate the expression of pro-inflammatory cytokines and chemokines in control and in patients with varicose veins and to test the effect of treating varicose vein patients with acetylsalicylic acid (ASA) on cytokine expression prior to removal of varices. Sections of vein were removed during operation from both patient groups, and ribonuclease protection assays (RPAs) were performed to assess the expression of chemokines. Group I included non-varicose saphenous veins from healthy patients undergoing amputation for trauma. Varicose veins were obtained from patients with primary varicose undergoing surgical treatment who received no drug (group II) or treatment with 300 mg day(-1) of ASA for 15 days before surgery (group III). Non-varicose veins constitutively expressed low levels of monocyte-chemoattractant protein (MCP-1) and interleukin (IL)-8 mRNA. Varicose veins had a distinct chemokine expression pattern, since significant up-regulation of MCP-1 and IL-8 and a marked expression of IP-10, RANTES, MIP-1alpha and MIP-1beta mRNA were detected. Removal of the endothelium did not alter this pattern. Varicose veins obtained from patients treated with ASA showed a consistent decrease in chemokine expression, although it did not reach statistical significance. Varicose veins showed increased expression of several chemokines compared to control veins. A non-significant reduction of activation was observed following treatment with ASA for 15 days.

  1. Biochemical methane potential tests of different autoclaved and microwaved lignocellulosic organic fractions of municipal solid waste.

    Science.gov (United States)

    Pecorini, Isabella; Baldi, Francesco; Carnevale, Ennio Antonio; Corti, Andrea

    2016-10-01

    The aim of this research was to enhance the anaerobic biodegradability and methane production of two synthetic Organic Fractions of Municipal Solid Waste with different lignocellulosic contents by assessing microwave and autoclave pre-treatments. Biochemical Methane Potential assays were performed for 21days. Changes in the soluble fractions of the organic matter (measured by soluble chemical oxygen demand, carbohydrates and proteins), the first order hydrolysis constant kh and the cumulated methane production at 21days were used to evaluate the efficiency of microwaving and autoclaving pretreatments on substrates solubilization and anaerobic digestion. Microwave treatment led to a methane production increase of 8.5% for both the tested organic fractions while autoclave treatment had an increase ranging from 1.0% to 4.4%. Results showed an increase of the soluble fraction after pre-treatments for both the synthetic organic fractions. Soluble chemical oxygen demand observed significant increases for pretreated substrates (up to 219.8%). In this regard, the mediocre results of methane's production led to the conclusion that autoclaving and microwaving resulted in the hydrolysis of a significant fraction of non-biodegradable organic substances recalcitrant to anaerobic digestion. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. Molecular genetic and biochemical analyses of a DNA repair gene from Serratia marcescens

    International Nuclear Information System (INIS)

    Murphy, K.E.

    1989-01-01

    In Escherichia coli, the SOS response and two 3-methyladenine DNA glycosylases (TagI and TagII) are required for repair of DNA damaged by alkylating agents such as methyl methanesulfonate (MMS). Mutations of the recA gene eliminate the SOS response. TagI and TagII are encoded by the tag and alkA genes, respectively. A gene (rpr) encoding 3-methyladenine DNA glycosylase activity was isolated from the Gram-negative bacterium Serratia marcescens. The gene, localized to a 1.5-kilobase pair SmaI-HindIII restriction fragment, was cloned into plasmid pUC18. The clone complemented E. coli tag alkA and recA mutations for MMS resistance. The rpr gene did not, however, complement recA mutations for resistance to ultraviolet light or the ability to perform homologous recombination reactions, nor did it complement E. coli ada or alkB mutations. Two proteins of molecular weights 42,000 and 16,000 were produced from the rpr locus. Analysis of deletion and insertion mutants of rpr suggested that the 42kD molecule is the active protein. The 16kD protein may either be a breakdown product of the 42kD species or may be encoded by another gene overlapping the reading frame of the rpr gene. Biochemical assays showed that the rpr gene product (Rpr) possesses 3-methyladenine DNA glycosylase activity

  3. Biochemical and structural properties of cyanases from Arabidopsis thaliana and Oryza sativa.

    Directory of Open Access Journals (Sweden)

    Dan Qian

    Full Text Available Cyanate is toxic to all organisms. Cyanase converts cyanate to CO₂ and NH₃ in a bicarbonate-dependent reaction. The biophysical functions and biochemical characteristics of plant cyanases are poorly studied, although it has been investigated in a variety of proteobacteria, cyanobacteria and fungi. In this study, we characterised plant cyanases from Arabidopsis thaliana and Oryza sativa (AtCYN and OsCYN. Prokaryotic-expressed AtCYN and OsCYN both showed cyanase activity in vitro. Temperature had a similar influence on the activity of both cyanases, but pH had a differential impact on AtCYN and OsCYN activity. Homology modelling provided models of monomers of AtCYN and OsCYN, and a coimmunoprecipitation assay and gel filtration indicated that AtCYN and OsCYN formed homodecamers. The analysis of single-residue mutants of AtCYN indicated that the conserved catalytic residues also contributed to the stability of the homodecamer. KCNO treatment inhibited Arabidopsis germination and early seedling growth. Plants containing AtCYN or OsCYN exhibited resistance to KCNO stress, which demonstrated that one role of cyanases in plants is detoxification. Transcription level of AtCYN was higher in the flower than in other organs of Arabidopsis. AtCYN transcription was not significantly affected by KCNO treatment in Arabidopsis, but was induced by salt stress. This research broadens our knowledge on plant detoxification of cyanate via cyanase.

  4. The biochemical characterization of two phosphate transport systems in Phytomonas serpens.

    Science.gov (United States)

    Vieira-Bernardo, Rodrigo; Gomes-Vieira, André Luiz; Carvalho-Kelly, Luiz Fernando; Russo-Abrahão, Thais; Meyer-Fernandes, José Roberto

    2017-02-01

    Inorganic phosphate (P i ) is an essential nutrient for all organisms because it is required for a variety of biochemical processes, such as signal transduction and the synthesis of phosphate-containing biomolecules. Assays of 32 P i uptake performed in the absence or in the presence of Na + indicated the existence of a Na + -dependent and a Na + -independent P i transporter in Phytomonas serpens. Phylogenetic analysis of two hypothetical protein sequences of Phytomonas (EM1) showed similarities to the high-affinity P i transporters of Saccharomyces cerevisiae: Pho84, a Na + -independent P i transporter, and Pho89, a Na + -dependent P i transporter. Plasma membrane depolarization by FCCP, an H + ionophore, strongly decreased P i uptake via both Na + -independent and Na + -dependent carriers, indicating that a membrane potential is essential for P i influx. In addition, the furosemide-sensitive Na + -pump activity in the cells grown in low P i conditions was found to be higher than the activity detected in the plasma membrane of cells cultivated at high P i concentration, suggesting that the up-regulation of the Na + -ATPase pump could be related to the increase of P i uptake by the Pho89p Na + :P i symporter. Here we characterize for the first time two inorganic phosphate transporters powered by Na + and H + gradients and activated by low P i availability in the phytopathogen P. serpens. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Structural and biochemical studies on ATP binding and hydrolysis by the Escherichia coli RNA chaperone Hfq.

    Directory of Open Access Journals (Sweden)

    Hermann Hämmerle

    Full Text Available In Escherichia coli the RNA chaperone Hfq is involved in riboregulation by assisting base-pairing between small regulatory RNAs (sRNAs and mRNA targets. Several structural and biochemical studies revealed RNA binding sites on either surface of the donut shaped Hfq-hexamer. Whereas sRNAs are believed to contact preferentially the YKH motifs present on the proximal site, poly(A(15 and ADP were shown to bind to tripartite binding motifs (ARE circularly positioned on the distal site. Hfq has been reported to bind and to hydrolyze ATP. Here, we present the crystal structure of a C-terminally truncated variant of E. coli Hfq (Hfq(65 in complex with ATP, showing that it binds to the distal R-sites. In addition, we revisited the reported ATPase activity of full length Hfq purified to homogeneity. At variance with previous reports, no ATPase activity was observed for Hfq. In addition, FRET assays neither indicated an impact of ATP on annealing of two model oligoribonucleotides nor did the presence of ATP induce strand displacement. Moreover, ATP did not lead to destabilization of binary and ternary Hfq-RNA complexes, unless a vast stoichiometric excess of ATP was used. Taken together, these studies strongly suggest that ATP is dispensable for and does not interfere with Hfq-mediated RNA transactions.

  6. Potential biochemical and genetic toxicity of triclosan as an emerging pollutant on earthworms (Eisenia fetida).

    Science.gov (United States)

    Lin, Dasong; Zhou, Qixing; Xie, Xiujie; Liu, Yao

    2010-11-01

    Triclosan as an important antimicrobial agent is increasingly detected in the terrestrial environment as sewage sludge and reclaimed water are applied on land, but little is known about its effect on non-target soil organisms. In this study, biochemical responses including changes in the activity of catalase (CAT), superoxide dismutase (SOD), glutathione-S-transferase (GST), and malondialdehyde (MDA) of the earthworm Eisenia fetida were examined in order to assess ecological toxicity of the chemical. The single-cell gel electrophoresis (SCGE) was also used to measure the potential genotoxicity of the chemical. The results showed that the activity of CAT and GST at the highest tested dose could be stimulated after a 2-d exposure, reaching 148% and 123% of that in the control, respectively. However, with prolonged exposure, the activity of CAT and GST at the highest tested dose was inhibited, falling to 47% and 33% of that in the control, respectively. Triclosan induced an increase in the activity of SOD, but no significant (p>0.05) changes were observed. The content of MDA was dependent both on the dose of triclosan and on the exposure duration. The comet assay demonstrated that triclosan treatments led to a dose-dependent DNA damage of E. fetida after exposures of 7 and 14 d. Our findings can suggest that triclosan has sublethal effects on E. fetida. Copyright © 2010 Elsevier Ltd. All rights reserved.

  7. Biochemical and cellular mechanisms regulating Acanthamoeba castellanii adherence to host cells.

    Science.gov (United States)

    Soto-Arredondo, K J; Flores-Villavicencio, L L; Serrano-Luna, J J; Shibayama, M; Sabanero-López, M

    2014-04-01

    Free-living amoebae belonging to the genus Acanthamoeba are the causative agents of infections such as amoebic keratitis (AK), granulomatous amoebic encephalitis (GAE) and cutaneous lesions. The mechanisms involved in the establishment of infection are unknown. However, it is accepted that the initial phase of pathogenesis involves adherence to the host tissue. In this work, we analysed surface molecules with an affinity for epithelial and neuronal cells from the trophozoites of Acanthamoeba castellanii. We also investigated the cellular mechanisms that govern the process of trophozoite adhesion to the host cells. We first used confocal and epifluorescence microscopy to examine the distribution of the A. castellanii actin cytoskeleton during interaction with the host cells. The use of drugs, as cytochalasin B (CB) and latrunculin B (LB), revealed the participation of cytoskeletal filaments in the adhesion process. In addition, to identify the proteins and glycoproteins on the surface of A. castellanii, the trophozoites were labelled with biotin and biotinylated lectins. The results revealed bands of surface proteins, some of which were glycoproteins with mannose and N-acetylglucosamine residues. Interaction assays of biotinylated amoebae proteins with epithelial and neuronal cells showed that some surface proteins had affinity for both cell types. The results of this study provide insight into the biochemical and cellular mechanisms of the Acanthamoeba infection process.

  8. Biochemical action of new complexes of ruthenium with quinolones as potential antitumor agents.

    Science.gov (United States)

    Gruia, Maria Iuliana; Negoita, Valentina; Vasilescu, Monica; Panait, Marieta; Gruia, Ion; Velescu, Bruno Stefan; Uivarosi, Valentina

    2015-06-01

    The aim of the present study paper was to identify the role of reactive oxygen species (ROS) in apoptosis signaling mechanisms. We used for this purpose two ruthenium complex compounds based on that overproduce these reactive species by their metabolism thus manifesting their antitumor activity too. In vivo studies were performed in Walker 256 carcinoma-bearing Wistar rats treated with two ruthenium (III) (Ru(III)) complexes with -fluoroquinolones norfloxacin and ofloxacin. The treatment started 7 days after tumor grafting. We assayed the dynamics of apoptosis by flow-cytometry and the biochemical oxidative status parameters. The biological samples used were serum and whole-tumor tissues; the results were compared to the untreated control group. The results showed an increase of apoptosis from 14.79% to 59.72% 14.79% to 59.72% in tumor cells treated with the most active combination, ruthenium complex with norfloxacin. We also noted an increase of the oxidative status and ROS production during treatment. The newly-synthesized complexes are less toxic and their activity is based on the induction of oxidative stress. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  9. Morphofunctional and Biochemical Approaches for Studying Mitochondrial Changes during Myoblasts Differentiation

    Directory of Open Access Journals (Sweden)

    Elena Barbieri

    2011-01-01

    Full Text Available This study describes mitochondrial behaviour during the C2C12 myoblast differentiation program and proposes a proteomic approach to mitochondria integrated with classical morphofunctional and biochemical analyses. Mitochondrial ultrastructure variations were determined by transmission electron microscopy; mitochondrial mass and membrane potential were analysed by Mitotracker Green and JC-1 stains and by epifluorescence microscope. Expression of PGC1 , NRF1 , and Tfam genes controlling mitochondrial biogenesis was studied by real-time PCR. The mitochondrial functionality was tested by cytochrome c oxidase activity and COXII expression. Mitochondrial proteomic profile was also performed. These assays showed that mitochondrial biogenesis and activity significantly increase in differentiating myotubes. The proteomic profile identifies 32 differentially expressed proteins, mostly involved in oxidative metabolism, typical of myotubes formation. Other notable proteins, such as superoxide dismutase (MnSOD, a cell protection molecule, and voltage-dependent anion-selective channel protein (VDAC1 involved in the mitochondria-mediated apoptosis, were found to be regulated by the myogenic process. The integration of these approaches represents a helpful tool for studying mitochondrial dynamics, biogenesis, and functionality in comparative surveys on mitochondrial pathogenic or senescent satellite cells.

  10. Blood biochemical parameters in male American mink (Neovison vison before and during the breeding season

    Directory of Open Access Journals (Sweden)

    Bogdan Lasota

    2014-05-01

    Full Text Available This study was aimed at selected blood biochemical parameters in the male American mink before (September-November and during the breeding (January-March season. Blood from 143 Black and Sapphire male mink at one year age was collected. The plasma was assayed for the concentration of glucose, total protein, cholesterol, HDL and LDL fractions, triacylglycerides (TG, and the activity of ALT and AST. Concentrations of glucose, protein, total and HDL/LDL cholesterol, and AST activity were generally slightly higher during the breeding season than during the non-breeding season, but remained within the reference range. In the case of ALT activity and TG concentration, the relations were reversed. The parameters studied in the Sapphire mink showed greater variation, both in- and out of the season. In Black and Sapphire males of the American mink, the studied parameters revealed slightly higher values during the breeding season than the non-breeding season. This will vary depending on the color variety. The decrease in TG concentration during the breeding season may indicate an increased energy demand due to ambient temperature falls, and/or may be a sign of increased energy consumption associated with physical exertion during mating. Consideration should be given to the nutrition of male mink during the breeding season.

  11. Polyamines as salinity biochemical marker in callus of eucalyptus urograndis

    Directory of Open Access Journals (Sweden)

    Giuseppina Lima Pace Pereira

    2003-01-01

    Full Text Available Biochemical markers have been used for the analysis of plant cells submitted to several types of stress, among them salinity. This work aimed at analyzing the effect of saline stress in callus of Eucalyptus urograndis on polyamine contents. Explants (hypocotyls obtained from seeds were inoculated in callus inductive medium, submitted to different levels of NaCl and analyzed at 10, 20 and 30 days after the inoculation. The free polyamines were extracted, isolated and quantified using TLC (Thin-Layer Chromatography. Putrescine content was higher and a fall in the spermidine content was observed in callus submitted to salinity condition. The results showed that polyamine accumulation is related to NaCl exposure in callus of Eucalyptus urograndis. The decrease in spermine content could be used as a biochemical marker for Eucalyptus callus subjected to salinity.

  12. The Biochemical Prognostic Factors of Subclinical Hypothyroidism

    Directory of Open Access Journals (Sweden)

    Myung Won Lee

    2014-06-01

    Full Text Available BackgroundPatients with subclinical hypothyroidism (SHT are common in clinical practice. However, the clinical significance of SHT, including prognosis, has not been established. Further clarifying SHT will be critical in devising a management plan and treatment guidelines for SHT patients. Thus, the aim of this study was to investigate the prognostic factors of SHT.MethodsWe reviewed the medical records of Korean patients who visited the endocrinology outpatient clinic of Severance Hospital from January 2008 to September 2012. Newly-diagnosed patients with SHT were selected and reviewed retrospectively. We compared two groups: the SHT maintenance group and the spontaneous improvement group.ResultsThe SHT maintenance group and the spontaneous improvement group had initial thyroid-stimulating hormone (TSH levels that were significantly different (P=0.035. In subanalysis for subjects with TSH levels between 5 to 10 µIU/mL, the spontaneous improvement group showed significantly lower antithyroid peroxidase antibody (anti-TPO-Ab titer than the SHT maintenance group (P=0.039. Regarding lipid profiles, only triglyceride level, unlike total cholesterol and low density lipoprotein cholesterol, was related to TSH level, which is correlated with the severity of SHT. Diffuse thyroiditis on ultrasonography only contributed to the severity of SHT, not to the prognosis. High sensitivity C-reactive protein and urine iodine excretion, generally regarded as possible prognostic factors, did not show any significant relation with the prognosis and severity of SHT.ConclusionOnly initial TSH level was a definite prognostic factor of SHT. TPO-Ab titer was also a helpful prognostic factor for SHT in cases with mildly elevated TSH. Other than TSH and TPO-Ab, we were unable to validate biochemical prognostic factors in this retrospective study for Korean SHT patients.

  13. The value of the indirect immunoradiometric assay of serum alpha - fetoprotein in detecting liver regeneration and neoplastic transformation in chronic liver disease. Part of a coordinated programme on in vitro assay techniques

    International Nuclear Information System (INIS)

    Voiculetz, N.

    1979-07-01

    To investigate the concentration of alphafetoprotein AFP in different liver diseases and above all in liver cancer the immunoradiometric assay was utilized. The results of AFP studies were compared with regeneration index, blastic T lymphocytes transformation as well as other morphological and biochemical data. The results of the investigations indicated that: 38% of chronic benign hepatopathies displayed the values of serum AFP in normal ranges, 54% were in the range of 41 - 200ng/ml, and 8% showed 200 and more ng/ml. The most important conclusion from the work performed was that the elevation of serum AFP level in the evaluation of chronic hepatopathies, especially in cirrhoses, appears as an index of malignancy

  14. A single nucleotide polymorphism (SNP) assay for population ...

    African Journals Online (AJOL)

    Therefore, we developed the first SNP assay to test stratification between Chinese and Japanese populations living in East Asia. The ancestry ... The SNP assay showed excellent promise as a highly potential application to test population stratification in case-control studies of association in Eastern Asians. Key words: ...

  15. Determination of antipsychotic drug in human serum by radioreceptor assay

    International Nuclear Information System (INIS)

    Wu Jinchang; Jiang Yimin

    1989-01-01

    Serum antipsychotic drug in 50 psychosis cases were measured by radioreceptor assay (RRA) and the values were compared in parallel with that by radioimmunoassay (RIA). The results showed that the RRA values were lower than the RIA values, but both assays gave significant correlation between the serum drug level and antipsychotic dose

  16. Paraprotein interference with turbidimetric gentamicin assay.

    Science.gov (United States)

    Dimeski, Goce; Bassett, Kendra; Brown, Nigel

    2015-01-01

    Gentamicin due to its low level of resistance and rapid bactericidal activity is commonly used to treat gram-negative bacteria. However, due to its toxic effects it needs to be monitored. To date, no interference has been reported with gentamicin assays. A patient with leg cellulitis and sepsis received a single dose of gentamicin and a sample was sent for gentamicin analysis. The sample showed high blank absorbance readings on Beckman DxC800 and DC800 analysers with various dilutions. A second sample was received and analysed on a Roche Cobas system to obtain a result. A third sample was received 107 hours later with the same results and this sample was then analysed neat and post ethanol precipitation on all the turbidimetric assays available on the DxC800 analyser. The high blank absorbance was observed upon addition of the reactive reagents due to protein precipitation. Although not obvious from the patient protein results, it was shown the presence of high IgM paraprotein, 18.9 g/L (reference range 0.4-2.3 g/L) was the cause of precipitation, giving high blank readings. Of all the other turbidimetric assays, only vancomicin and valproate showed similar high blank absorbance readings. To be able to provide more rapid results it was shown ethanol could be used as a precipitant of proteins in both calibrators and patient samples with acceptable recovery. IgM paraprotein was identified as the cause of interference with the gentamicin, vancomicin and valproate assays. Protein interference in these assays can be overcome by precipitation with ethanol.

  17. The fluorometric microculture cytotoxicity assay.

    Science.gov (United States)

    Lindhagen, Elin; Nygren, Peter; Larsson, Rolf

    2008-01-01

    The fluorometric microculture cytotoxicity assay (FMCA) is a nonclonogenic microplate-based cell viability assay used for measurement of the cytotoxic and/or cytostatic effect of different compounds in vitro. The assay is based on hydrolysis of the probe, fluorescein diacetate (FDA) by esterases in cells with intact plasma membranes. The assay is available as both a semiautomated 96-well plate setup and a 384-well plate version fully adaptable to robotics. Experimental plates are prepared with a small amount of drug solution and can be stored frozen. Cells are seeded on the plates and cell viability is evaluated after 72 h. The protocol described here is applicable both for cell lines and freshly prepared tumor cells from patients and is suitable both for screening in drug development and as a basis for a predictive test for individualization of anticancer drug therapy.

  18. Modeling error in experimental assays using the bootstrap principle: understanding discrepancies between assays using different dispensing technologies

    Science.gov (United States)

    Hanson, Sonya M.; Ekins, Sean; Chodera, John D.

    2015-12-01

    All experimental assay data contains error, but the magnitude, type, and primary origin of this error is often not obvious. Here, we describe a simple set of assay modeling techniques based on the bootstrap principle that allow sources of error and bias to be simulated and propagated into assay results. We demonstrate how deceptively simple operations—such as the creation of a dilution series with a robotic liquid handler—can significantly amplify imprecision and even contribute substantially to bias. To illustrate these techniques, we review an example of how the choice of dispensing technology can impact assay measurements, and show how large contributions to discrepancies between assays can be easily understood and potentially corrected for. These simple modeling techniques—illustrated with an accompanying IPython notebook—can allow modelers to understand the expected error and bias in experimental datasets, and even help experimentalists design assays to more effectively reach accuracy and imprecision goals.

  19. Assessment and reduction of comet assay variation in relation to DNA damage: studies from the European Comet Assay Validation Group

    DEFF Research Database (Denmark)

    Møller, Peter; Möller, Lennart; Godschalk, Roger W L

    2010-01-01

    The alkaline single cell gel electrophoresis (comet) assay has become a widely used method for the detection of DNA damage and repair in cells and tissues. Still, it has been difficult to compare results from different investigators because of differences in assay conditions and because the data...... are reported in different units. The European Comet Assay Validation Group (ECVAG) was established for the purpose of validation of the comet assay with respect to measures of DNA damage formation and its repair. The results from this inter-laboratory validation trail showed a large variation in measured level...... reliability for the measurement of DNA damage by the comet assay but there is still a need for further validation to reduce both assay and inter-laboratory variation....

  20. Biochemical Removal of HAP Precursors From Coal

    Energy Technology Data Exchange (ETDEWEB)

    Olson, G.; Tucker, L.; Richards, J.

    1997-07-01

    This project addresses DOE`s interest in advanced concepts for controlling emissions of air toxics from coal-fired utility boilers. We are determining the feasibility of developing a biochemical process for the precombustion removal of substantial percentages of 13 inorganic hazardous air pollutant (HAP) precursors from coal. These HAP precursors are Sb, As, Be, Cd, Cr, Cl, Co, F, Pb, Hg, Mn, Ni, and Se. Although rapid physical coal cleaning is done routinely in preparation plants, biochemical processes for removal of HAP precursors from coal potentially offer advantages of deeper cleaning, more specificity, and less coal loss. Compared to chemical processes for coal cleaning, biochemical processes potentially offer lower costs and milder process conditions. Pyrite oxidizing bacteria, most notably Thiobacillusferrooxidans, are being evaluated in this project for their ability to remove HAP precursors from U.S. coals.

  1. Biochemical Removal of HAP Precursors From Coal

    International Nuclear Information System (INIS)

    Olson, G.; Tucker, L.; Richards, J.

    1997-07-01

    This project addresses DOE's interest in advanced concepts for controlling emissions of air toxics from coal-fired utility boilers. We are determining the feasibility of developing a biochemical process for the precombustion removal of substantial percentages of 13 inorganic hazardous air pollutant (HAP) precursors from coal. These HAP precursors are Sb, As, Be, Cd, Cr, Cl, Co, F, Pb, Hg, Mn, Ni, and Se. Although rapid physical coal cleaning is done routinely in preparation plants, biochemical processes for removal of HAP precursors from coal potentially offer advantages of deeper cleaning, more specificity, and less coal loss. Compared to chemical processes for coal cleaning, biochemical processes potentially offer lower costs and milder process conditions. Pyrite oxidizing bacteria, most notably Thiobacillusferrooxidans, are being evaluated in this project for their ability to remove HAP precursors from U.S. coals

  2. eQuilibrator—the biochemical thermodynamics calculator

    Science.gov (United States)

    Flamholz, Avi; Noor, Elad; Bar-Even, Arren; Milo, Ron

    2012-01-01

    The laws of thermodynamics constrain the action of biochemical systems. However, thermodynamic data on biochemical compounds can be difficult to find and is cumbersome to perform calculations with manually. Even simple thermodynamic questions like ‘how much Gibbs energy is released by ATP hydrolysis at pH 5?’ are complicated excessively by the search for accurate data. To address this problem, eQuilibrator couples a comprehensive and accurate database of thermodynamic properties of biochemical compounds and reactions with a simple and powerful online search and calculation interface. The web interface to eQuilibrator (http://equilibrator.weizmann.ac.il) enables easy calculation of Gibbs energies of compounds and reactions given arbitrary pH, ionic strength and metabolite concentrations. The eQuilibrator code is open-source and all thermodynamic source data are freely downloadable in standard formats. Here we describe the database characteristics and implementation and demonstrate its use. PMID:22064852

  3. Biochemical and serological properties of Streptococcus uberis.

    Science.gov (United States)

    Lämmler, C

    1991-12-01

    The Strep-Zym identification system, a combination of 23 enzymatic tests, allowed a rapid biochemical characterization of Streptococcus uberis. The biochemical profiles of the S. uberis cultures clearly differed from those of S. agalactiae and S. dysgalactiae. Serological grouping of S. uberis revealed polysaccharide antigens of groups E, G, P and U. Some cultures of S. uberis demonstrated CAMP-like synergistic hemolytic activities on sheep blood agar and reacted specifically with the lectins of Helix pomatia and Dolichos biflorus. The occurrence of group polysaccharides, CAMP-like reactivities, and the lectin agglutination reactions were obviously not related to each other or to any of the biochemical properties. These reactions, possibly of importance as virulence factors, might serve as epidemiological markers.

  4. Radioligand assay in reproductive biology

    International Nuclear Information System (INIS)

    Korenman, S.G.; Sherman, B.M.

    1975-01-01

    Radioligand assays have been developed for the principal reproductive steroids and peptide hormones. Specific binding reagents have included antibodies, plasma binders, and intracellular receptors. In each assay, problems of specificity, sensitivity, and nonspecific inhibitors were encountered. Many features of the endocrine physiology in childhood, during puberty, and in adulthood have been characterized. Hormonal evaluations of endocrine disorders of reproduction are characterized on the basis of their characteristic pathophysiologic alterations. (U.S.)

  5. Nondestructive assay of HTGR fuel rods

    International Nuclear Information System (INIS)

    Menlove, H.O.

    1974-01-01

    Performance characteristics of three different radioactive source NDA systems are compared for the assay of HTGR fuel rods and stacks of rods. These systems include the fast neutron Sb-Be assay system, the 252 Cf ''Shuffler,'' and the thermal neutron PAPAS assay system. Studies have been made to determinethe perturbation on the measurements from particle size, kernel Th/U ratio, thorium content, and hydrogen content. In addition to the total 235 U determination, the pellet-to-pellet or rod-to-rod uniformity of HTGR fuel rod stacks has been measured by counting the delayed gamma rays with a NaI through-hole in the PAPAS system. These measurements showed that rod substitutions can be detected easily in a fuel stack, and that detailed information is available on the loading variations in a uniform stack. Using a 1.0 mg 252 Cf source, assay rates of 2 to 4 rods/s are possible, thus facilitating measurement of 100 percent of a plant's throughput. (U.S.)

  6. Scalable Parameter Estimation for Genome-Scale Biochemical Reaction Networks

    Science.gov (United States)

    Kaltenbacher, Barbara; Hasenauer, Jan

    2017-01-01

    Mechanistic mathematical modeling of biochemical reaction networks using ordinary differential equation (ODE) models has improved our understanding of small- and medium-scale biological processes. While the same should in principle hold for large- and genome-scale processes, the computational methods for the analysis of ODE models which describe hundreds or thousands of biochemical species and reactions are missing so far. While individual simulations are feasible, the inference of the model parameters from experimental data is computationally too intensive. In this manuscript, we evaluate adjoint sensitivity analysis for parameter estimation in large scale biochemical reaction networks. We present the approach for time-discrete measurement and compare it to state-of-the-art methods used in systems and computational biology. Our comparison reveals a significantly improved computational efficiency and a superior scalability of adjoint sensitivity analysis. The computational complexity is effectively independent of the number of parameters, enabling the analysis of large- and genome-scale models. Our study of a comprehensive kinetic model of ErbB signaling shows that parameter estimation using adjoint sensitivity analysis requires a fraction of the computation time of established methods. The proposed method will facilitate mechanistic modeling of genome-scale cellular processes, as required in the age of omics. PMID:28114351

  7. BIOCHEMICAL NUTRITIONAL PROFILE OF LIVER CIRRHOSIS PATIENTS WITH HEPATOCELLULAR CARCINOMA

    Directory of Open Access Journals (Sweden)

    Gabriela Zanatta PORT

    2014-03-01

    Full Text Available Context Liver cirrhosis patients with hepatocellular carcinoma present nutritional alterations and metabolic disorders that negatively impact the prognosis. Objective The objective is to identify alterations in the metabolism of macro and micronutrients among liver cirrhosis patients with and without hepatocellular carcinoma and their relation to the Child-Turcote-Pugh score and Barcelona Clinic Liver Cancer staging. Methods Analytical transversal study, with 31 hepatocellular carcinoma patients and 48 liver cirrhosis patients. Laboratorial exams were carried out. The existence of an association between the biochemical parameters and the disease severity as well as the presence of hepatocellular carcinoma was assessed. Results The metabolic-nutritional profile of liver cirrhosis patients caused by the hepatitis C virus and hepatocellular carcinoma showed alterations, specifically the lipid (total cholesterol, HDL and triglycerides, protein (albumin, creatinine and uric acid, iron (transferrin, iron and ferritin saturation, hematocrit and hemoglobin, zinc and B12 vitamin profiles. There is a relation between nutritional biochemical markers and the Child-Turcote-Pugh, as well as Barcelona Clinic Liver Cancer staging. Conclusions Considering the existence of alterations in the metabolism of nutrients in liver cirrhosis patients with and without hepatocellular carcinoma, and also that conventional nutritional assessment methods present limitations for this population, the biochemical laboratorial exams are valid to complement the diagnosis of the nutritional state in a quick and practical manner.

  8. DMPD: The oxidation of lipoproteins by monocytes-macrophages. Biochemical andbiological mechanisms. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 10473535 The oxidation of lipoproteins by monocytes-macrophages. Biochemical andbio.... (.png) (.svg) (.html) (.csml) Show The oxidation of lipoproteins by monocytes-macrophages. Biochemical and...biological mechanisms. PubmedID 10473535 Title The oxidation of lipoproteins by m

  9. Accelerated maximum likelihood parameter estimation for stochastic biochemical systems

    Directory of Open Access Journals (Sweden)

    Daigle Bernie J

    2012-05-01

    Full Text Available Abstract Background A prerequisite for the mechanistic simulation of a biochemical system is detailed knowledge of its kinetic parameters. Despite recent experimental advances, the estimation of unknown parameter values from observed data is still a bottleneck for obtaining accurate simulation results. Many methods exist for parameter estimation in deterministic biochemical systems; methods for discrete stochastic systems are less well developed. Given the probabilistic nature of stochastic biochemical models, a natural approach is to choose parameter values that maximize the probability of the observed data with respect to the unknown parameters, a.k.a. the maximum likelihood parameter estimates (MLEs. MLE computation for all but the simplest models requires the simulation of many system trajectories that are consistent with experimental data. For models with unknown parameters, this presents a computational challenge, as the generation of consistent trajectories can be an extremely rare occurrence. Results We have developed Monte Carlo Expectation-Maximization with Modified Cross-Entropy Method (MCEM2: an accelerated method for calculating MLEs that combines advances in rare event simulation with a computationally efficient version of the Monte Carlo expectation-maximization (MCEM algorithm. Our method requires no prior knowledge regarding parameter values, and it automatically provides a multivariate parameter uncertainty estimate. We applied the method to five stochastic systems of increasing complexity, progressing from an analytically tractable pure-birth model to a computationally demanding model of yeast-polarization. Our results demonstrate that MCEM2 substantially accelerates MLE computation on all tested models when compared to a stand-alone version of MCEM. Additionally, we show how our method identifies parameter values for certain classes of models more accurately than two recently proposed computationally efficient methods

  10. Quantifiable Lateral Flow Assay Test Strips

    Science.gov (United States)

    2003-01-01

    As easy to read as a home pregnancy test, three Quantifiable Lateral Flow Assay (QLFA) strips used to test water for E. coli show different results. The brightly glowing control line on the far right of each strip indicates that all three tests ran successfully. But the glowing test line on the middle left and bottom strips reveal their samples were contaminated with E. coli bacteria at two different concentrations. The color intensity correlates with concentration of contamination.

  11. A luminescence assay for natural product inhibitors of the Mycobacterium tuberculosis proteasome.

    Science.gov (United States)

    Gunderwala, Amber; Porter, John

    2016-01-01

    Mycobacterium tuberculosis (Mtb) causes a large global burden of disease, with a high mortality rate in healthy and immuno-compromised patients. A number of molecular targets have been identified for treatment of this disease, including the Mtb proteasome. The Mtb proteasome enhances Mtb survival during nitrosative and oxidative stress in the latent, non-replicative phase. Therefore, Mtb proteasome inhibition could help to combat Mtb infections that do not respond to current therapies. To develop and validate a novel biochemical assay to assess Mtb proteasome activity in the presence of organic and aqueous plant test extracts. Fluorescence (photoluminescence) and luminescence (chemiluminescence) assays were investigated as potential methods to determine the robustness and repeatability for use in screening natural product extracts for Mtb proteasome inhibitors. The fluorescence assay, used widely for Mtb proteasome activity assays, was subject to interference due to the natural fluorescence of compounds in many of the extracts; there is little interference with the luminescence approach. As proof of principle, we used the luminescence assay to screen a small set of plant test extracts. Luminescence is the more suitable assay for assay of plant natural product extracts. The sensitivities of the luminescence and fluorescence assays are comparable. A Z'-factor of 0.58 for the luminescence assay makes it suitable for medium-to-high throughput screening efforts. Copyright © 2016 John Wiley & Sons, Ltd.

  12. 6-sulfatoxymelatonin collected from infant diapers: feasibility and implications for urinary biochemical markers.

    Science.gov (United States)

    Thomas, Karen A

    2010-01-01

    The purpose of this study was to assess feasibility and acceptability of using a diaper pad for collection of in-home infant urinary samples and to test the accuracy of diaper pad extraction for 6-sulfatoxymelatonin and creatinine, which was used to correct assay results for urinary volume. To assess feasibility and acceptability, urine samples from 20 infants were collected over a 24-hr day using a cotton pad inserted in the diaper. The accuracy of diaper pad extraction was evaluated in the laboratory using urine samples collected from 11 adult volunteers and assayed using enzyme immunosorbent assay (EIA). Urine samples were divided, one aliquot was assayed without extraction, and one aliquot was instilled into a diaper pad, extracted, and assayed. Mothers found diaper pad collection acceptable and easy to perform. Of 144 infant urinary samples obtained in the home environment, 59% were usable for assay purposes, and the remaining either were contaminated with stool or were of insufficient volume. While creatinine values from diaper pad extracted and nonextracted samples were highly correlated (r(2) = .947), those of creatinine-corrected 6-sulfatoxymelatonin were not (r(2) = .216). Diaper pad collection procedures altered 6-sulfatoxymelatonin values. Implications for measurement of urinary biochemical substances and statistical analysis are discussed.

  13. Correlation between the genotoxicity endpoints measured by two different genotoxicity assays: comet assay and CBMN assay

    Directory of Open Access Journals (Sweden)

    Carina Ladeira

    2015-06-01

    The results concerning of positive findings by micronuclei and non significant ones by comet assay, are corroborated by Deng et al. (2005 study performed in workers occupationally exposed to methotrexate, also a cytostatic drug. According to Cavallo et al. (2009, the comet assay seems to be more suitable for the prompt evaluation of the genotoxic effects, for instance, of polycyclic aromatic hydrocarbons mixtures containing volatile substances, whereas the micronucleus test seems more appropriate to evaluate the effects of exposure to antineoplastic agents. However, there are studies that observed an increase in both the comet assay and the micronucleus test in nurses handling antineoplastic drugs, although statistical significance was only seen in the comet assay, quite the opposite of our results (Maluf & Erdtmann, 2000; Laffon et al. 2005.

  14. Biochemical characterization of MLH3 missense mutations does not reveal an apparent role of MLH3 in Lynch syndrome

    DEFF Research Database (Denmark)

    Ou, Jianghua; Rasmussen, Merete; Westers, Helga

    2009-01-01

    for eight of these MLH3 UVs identified in suspected Lynch syndrome patients, we performed several biochemical tests. We determined the protein expression and stability, protein localization and interaction of the mutant MLH3 proteins with wildtype MLH1. All eight MLH3 UVs gave protein expression levels...... comparable with wildtype MLH3. Furthermore, the UV-containing proteins, in contrast to previous studies, were all localized normally in the nucleus and they interacted normally with wildtype MLH1. Our different biochemical assays yielded no evidence that the eight MLH3 UVs tested are the cause of hereditary...

  15. Comparison of the biochemical components and characteristic of ...

    African Journals Online (AJOL)

    The nutrient composition, enzyme activities, the profile of free amino acids (FAAs) and protein compositions in Tibetan sheep milk were assayed and compared with those in Chengdu Ma goat. The results showed that the total solids, milk fat and activity of alkaline phosphatase (AKP) in Tibetan sheep milk were significantly ...

  16. Biochemical diagnosis in prenatal uropathy.

    Science.gov (United States)

    Lun, A; Lenz, F; Priem, F; Brux, B; Gross, J; Bollmann, R; Hartung, J; Bartho, S; Kirchmaier, F; Reisinger, I

    1994-08-01

    Thirty-two fetuses, six with prune-belly syndrome, seven with renal cyst, 19 with obstructive uropathy, underwent intrauterine fluid aspiration during weeks 15-37 of gestation. Fluid samples were analysed for Na, K, creatinine, urea, alpha 1-, and beta 2-microglobulin. Aspirate concentrations of sodium below 130 mmol/L and creatinine above 115 mumol/L indicate an active kidney and exclude a renal cyst. However, aspirates from fetal cysts or fetuses with obstructive uropathy showed analyte concentrations for sodium, potassium, creatinine, and urea corresponding to extracellular fluid (ECF). In conclusion fluid aspirates of fetuses with ultrasonographically detectable cystic cavities in the abdomen should be examined for sodium and creatinine to assess remaining renal function for planning of obstetric management.

  17. Towards new antituberculotic targets: biochemical characterisation of mycobacterial RNase E/G

    Directory of Open Access Journals (Sweden)

    Agnes Csanadi

    2008-06-01

    Full Text Available The World Health Organization estimates that each year 3 million people die from tuberculosis (TB and 8 million people become infected. No new anti-TB drugs have been introduced in the past 30 years, even though their development becomes increasingly important to face new challenges posed by multidrug-resistant and extensively drug-resistant strains and by acute infection with M. tuberculosis of HIV positive patients. Owing to its apparently important role in RNA metabolism, the RNase E/G family of endoribonucleases can be considered as a promising target for antimicrobial drugs. This consideration promted us to characterise biochemical properties of the M. tuberculosis RNase E/G homologue. To learn more about specific properties of RNase E/G homologues a M. tuberculosis RNase E/G (MycRne was overexpressed in E. coli and purified as a 6His-tagged polypeptide. To characterise MycRne, we used in vitro cleavage assays and primer extension analysis of total RNA extracted from mycobacteria. We show that affinity purified MycRne has an endoribonucleolytic activity, which is dependent on the 5'-phosphorylation status of RNA. We could also show that RNase E/G has Mg2+ dependent activity and similar to E. coli RNase E, MycRne was able to cleave in an intercistronic region of the putative 9S precursor of 5S rRNA. Although, similar to E. coli RNase E, the mycobacterial RNase E/G homologue plays a role in rRNA processing, the substrate specificities of these enzymes show differences. This suggests that RNase E/G can be used as a promising target for antimicrobial drugs that can be optimized to specifically target pathogenic species.

  18. Comparison of the Hershberger assay and androgen receptor binding assay of twelve chemicals.

    Science.gov (United States)

    Yamasaki, Kanji; Sawaki, Masakuni; Noda, Shoji; Muroi, Takako; Takakura, Saori; Mitoma, Hideo; Sakamoto, Satoko; Nakai, Makoto; Yakabe, Yoshikuni

    2004-02-15

    We performed the Hershberger assay of 12 chemicals based on the OECD draft protocol. The chemicals tested by the Hershberger assay were phthalic acid di-n-hexyl ester, phthalic acid di-n-amyl ester, phthalic acid di-n-propyl ester, diethylstilbestrol, 17beta-estradiol, tamoxifen, 5alpha-dihydrotestosterone, dichlorodiphenyldichloroethane, cyproterone acetate, 6alpha-methyl-17alpha-hydroxy-progesterone, atrazine, and spironolactone. Phthalic acid di-n-hexyl ester, phthalic acid di-n-amyl ester, and phthalic acid di-n-propyl ester are phthalates; diethylstilbestrol and 17beta-estradiol are estrogenic chemicals; tamoxifen is partial estrogen receptor antagonist with mainly estrogenic properties; 5alpha-dihydrotestosterone is an androgen derivatives; dichlorodiphenyldichloroethane is a reference androgen antagonistic chemical; cyproterone acetate, 6alpha-methyl-17alpha-hydroxy-progesterone, and spironolactone have an androgenic steroid structure and are known as androgen antagonistic chemicals; and atrazine is a reference endocrine disruptor. We also subjected these chemicals to the receptor binding assay for androgen. A clear androgen agonistic effect was detected in 5alpha-dihydrotestosterone, and an androgen antagonistic effect was observed in five chemicals: cyproterone acetate, spironolactone, 6alpha-methyl-17alpha-hydroxy-progesterone, phthalic acid di-n-amyl ester, and dichlorodiphenyldichloroethane. By contrast, diethylstilbestrol, 17beta-estradiol, tamoxifen, 5alpha-dihydrotestosterone, dichlorodiphenyldichloroethane, cyproterone acetate, 6alpha-methyl-17alpha-hydroxy-progesterone, and spironolactone were positive in the receptor binding assay for androgen. Three estrogenic chemicals, diethylstilbestrol, 17beta-estradiol, and tamoxifen, were negative in the Hershberger assay with receptor binding affinity. On the other hand, the Hershberger assays of three phthalates were performed at the same dosages, and the results showed androgen antagonistic affinity only

  19. Pathological and biochemical changes in rat eyes exposed to gamma irradiation and benzo(A) pyrene and the protective role of glutathione and oltipraze

    International Nuclear Information System (INIS)

    Abd Elmaguid, A.; Naguib, N.I.; Saad, T.M.M.

    2007-01-01

    This study aims to evaluate the effect of exposure to carcinogenic compounds as benzo(a)pyrene in combination with other risk factor which is gamma irradiation on different eye tissues. The study was also conducted to evaluate the protective role of antioxidants such as glutathione and oltipraze before and during exposure to the risk factors. The first group of rats was kept as normal untreated control group. The second group was treated with oltipraze and glutathione for 14 days (positive control group). The third group was injected (i.p) with benzo(a)pyrene in three successive doses parallel with exposure to whole body gamma irradiation of 6 Gy divided in three successive doses ( 2 Gy/ day). The fourth group was treated with oltipraze and glutathione for 14 days then injected (i.p) with benzo(a)pyrene in the last 3 days of treatment in three successive doses parallel with exposure to the same whole body gamma irradiation as third group (6 Gy). Rat eyes were examined clinically every week. For histopathological and biochemical examinations, all groups were sacrificed at 1 month and 2 months after irradiation exposure and the eye tissues were examined by light microscope. The biochemical parameters such as lipid peroxides, SOD, GSH, GSH reductase and GSH peroxidase were estimated in blood and lens. Soluble and insoluble proteins were measured in lens only.The results showed that i.p injection of rats with benzo(a)pyrene and exposure to gamma irradiation caused alterations in eyes of rats clinically, histologically and biochemically. Animals that received glutathione and oltipraze and subjected to benzo(a)pyrene and radiation showed noticeable amelioration in the assayed parameters indicating their protective role as promising agents

  20. Biochemical responses of ecological importance in males of the austral South America amphipod Hyalella curvispina Shoemaker, 1942 exposed to waterborne cadmium and copper.

    Science.gov (United States)

    Giusto, Anabella; Ferrari, Lucrecia

    2014-02-01

    The use of physiological parameters as sensitive indicators of toxic stress from exposure to different pollutants is an important issue to be studied. Hyalella curvispina is a Neotropical amphipod often used in ecotoxicological evaluations. This work aimed to quantify biochemical responses of ecological importance in H. curvispina males under stress exposure to sublethal concentrations of waterborne copper (Cu) and cadmium (Cd); in order to obtain basic physiological data as indicators of early effect on this species, on track to its standardization. In order to evaluate the physiological, biochemical and energetic status of the exposed animals, the following endpoints were selected: content of glycogen, total proteins, total lipids, triglycerides, glycerol, arginine, arginine phosphate, levels of lipid peroxidation (TBARS), and Na(+)/K(+)ATPase, catalase (CAT) and superoxide dismutase (SOD) activities. Our results show that the concentrations of Cu (135 and 175 µg/L) and Cd (6.5 and 10.5 µg/L) tested altered most of the biochemical variables measured (glycogen, total proteins, total lipids, triglycerides, arginine phosphate, TBARS, and SOD and Na(+)/K(+)ATPase activities). In addition, neither the levels of glycerol and arginine nor CAT activity were affected by exposure to either metal. Energy metabolism was similarly affected both by exposure to Cu and exposure to Cd. The results obtained show the existence of an energy imbalance associated with oxidative damage, suggesting a comprehensive response. This work represents a first contribution of the evaluation of the effect of two heavy metals in some parameters of oxidative stress and energy metabolism of H. curvispina males. The results indicate these parameters can provide a sensitive criterion for the assessment of early ecotoxicological effects of Cu and Cd in laboratory assays, on a native species representative of the zoobenthic and epiphytic communities of South America. © 2013 Elsevier Inc. All rights

  1. Tolerability of a fully maturated cheese in cow's milk allergic children: biochemical, immunochemical, and clinical aspects.

    Directory of Open Access Journals (Sweden)

    Claudia Alessandri

    Full Text Available BACKGROUND: From patients' reports and our preliminary observations, a fully maturated cheese (Parmigiano-Reggiano; PR seems to be well tolerated by a subset of cow's milk (CM allergic patients. OBJECTIVE AND METHODS: To biochemically and immunologically characterize PR samples at different maturation stage and to verify PR tolerability in CM allergic children. Seventy patients, with suspected CM allergy, were enrolled. IgE to CM, α-lactalbumin (ALA, β-lactoglobulin (BLG and caseins (CAS were tested using ImmunoCAP, ISAC103 and skin prick test. Patients underwent a double-blind, placebo-controlled food challenge with CM, and an open food challenge with 36 months-maturated PR. Extracts obtained from PR samples were biochemically analyzed in order to determine protein and peptide contents. Pepsin and trypsin-chymotrypsin-pepsin simulated digestions were applied to PR extracts. Each PR extract was investigated by IgE Single Point Highest Inhibition Achievable assay (SPHIAa. The efficiency analysis was carried out using CM and PR oral challenges as gold standards. RESULTS: The IgE binding to milk allergens was 100% inhibited by almost all PR preparations; the only difference was for CAS, mainly α(S1-CAS. Sixteen patients sensitized to CM tolerated both CM and PR; 29 patients tolerated PR only; 21 patients, reacted to both CM and PR, whereas 4 patients reactive to CM refused to ingest PR. ROC analysis showed that the absence of IgE to BLG measured by ISAC could be a good marker of PR tolerance. The SPHIAa using digested PR preparations showed a marked effect on IgE binding to CAS and almost none on ALA and BLG. CONCLUSIONS: 58% of patients clinically reactive to CM tolerated fully maturated PR. The preliminary digestion of CAS induced by PR maturation process, facilitating a further loss of allergenic reactivity during gut digestion, might explain the tolerance. This hypothesis seems to work when no IgE sensitization to ISAC BLG is detected.

  2. Haematological And Biochemical Effects Of Sulphadimidine In ...

    African Journals Online (AJOL)

    Haematological and biochemical efects of sulphadmidine were studied in Nigerian mongrel dogs. Five Nigerian mongrel dogs of either sex weighing between 7 and 12 kg were used for the study. The pretreatment blood and serum samples were collected and the weight of animals taken before the administraton of 100 ...

  3. Haematological and biochemical responses of starter broiler ...

    African Journals Online (AJOL)

    A study was conducted to investigate the haematological and biochemical responses of starter broiler chickens fed copper and probiotics supplemented diets. A total of 180-day old Marshal broiler chicks were randomly allotted to six treatment groups of 30 birds each. The treatments were divided into three replicates of ten ...

  4. Estimation of biochemical variables using quantumbehaved particle ...

    African Journals Online (AJOL)

    Due to the difficulties in the measurement of biochemical variables in fermentation process, softsensing model based on radius basis function neural network had been established for estimating the variables. To generate a more efficient neural network estimator, we employed the previously proposed quantum-behaved ...

  5. Biochemical Markers of Joint Tissue Turnover

    DEFF Research Database (Denmark)

    Bay-Jensen, Anne-Christine; Sondergaard, Bodil Cecilie; Christiansen, Claus

    2009-01-01

    Recent disappointments in late stage developments of anti-osteoarthritic drugs have reinforced efforts to develop better biomarkers for application in both the drug development process as well as in the routine management of these patients. Here we provide a brief review of biochemical tests...

  6. EDITORIAL A CASCADE OF BIOCHEMICAL AND PHYSIOLOGICAL ...

    African Journals Online (AJOL)

    pharm-chem

    A CASCADE OF BIOCHEMICAL AND PHYSIOLOGICAL MARKERS IN PATHOLOGICAL. DISORDERS. A few years ago, a neurophysiology lecturer in the Department of Medical Physiology, University of. Nairobi, was doing ... from haemolysis of RBC is rapidly distributed into cells and excreted in kidney. It would have been.

  7. Isolation, molecular and biochemical characterization of oil ...

    African Journals Online (AJOL)

    Biochemical and physiological characterization performed on the 34 bacterial isolates, revealed the presence of oil biodegrading bacterial genera and species of Pseudomonas Acidovorans, P. aerugi-nosa, P. alcaligenes, P. fluorescens, P. cepacia, P. mallei, P. maltophilia, P. oleovorans, P. putida, P. stutzeri P. vesicularis, ...

  8. Some haematological, biochemical and zootechnical parameters of ...

    African Journals Online (AJOL)

    biochemical and zootechnical parameters of a colony of fruit eating bats (Eidolon helvum) in Morogoro region, Tanzania. A total of 50 bats were captured using hand held hooks and quickly transported to the laboratory where the various parameters were measured. Zootechnical parameters (body length, wing span and ...

  9. Biochemical Applications in the Analytical Chemistry Lab

    Science.gov (United States)

    Strong, Cynthia; Ruttencutter, Jeffrey

    2004-01-01

    An HPLC and a UV-visible spectrophotometer are identified as instruments that helps to incorporate more biologically-relevant experiments into the course, in order to increase the students understanding of selected biochemistry topics and enhances their ability to apply an analytical approach to biochemical problems. The experiment teaches…

  10. Reference Physiological Ranges for Serum Biochemical Parameters ...

    African Journals Online (AJOL)

    clinical trials and scaling up of ARV drugs among AIDS patients. ... parametric and non parametric statistics for analyses with 2.5 and 97.5 percentiles considered ..... Physiological Ranges for Plasma Biochemical Parameters among Adult Healthy Cameroonians. Urban. Rural. Males. Females. Males. Females. Parameter.

  11. Biochemical and Kinetic Characterization of Geranylgeraniol 18 ...

    African Journals Online (AJOL)

    This enzyme and its gene are an attractive target for development of plaunotol production and its detailed biochemical properties need to be understood. Recently, even though the gene (CYP97C27) coding for GGOH 18-hydroxylase has been identified, cloned, and expressed in Escherichia coli system, the enzyme activity ...

  12. Characterizing multistationarity regimes in biochemical reaction networks.

    Directory of Open Access Journals (Sweden)

    Irene Otero-Muras

    Full Text Available Switch like responses appear as common strategies in the regulation of cellular systems. Here we present a method to characterize bistable regimes in biochemical reaction networks that can be of use to both direct and reverse engineering of biological switches. In the design of a synthetic biological switch, it is important to study the capability for bistability of the underlying biochemical network structure. Chemical Reaction Network Theory (CRNT may help at this level to decide whether a given network has the capacity for multiple positive equilibria, based on their structural properties. However, in order to build a working switch, we also need to ensure that the bistability property is robust, by studying the conditions leading to the existence of two different steady states. In the reverse engineering of biological switches, knowledge collected about the bistable regimes of the underlying potential model structures can contribute at the model identification stage to a drastic reduction of the feasible region in the parameter space of search. In this work, we make use and extend previous results of the CRNT, aiming not only to discriminate whether a biochemical reaction network can exhibit multiple steady states, but also to determine the regions within the whole space of parameters capable of producing multistationarity. To that purpose we present and justify a condition on the parameters of biochemical networks for the appearance of multistationarity, and propose an efficient and reliable computational method to check its satisfaction through the parameter space.

  13. Probabilistic sensitivity analysis of biochemical reaction systems.

    Science.gov (United States)

    Zhang, Hong-Xuan; Dempsey, William P; Goutsias, John

    2009-09-07

    Sensitivity analysis is an indispensable tool for studying the robustness and fragility properties of biochemical reaction systems as well as for designing optimal approaches for selective perturbation and intervention. Deterministic sensitivity analysis techniques, using derivatives of the system response, have been extensively used in the literature. However, these techniques suffer from several drawbacks, which must be carefully considered before using them in problems of systems biology. We develop here a probabilistic approach to sensitivity analysis of biochemical reaction systems. The proposed technique employs a biophysically derived model for parameter fluctuations and, by using a recently suggested variance-based approach to sensitivity analysis [Saltelli et al., Chem. Rev. (Washington, D.C.) 105, 2811 (2005)], it leads to a powerful sensitivity analysis methodology for biochemical reaction systems. The approach presented in this paper addresses many problems associated with derivative-based sensitivity analysis techniques. Most importantly, it produces thermodynamically consistent sensitivity analysis results, can easily accommodate appreciable parameter variations, and allows for systematic investigation of high-order interaction effects. By employing a computational model of the mitogen-activated protein kinase signaling cascade, we demonstrate that our approach is well suited for sensitivity analysis of biochemical reaction systems and can produce a wealth of information about the sensitivity properties of such systems. The price to be paid, however, is a substantial increase in computational complexity over derivative-based techniques, which must be effectively addressed in order to make the proposed approach to sensitivity analysis more practical.

  14. Histopathological and biochemical disrupting effects of Escravos ...

    African Journals Online (AJOL)

    user

    2014-02-27

    Feb 27, 2014 ... The aim of this study was to investigate the histological and biochemical disrupting effects of Escravos ... rupturing or leaking of production infrastructures that are described as, “very old and lack regular ... crude oil were measured in weight on an electronic weighing balance and given orally (oral gavage) ...

  15. Biochemical and serological characterization of Escherichia coli ...

    African Journals Online (AJOL)

    This study was designed to determine the isolation rate, serotypes and biochemical profiles of E. coli from colibacillosis and dead-in-shell embryos in Zaria, Northern-Nigeria. The isolation rate of E. coli from hatcheries studied were 4.67% and 7.50% from farms of Simtu Agricultural Company and National Animal Production ...

  16. Growth, haematological and biochemical responses of growing ...

    African Journals Online (AJOL)

    Growth, haematological and biochemical responses of growing lambs injected. A.N.M Nour El-Din, S.Z El-Zarkouny, S.Z El-Zarkouny, H Ghobashy, H Ghobashy, E.I Abdel-Gawad, E.I Abdel-Gawad, D.J Kesler, D.J Kesler ...

  17. Reference Physiological Ranges for Serum Biochemical Parameters ...

    African Journals Online (AJOL)

    Background: A valid scientific evaluation of the efficacy of HIV vaccines or antiretroviral drugs includes measurement of changes in physiological parameters of subjects from known established baseline reference ranges. This study was designed to establish reference ranges for biochemical parameters among healthy ...

  18. Biochemical analysis of Brachystegia aurycoma harms seeds ...

    African Journals Online (AJOL)

    A biochemical analysis of Brachystegia aurycoma harm seeds was conducted. Brachystegia, of the family Leguminosae sub-family Caesalpinioidae is a timber tree whose seeds “achi”, are relished as soup condiment by the lgbo-speaking people of Nigeria. The seed were subjected to oil extraction using Soxhlet method.

  19. Haematological and Biochemical Studies on some Ruminants ...

    African Journals Online (AJOL)

    Michael Horsfall

    CHUKU, L C; *UWAKWE, A A. Department of Biochemistry, University of Port Harcourt, Rivers State, Nigeria. ABSTRACT: Haematological studies of ruminant vertebrates (cow, sheep and goat) present one acceptable method of understanding the ecological biochemical and physiological relationship between lower and ...

  20. Some hematological and biochemical parameters in smokeless ...

    African Journals Online (AJOL)

    The effect of Jharda powder (smokeless tobacco) on some hematological and biochemical parameters in consumers was investigated. Hematological parameters including hemoglobin content and white blood cell and leukocyte counts were higher in jharda powder consumers, while monocytes and basophiles counts were ...

  1. Some hematological and biochemical parameters in smokeless ...

    African Journals Online (AJOL)

    AJB SERVER

    2007-01-04

    Jan 4, 2007 ... The effect of Jharda powder (smokeless tobacco) on some hematological and biochemical parameters in consumers was investigated. Hematological parameters including hemoglobin content and white blood cell and leukocyte counts were higher in jharda powder consumers, while monocytes and.

  2. Biochemical reference values in elderly black subjects

    African Journals Online (AJOL)

    1990-09-01

    Sep 1, 1990 ... the resulting estimate of normal range. Clin Chem 1971; 17: 275-284. 13. Wooton IDO, King E]. Normal values for blood constituents: inter-hospital differences. Lancer 1953; I: 70. 14. Chen FWK, Millard PH. The effect of ageing on certain biochemical values. Mod Geriarrics 1972; 2: 92. IS. Forbes GB, Reina ...

  3. Biochemical applications of FT-IR spectroscopy

    NARCIS (Netherlands)

    Pistorius, A.M.A.

    1996-01-01

    This thesis describes the use of (FT-)IR spectroscopy in general biochemical research. In chapter 3, IR spectroscopy is used in the quantitation of residual detergent after reconstitution of an integral membrane protein in a pre-defined lipid matrix. This chapter discusses the choice of the

  4. Histopathological and biochemical disrupting effects of Escravos ...

    African Journals Online (AJOL)

    The histological findings include: lymphocytic infiltration, cirrhosis, fibrosis, hemosiderin, oedema, mild tissue scaring and tissue necrosis. Thus, this result suggests that Escravos crude oil is a potential biochemical disruptor and can also affect the micro-architecture of liver and heart. Key words: Escravos crude oil, liver, ...

  5. Selected plasma biochemical parameters in improved indigenous ...

    African Journals Online (AJOL)

    This study was to assess the biochemical parameters of apparently healthy NIGERHYB pigs and comparison of age and sex related differences in these parameters. One hundred and thirty five NIGERHYB pigs (35 boar, 35 sow, 30 weaned boar piglets and 35 weaned gilt piglets) obtained from intensively managed pig ...

  6. 2009 Biochemical Conversion Platform Review Report

    Energy Technology Data Exchange (ETDEWEB)

    Ferrell, John [Office of Energy Efficiency and Renewable Energy (EERE), Washington, DC (United States)

    2009-12-01

    This document summarizes the recommendations and evaluations provided by an independent external panel of experts at the U.S. Department of Energy Biomass Program’s Biochemical Conversion platform review meeting, held on April 14-16, 2009, at the Sheraton Denver Downtown, Denver, Colorado.

  7. Characterization of biochemical behavior of sorghum ( Sorghum ...

    African Journals Online (AJOL)

    The aim of this research was to characterize the biochemical behavior of sorghum plants under saline stress using multivariate statistical analysis methods for efficient management of Sorghum bicolor [Moench.]). The experimental design was completely randomized design composed of three saline concentrations (0, 1.5 ...

  8. Drinking patterns: biochemical and haematological findings in ...

    African Journals Online (AJOL)

    Administrator

    drinking patterns on biochemical and haematological parameters in alcohol consumers in Ile-Ife. Conigrave et al. (1995) stated that a variety of blood tests have been used to aid the assessment of drinking history and that more recently, laboratory tests based on urine, breath and sweat analyses have been investigated.

  9. Biochemical characterization of sea buckthorn ( Hippophae ...

    African Journals Online (AJOL)

    Sea buckthorn is a valuable medicinal plant, cultivated and naturally grown in northern Pakistan. The plant produces berry with small hard seed in the centre. The seed is the source of all nutrients and phytochemcials. However, there is lack of literature regarding the biochemical and physico-chemical quality of the seed.

  10. Effects Of Ceftriaxone On Haematological And Biochemical ...

    African Journals Online (AJOL)

    Short-term effects of ceftriaxone on haematological and biochemical parameters of Nigerian local turkey poults were studied. The pre-treatment blood and serum samples were collected and the weight of animals taken before the administration of body weight for a period of 4 days. The animals were weighed daiy.

  11. Growth performance, hematological and serum biochemical ...

    African Journals Online (AJOL)

    A twelve (12) weeks feeding trial was carried out to determine the effect of varying dietary inclusions of Microdesmis puberula leaf meal (MPLM) on the growth performance, hematological indices and serum biochemical constituents of growing rabbits. Four grower rabbit diets were formulated to contain the leaf meal at ...

  12. Evaluation of Haematological and Biochemical Parameters of ...

    African Journals Online (AJOL)

    ADOWIE PERE

    Evaluation of Haematological and Biochemical Parameters of Juvenile Oreochromis niloticus after Exposure to Water Soluble Fractions of ... niloticus were evaluated. After a preliminary determination of the 96 h-LC50 of ... evaporation, dissolution, emulsion, photolysis and biodegradation which generate a water soluble.

  13. Biochemical and microstructural characteristics of meat samples ...

    African Journals Online (AJOL)

    This study was conducted to compare the efficiency of different plant proteases for changing biochemical and microstructural characteristics in muscle foods. The meat samples from chicken, giant catfish, pork and beef were treated with four types of proteolytic enzymes: Calotropis procera latex proteases, papaya latex ...

  14. First results with a radioreceptor-assay (TRAK-Assay) for TSH-receptor-autoantibodies

    International Nuclear Information System (INIS)

    Becker, W.; Reiners, C.; Boerner, W.

    1983-01-01

    A new radioreceptor-assay (TRAK-assay) for autoantibodies against TSH-receptors was tested in 48 untreated thyrotoxic patients (26 regional autonomies, 22 toxic diffuse goiters). None of the 26 patients with regional autonomy showed positive autoantibody-titers. 4 patients with toxic diffuse goiter and thyrotoxic exophthalmos were TRAK-positive. Positive titers of microsomal and thyreoglobulin autoantibodies could be seen in 8 of 9 patients with positive TRAK-titers. In accordance with the conventional methods for detecting thyroid-stimulating immunoglobulins the new TRAK-assay seems to be suited for differentiating between immunogenic toxic diffuse goiter (Graves' disease) and goiter with disseminated autonomy as well as for prediction of relapse. (orig.) [de

  15. Radiation effects on biochemical systems

    International Nuclear Information System (INIS)

    Seddon, G.M.

    2000-04-01

    Xanthine oxidase catalyses the oxidative hydroxylation of hypoxanthine, xanthine and a wide range of carbonyl compounds. The enzyme exists as an oxidase and a dehydrogenase; both catalyze the oxidation of the same substrates. Steady state radiolysis and pulse radiolysis were used to generate oxidative and reductive free radicals. Their effects on the enzymatic activity of xanthine oxidase were determined. Initially inactivation studies were carried out to evaluate the extent to which radiolysis in aqueous solution affects the enzyme activity. Values of D 37 and G inactivation were calculated following irradiation in the presence of free radical scavengers and in the presence of catalase and superoxide dismutase. The kinetic constants Vmax and Km were also determined following radiolysis. The effect of ionising radiation on the iron content of xanthine oxidase was measured using atomic absorption spectrometry. Native gel electrophoresis and iso-electric focussing were performed in an attempt to demonstrate changes in the overall structure of the enzyme. The binding of xanthine oxidase to heparin was carried out by measuring, (1) the displacement of methylene blue (MB + ) from a heparin-MB + complex, (2) affinity chromatography and, (3) pulse radiolysis. The effect of irradiation on the binding process was investigated using techniques (1) and (2). Finally the radiation-induced conversion of xanthine oxidase to dehydrogenase was established. The results indicate that xanthine oxidase is inactivated greatest in the presence of air and irradiation causes Vmax to he reduced and Km to increase. The iron content of irradiated xanthine oxidase is unaffected. Electrophoresis shows the enzyme becomes fragmented and the isoelectric points of the fragments vary over a wide range of pH. Binding of xanthine oxidase to heparin as measured by displacement of MB + from a heparin-MB + complex suggests that irradiation increases the affinity of the enzyme for the polyanion, whereas

  16. Improved analysis of C4 and C3 photosynthesis via refined in vitro assays of their carbon fixation biochemistry

    Science.gov (United States)

    Sharwood, Robert E.; Sonawane, Balasaheb V.; Ghannoum, Oula; Whitney, Spencer M.

    2016-01-01

    Plants operating C3 and C4 photosynthetic pathways exhibit differences in leaf anatomy and photosynthetic carbon fixation biochemistry. Fully understanding this underpinning biochemical variation is requisite to identifying solutions for improving photosynthetic efficiency and growth. Here we refine assay methods for accurately measuring the carboxylase and decarboxylase activities in C3 and C4 plant soluble protein. We show that differences in plant extract preparation and assay conditions are required to measure NADP-malic enzyme and phosphoenolpyruvate carboxylase (pH 8, Mg2+, 22 °C) and phosphoenolpyruvate carboxykinase (pH 7, >2mM Mn2+, no Mg2+) maximal activities accurately. We validate how the omission of MgCl2 during leaf protein extraction, lengthy (>1min) centrifugation times, and the use of non-pure ribulose-1,5-bisphosphate (RuBP) significantly underestimate Rubisco activation status. We show how Rubisco activation status varies with leaf ontogeny and is generally lower in mature C4 monocot leaves (45–60% activation) relative to C3 monocots (55–90% activation). Consistent with their >3-fold lower Rubisco contents, full Rubisco activation in soluble protein from C4 leaves (<5min) was faster than in C3 plant samples (<10min), with addition of Rubisco activase not required for full activation. We conclude that Rubisco inactivation in illuminated leaves primarily stems from RuBP binding to non-carbamylated enzyme, a state readily reversible by dilution during cellular protein extraction. PMID:27122573

  17. Identification of irradiated pepper with comet assay

    International Nuclear Information System (INIS)

    Prieto Miranda, Enrique Fco.; Moreno Alvarez, Damaris L.; Carro Palacio, Sandra; Iglesia Enriquez, Isora

    2007-01-01

    The treatment of foods with ionizing radiations is a technological process utilized in order to increase the hygienic quality and the storage time of the foods. Several methods of detection of irradiated foods have been recommended. The comet assay of DNA is one fast and economical technique for the qualitative identification of irradiated foods. The objective of the present paper was to identify with the comet assay technique the modifications of the DNA molecule of irradiated pepper storage at environment and refrigeration temperatures and different post-irradiation times for different absorbed dose values, (0.1, 0.3 and 0.5 kGy). It was demonstrated that for the high absorbed dose values was observed a greater break into fragments of the DNA molecule, which shows the application of this technique for the identification of irradiated foods. (author)

  18. Radiation effects on biochemical systems

    Energy Technology Data Exchange (ETDEWEB)

    Seddon, G.M

    2000-04-01

    Xanthine oxidase catalyses the oxidative hydroxylation of hypoxanthine, xanthine and a wide range of carbonyl compounds. The enzyme exists as an oxidase and a dehydrogenase; both catalyze the oxidation of the same substrates. Steady state radiolysis and pulse radiolysis were used to generate oxidative and reductive free radicals. Their effects on the enzymatic activity of xanthine oxidase were determined. Initially inactivation studies were carried out to evaluate the extent to which radiolysis in aqueous solution affects the enzyme activity. Values of D{sub 37} and G{sub inactivation} were calculated following irradiation in the presence of free radical scavengers and in the presence of catalase and superoxide dismutase. The kinetic constants Vmax and Km were also determined following radiolysis. The effect of ionising radiation on the iron content of xanthine oxidase was measured using atomic absorption spectrometry. Native gel electrophoresis and iso-electric focussing were performed in an attempt to demonstrate changes in the overall structure of the enzyme. The binding of xanthine oxidase to heparin was carried out by measuring, (1) the displacement of methylene blue (MB{sup +}) from a heparin-MB{sup +} complex, (2) affinity chromatography and, (3) pulse radiolysis. The effect of irradiation on the binding process was investigated using techniques (1) and (2). Finally the radiation-induced conversion of xanthine oxidase to dehydrogenase was established. The results indicate that xanthine oxidase is inactivated greatest in the presence of air and irradiation causes Vmax to he reduced and Km to increase. The iron content of irradiated xanthine oxidase is unaffected. Electrophoresis shows the enzyme becomes fragmented and the isoelectric points of the fragments vary over a wide range of pH. Binding of xanthine oxidase to heparin as measured by displacement of MB{sup +} from a heparin-MB{sup +} complex suggests that irradiation increases the affinity of the enzyme

  19. A radioreceptor assay for benzodiazepines

    International Nuclear Information System (INIS)

    Hunt, P.; Husson, J.-M.; Raynaud, J.-P.

    1979-01-01

    A simple, rapid and sensitive radioreceptor assay for determining benzodiazepines in serum is based on the displacement by the drug specific [ 3 H] diazepam binding to a membrane fraction from rat brain. The limit of detection of the more active benzodiazepines is about 0.5 ng. Diazepam, nitrazepam, clobazam and HR 458 have been assayed in human serum after a single oral clinical dose. The results can be used for determining pharmacokinetic parameters. The technique measures not only the parent benzodiazepine but also clinically active metabolites. (author)

  20. Calixarene methylene bisphosphonic acids as promising effectors of biochemical processes

    Directory of Open Access Journals (Sweden)

    S. V. Komisarenko

    2013-12-01

    rim have been studied with respect to their effects on fibrin polymerization. The most potent inhibitor proved to be calix[4]arene tetrakis-methylene-bis-phosphonic acid (C-192, in which case the maximum rate of fibrin polymerization in the fibrinogen + thrombin reaction decreased by 50% at concentrations of 0.52·10-6 M (IC50. At this concentration, the molar ratio of the compound to fibrinogen was 1.7 : 1. For the case of desAB fibrin polymerization, the IC50 was 1.26·10-6 M at a molar ratio of C-192 to fibrin monomer of 4 : 1. Dipropoxycalix[4]-arene bis-methylene-bis-phosphonic acid (C-98 inhibited fibrin desAB polymerization with an IC50 = 1.31·10-4 M. We hypothesized that C-192 blocks fibrin formation by combining with polymerization site ‘A’ (Aa17–19, which ordinarily initiates protofibril formation in a ‘knob-hole’ manner. This suggestion was confirmed by an HPLC assay, which showed a host–guest inclusion complex of C-192 with the synthetic peptide Gly-Pro-Arg-Pro, an analogue of site ‘A’. Further confirmation that the inhibitor was acting at the initial step of the reaction was obtained by electron microscopy, with no evidence of protofibril formation being evident. Calixarene C-192 also doubled both the prothrombin time and the activated partial thromboplastin time in normal human blood plasma at concentrations of 7.13·10-5 and 1.10·10-5 M, respectively. These experiments demonstrate that C-192 is a specific inhibitor of fibrin polymerization and blood coagulation and can be used for the design of a new class of antithrombotic agents.

  1. Development of loop-mediated isothermal amplification and PCR assays for rapid and simple detection of Campylobacter fetus subsp. venerealis.

    Science.gov (United States)

    Yamazaki, Wataru; Taguchi, Masumi; Misawa, Naoaki

    2010-07-01

    Campylobacter fetus is divided into CFV and CFF. Because CFV causes bovine genital campylobacteriosis, differentiation of the two subspecies is essential to the implementation of efficient CFV control and eradication programs. We have developed LAMP and duplex PCR assays for rapid and simple detection of CFV. The LAMP assay correctly detected 7 CFV strains and did not detect 53 CFF, 35 non-fetus Campylobacter and 25 non-Campylobacter strains. The PCR assay successfully differentiated the two subspecies. The LAMP and PCR assays were faster than conventional biochemical assays, requiring for detection less than 50 min and less than 4 hr, respectively, from the beginning of DNA extraction from a single colony on blood agar to final determination. Our LAMP and PCR assays are rapid and practical tools for detection of CFV.

  2. A radioactive assay for the degradation of neuropeptide Y

    International Nuclear Information System (INIS)

    Ludwig, R.; Lucius, R.; Mentlein, R.

    1995-01-01

    Neuropeptide Y (NPY) is one of the most abundant neuropeptides in the mammalian central nervous system. Like other neuropeptides, NPY is inactivated by specialized neuro-peptidases. To trace the degradation of NPY, an assay was established using biotinylated NPY. Biotinyl-NPY was radiolabeled with Na 125 I by the chloramine-T method and bound to a streptavidin-agarose matrix. The amount of radiolabeling was analyzed by reverse-phase HPLC. The assay was carried out with five peptidases and inhibitors to demonstrate different specific activity. Measurable amounts of radioactivity were released by treatment with endopeptidase-24.18, plasmin, and trypsin, whereas dipetidylpeptidase IV (DPPIV) and angiotensin-converting enzyme (ACE) showed no activity in this assay. In the case of DPPIV this is due to a resistance of the assay to aminopeptidase attack. The assay is useful to study the specific degradation of NPY particularly by endopeptidases in all kinds of biological samples. (authors). 31 refs., 6 figs

  3. Biochemical mechanisms of insecticide resistance in field population of Dengue vector Aedes aegypti (Diptera: Culicidae.

    Directory of Open Access Journals (Sweden)

    R. Muthusamy

    2014-03-01

    Full Text Available Insecticide resistance has been known to be prevalent in several insect species including mosquito. It has become a major problem in vector control programme due to pesticide resistance through detoxification enzymes. The present study investigated the toxicity of Ae. aegypti to organophosphates and pyrethroid insecticide and biochemical mechanisms involved in insecticide resistance in larval population. Larval bioassay revealed an LC50 value of 0.734 ppm for dichlorvos and 1.140 ppm for λ-cyhalothrin exposure. Biochemical assay revealed increased activity of AChE (0.3 µmole/mg protein and GST in dichlorvos (1-1.5 µmole/mg protein treatment and esterase activity in λ-cyhalothrin treated compared to control activity. These studies suggest that AChE and GST is associated with organophosphate and esterase associated with pyrethroid resistance in Ae. aegypti.

  4. Detection of Misfolded Aβ Oligomers for Sensitive Biochemical Diagnosis of Alzheimer’s Disease

    Directory of Open Access Journals (Sweden)

    Natalia Salvadores

    2014-04-01

    Full Text Available Alzheimer's disease (AD diagnosis is hampered by the lack of early, sensitive, and objective laboratory tests. We describe a sensitive method for biochemical diagnosis of AD based on specific detection of misfolded Aβ oligomers, which play a central role in AD pathogenesis. The protein misfolding cyclic amplification assay (Aβ-PMCA, exploits the functional property of Aβ oligomers to seed the polymerization of monomeric Aβ. Aβ-PMCA allowed detection of as little as 3 fmol of Aβ oligomers. Most importantly, using cerebrospinal fluid, we were able to distinguish AD patients from control individuals affected by a variety of other neurodegenerative disorders or nondegenerative neurological diseases with overall sensitivity of 90% and specificity of 92%. These findings provide the proof-of-principle basis for developing a highly sensitive and specific biochemical test for AD diagnosis.

  5. A simple guide to biochemical approaches for analyzing protein–lipid interactions

    Science.gov (United States)

    Zhao, Hongxia; Lappalainen, Pekka

    2012-01-01

    Eukaryotic cells contain many different membrane compartments with characteristic shapes, lipid compositions, and dynamics. A large fraction of cytoplasmic proteins associate with these membrane compartments. Such protein–lipid interactions, which regulate the subcellular localizations and activities of peripheral membrane proteins, are fundamentally important for a variety of cell biological processes ranging from cytoskeletal dynamics and membrane trafficking to intracellular signaling. Reciprocally, many membrane-associated proteins can modulate the shape, lipid composition, and dynamics of cellular membranes. Determining the exact mechanisms by which these proteins interact with membranes will be essential to understanding their biological functions. In this Technical Perspective, we provide a brief introduction to selected biochemical methods that can be applied to study protein–lipid interactions. We also discuss how important it is to choose proper lipid composition, type of model membrane, and biochemical assay to obtain reliable and informative data from the lipid-interaction mechanism of a protein of interest. PMID:22848065

  6. A simple guide to biochemical approaches for analyzing protein-lipid interactions.

    Science.gov (United States)

    Zhao, Hongxia; Lappalainen, Pekka

    2012-08-01

    Eukaryotic cells contain many different membrane compartments with characteristic shapes, lipid compositions, and dynamics. A large fraction of cytoplasmic proteins associate with these membrane compartments. Such protein-lipid interactions, which regulate the subcellular localizations and activities of peripheral membrane proteins, are fundamentally important for a variety of cell biological processes ranging from cytoskeletal dynamics and membrane trafficking to intracellular signaling. Reciprocally, many membrane-associated proteins can modulate the shape, lipid composition, and dynamics of cellular membranes. Determining the exact mechanisms by which these proteins interact with membranes will be essential to understanding their biological functions. In this Technical Perspective, we provide a brief introduction to selected biochemical methods that can be applied to study protein-lipid interactions. We also discuss how important it is to choose proper lipid composition, type of model membrane, and biochemical assay to obtain reliable and informative data from the lipid-interaction mechanism of a protein of interest.

  7. A novel miRNA-based predictive model for biochemical failure following post-prostatectomy salvage radiation therapy.

    Directory of Open Access Journals (Sweden)

    Erica Hlavin Bell

    Full Text Available To develop a microRNA (miRNA-based predictive model for prostate cancer patients of 1 time to biochemical recurrence after radical prostatectomy and 2 biochemical recurrence after salvage radiation therapy following documented biochemical disease progression post-radical prostatectomy.Forty three patients who had undergone salvage radiation therapy following biochemical failure after radical prostatectomy with greater than 4 years of follow-up data were identified. Formalin-fixed, paraffin-embedded tissue blocks were collected for all patients and total RNA was isolated from 1mm cores enriched for tumor (>70%. Eight hundred miRNAs were analyzed simultaneously using the nCounter human miRNA v2 assay (NanoString Technologies; Seattle, WA. Univariate and multivariate Cox proportion hazards regression models as well as receiver operating characteristics were used to identify statistically significant miRNAs that were predictive of biochemical recurrence.Eighty eight miRNAs were identified to be significantly (p36 months. Nine miRNAs were identified to be significantly (p<0.05 associated by multivariate analysis with biochemical failure after salvage radiation therapy. A new predictive model for biochemical recurrence after salvage radiation therapy was developed; this model consisted of miR-4516 and miR-601 together with, Gleason score, and lymph node status. The area under the ROC curve (AUC was improved to 0.83 compared to that of 0.66 for Gleason score and lymph node status alone.miRNA signatures can distinguish patients who fail soon after radical prostatectomy versus late failures, giving insight into which patients may need adjuvant therapy. Notably, two novel miRNAs (miR-4516 and miR-601 were identified that significantly improve prediction of biochemical failure post-salvage radiation therapy compared to clinico-histopathological factors, supporting the use of miRNAs within clinically used predictive models. Both findings warrant further

  8. Assessment of process control parameters in the biochemical methane potential of sunflower oil cake

    Energy Technology Data Exchange (ETDEWEB)

    Raposo, F.; Borja, R.; Rincon, B. [Instituto de la Grasa (CSIC), Avda. Padre Garcia Tejero 4, 41012 Seville (Spain); Jimenez, A.M. [Departamento de Ciencias Ambientales, Universidad Pablo de Olavide, Ctra. De Utrera, km 1, 41013 Sevilla (Spain)

    2008-12-15

    A laboratory-scale study was conducted on the batch anaerobic digestion of sunflower oil cake (SuOC), solid waste derived from the extraction process of sunflower oil. A multi-reactor system was used to compare methane production from this waste at inoculum to substrate ratios (ISRs) of 3.0, 2.0, 1.5, 1.0, 0.8 and 0.5 (expressed as volatile solids (VS) basis). The tests were carried out at mesophilic temperature (35 C) and run against a control of inoculum without substrate. The results obtained in the biochemical methane potential (BMP) tests showed that the ultimate methane yield (Y{sub M,ult}) decreased considerably from 227{+-}23 to 107{+-}11 ml CH{sub 4} at standard temperature and pressure (STP) conditions g{sup -1} VS{sub added} when the ISR decreased from 3.0 to 0.5, showing a clear influence of the ISR on the methane yield coefficient. The biodegradability (BD) of the waste also decreased from 86% to 41% when the ISR varied from 3.0 to 0.5. A net total ammonia nitrogen (TAN) yield of 39.2 mg N g{sup -1} VS{sub added} was obtained, and this value was not influenced by the ISRs assayed, which demonstrated the appropriate operation of the hydrolytic-acidogenic stage of the overall digestion process. A clear imbalance of the methanogenic process was observed at the lowest ISRs studied (0.5 and 0.8) due to a considerable increase in CODs and TVFA in the digestates. The profile of VFA was also influenced by the ISR, typical of the proteinaceous substrates. (author)

  9. Assessment of process control parameters in the biochemical methane potential of sunflower oil cake

    International Nuclear Information System (INIS)

    Raposo, F.; Borja, R.; Rincon, B.; Jimenez, A.M.

    2008-01-01

    A laboratory-scale study was conducted on the batch anaerobic digestion of sunflower oil cake (SuOC), solid waste derived from the extraction process of sunflower oil. A multi-reactor system was used to compare methane production from this waste at inoculum to substrate ratios (ISRs) of 3.0, 2.0, 1.5, 1.0, 0.8 and 0.5 (expressed as volatile solids (VS) basis). The tests were carried out at mesophilic temperature (35 deg. C) and run against a control of inoculum without substrate. The results obtained in the biochemical methane potential (BMP) tests showed that the ultimate methane yield (Y M,ult ) decreased considerably from 227±23 to 107±11 ml CH 4 at standard temperature and pressure (STP) conditions g -1 VS added when the ISR decreased from 3.0 to 0.5, showing a clear influence of the ISR on the methane yield coefficient. The biodegradability (BD) of the waste also decreased from 86% to 41% when the ISR varied from 3.0 to 0.5. A net total ammonia nitrogen (TAN) yield of 39.2 mg N g -1 VS added was obtained, and this value was not influenced by the ISRs assayed, which demonstrated the appropriate operation of the hydrolytic-acidogenic stage of the overall digestion process. A clear imbalance of the methanogenic process was observed at the lowest ISRs studied (0.5 and 0.8) due to a considerable increase in CODs and TVFA in the digestates. The profile of VFA was also influenced by the ISR, typical of the proteinaceous substrates

  10. Management Options for Biochemically Recurrent Prostate Cancer.

    Science.gov (United States)

    Fakhrejahani, Farhad; Madan, Ravi A; Dahut, William L

    2017-05-01

    Prostate cancer is the most common solid tumor malignancy in men worldwide. Treatment with surgery and radiation can be curative in organ-confined disease. Unfortunately, about one third of men develop biochemically recurrent disease based only on rising prostate-specific antigen (PSA) in the absence of visible disease on conventional imaging. For these patients with biochemical recurrent prostate cancer, there is no uniform guideline for subsequent management. Based on available data, it seems prudent that biochemical recurrent prostate cancer should initially be evaluated for salvage radiation or prostatectomy, with curative intent. In selected cases, high-intensity focused ultrasound and cryotherapy may be considered in patients that meet very narrow criteria as defined by non-randomized trials. If salvage options are not practical or unsuccessful, androgen deprivation therapy (ADT) is a standard option for disease control. While some patients prefer ADT to manage the disease immediately, others defer treatment because of the associated toxicity. In the absence of definitive randomized data, patients may be followed using PSA doubling time as a trigger to initiate ADT. Based on retrospective data, a PSA doubling time of less than 3-6 months has been associated with near-term development of metastasis and thus could be used signal to initiate ADT. Once treatment is begun, patients and their providers can choose between an intermittent and continuous ADT strategy. The intermittent approach may limit side effects but in patients with metastatic disease studies could not exclude a 20% greater risk of death. In men with biochemical recurrence, large studies have shown that intermittent therapy is non-inferior to continuous therapy, thus making this a reasonable option. Since biochemically recurrent prostate cancer is defined by technological limitations of radiographic detection, as new imaging (i.e., PSMA) strategies are developed, it may alter how the disease is

  11. Biochemical adaptation of camelids during periods where feed is withheld

    Directory of Open Access Journals (Sweden)

    J. Wensvoort

    2001-03-01

    Full Text Available Biochemical changes during fasting or the withholding of feed for 5 day were studied in serum of camelids (dromedary camel, llama and ruminants (sheep, steers. Camels maintained low levels of 13-hydroxybutyrate (BHB and high levels of glucose but showed some increased levels of non-esterified fatty acid (NEFA and urea when fasting. Sheep and steers showed a rise in serum BHB and much higher increases of NEFA than camels and llamas. Sheep showed decreased serum glucose. The llama showed some increase in BHB but NEFA was lower than the other three species. The results indicate that camelids have a unique ability to control lipolytic and gluconeogenic activity to prevent or postpone the state of ketosis. Understanding and manipulation of these metabolic mechanisms in cattle and sheep could have great benefit to the livestock industry.

  12. A quantitative assay to measure the interaction between immunogenic peptides and purified class I major histocompatibility complex molecules

    DEFF Research Database (Denmark)

    Olsen, A C; Pedersen, L O; Hansen, A S

    1994-01-01

    A direct and sensitive biochemical assay to measure the interaction in solution between peptides and affinity-purified major histocompatibility complex (MHC) class I molecules has been generated. Specific binding reflecting the known class I restriction of cytotoxic T cell responses was obtained...

  13. Suppressive effect of Spirulina fusiformis on diclofenac-induced hepato-renal injury and gastrointestinal ulcer in Wistar albino rats: A biochemical and histological approach.

    Science.gov (United States)

    Peter S, Jerine; Basha S, Kadar; Giridharan, R; Lavinya B, Udhaya; Sabina, Evan Prince

    2017-04-01

    The non-steroidal anti-inflammatory drug (NSAID), diclofenac causes hepato-renal toxicity and gastric ulcer. The aim of this study was to investigate the protective effect of Spirulina fusiformis on Diclofenac-induced toxicity in Wistar albino rats. Rats were treated as follows: normal control (group I); diclofenac (50mg/kgb.w., i.p.) treated rats (group II); diclofenac-induced (50mg/kgb.w., i.p.) rats treated with Spirulina fusiformis (400mg/kgb.w., p.o.) (group III); diclofenac-induced (50mg/kgb.w., i.p.) rats treated with silymarin (25mg/kgb.w., p.o.) (group IV); Spirulina fusiformis (400mg/kgb.w., p.o.) alone treated rats (groupV). Biochemical (liver and kidney functional markers) and antioxidant parameters (enzymic and non-enzymic antioxidants) were measured in the blood and tissue homogenates of the rats. Evaluation of intestinal ulcer score and assessment of liver and kidney histology were also done. Alterations in the levels of biochemical and antioxidant assays and histopathological changes in liver and kidney proved the toxic effect of diclofenac. The ulcer score was significantly increased in the diclofenac treated rats. Spirulina fusiformis showed to reduce such changes and was able to restore normal antioxidant status in the rats. Our study proves the hepato-renal and gastroprotective activity of Spirulina fusiformis in diclofenac-treated rats. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  14. Hepato-protective potential of methanol extract of leaf of Ziziphus mucronata (ZMLM) against dimethoate toxicity: biochemical and histological approach.

    Science.gov (United States)

    Kwape, T E; Chaturvedi, P; Kamau, J M; George, S

    2013-09-01

    The leaves of Ziziphus mucronata are used locally as food and a health drink; the leaf paste can also be used in the treatment of boils. The root of the plant is usually used in the treatment of a wide range of pains. The study was carried out to evaluate the hepatoprotective potential of the methanol leaf extract of Ziziphus mucronata (ZMLM). The extract was prepared by soaking in 70% methanol/water and rotary evaporation. The phenol content of extract was then estimated. Twenty five adult male Sprague dawley rats (aged 21 weeks) were divided into five groups of five rats each and treated as follows; normal control (NC) received distilled water. Dimethoate control (DC) (received 6 mg/kg.bw.day(-1) dimethoate dissolved in distilled water). Experimental Groups (E1) received dimethoate (6mg/kg.bw) + ZMLM (100 mg/kg.bw(-1)); (E2) received dimethoate (6 mg/kg.bw) + ZMLM (200 mg/kg.bw(-1)) and (E3) received dimethoate (6 mg/kg.bw) + ZMLM (300 mg/kg.bw(-1)). In both the cases a normal control and dimethoate control were kept to compare the results. After 90 days, blood was collected and rats were sacrificed to collect the liver tissue for biochemical assays and histological estimations. The results of E1 did not show much change from the normal control group but was significantly different from the dimethoate control group (P≤ 0.05). The preventive effect which was tested in E2 and E3 proved that the extract could almost retain the normal condition in 90 days time. Histological observations also agreed with the results obtained in biochemical parameters. Ziziphus mucronata methanol leaf extract possesses a preventive effect against dimethoate induced oxidative stress as observed in male albino Sprague Dawley rats.

  15. Problems with the PTH assays.

    Science.gov (United States)

    Cavalier, Etienne; Delanaye, Pierre; Nyssen, Laurent; Souberbielle, Jean-Claude

    2015-05-01

    Even if the first assay for parathyroid hormone (PTH) was published in the early 1960s, its determination remains a challenge even today. Indeed, in the circulation, PTH is present in its active form (PTH 1-84), but many PTH fragments can also be present. These fragments accumulate when renal function declines and are recognized, at different extents, by the 2nd generation ("intact") PTH assays that are widely used in the clinical laboratories. Some assays, called "3rd generation PTH" do not recognize these fragments, but are not available everywhere. Hence, different problems are also linked with PTH determination. Among them, one can cite the lack of a reference method, the lack of standardization of the assays and, sometimes, the lack of consistent reference range. We can also point out stability problems and a large intra-individual variation. A workgroup is working on these problems under the auspices of the IFCC and we hope that some of these problems will be resolved in the next years. In this article, we will discuss all the possible issues of PTH determination. Copyright © 2015. Published by Elsevier Masson SAS.

  16. semen by MTT reduction assay

    African Journals Online (AJOL)

    user

    the concentration of sperm in each semen sample, sperm motility, plasma integrity of sperm in terms of hypo-osmotic swelling (HOS) test, live and dead ratio of sperm and MTT reduction assay of each ejaculate were determined. Plasma membrane integrity of fresh sperm was assessed using a hypo-osmotic swelling (HOS) ...

  17. Biochemical evaluation of antioxidant activity and polysaccharides fractions in seaweeds

    Directory of Open Access Journals (Sweden)

    A. Tariq

    2015-01-01

    Full Text Available In the present study ethanol and water extracts of 15 seaweeds, Dictyota dichotoma var. velutricata, Dictyota indica, Iyengaria stellata, Padina pavonia, Sargassum swartzii, Sargassum variegatum, Stoechospermum marginatum, Stokeyia indica, Jolyna laminarioides, Caulerpa taxifolia, Halimeda tuna, Ulva fasciata, Ulva lactuca, Solieria robusta, and Melanothamnus afaqhusainii, were evaluated for their antioxidant potential by ABTS or 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid, superoxide and total antioxidant capacity (TAC assays.  The activity was concentration dependent and the variation in antioxidant potential was also observed by different assays in both extracts.  Ethanol extract of D. dichotoma var. velutricata, D. indica and S. marginatum demonstrated highest activity by TAC assay.  The antioxidant potential in organic solvent fractions of seaweeds namely P. pavonia, S. swartzii, S. marginatum and M. afaqhusainii was also determined and chloroform fraction of all the four seaweeds showed highest activity by superoxide assay.  Antioxidant activity of extracted fractions of polysaccharides from S. indica, C. taxifolia and D. dichotoma var. velutricata was also evaluated by superoxide method.  Polysaccharide fractions of S. indica obtained from HCl (at 700C and room temperature and water extract demonstrated highest activity respectively.  All the polysaccharide fractions of C. taxifolia showed excellent activity except CaClF70°C. Polysaccharide fractions of D. dichotoma var. velutricata also exhibited very good activity.

  18. High frequency lateral flow affinity assay using superparamagnetic nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Lago-Cachón, D., E-mail: dlagocachon@gmail.com [Dpto. de Física, Universidad de Oviedo, Edificio Departamental Este, Campus de Viesques, 33204 Gijón (Spain); Rivas, M., E-mail: rivas@uniovi.es [Dpto. de Física, Universidad de Oviedo, Edificio Departamental Este, Campus de Viesques, 33204 Gijón (Spain); Martínez-García, J.C., E-mail: jcmg@uniovi.es [Dpto. de Física, Universidad de Oviedo, Edificio Departamental Este, Campus de Viesques, 33204 Gijón (Spain); Oliveira-Rodríguez, M., E-mail: oliveiramyriam@uniovi.es [Dpto. de Química Física y Analítica, Universidad de Oviedo, C/Julián Clavería 8, 33006 Oviedo (Spain); Blanco-López, M.C., E-mail: cblanco@uniovi.es [Dpto. de Química Física y Analítica, Universidad de Oviedo, C/Julián Clavería 8, 33006 Oviedo (Spain); García, J.A., E-mail: joseagd@uniovi.es [Dpto. de Física, Universidad de Oviedo, Escuela de Marina, Campus de Viesques, 33204 Gijón (Spain)

    2017-02-01

    Lateral flow assay is one of the simplest and most extended techniques in medical diagnosis for point-of-care testing. Although it has been traditionally a positive/negative test, some work has been lately done to add quantitative abilities to lateral flow assay. One of the most successful strategies involves magnetic beads and magnetic sensors. Recently, a new technique of superparamagnetic nanoparticle detection has been reported, based on the increase of the impedance induced by the nanoparticles on a RF-current carrying copper conductor. This method requires no external magnetic field, which reduces the system complexity. In this work, nitrocellulose membranes have been installed on the sensor, and impedance measurements have been carried out during the sample diffusion by capillarity along the membrane. The impedance of the sensor changes because of the presence of magnetic nanoparticles. The results prove the potentiality of the method for point-of-care testing of biochemical substances and nanoparticle capillarity flow studies. - Highlights: • A method for quantification of Lateral Flow Assays is proposed. • MNP induce an increase of the impedance on a RF-current carrying copper sensor. • Magnetic nanoparticles (MNP) can be detected flowing over the sensing element.

  19. Biochemical changes during aging of soybean seed

    Directory of Open Access Journals (Sweden)

    Balešević-Tubić Svetlana

    2009-01-01

    Full Text Available Biochemical changes that occur in the seed as a result of ageing are very significant for seed quality and longevity. Because of its characteristic composition, processes occurring in the seed of oil crops during storage will be typical as well. Six soybean varieties developed in Institute of field and vegetable crops Novi Sad, submitted to accelerated and natural aging, under controlled and conventional storage conditions were used in these trials. The content of malondialdehyde, superoxide dismutase and peroxidase activities were studied. The biochemical processes i.e. lipid peroxidation, as well as the decrease in supeoxide dismutase and peroxidase activities (especially pronounced by applied accelerated aging were caused by both type of aging. The degree of seed damage and the ability of seed to resist the negative consequences of aging were influenced, beside duration of aging period, by type of storage and characteristics of soybean varieties. .

  20. Design of radiation dose tumor response assays

    International Nuclear Information System (INIS)

    Suit, H.D.; Hwang, T.; Hsieh, C.; Thames, H.

    1985-01-01

    The efficient utilization of animals in a radiation dose response assay for tumor control requires a definition of the goal, e.g., TCD50 or slope. A series of computer modelled ''experiments'' have been performed for each of a number of allocations of dose levels (DL) and number of animals/DL. The authors stipulated that the assumed TCD50 was .85 of true value; assumed slope was correct. They stipulated a binominal distribution of observed tumor control results at each dose level. A pilot assay used 6 tumors at 7 DL (from TCD1-TCD97). The second assay used 30 tumors assigned to 2,3,5 or 9 DL and to selected tumor control probabilities (TCP derived from the pilot run. Results from 100 test runs were combined with the pilot run for each of the combination of DL and TCP values. Logit regression lines were fitted through these ''data'' and the 95% CL around the TCD50 and the TCD37 values and the variances of the slopes were computed. These experiments were repeated using the method suggested by Porter (1980). Results show that a different strategy is needed depending upon the goal, viz. TCD50 or TCD37 vs slope. The differences between the two approaches are discussed

  1. Automation of the dicentric chromosome assay and related assays

    International Nuclear Information System (INIS)

    Balajee, Adayabalam S.; Dainiak, Nicholas

    2016-01-01

    Dicentric Chromosome Assay (DCA) is considered to be the 'gold standard' for personalized dose assessment in humans after accidental or incidental radiation exposure. Although this technique is superior to other cytogenetic assays in terms of specificity and sensitivity, its potential application to radiation mass casualty scenarios is highly restricted because DCA is time consuming and labor intensive when performed manually. Therefore, it is imperative to develop high throughput automation techniques to make DCA suitable for radiological triage scenarios. At the Cytogenetic Biodosimetry Laboratory in Oak Ridge, efforts are underway to develop high throughput automation of DCA. Current status on development of various automated cytogenetic techniques in meeting the biodosimetry needs of radiological/nuclear incident(s) will be discussed

  2. Correlation between 31P NMR phosphomonoester and biochemically determined phosphorylethanolamine and phosphatidylethanolamine during development of the rat brain.

    Science.gov (United States)

    Burri, R; Lazeyras, F; Aue, W P; Straehl, P; Bigler, P; Althaus, U; Herschkowitz, N

    1988-01-01

    Phosphomonoesters were measured in the developing rat brain by in vivo and in vitro 31P nuclear magnetic resonance (NMR) spectroscopy and by classical biochemical methods. In vitro NMR showed that the main component of the phosphomonoester peak is phosphorylethanolamine. Phosphomonoesters measured by in vivo NMR decreased during development at the same rate as the biochemically estimated phosphorylethanolamine. Phosphorylethanolamine, a precursor of the membrane lipid phosphatidylethanolamine, decreased during development parallel to an increase of the lipid phosphatidylethanolamine, which was measured biochemically. These studies show that 31P NMR can be used to monitor brain development in vivo.

  3. Growth-based determination and biochemical confirmation of genetic requirements for protein degradation in Saccharomyces cerevisiae.

    Science.gov (United States)

    Watts, Sheldon G; Crowder, Justin J; Coffey, Samuel Z; Rubenstein, Eric M

    2015-02-16

    Regulated protein degradation is crucial for virtually every cellular function. Much of what is known about the molecular mechanisms and genetic requirements for eukaryotic protein degradation was initially established in Saccharomyces cerevisiae. Classical analyses of protein degradation have relied on biochemical pulse-chase and cycloheximide-chase methodologies. While these techniques provide sensitive means for observing protein degradation, they are laborious, time-consuming, and low-throughput. These approaches are not amenable to rapid or large-scale screening for mutations that prevent protein degradation. Here, a yeast growth-based assay for the facile identification of genetic requirements for protein degradation is described. In this assay, a reporter enzyme required for growth under specific selective conditions is fused to an unstable protein. Cells lacking the endogenous reporter enzyme but expressing the fusion protein can grow under selective conditions only when the fusion protein is stabilized (i.e. when protein degradation is compromised). In the growth assay described here, serial dilutions of wild-type and mutant yeast cells harboring a plasmid encoding a fusion protein are spotted onto selective and non-selective medium. Growth under selective conditions is consistent with degradation impairment by a given mutation. Increased protein abundance should be biochemically confirmed. A method for the rapid extraction of yeast proteins in a form suitable for electrophoresis and western blotting is also demonstrated. A growth-based readout for protein stability, combined with a simple protocol for protein extraction for biochemical analysis, facilitates rapid identification of genetic requirements for protein degradation. These techniques can be adapted to monitor degradation of a variety of short-lived proteins. In the example presented, the His3 enzyme, which is required for histidine biosynthesis, was fused to Deg1-Sec62. Deg1-Sec62 is targeted for

  4. Multidimensional biochemical information processing of dynamical patterns.

    Science.gov (United States)

    Hasegawa, Yoshihiko

    2018-02-01

    Cells receive signaling molecules by receptors and relay information via sensory networks so that they can respond properly depending on the type of signal. Recent studies have shown that cells can extract multidimensional information from dynamical concentration patterns of signaling molecules. We herein study how biochemical systems can process multidimensional information embedded in dynamical patterns. We model the decoding networks by linear response functions, and optimize the functions with the calculus of variations to maximize the mutual information between patterns and output. We find that, when the noise intensity is lower, decoders with different linear response functions, i.e., distinct decoders, can extract much information. However, when the noise intensity is higher, distinct decoders do not provide the maximum amount of information. This indicates that, when transmitting information by dynamical patterns, embedding information in multiple patterns is not optimal when the noise intensity is very large. Furthermore, we explore the biochemical implementations of these decoders using control theory and demonstrate that these decoders can be implemented biochemically through the modification of cascade-type networks, which are prevalent in actual signaling pathways.

  5. Multidimensional biochemical information processing of dynamical patterns

    Science.gov (United States)

    Hasegawa, Yoshihiko

    2018-02-01

    Cells receive signaling molecules by receptors and relay information via sensory networks so that they can respond properly depending on the type of signal. Recent studies have shown that cells can extract multidimensional information from dynamical concentration patterns of signaling molecules. We herein study how biochemical systems can process multidimensional information embedded in dynamical patterns. We model the decoding networks by linear response functions, and optimize the functions with the calculus of variations to maximize the mutual information between patterns and output. We find that, when the noise intensity is lower, decoders with different linear response functions, i.e., distinct decoders, can extract much information. However, when the noise intensity is higher, distinct decoders do not provide the maximum amount of information. This indicates that, when transmitting information by dynamical patterns, embedding information in multiple patterns is not optimal when the noise intensity is very large. Furthermore, we explore the biochemical implementations of these decoders using control theory and demonstrate that these decoders can be implemented biochemically through the modification of cascade-type networks, which are prevalent in actual signaling pathways.

  6. The use of microorganisms with broad range substrate utilisation for the ferricyanide-mediated rapid determination of biochemical oxygen demand.

    Science.gov (United States)

    Catterall, K; Morris, K; Gladman, C; Zhao, H; Pasco, N; John, R

    2001-12-24

    The feasibility of replacing oxygen with a synthetic electron acceptor in microbial catabolism was investigated as a rapid method for the determination of biochemical oxygen demand (BOD). Microorganisms known for their broad range organic substrate utilisation were investigated. It was shown that Trichosporon cutaneum, Pseudomonas putida and Bacillus licheniformis could utilize the ferricyanide ion as an alternative electron acceptor, in place of oxygen, for the catabolic oxidation of a range of simple organic compounds. The biochemical reactions were monitored by measuring the amount of microbially produced ferrocyanide using amperometry at a Pt disk microelectrode. Catabolic degradation efficiencies approaching those of the conventional 5-day assay were achieved in 1 h. BOD(5) equivalent values for a range of simple organic solutions were determined for each of the microorganisms. The effect of increased incubation time and the choice of appropriate calibration standards for rapid BOD assays were also considered.

  7. A radioreceptor assay for catecholamines, 2

    International Nuclear Information System (INIS)

    Ohmori, Yoshiaki; Nakano, Ryuichi; Lie, Shozo; Imura, Hiroo; Shimbo, Shinichiro.

    1980-01-01

    We have already reported on a radioreceptor assay for catecholamines utilizing the microsome fraction of bovine myocardium as a catecholamine (CA) receptor and 3 H-norepinephrine as a labelled CA. In order to increase the sensitivity of the radioreceptor assay, we used 4-(2-iodoethyl) pyrocatechol ( 125 I-CA) as a ligand instead of 3 H-norepinephrine and performed a radioreceptor assay for CA. The following results were obtained: 1) 125 I-CA was able to bind alpha-receptors prepared from bovine myocardium. 2) The optimal amount of the microsomal fraction was 250 μg/tube, when 125 I-CA of 50,000 c.p.m. was used. The appropriate conditions for incubation were 90 minutes at 20 0 C in a pH 7.0 sucrose solution. 3) By this method utilizing 125 I-CA, norepinephrine was detectable in a range from 500 pg to 10 ng/tube. 4) Various compounds with a catechol nucleus showed cross-reaction in this radioreceptor assay system. 5) Whereas beta-adrenergic blocking agents did not inhibit the binding of 125 I-CA, phentholamine, a short acting type of alpha-adrenergic blocking agents, was effective in inhibiting the binding. However, dibenamine and phenoxybenzamine, long acting types of alpha-adrenergic blocking agents, increased the binding of 125 I-CA to the microsomal fraction. 6) Utilizing this phenomenon, norepinephrine was detectable in the range from 100 pg to 5 ng/tube. (author)

  8. Biological and Biochemical Potential of Sea Snake Venom and Characterization of Phospholipase A2 and Anticoagulation Activity.

    Science.gov (United States)

    Damotharan, Palani; Veeruraj, Anguchamy; Arumugam, Muthuvel; Balasubramanian, Thangavel

    2016-03-01

    This study is designed to isolate and purify a novel anti-clotting protein component from the venom of Enhydrina schistosa, and explore its biochemical and biological activities. The active protein was purified from the venom of E. schistosa by ion-exchange chromatography using DEAE-cellulose. The venom protein was tested by various parameters such as, proteolytic, haemolytic, phospholipase and anti-coagulant activities. 80 % purity was obtained in the final stage of purification and the purity level of venom was revealed as a single protein band of about 44 kDa in SDS-polyacrylamide electrophoresis under reducing conditions. The results showed that the Potent hemolytic activity was observed against cow, goat, chicken and human (A, B and O positive) erythrocytes. Furthermore, the clotting assays showed that the venom of E. schistosa significantly prolonged in activated partial thromboplastin time, thrombin time, and prothrombin time. Venomous enzymes which hydrolyzed casein and gelatin substrate were found in this venom protein. Gelatinolytic activity was optimal at pH 5-9 and (1)H NMR analysis of purified venom was the base line information for the structural determination. These results suggested that the E. schistosa venom holds good promise for the development of novel lead compounds for pharmacological applications in near future.

  9. Anaerobic co-digestion of commercial food waste and dairy manure: Characterizing biochemical parameters and synergistic effects.

    Science.gov (United States)

    Ebner, Jacqueline H; Labatut, Rodrigo A; Lodge, Jeffrey S; Williamson, Anahita A; Trabold, Thomas A

    2016-06-01

    Anaerobic digestion of commercial food waste is a promising alternative to landfilling commercial food waste. This study characterized 11 types of commercial food wastes and 12 co-digestion blends. Bio-methane potential, biodegradable fraction, and apparent first-order hydrolysis rate coefficients were reported based upon biochemical methane potential (BMP) assays. Food waste bio-methane potentials ranged from 165 to 496 mL CH4/g VS. Substrates high in lipids or readily degradable carbohydrates showed the highest methane production. Average bio-methane potential observed for co-digested substrates was -5% to +20% that of the bio-methane potential of the individual substrates weighted by VS content. Apparent hydrolysis rate coefficients ranged from 0.19d(-1) to 0.65d(-1). Co-digested substrates showed an accelerated apparent hydrolysis rate relative to the weighted average of individual substrate rates. These results provide a database of key bio-digestion parameters to advance modeling and utilization of commercial food waste in anaerobic digestion. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Biochemical and Molecular Study of Carpobrotus edulis Bioactive Properties and Their Effects on Dugesia sicula (Turbellaria, Tricladida) Regeneration.

    Science.gov (United States)

    Meddeb, Emna; Charni, Mohamed; Ghazouani, Tesnime; Cozzolino, Autilia; Fratianni, Florinda; Raboudi, Faten; Nazzaro, Filomena; Fattouch, Sami

    2017-07-01

    The traditional medicinal properties of Carpobrotus edulis are well recognized, particularly in Tunisia where it is used for wound healing. Thus, in this study, biochemical and molecular properties of its leaves' bioactive aqueous-acetone extract were investigated. The total phenolic content (TPC) of the extract was estimated to be 184 ± 5 mg/100 g of fresh matter (FM). The qualitative and quantitative polyphenolic profile was determined by ultra performance liquid chromatography with diode array detection (UPLC-DAD) and showed that chlorogenic acid was the major compound (43.7%). The extract exhibits potent antioxidant capacities with IC50 = 56.19 and 58.91 μg/ml, as accessed via the anionic DPPH and cationic ABTS radical scavenging assays, respectively. The extract has high antibacterial properties, especially against the Gram+ Staphylococcus aureus and Bacillus cereus strains. To investigate the extract effect on regeneration, the flatworm Dugesia sicula Lepori, 1948, was used as a model. The macroscopic analysis of planarian cultures in ordinary medium containing phenolic extract at non-toxic concentrations illustrated that the extract caused morphological changes. Additionally, the molecular study through the fluorescence-activated cell sorting (FACS) technique showed that C. edulis polyphenols can harm the stem cells' development. These results emphasize the ecotoxicological impact of phenolic rejections in the environment on flatworms' physiology.

  11. Structural and Biochemical Characterization of Xylella fastidiosa DsbA Family Members: New insightsinto the Enzyme-Substrate Interaction

    Energy Technology Data Exchange (ETDEWEB)

    Rinaldi, F.; Meza, A; Gulmarges, B

    2009-01-01

    Disulfide oxidoreductase DsbA catalyzes disulfide bond formation in proteins secreted to the periplasm and has been related to the folding process of virulence factors in many organisms. It is among the most oxidizing of the thioredoxin-like proteins, and DsbA redox power is understood in terms of the electrostatic interactions involving the active site motif CPHC. The plant pathogen Xylella fastidiosa has two chromosomal genes encoding two oxidoreductases belonging to the DsbA family, and in one of them, the canonical motif CPHC is replaced by CPAC. Biochemical assays showed that both X. fastidiosa homologues have similar redox properties and the determination of the crystal structure of XfDsbA revealed substitutions in the active site of X. fastidiosa enzymes, which are proposed to compensate for the lack of the conserved histidine in XfDsbA2. In addition, electron density maps showed a ligand bound to the XfDsbA active site, allowing the characterization of the enzyme interaction with an 8-mer peptide. Finally, surface analysis of XfDsbA and XfDsbA2 suggests that X. fastidiosa enzymes may have different substrate specificities.

  12. BIOCHEMICAL AND BIOPHYSICAL CHARACTERIZATION OF RECOMBINANT YEAST PROTEASOME MATURATION FACTOR UMP1

    Directory of Open Access Journals (Sweden)

    Bebiana Sá-Moura

    2013-04-01

    We show that recombinant Ump1 is purified as a mixture of different oligomeric species and thatoligomerization is mediated by intermolecular disulfide bond formation involving the only cysteine residue present in the protein.Furthermore, a combination of bioinformatic, biochemical and structural analysis revealed that Ump1 shows characteristics of anintrinsically disordered protein, which might become structured only upon interaction with the proteasomesubunits.

  13. Assay strategies and methods for phospholipases

    International Nuclear Information System (INIS)

    Reynolds, L.J.; Washburn, W.N.; Deems, R.A.; Dennis, E.A.

    1991-01-01

    Of the general considerations discussed, the two issues which are most important in choosing an assay are (1) what sensitivity is required to assay a particular enzyme and (2) whether the assay must be continuous. One can narrow the options further by considering substrate availability, enzyme specificity, assay convenience, or the presence of incompatible side reactions. In addition, the specific preference of a particular phospholipase for polar head group, micellar versus vesicular substrates, and anionic versus nonionic detergents may further restrict the options. Of the many assays described in this chapter, several have limited applicability or serious drawbacks and are not commonly employed. The most commonly used phospholipase assays are the radioactive TLC assay and the pH-stat assay. The TLC assay is probably the most accurate, sensitive assay available. These aspects often outweigh the disadvantages of being discontinuous, tedious, and expensive. The radioactive E. coli assay has become popular recently as an alternative to the TLC assay for the purification of the mammalian nonpancreatic phospholipases. The assay is less time consuming and less expensive than the TLC assay, but it is not appropriate when careful kinetics are required. Where less sensitivity is needed, or when a continuous assay is necessary, the pH-stat assay is often employed. With purified enzymes, when free thiol groups are not present, a spectrophotometric thiol assay can be used. This assay is ∼ as sensitive as the pH-stat assay but is more convenient and more reproducible, although the substrate is not available commercially. Despite the many assay choices available, the search continues for a convenient, generally applicable assay that is both sensitive and continuous

  14. 21 CFR 225.158 - Laboratory assays.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 4 2010-04-01 2010-04-01 false Laboratory assays. 225.158 Section 225.158 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) DRUGS... Laboratory assays. Where the results of laboratory assays of drug components, including assays by State feed...

  15. 40 CFR 158.2000 - Biochemical pesticides definition and applicability.

    Science.gov (United States)

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Biochemical pesticides definition and applicability. 158.2000 Section 158.2000 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2000 Biochemical pesticides...

  16. Biochemical analysis of the crude extract of Momordica charantia (L.).

    Science.gov (United States)

    Dar, Ume Kalsoom; Owais, Farah; Ahmad, Manzoor; Rizwani, Ghazala H

    2014-11-01

    Momordica charantia (L.) commonly referred as bitter gourd, karela and balsam pear. Its fruit is used for the treatment of diabetes and related conditions amongst the indigenous populations of Asia, South America, India and East Africa. The study was conducted to find out the biochemical aspects of crude extract of whole fruit of M. charantia including seeds which includes blood test (Hemoglobin, RBC, Total leukocyte count, platelets count, HbA1C (Glycocylated heamoglobin Type A1C)), Lipid profile test and electrolyte balance. Hemoglobin (7.1±0.14), platelets count (827 ×109±1.95), Cholesterol level (111±2), HDL (high density lipoproteins) (20±1.22) at 10mg shows marked increase in values as compared to control. While 25 mg dose shows insignificant result. Electrolyte balance are found significant at 10mg and 25mg except bicarbonates (Na(+¬)=143±1.87, K-=3.45±0.35, Cl(-) =108±1.48). Another important property of M. charantia is the elevation of platelet counts, heamoglobin and specifically high-density lipoproteins (HDL). It also controls cholesterol, triglycerides, HDL, LDL and VLDL at low dosage (10mg). Further studies can be conducted to find out which phytochemical components acts on specific biochemical activity.

  17. A Sensitive Chemotaxis Assay Using a Novel Microfluidic Device

    Directory of Open Access Journals (Sweden)

    Chen Zhang

    2013-01-01

    Full Text Available Existing chemotaxis assays do not generate stable chemotactic gradients and thus—over time—functionally measure only nonspecific random motion (chemokinesis. In comparison, microfluidic technology has the capacity to generate a tightly controlled microenvironment that can be stably maintained for extended periods of time and is, therefore, amenable to adaptation for assaying chemotaxis. We describe here a novel microfluidic device for sensitive assay of cellular migration and show its application for evaluating the chemotaxis of smooth muscle cells in a chemokine gradient.

  18. Radioimmunoassay and radioenzymatic assay of a new aminoglycoside antibiotic, netilmicin

    International Nuclear Information System (INIS)

    Broughton, A.; Strong, J.E.; Pickering, L.K.; Knight, J.; Bodey, G.P.

    1978-01-01

    A radioimmunoassay and a radioenzymatic assay for netilmicin, a new aminoglycoside, were developed in our laboratories to assist in the study of the pharmacology of the drug and establish values for use in its monitoring. The assays are sensitive, precise, and rapid, giving results that correlate (r = 0.90) with each other and with those of a microbiological assay in which Klebsiella pneumoniae is used as the test organism. Preliminary pharmacological studies show the drug to have a biological half-life of 135 min, which is comparable to that for other aminoglycosides

  19. Biochemical methane potential of kraft bleaching effluent and codigestion with other in-mill streams

    DEFF Research Database (Denmark)

    Fitamo, Temesgen Mathewos; Dahl, Olli; Master, Emma

    2016-01-01

    A biochemical methane potential assay was conducted to investigate the anaerobic digestibility of bleaching effluent from hardwood kraft pulping and the potential of codigestion with other effluents from an integrated pulp and paper mill. Four in-mill streams were tested individually...... degradation and methane generation. Chemical oxygen demand (COD) removal ranged from 57%-76%, and methane generation was 220-280 mL/g COD contained in the wastewater, depending on the degree of dilution. When codigestion was tested, the composite consisting of total bleaching effluent, chemithermomechanical...

  20. Rapid biochemical methane potential prediction of urban organic waste with near-infrared reflectance spectroscopy

    DEFF Research Database (Denmark)

    Fitamo, Temesgen Mathewos; Triolo, Jin Mi; Boldrin, Alessio

    2017-01-01

    The anaerobic digestibility of various biomass feedstocks in biogas plants is determined with biochemical methane potential (BMP) assays. However, experimental BMP analysis is time-consuming, costly and challenging to optimise stock management and feeding to achieve improved biogas production....... The aim of the present study is to develop a fast and reliable model based on near-infrared reflectance spectroscopy (NIRS) for the BMP prediction of urban organic waste (UOW). The model comprised 87 UOW samples. Additionally, 88 plant biomass samples were included, to develop a combined model predicting...

  1. Clinico-hemato-biochemical profile of dogs with liver cirrhosis

    Directory of Open Access Journals (Sweden)

    M. A. Elhiblu

    2015-04-01

    Full Text Available Aim: The aim of this study was to determine the relevant tools in the diagnosis of liver cirrhosis in dogs. Material and Methods: A total of 140 dogs presented at Veterinary Teaching Hospital, Guru Angad Dev Veterinary and Animal Sciences University, Ludhiana, showing clinical signs of hepatic insufficiency were subjected to clinico-hemato biochemical, urological, ultrasonographic (USG, and USG guided fine-needle biopsy examinations by standard methods. On the basis of these results, 6 dogs out of 140 dogs were found to be suffering from liver cirrhosis. Six clinically healthy dogs constituted the control group. Results: The dogs suffering from liver cirrhosis manifested inappetence, halitosis, abdominal distension, weight loss, melena, icterus, anemia, and neutrophilic leukocytosis with the left shift. Levels of hemoglobin, lymphocytes, packed cell volume, mean corpuscular volume, mean corpuscular Hb (MCH, and platelet count were significantly lower in liver cirrhosis group than control group while total leukocyte count, neutrophils, and MCH concentration were significantly higher. Glucose, total protein, albumin, A/G ratio, and fibrinogen were significantly lower, and creatinine, alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, prothrombin time, and APTT were significantly higher than the control values. Ultrasound revealed diffuse increase in echogenicity with rounded and irregular liver margins. Cytological examination of the ascitic fluid and fine-needle aspiration biopsy of liver was not fruitful in the diagnosis of liver cirrhosis. Conclusions: Liver cirrhosis causes clinical and hemo-biochemical alterations, which require special consideration when treating diseased animals. USG, diffuse increase in echogenicity of liver, rounding and irregularity of liver margins and microhepatica were the consistent findings. It is suggested that USG along with hemo-biochemical alterations may be used as a diagnostic tool for

  2. Nanostar Clustering Improves the Sensitivity of Plasmonic Assays.

    Science.gov (United States)

    Park, Yong Il; Im, Hyungsoon; Weissleder, Ralph; Lee, Hakho

    2015-08-19

    Star-shaped Au nanoparticles (Au nanostars, AuNS) have been developed to improve the plasmonic sensitivity, but their application has largely been limited to single-particle probes. We herein describe a AuNS clustering assay based on nanoscale self-assembly of multiple AuNS and which further increases detection sensitivity. We show that each cluster contains multiple nanogaps to concentrate electric fields, thereby amplifying the signal via plasmon coupling. Numerical simulation indicated that AuNS clusters assume up to 460-fold higher field density than Au nanosphere clusters of similar mass. The results were validated in model assays of protein biomarker detection. The AuNS clustering assay showed higher sensitivity than Au nanosphere. Minimizing the size of affinity ligand was found important to tightly confine electric fields and improve the sensitivity. The resulting assay is simple and fast and can be readily applied to point-of-care molecular detection schemes.

  3. Biochemical and pharmacological characterization of the human lymphocyte antigen B-associated transcript 5 (BAT5/ABHD16A.

    Directory of Open Access Journals (Sweden)

    Juha R Savinainen

    Full Text Available Human lymphocyte antigen B-associated transcript 5 (BAT5, also known as ABHD16A is a poorly characterized 63 kDa protein belonging to the α/β-hydrolase domain (ABHD containing family of metabolic serine hydrolases. Its natural substrates and biochemical properties are unknown.Amino acid sequence comparison between seven mammalian BAT5 orthologs revealed that the overall primary structure was highly (≥95% conserved. Activity-based protein profiling (ABPP confirmed successful generation of catalytically active human (h and mouse (m BAT5 in HEK293 cells, enabling further biochemical characterization. A sensitive fluorescent glycerol assay reported hBAT5-mediated hydrolysis of medium-chain saturated (C14:0, long-chain unsaturated (C18:1, C18:2, C20:4 monoacylglycerols (MAGs and 15-deoxy-Δ12,14-prostaglandin J2-2-glycerol ester (15d-PGJ2-G. In contrast, hBAT5 possessed only marginal diacylglycerol (DAG, triacylglycerol (TAG, or lysophospholipase activity. The best MAG substrates were 1-linoleylglycerol (1-LG and 15d-PGJ2-G, both exhibiting low-micromolar Km values. BAT5 had a neutral pH optimum and showed preference for the 1(3- vs. 2-isomers of MAGs C18:1, C18:2 and C20:4. Inhibitor profiling revealed that β-lactone-based lipase inhibitors were nanomolar inhibitors of hBAT5 activity (palmostatin B > tetrahydrolipstatin > ebelactone A. Moreover, the hormone-sensitive lipase inhibitor C7600 (5-methoxy-3-(4-phenoxyphenyl-3H-[1], [3], [4]oxadiazol-2-one was identified as a highly potent inhibitor (IC50 8.3 nM. Phenyl and benzyl substituted analogs of C7600 with increased BAT5 selectivity were synthesized and a preliminary SAR analysis was conducted to obtain initial insights into the active site dimensions.This study provides an initial characterization of BAT5 activity, unveiling the biochemical and pharmacological properties with in vitro substrate preferences and inhibitor profiles. Utilization of glycerolipid substrates and sensitivity to

  4. Histological vis-a-vis biochemical assessment on the toxic level and antineoplastic efficacy of a synthetic drug Pt-ATP on experimental animal models.

    Science.gov (United States)

    Pal, Shipra; Sadhu, Arpita Sengupta; Patra, Swarup; Mukherjea, Kalyan K

    2008-11-12

    Cisplatin, a platinum based anticancer drug has played a vital role in the treatment of cancers by chemical agents, but in view of the serious toxicity including nephrotoxicity of cisplatin, various other platinum based drugs have been synthesized and screened to overcome its toxicity. A Pt-ATP compound was prepared in our laboratory hoping to have reduced or no toxicity along with the potentiality of reducing neoplasm growth. A Pt-ATP compound was prepared. It was first screened for its antineoplastic efficacy. Confirming that, subsequent experiments were carried on to test its toxicity on animals, viz. Albino Swiss mice. The animals were randomly divided into four sets--Set I: Erhlich Ascites Carcinoma (EAC) challenged mice; Set II: Normal mice; Set III: Drug treated mice, Set IVA Cisplatin (CDDP) treated mice, Set IV B EAC challenged Cisplatin treated mice. Set I was used to test antineoplasticity of the drug, Set II and Set III for studying drug toxicity and Set IV was treated with CDDP. Set II was used as a control. Animals were sacrificed after 5 days, 10 days 15 days and 20 days of drug administration on the 6th, 11th, 16th and 21st days respectively for Set I, II and III. Set IVA was sacrificed only on the 16th day and Set IV B on 6th and 11th days. For Set I only tumor cell count and packed cell volume (PCV) of tumor cells were recorded. For Set II and III, aspartate aminotransferase (AST), alanine aminotransferase (ALT) assays were done using serum while blood creatinine and creatine were assayed from blood filtrate. For cytotoxicity assessment liver, spleen and kidney tissues were collected and subjected to scanning electron microscopy (SEM) after extensive treatment. Set IV A was only studied for the biochemical parameters viz. aspartate aminotransferase (AST), alanine aminotransferase (ALT) assays were done using serum while blood creatinine and creatine were assayed from blood filtrate. Set IV B was studied for tumor cell count after treatment with

  5. Assay of vitamin B12

    International Nuclear Information System (INIS)

    Tovey, K.C.; Carrick, D.T.

    1982-01-01

    A radioassay is described for vitamin B12 which involves denaturing serum protein binding proteins with alkali. In the denaturation step a dithiopolyol and cyanide are used and in the intrinsic factor assay step a vitamin B12 analogue such as cobinamide is used to bind with any remaining serum proteins. The invention also includes a kit in which the dithiopolyol is provided in admixture with the alkali. The dithiopolyol may be dithiothreitol or dithioerythritol. (author)

  6. Development of a loop-mediated Isothermal amplification assay for sensitive and rapid detection of Vibrio parahaemolyticus

    Directory of Open Access Journals (Sweden)

    Kawahara Ryuji

    2008-09-01

    Full Text Available Abstract Background Vibrio parahaemolyticus is a marine seafood-borne pathogen causing gastrointestinal disorders in humans. Thermostable direct hemolysin (TDH and TDH-related hemolysin (TRH are known as major virulence determinants of V. parahaemolyticus. Most V. parahaemolyticus isolates from the environment do not produce TDH or TRH. Total V. parahaemolyticus has been used as an indicator for control of seafood contamination toward prevention of infection. Detection of total V. parahaemolyticus using conventional culture- and biochemical-based assays is time-consuming and laborious, requiring more than three days. Thus, we developed a novel and highly specific loop-mediated isothermal amplification (LAMP assay for the sensitive and rapid detection of Vibrio parahaemolyticus. Results The assay provided markedly more sensitive and rapid detection of V. parahaemolyticus strains than conventional biochemical and PCR assays. The assay correctly identified 143 V. parahaemolyticus strains, but did not detect 33 non-parahaemolyticus Vibrio and 56 non-Vibrio strains. Sensitivity of the LAMP assay for direct detection of V. parahaemolyticus in pure cultures and in spiked shrimp samples was 5.3 × 102 CFU per ml/g (2.0 CFU per reaction. The sensitivity of the LAMP assay was 10-fold more sensitive than that of the conventional PCR assay. The LAMP assay was markedly faster, requiring for amplification 13–22 min in a single colony on TCBS agar from each of 143 V. parahaemolyticus strains and less than 35 min in spiked shrimp samples. The LAMP assay for detection of V. parahaemolyticus required less than 40 min in a single colony on thiosulfate citrate bile salt sucrose (TCBS agar and 60 min in spiked shrimp samples from the beginning of DNA extraction to final determination. Conclusion The LAMP assay is a sensitive, rapid and simple tool for the detection of V. parahaemolyticus and will facilitate the surveillance for control of contamination of V

  7. Assay of ribulose bisphosphate carboxylase

    International Nuclear Information System (INIS)

    Pike, C.; Berry, J.

    1987-01-01

    Assays of ribulose bisphosphate carboxylase (rubisco) can be used to illustrate many properties of photosynthetic systems. Many different leaves have been assayed with this standard procedure. The tissue is ground with a mortar and pestle in extraction buffer. The supernatant after centrifugation is used as the source of enzyme. Buffer, RuBP, [ 14 C]-NaHCO 3 , and enzyme are combined in a scintillation vial; the reaction is run for 1 min at 30 0 . The acid-stable products are counted. Reproducibility in student experiments has been excellent. The assay data can be combined with analyses of leaf properties such as fresh and dry weight, chlorophyll and protein content, etc. Students have done projects such as the response of enzyme to temperature and to various inhibitors. They also report on the use of a transition state analog, carboxyarabinitol bisphosphate, to titrate the molar concentration of rubisco molecules (active sites) in an enzyme sample. Thus, using crude extracts the catalytic activity of a sample can be compared to the absolute quantity of enzyme or to the turnover number

  8. Changes of hematological and biochemical parameters revealed genotoxicity and immunotoxicity of neonicotinoids on Chinese rare minnows (Gobiocypris rarus).

    Science.gov (United States)

    Hong, Xiangsheng; Zhao, Xu; Tian, Xue; Li, Jiasu; Zha, Jinmiao

    2018-02-01

    Adverse impacts of immunity in terrestrial non-target organisms exposed to neonicotinoid insecticides have been reported, but the causal link between insecticide exposure and possible immune alterations in fish remains limited. In the present study, the potential genotoxicity and immunotoxicity of three neonicotinoids (imidacloprid, nitenpyram, and dinotefuran) were assessed in Chinese rare minnows by using a 60-day chronic toxicity test. The hematological and biochemical parameters of juvenile Chinese rare minnows and changes in the transcription of six inflammation-related genes were determined after exposure to neonicotinoids at 0.1, 0.5, or 2.0 mg/L. A clear difference in the frequency of erythrocytes with micronuclei (MN) was observed after treatment with 2.0 mg/L imidacloprid (p < .05). Additionally, exposure to 0.5 or 2.0 mg/L imidacloprid significantly increased the binucleated (BN) erythrocytes and those with notched nuclei (NT) (p < .05). A serum protein electrophoresis (SPE) assay showed significant alterations in the serum protein in all treatments (p < .05), and further analysis indicated decreases in immunoglobulin (Ig) in treatments with 0.5 or 2.0 mg/L imidacloprid or dinotefuran or with 0.1 mg/L nitenpyram (p < .05). Moreover, a biochemical assay confirmed that immunoglobulin M (IgM) levels were indeed significantly decreased upon treatment with imidacloprid or dinotefuran at 0.5 or 2.0 mg/L (p < .05). In addition, the transcriptional levels of the inflammatory cytokines IL-6, INF-α, TNF-α, and IL-1β were markedly down-regulated after all imidacloprid treatments (p < .05), whereas the expression levels of only TNF-α and IL-1β were significantly down-regulated following the 0.5 and 2.0 mg/L dinotefuran treatments (p < .05). Taken together, our results clearly demonstrate that imidacloprid, rather than nitenpyram and dinotefuran, can induce genotoxicity. The responsiveness of these immune indicators

  9. Biocompatibility of a novel cyanoacrylate based tissue adhesive: cytotoxicity and biochemical property evaluation.

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    Young Ju Lee

    Full Text Available Cyanoacrylate (CA is most widely used as a medical and commercial tissue adhesive because of easier wound closure, good cosmetic results and little discomfort. But, CA-based tissue adhesives have some limitations including the release of cytotoxic chemicals during biodegradation. In previous study, we made prepolymerized allyl 2-CA (PACA based tissue adhesive, resulting in longer chain structure. In this study, we investigated a biocompatibility of PACA as alternative tissue adhesive for medical application, comparing with that of Dermabond® as commercial tissue adhesive. The biocompatibility of PACA was evaluated for short-term (24 hr and long-term (3 and 7 days using conventional cytotoxicity (WST, neutral red, LIVE/DEAD and TUNEL assays, hematoxylin-eosin (H&E and Masson trichrome (MT staining. Besides we examined the biochemical changes in cells and DNA induced by PACA and Dermabond® utilizing Raman spectroscopy which could observe the denaturation and conformational changes in protein, as well as disintegration of the DNA/RNA by cell death. In particular, we analyzed Raman spectrum using the multivariate statistical methods including principal component analysis (PCA and support vector machine (SVM. As a result, PACA and Dermabond® tissue adhesive treated cells and tissues showed no difference of the cell viability values, histological analysis and Raman spectral intensity. Also, the classification analysis by means of PCA-SVM classifier could not discriminate the difference between the PACA and Dermabond® treated cells and DNA. Therefore we suggest that novel PACA might be useful as potential tissue adhesive with effective biocompatibility.

  10. Biochemical Characterization of the Split Class II Ribonucleotide Reductase from Pseudomonas aeruginosa.

    Directory of Open Access Journals (Sweden)

    Mikael Crona

    Full Text Available The opportunistic pathogen Pseudomonas aeruginosa can grow under both aerobic and anaerobic conditions. Its flexibility with respect to oxygen load is reflected by the fact that its genome encodes all three existing classes of ribonucleotides reductase (RNR: the oxygen-dependent class I RNR, the oxygen-indifferent class II RNR, and the oxygen-sensitive class III RNR. The P. aeruginosa class II RNR is expressed as two separate polypeptides (NrdJa and NrdJb, a unique example of a split RNR enzyme in a free-living organism. A split class II RNR is also found in a few closely related γ-Proteobacteria. We have characterized the P. aeruginosa class II RNR and show that both subunits are required for formation of a biologically functional enzyme that can sustain vitamin B12-dependent growth. Binding of the B12 coenzyme as well as substrate and allosteric effectors resides in the NrdJa subunit, whereas the NrdJb subunit mediates efficient reductive dithiol exchange during catalysis. A combination of activity assays and activity-independent methods like surface plasmon resonance and gas phase electrophoretic macromolecule analysis suggests that the enzymatically active form of the enzyme is a (NrdJa-NrdJb2 homodimer of heterodimers, and a combination of hydrogen-deuterium exchange experiments and molecular modeling suggests a plausible region in NrdJa that interacts with NrdJb. Our detailed characterization of the split NrdJ from P. aeruginosa provides insight into the biochemical function of a unique enzyme known to have central roles in biofilm formation and anaerobic growth.

  11. Biochemical and molecular characterization of the venom from the Cuban scorpion Rhopalurus junceus.

    Science.gov (United States)

    García-Gómez, B I; Coronas, F I V; Restano-Cassulini, R; Rodríguez, R R; Possani, L D

    2011-07-01

    This communication describes the first general biochemical, molecular and functional characterization of the venom from the Cuban blue scorpion Rhopalurus junceus, which is often used as a natural product for anti-cancer therapy in Cuba. The soluble venom of this arachnid is not toxic to mice, injected intraperitoneally at doses up to 200 μg/20 g body weight, but it is deadly to insects at doses of 10 μg per animal. The venom causes typical alpha and beta-effects on Na+ channels, when assayed using patch-clamp techniques in neuroblastoma cells in vitro. It also affects K+ currents conducted by ERG (ether-a-go-go related gene) channels. The soluble venom was shown to display phospholipase, hyaluronidase and anti-microbial activities. High performance liquid chromatography of the soluble venom can separate at least 50 components, among which are peptides lethal to crickets. Four such peptides were isolated to homogeneity and their molecular masses and N-terminal amino acid sequence were determined. The major component (RjAa12f) was fully sequenced by Edman degradation. It contains 64 amino acid residues and four disulfide bridges, similar to other known scorpion toxins. A cDNA library prepared from the venomous glands of one scorpion allowed cloning 18 genes that code for peptides of the venom, including RjA12f and eleven other closely related genes. Sequence analyses and phylogenetic reconstruction of the amino acid sequences deduced from the cloned genes showed that this scorpion contains sodium channel like toxin sequences clearly segregated into two monophyletic clusters. Considering the complex set of effects on Na+ currents verified here, this venom certainly warrant further investigation. Copyright © 2011 Elsevier Ltd. All rights reserved.

  12. A Molecular Biological and Biochemical Investigation on Mycobacterium tuberculosis MutT Protein.

    Science.gov (United States)

    Huang, Hsiu-Lin; Su, Ho-Ting; Wu, Chung-Hsiun Herbert; Tsai-Wu, Jyy-Jih

    2014-03-01

    Mycobacterium tuberculosis is a vicious microbe co-existing with the infected host. This pathogen exploited opportunities to spread during periods of urbanization and social upheaval, and got retreated with improved hygiene. This investigation was designed to clone and characterize M. tuberculosis mutT gene, a homologue of a DNA repair protein in Escherichia coli. The aim was to depict the possible role of this homologue in the virulent microbe. A DNA fragment of the mutT gene was amplified with PCR from the genomic DNA of strain H37Rv M. tuberculosis. The expression vector was transformed into E. coli strains BL21 (DE3) and MK602 (DE3) (mutT-). The protein activity assay was performed by biochemical methods. M. tuberculosis MutT shares 23% identity with the E. coli MutT protein. The mutT gene DNA fragment was subcloned into the expression vector pET28a(+) and the recombinant plasmid was overexpressed in E. coli. Purified and refolded M. tuberculosis MutT possesses a dGTPase activity, which is one of the most well-known preference nucleotidase activities of MutT in E. coli. This study also showed that the dGTPase activity of M. tuberculosis MutT was enhanced by magnesium and inhibited by Ni(2+) or EDTA. Endogenous MutT protein in M. tuberculosis lysate displayed a smear pattern in the Western blot, suggesting instability of this protein in the bacteria similar to the important proteins, such as P53 protein, tightly regulated by protein degradation. The cloned M. tuberculosis mutT gene and MutT protein were characterized. M. tuberculosis MutT has a dGTPase activity, which is one of the most well-known preference nucleotidase activities of MutT in E. coli. These findings provide further understanding about the vicious bacterium.

  13. Acute effects of ginger extract on biochemical and functional symptoms of delayed onset muscle soreness.

    Science.gov (United States)

    Hoseinzadeh, Khadijeh; Daryanoosh, Farhad; Baghdasar, Parvin Javad; Alizadeh, Hamid

    2015-01-01

    Inflammation and pain induced by delayed onset muscle soreness (DOMS) as a result of eccentric exercise (EE) or unaccustomed activity cause some difficulties in exercise for athletes. The purpose of this study was to survey the effect of ginger extract on biochemical and functional symptom of delayed onset muscle soreness. In a quasi-experimental study, 36 healthy female subjects, who were recruited by intra dormitory calls, randomly divided into 3 groups, including: ginger intake 1 hour before exercise (GIBE), ginger intake immediately after exercise (GIAE) and placebo group (PL). Subjects consumed capsules contain 60 mg of ginger extract (equivalent of 2 g dried ginger powder) or placebo before and after exercise. The exercise protocol consisted of a 20 minute step test using a 46cm step at a rate of 15 steps per minute. The blood samples were taken before, 1, 24 and 48 hour after exercise to assay creatine kinase (CK) and interleukin-6 (IL-6). Muscle pain scores, isometric strength and circumference of thigh muscle, and hip range of motion were recorded at mentioned times. The analysis of variance (ANOVA) with repeated measure was used to determine the differences between groups. The results showed a significant reduction of pain in GIBE compared to GIAE after 24 and 48h of EE and GIAE compared to PL (p<0.05). IL-6 changed significantly in GIBE compared to PL (p<0.05) after 1, 24, and 48h after EE. The other factors didn't change meaningfully. The finding of this study suggests that 2 grams of ginger may have anti-inflammation and analgesic effect on DOMS.

  14. CLSI-based transference and verification of CALIPER pediatric reference intervals for 29 Ortho VITROS 5600 chemistry assays.

    Science.gov (United States)

    Higgins, Victoria; Truong, Dorothy; Woroch, Amy; Chan, Man Khun; Tahmasebi, Houman; Adeli, Khosrow

    2018-03-01

    Evidence-based reference intervals (RIs) are essential to accurately interpret pediatric laboratory test results. To fill gaps in pediatric RIs, the Canadian Laboratory Initiative on Pediatric Reference Intervals (CALIPER) project developed an age- and sex-specific pediatric RI database based on healthy pediatric subjects. Originally established for Abbott ARCHITECT assays, CALIPER RIs were transferred to assays on Beckman, Roche, Siemens, and Ortho analytical platforms. This study provides transferred reference intervals for 29 biochemical assays for the Ortho VITROS 5600 Chemistry System (Ortho). Based on Clinical Laboratory Standards Institute (CLSI) guidelines, a method comparison analysis was performed by measuring approximately 200 patient serum samples using Abbott and Ortho assays. The equation of the line of best fit was calculated and the appropriateness of the linear model was assessed. This equation was used to transfer RIs from Abbott to Ortho assays. Transferred RIs were verified using 84 healthy pediatric serum samples from the CALIPER cohort. RIs for most chemistry analytes successfully transferred from Abbott to Ortho assays. Calcium and CO 2 did not meet statistical criteria for transference (r 2 CALIPER pediatric RI database to laboratories using Ortho VITROS 5600 biochemical assays. Clinical laboratories should verify CALIPER reference intervals for their specific analytical platform and local population as recommended by CLSI. Copyright © 2018 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  15. Classification and fingerprinting of different berries based on biochemical profiling and antioxidant capacity

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    Jasminka Milivojević

    2013-09-01

    Full Text Available The objective of this work was to evaluate the biochemical composition of six berry types belonging to Fragaria, Rubus, Vaccinium and Ribes genus. Fruit samples were collected in triplicate (50 fruit each from 18 different species or cultivars of the mentioned genera, during three years (2008 to 2010. Content of individual sugars, organic acids, flavonols, and phenolic acids were determined by high performance liquid chromatography (HPLC analysis, while total phenolics (TPC and total antioxidant capacity (TAC, by using spectrophotometry. Principal component analysis (PCA and hierarchical cluster analysis (CA were performed to evaluate the differences in fruit biochemical profile. The highest contents of bioactive components were found in Ribes nigrum and in Fragaria vesca, Rubus plicatus, and Vaccinium myrtillus. PCA and CA were able to partially discriminate between berries on the basis of their biochemical composition. Individual and total sugars, myricetin, ellagic acid, TPC and TAC showed the highest impact on biochemical composition of the berry fruits. CA separated blackberry, raspberry, and blueberry as isolate groups, while classification of strawberry, black and red currant in a specific group has not occurred. There is a large variability both between and within the different types of berries. Metabolite fingerprinting of the evaluated berries showed unique biochemical profiles and specific combination of bioactive compound contents.

  16. Optical Assay of Erythrocyte Function in Banked Blood

    Science.gov (United States)

    Bhaduri, Basanta; Kandel, Mikhail; Brugnara, Carlo; Tangella, Krishna; Popescu, Gabriel

    2014-09-01

    Stored red blood cells undergo numerous biochemical, structural, and functional changes, commonly referred to as storage lesion. How much these changes impede the ability of erythrocytes to perform their function and, as result, impact clinical outcomes in transfusion patients is unknown. In this study we investigate the effect of the storage on the erythrocyte membrane deformability and morphology. Using optical interferometry we imaged red blood cell (RBC) topography with nanometer sensitivity. Our time-lapse imaging quantifies membrane fluctuations at the nanometer scale, which in turn report on cell stiffness. This property directly impacts the cell's ability to transport oxygen in microvasculature. Interestingly, we found that cells which apparently maintain their normal shape (discocyte) throughout the storage period, stiffen progressively with storage time. By contrast, static parameters, such as mean cell hemoglobin content and morphology do not change during the same period. We propose that our method can be used as an effective assay for monitoring erythrocyte functionality during storage time.

  17. On the Adaptive Design Rules of Biochemical Networks in Evolution

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    Bor-Sen Chen

    2007-01-01

    Full Text Available Biochemical networks are the backbones of physiological systems of organisms. Therefore, a biochemical network should be sufficiently robust (not sensitive to tolerate genetic mutations and environmental changes in the evolutionary process. In this study, based on the robustness and sensitivity criteria of biochemical networks, the adaptive design rules are developed for natural selection in the evolutionary process. This will provide insights into the robust adaptive mechanism of biochemical networks in the evolutionary process. We find that if a mutated biochemical network satisfies the robustness and sensitivity criteria of natural selection, there is a high probability for the biochemical network to prevail during natural selection in the evolutionary process. Since there are various mutated biochemical networks that can satisfy these criteria but have some differences in phenotype, the biochemical networks increase their diversities in the evolutionary process. The robustness of a biochemical network enables co-option so that new phenotypes can be generated in evolution. The proposed robust adaptive design rules of natural selection gain much insight into the evolutionary mechanism and provide a systematic robust biochemical circuit design method of biochemical networks for biotechnological and therapeutic purposes in the future.

  18. Improving Precision of Proximity Ligation Assay by Amplified Single Molecule Detection

    OpenAIRE

    Ke, Rongqin; Nong, Rachel Yuan; Fredriksson, Simon; Landegren, Ulf; Nilsson, Mats

    2013-01-01

    Proximity ligation assay (PLA) has been proven to be a robust protein detection method. The technique is characterized by high sensitivity and specificity, but the assay precision is probably limited by the PCR readout. To investigate this potential limitation and to improve precision, we developed a digital proximity ligation assay for protein measurement in fluids based on amplified single molecule detection. The assay showed significant improvements in precision, and thereby also detection...

  19. Automated saccharification assay for determination of digestibility in plant materials

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    Halpin Claire

    2010-10-01

    Full Text Available Abstract Background Cell wall resistance represents the main barrier for the production of second generation biofuels. The deconstruction of lignocellulose can provide sugars for the production of fuels or other industrial products through fermentation. Understanding the biochemical basis of the recalcitrance of cell walls to digestion will allow development of more effective and cost efficient ways to produce sugars from biomass. One approach is to identify plant genes that play a role in biomass recalcitrance, using association genetics. Such an approach requires a robust and reliable high throughput (HT assay for biomass digestibility, which can be used to screen the large numbers of samples involved in such studies. Results We developed a HT saccharification assay based on a robotic platform that can carry out in a 96-well plate format the enzymatic digestion and quantification of the released sugars. The handling of the biomass powder for weighing and formatting into 96 wells is performed by a robotic station, where the plant material is ground, delivered to the desired well in the plates and weighed with a precision of 0.1 mg. Once the plates are loaded, an automated liquid handling platform delivers an optional mild pretreatment ( Conclusions The automated assay systems are sensitive, robust and reliable. The system can reliably detect differences in the saccharification of plant tissues, and is able to process large number of samples with a minimum amount of human intervention. The automated system uncovered significant increases in the digestibility of certain lignin modified lines in a manner compatible with known effects of lignin modification on cell wall properties. We conclude that this automated assay platform is of sufficient sensitivity and reliability to undertake the screening of the large populations of plants necessary for mutant identification and genetic association studies.

  20. Biochemical Network Stochastic Simulator (BioNetS: software for stochastic modeling of biochemical networks

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    Elston Timothy C

    2004-03-01

    Full Text Available Abstract Background Intrinsic fluctuations due to the stochastic nature of biochemical reactions can have large effects on the response of biochemical networks. This is particularly true for pathways that involve transcriptional regulation, where generally there are two copies of each gene and the number of messenger RNA (mRNA molecules can be small. Therefore, there is a need for computational tools for developing and investigating stochastic models of biochemical networks. Results We have developed the software package Biochemical Network Stochastic Simulator (BioNetS for efficientlyand accurately simulating stochastic models of biochemical networks. BioNetS has a graphical user interface that allows models to be entered in a straightforward manner, and allows the user to specify the type of random variable (discrete or continuous for each chemical species in the network. The discrete variables are simulated using an efficient implementation of the Gillespie algorithm. For the continuous random variables, BioNetS constructs and numerically solvesthe appropriate chemical Langevin equations. The software package has been developed to scale efficiently with network size, thereby allowing large systems to be studied. BioNetS runs as a BioSpice agent and can be downloaded from http://www.biospice.org. BioNetS also can be run as a stand alone package. All the required files are accessible from http://x.amath.unc.edu/BioNetS. Conclusions We have developed BioNetS to be a reliable tool for studying the stochastic dynamics of large biochemical networks. Important features of BioNetS are its ability to handle hybrid models that consist of both continuous and discrete random variables and its ability to model cell growth and division. We have verified the accuracy and efficiency of the numerical methods by considering several test systems.

  1. THE IMPORTANCE OF DIAGNOSTICS OF THE SPORTSMENS BIOCHEMICAL STATUS

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    Franja Fratrić

    2008-08-01

    Full Text Available Widespread opinion that all the sportsmen nowadays “are sick”, as well as the results of the analysis of the random sample from all over Europe, show that very few of them are not under some kind of treatment or doctor’s supervision. This should be a warning for experts and scientists in sports, for making a strategy to prevent this trend. Sickness is a huge individual and socio-economical problem. Sportsmen nowadays are under extreme pressure, without control of biochemical processes. There is a risk of becoming an “organism dump” full of wasteful and harmful substances, that are sediment. These are: - acids - medicaments - chemicals - proteins, etc. The cause of an error is possible resolve only by knowing its origin. This is precisely the most important function of the diagnostics in sport. In this paper we are presenting the strategy of the diagnostics of the biochemical status of the sportsman’s organism. Moreover, we are bringing out the key problems and suggesting concrete prevention measures.

  2. Dust Fertilization of the Western Atlantic Biota: a Biochemical Model

    Science.gov (United States)

    Holmes, C. W.

    2017-12-01

    Every year an estimated 50 million tons of African dust reaches the Western Atlantic. This dust is composed of quartz sand, clay, and a mixture of quartz and clay particles agglutinated with micronutrient enriched ferruginous cement. However, whether it is friend or foe to biochemical systems is a matter of conjecture. Corals are ideal recorders of changing conditions as the layers can be dated so that the record of chemical changes is easily assessed. There is extensive shallow-and deep water coral development bordering the Florida Straits. The changes in trace element chemistry within these corals show a positive relationship with the African dust record. Recently, it has been demonstrated that many of the metals contained within the dust are necessary micronutrients in the fertilization of plankton. Using the results of these studies, a biochemical model has been constructed. This model suggests a path from inorganic dust through microbial transformation to micronutrient enzymes (i.e. Cd-enriched carbonic anahydrase) and carbonate precipitation on the Bahamian Banks. It is estimated that more than ten million metric tons of this fine, metal-rich sediment is formed each year. However, for much of this sediment, its deposition is temporary, as it is transported into the Florida Straits yearly by tropical cyclones. This metal-enriched fine carbonate becomes nutrients for phytoplankton, providing food for the corals, both shallow and deep.

  3. Physiological and biochemical performances of menthol-induced aposymbiotic corals.

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    Jih-Terng Wang

    Full Text Available The unique mutualism between corals and their photosynthetic zooxanthellae (Symbiodinium spp. is the driving force behind functional assemblages of coral reefs. However, the respective roles of hosts and Symbiodinium in this endosymbiotic association, particularly in response to environmental challenges (e.g., high sea surface temperatures, remain unsettled. One of the key obstacles is to produce and maintain aposymbiotic coral hosts for experimental purposes. In this study, a simple and gentle protocol to generate aposymbiotic coral hosts (Isopora palifera and Stylophora pistillata was developed using repeated incubation in menthol/artificial seawater (ASW medium under light and in ASW in darkness, which depleted more than 99% of Symbiodinium from the host within 4∼8 days. As indicated by the respiration rate, energy metabolism (by malate dehydrogenase activity, and nitrogen metabolism (by glutamate dehydrogenase activity and profiles of free amino acids, the physiological and biochemical performances of the menthol-induced aposymbiotic corals were comparable to their symbiotic counterparts without nutrient supplementation (e.g., for Stylophora or with a nutrient supplement containing glycerol, vitamins, and a host mimic of free amino acid mixture (e.g., for Isopora. Differences in biochemical responses to menthol-induced bleaching between Stylophora and Isopora were attributed to the former digesting Symbiodinium rather than expelling the algae live as found in the latter species. Our studies showed that menthol could successfully bleach corals and provided aposymbiotic corals for further exploration of coral-alga symbioses.

  4. [Development of a new biosensor for biochemical oxygen demand].

    Science.gov (United States)

    Chang, Dong; Du, Xiao-yan; Pan, Hong-zhi; Jia, Jian-bo; Wang, Bing-quan; Cheng, Guang-jin; Dong, Shao-jun

    2003-03-01

    To use a new kind of fixing material, i.e. Sol-Gel organic-inorganic hybridized material to immobilize bacterium to detect Biochemical oxygen demand quickly. The biosensor was fabricated using a thin film in which Hansenula anomala was immobilized by sol-gel and an oxygen electrode. The optimum measurement for biochemical oxygen demand was at pH 7.0; 28 degrees C; response time 3 - 12 min. Pure organic compound, sewage and rate of recovery were detected with the biosensor. It shows that the BOD biosensor can be used to detect many organic compounds such as amino acid, glucide. It is suitable to monitor sewage and industrial waste water which has low level alcohols and phenols. The microbial membrane can work 3 months and remain its 70% activity. It is measured that the rate of recovery of BOD is between 90% to 105% in sewage. The study confirmed the effectiveness and usefulness of BOD sensor, which is quick, convenient, low cost and reliable with little interference.

  5. Biochemical oxygen demand sensor using Serratia marcescens LSY 4.

    Science.gov (United States)

    Kim, M N; Kwon, H S

    1999-01-01

    A microbial biochemical oxygen demand (BOD) sensor consisting of Serratia marcescens LSY 4 and an oxygen electrode was prepared for estimation of the biochemical oxygen demand. The response of the BOD sensor was insensitive to pH in the range of pH 6.0-8.0, and the baseline drift of the signal was nearly absent even in unbuffered aqueous solution. Because heavy metal ions were precipitated from the phosphate buffer solution, unbuffered solution was used to investigate the effect of the concentration of heavy metal ions on the sensor response. Contrary to previous studies, not only Cu2+ and Ag+ but also Cd2+ and Zn2+ significantly decreased the response of the BOD sensor in unbuffered solution. Graft polymerization of sodium styrene sulfonate on the surface of the porous teflon membrane was carried out to absorb the heavy metal ions permeating through the membrane. Tolerance against Zn2+ was induced for S. marcescens LSY 4 to make the cells less sensitive to the presence of heavy metal ions. The membrane modification and the Zn2+ tolerance induction showed some positive effects in such a way that they reduced the inhibitory effects of Zn2+ and Cd2+ on the sensitivity of the BOD sensor. However, they had no effect on the protection of the cells against the interference of Cu2+ and Ag+ on the performance of the sensor.

  6. A biochemical basis for induction of retina regeneration by antioxidants.

    Science.gov (United States)

    Echeverri-Ruiz, Nancy; Haynes, Tracy; Landers, Joseph; Woods, Justin; Gemma, Michael J; Hughes, Michael; Del Rio-Tsonis, Katia

    2018-01-15

    The use of antioxidants in tissue regeneration has been studied, but their mechanism of action is not well understood. Here, we analyze the role of the antioxidant N-acetylcysteine (NAC) in retina regeneration. Embryonic chicks are able to regenerate their retina after its complete removal from retinal stem/progenitor cells present in the ciliary margin (CM) of the eye only if a source of exogenous factors, such as FGF2, is present. This study shows that NAC modifies the redox status of the CM, initiates self-renewal of the stem/progenitor cells, and induces regeneration in the absence of FGF2. NAC works as an antioxidant by scavenging free radicals either independently or through the synthesis of glutathione (GSH), and/or by reducing oxidized proteins through a thiol disulfide exchange activity. We dissected the mechanism used by NAC to induce regeneration through the use of inhibitors of GSH synthesis and the use of other antioxidants with different biochemical structures and modes of action, and found that NAC induces regeneration through its thiol disulfide exchange activity. Thus, our results provide, for the first time, a biochemical basis for induction of retina regeneration. Furthermore, NAC induction was independent of FGF receptor signaling, but dependent on the MAPK (pErk1/2) pathway. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  7. Clinical and Biochemical Characteristics of Children with Juvenile Idiopathic Arthritis

    International Nuclear Information System (INIS)

    Ahmed, S.; Ali, S. R.; Ishaque, S.

    2014-01-01

    Objective: To determine the clinical and biochemical characteristics of children with Juvenile Idiopathic Arthritis (JIA) at a tertiary care centre in Karachi, Pakistan. Study Design: A descriptive study. Place and Duration of Study: Paediatric Rheumatology Clinic of The Aga Khan University Hospital (AKUH), Karachi, from January 2008 to December 2011. Methodology: Clinical and laboratory profile and outcome of children less than 15 years of age attending the Paediatric Rheumatology Clinic of the Aga Khan University, Karachi with the diagnosis of Juvenile Idiopathic Arthritis according to International League against Rheumatism were studied. These children were classified into different types of JIA; their clinical and laboratory characteristics, response to therapy and outcome was evaluated. Results: Sixty eight patients satisfying the criteria of International League against Rheumatism (ILAR) for Juvenile Idiopathic Arthritis were enrolled during the study period of four consecutive years, their age ranged from 9 months to 15 years. Mean age at onset was 6.45 +- 4.03 years while mean age at diagnosis was 7.60 +- 3.93 years. Polyarticular was the most predominant subtype with 37 (54%) patients, out of these, 9 (24%) were rheumatoid factor positive. An almost equal gender predisposition was observed. Fever and arthritis were the most common presenting symptoms, with only 2 patients presenting with uveitis. Conclusion: The clinico-biochemical characteristics of JIA at the study centre showed a pattern distinct with early onset of disease, high frequency of polyarticular type and a higher rheumatoid factor (QRA) and ANA positivity in girls. (author)

  8. Coarse-graining stochastic biochemical networks: adiabaticity and fast simulations

    Energy Technology Data Exchange (ETDEWEB)

    Nemenman, Ilya [Los Alamos National Laboratory; Sinitsyn, Nikolai [Los Alamos National Laboratory; Hengartner, Nick [Los Alamos National Laboratory

    2008-01-01

    We propose a universal approach for analysis and fast simulations of stiff stochastic biochemical kinetics networks, which rests on elimination of fast chemical species without a loss of information about mesoscoplc, non-Poissonian fluctuations of the slow ones. Our approach, which is similar to the Born-Oppenhelmer approximation in quantum mechanics, follows from the stochastic path Integral representation of the cumulant generating function of reaction events. In applications with a small number of chemIcal reactions, It produces analytical expressions for cumulants of chemical fluxes between the slow variables. This allows for a low-dimensional, Interpretable representation and can be used for coarse-grained numerical simulation schemes with a small computational complexity and yet high accuracy. As an example, we derive the coarse-grained description for a chain of biochemical reactions, and show that the coarse-grained and the microscopic simulations are in an agreement, but the coarse-gralned simulations are three orders of magnitude faster.

  9. Thiamethoxam resistance selected in the western flower thrips Frankliniella occidentalis (Thysanoptera: Thripidae): cross-resistance patterns, possible biochemical mechanisms and fitness costs analysis.

    Science.gov (United States)

    Gao, Cong-Fen; Ma, Shao-Zhi; Shan, Cai-Hui; Wu, Shun-Fan

    2014-09-01

    The western flower thrips (WFT) Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), an important pest of various crops in the world, has invaded China since 2003. To understand the risks and to determine possible mechanisms of resistance to thiamethoxam in WFT, a resistant strain was selected under the laboratory conditions. Cross-resistance and the possible biochemical resistance mechanisms were investigated in this study. A 15.1-fold thiamethoxam-resistant WFT strain (TH-R) was established after selection for 55 generations. Compared with the susceptible strain (TH-S), the selected TH-R strain showed extremely high level cross-resistance to imidaclothiz (392.1-fold) and low level cross-resistance to dinotefuran (5.7-fold), acetamiprid (2.9-fold) and emamectin benzoate (2.1-fold), respectively. No cross-resistance to other fourteen insecticides was detected. Synergism tests showed that piperonyl butoxide (PBO) and triphenyl phosphate (TPP) produced a high synergism of thiamethoxam effects in the TH-R strain (2.6- and 2.6-fold respectively). However, diethyl maleate (DEM) did not act synergistically with thiamethoxam. Biochemical assays showed that mixed function oxidase (MFO) activities and carboxylesterase (CarE) activity of the TH-R strain were 2.8- and 1.5-fold higher than that of the TH-S strain, respectively. When compared with the TH-S strain, the TH-R strain had a relative fitness of 0.64. The results show that WFT develops resistance to thiamethoxam after continuous application and thiamethoxam resistance had considerable fitness costs in the WFT. It appears that enhanced metabolism mediated by cytochrome P450 monooxygenases and CarE was a major mechanism for thiamethoxam resistance in the WFT. The use of cross-resistance insecticides, including imidaclothiz and dinotefuran, should be avoided for sustainable resistance management. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Conservation Laws in Biochemical Reaction Networks

    DEFF Research Database (Denmark)

    Mahdi, Adam; Ferragut, Antoni; Valls, Claudia

    2017-01-01

    We study the existence of linear and nonlinear conservation laws in biochemical reaction networks with mass-action kinetics. It is straightforward to compute the linear conservation laws as they are related to the left null-space of the stoichiometry matrix. The nonlinear conservation laws...... are difficult to identify and have rarely been considered in the context of mass-action reaction networks. Here, using the Darboux theory of integrability, we provide necessary structural (i.e., parameterindependent) conditions on a reaction network to guarantee the existence of nonlinear conservation laws...

  11. The biochemical womb of schizophrenia: A review.

    Science.gov (United States)

    Gaur, N; Gautam, S; Gaur, M; Sharma, P; Dadheech, G; Mishra, S

    2008-10-01

    The conclusive identification of specific etiological factors or pathogenic processes in the illness of schizophrenia has remained elusive despite great technological progress. The convergence of state-of-art scientific studies in molecular genetics, molecular neuropathophysiology, in vivo brain imaging and psychopharmacology, however, indicates that we may be coming much closer to understanding the genesis of schizophrenia. In near future, the diagnosis and assessment of schizophrenia using biochemical markers may become a "dream come true" for the medical community as well as for the general population. An understanding of the biochemistry/ visa vis pathophysiology of schizophrenia is essential to the discovery of preventive measures and therapeutic intervention.

  12. Simplifying biochemical models with intermediate species

    DEFF Research Database (Denmark)

    Feliu, Elisenda; Wiuf, Carsten

    2013-01-01

    Mathematical models are increasingly being used to understand complex biochemical systems, to analyse experimental data and make predictions about unobserved quantities. However, we rarely know how robust our conclusions are with respect to the choice and uncertainties of the model. Using algebraic......-state concentrations of the species in the core model, after suitable matching of parameters. Importantly, our results provide guidelines to the modeller in choosing between models and in distinguishing their properties. Further, our work provides a formal way of comparing models that share a common skeleton....

  13. Radiation treatment of drugs, biochemicals and vaccines

    International Nuclear Information System (INIS)

    Nordheim, W.; Braeuniger, S.; Kirsch, B.; Kotowski, H.; Teupel, D.

    1984-12-01

    The concise and tabulated review reports experimental results on the effects of radiation treatment on drugs, vaccines, biochemicals and adjuvants including enzymes as well. Irradiation was mostly performed by γ-radiation using 60 Co and to a lesser extent by 137 Cs, 182 Ta, X-rays and accelerators. Ionizing radiation proved to be a useful tool for sterilization and inactivation in producing drugs, vaccines, and bioactive agents and will contribute to realize procedures difficultly solvable as to engineering and economy, respectively. 124 refs

  14. Dosimetric response of some biochemicals used as lyoluminescent dosimeters

    International Nuclear Information System (INIS)

    Ettinger, K.V.; Rowe, R.W.; Mallard, J.R.; Takavar, A.; Sephton, J.

    1977-01-01

    It has been found recently that a whole variety of biochemicals exhibit lyoluminescent response to ionizing and UV radiation, which can be used for the purpose of dosimetry. Among the amino acids, glutamine, glutamic acid and valine are showing good response and satisfactory stability of the stored energy i.e. stability of 'frozen' free radicals. Actually, all amino acids involved in naturally occuring proteins, which were investigated (20 compounds) show lyoluminescence to smaller or greater extent. The response is proportional to the dose in the region of 50 rad to 100 kilorad. The mechanism of lyoluminescence in amino acids is probably the same as in the saccharides; formation of free radicals in the solid, conversion to peroxy radicals and, finally generation of excited oxygen dimers on dissolution. Other categories of biochemicals which exhibit lyoluminescence (LL) are DNA, RNA and their salts, as well as some antibiotics like streptomycin, gentamycin and oxytetracycline. Those of proteins that are easily soluble in water, show a good LL response. The half-life of free radicals responsible for LL in albumins (egg, horse and human) is of an order of 24 h. However, in salmine (protamine sulphate) the decay is very slow, of an order of few month, so this material can be used as a 'protein equivalent' dosemeter. In all experiments a 60 Co source was used for irradiations. Proteins, RNA and DNA show a considerable response to UV. In experiments with UV radiation, mostly 2547 A wavelength, the response to radiation in terms of energy deposited in gramme of material was almost twice that for gamma rays of 1.33 and 1.17MeV

  15. Genetic and biochemical analysis of a laboratory-selected spirodiclofen-resistant strain of Tetranychus urticae Koch (Acari: Tetranychidae).

    Science.gov (United States)

    Van Pottelberge, Steven; Van Leeuwen, Thomas; Khajehali, Jahangir; Tirry, Luc

    2009-04-01

    Spirodiclofen is a selective, non-systemic acaricide from the new chemical class of tetronic acid derivatives. In order to develop strategies to minimise resistance in the field, a laboratory-selected spirodiclofen-resistant strain of the two-spotted spider mite, Tetranychus urticae Koch, was used to determine genetic, toxicological, biochemical and cross-resistance data. Selecting for spirodiclofen resistance in the laboratory yielded a strain (SR-VP) with a resistance ratio of 274, determined on the larval stage. The egg stage remained far more susceptible. No cross-resistance was found against other established acaricides, except for spiromesifen. Based on synergist experiments and enzyme assays, it appeared that especially P450 monooxygenases, but also esterases and glutathione-S-transferases, could be involved in the metabolic detoxification of spirodiclofen. Genetic analysis showed that the resistance is inherited as an intermediate trait under control of more than one gene. Resistance to spirodiclofen exceeded by far the recommended field rate. A good acaricide resistance management programme is necessary to prevent fast resistance build-up in the field. Spirodiclofen can be used in alternation with most established acaricides, except for other tetronic acid derivatives. Without selection pressure, resistance tends to be unstable and can decrease in the presence of susceptible individuals owing to the intermediate, polygenic inheritance mode. Copyright (c) 2009 Society of Chemical Industry.

  16. Biochemical and genetic characterization of a novel metallo-β-lactamase from marine bacterium Erythrobacter litoralis HTCC 2594.

    Science.gov (United States)

    Jiang, Xia-Wei; Cheng, Hong; Huo, Ying-Yi; Xu, Lin; Wu, Yue-Hong; Liu, Wen-Hong; Tao, Fang-Fang; Cui, Xin-Jie; Zheng, Bei-Wen

    2018-01-16

    Metallo-β-lactamases (MBLs) are a group of enzymes that can inactivate most commonly used β-lactam-based antibiotics. Among MBLs, New Delhi metallo-β-lactamase-1 (NDM-1) constitutes an urgent threat to public health as evidenced by its success in rapidly disseminating worldwide since its first discovery. Here we report the biochemical and genetic characteristics of a novel MBL, ElBla2, from the marine bacterium Erythrobacter litoralis HTCC 2594. This enzyme has a higher amino acid sequence similarity to NDM-1 (56%) than any previously reported MBL. Enzymatic assays and secondary structure alignment also confirmed the high similarity between these two enzymes. Whole genome comparison of four Erythrobacter species showed that genes located upstream and downstream of elbla2 were highly conserved, which may indicate that elbla2 was lost during evolution. Furthermore, we predicted two prophages, 13 genomic islands and 25 open reading frames related to insertion sequences in the genome of E. litoralis HTCC 2594. However, unlike NDM-1, the chromosome encoded ElBla2 did not locate in or near these mobile genetic elements, indicating that it cannot transfer between strains. Finally, following our phylogenetic analysis, we suggest a reclassification of E. litoralis HTCC 2594 as a novel species: Erythrobacter sp. HTCC 2594.

  17. Morphological and biochemical characterization of xanthomonas campestris (pammel) dawson pv. Sesame and it's management by bacterial antagonists

    International Nuclear Information System (INIS)

    Naqvi, S.F.; Haq, M.I.U.; Tahir, M.I.; Khan, M.A.; Ali, Z.

    2013-01-01

    Isolates of Xanthomonas campestris (Pammel) Dawson pv. sesami (Xcs) were collected from various locations in Pakistan and were characterized using several biochemical tests including; (KOH) string test, gram staining, H/sub 2/S production, catalase activity, oxidation/fermentation of glucose, oxidase activity and nitrate reduction. The isolates were positive in tests for KOH string assay, H/sub 2/S, catalase and showed negative reactions in gram staining, oxidase test and nitrate reduction while oxidative utilization of glucose was observed. To develop biological control for the pathogen we isolated numerous bacteria from the rhizosphere of sesame plants and screened them for antagonism against the pathogen using the zone inhibition method. Out of 87 isolates, 9 isolates were inhibitory to the pathogen in vitro. Pseudomonas fluorescens ID-3 exhibited the highest zone of inhibition i.e. 7.97mm. All antagonistic isolates were tested for disease control under greenhouse conditions. Four sesame lines that were found to be moderately resistant in previous studies (i.e. 95001, 96007, 96019, and 20003) were selected for evaluating biocontrol efficacy. The isolates were found to control the disease and reduced severity to 3% compared to untreated plants. The isolate P. fluorescens ID-3 was considered to provide the best disease control with each cultivar. Average biocontrol efficiency (BE) of ID-3 was 78.25% while the best BE was obtained in line 96019, i.e. 81%. (author)

  18. Radiosotopic assay and binder therefor

    International Nuclear Information System (INIS)

    Caston, J.D.; Kamen, B.A.

    1976-01-01

    A rapid and less costly radioisotopic assay for measuring the concentration of folate in blood serum is described. This procedure utilizes 3 H-pteroylmonoglutamate, unlabeled 5-methyltetrahydrofolic acid, and a partially purified folate binder, such as for example a folate binder extracted from hog kidney. The procedure involves radioisotopically relating the bound amounts of a labeled folate and a known folate at various concentrations of the known folate in a system containing a predetermined amount of the labeled folate, a predetermined amount of the binder factor for the folates, and a predetermined amount of defolated test serum. 16 claims, 8 drawing figures

  19. Fluorescence-based assay as a new screening tool for toxic chemicals

    Science.gov (United States)

    Moczko, Ewa; Mirkes, Evgeny M.; Cáceres, César; Gorban, Alexander N.; Piletsky, Sergey

    2016-09-01

    Our study involves development of fluorescent cell-based diagnostic assay as a new approach in high-throughput screening method. This highly sensitive optical assay operates similarly to e-noses and e-tongues which combine semi-specific sensors and multivariate data analysis for monitoring biochemical processes. The optical assay consists of a mixture of environmental-sensitive fluorescent dyes and human skin cells that generate fluorescence spectra patterns distinctive for particular physico-chemical and physiological conditions. Using chemometric techniques the optical signal is processed providing qualitative information about analytical characteristics of the samples. This integrated approach has been successfully applied (with sensitivity of 93% and specificity of 97%) in assessing whether particular chemical agents are irritating or not for human skin. It has several advantages compared with traditional biochemical or biological assays and can impact the new way of high-throughput screening and understanding cell activity. It also can provide reliable and reproducible method for assessing a risk of exposing people to different harmful substances, identification active compounds in toxicity screening and safety assessment of drugs, cosmetic or their specific ingredients.

  20. A Non-Isotopic In Vitro Assay for Histone Acetylation

    Science.gov (United States)

    Kuninger, David; Lundblad, James; Semirale, Anthony; Rotwein, Peter

    2007-01-01

    We describe a simple, robust, and relatively inexpensive non-radioactive in vitro assay for measuring histone acetyl-transferase activity. The assay takes advantage of easy to purify recombinant E. coli-derived fusion proteins containing the NH2-terminal tails of histones H3 and H4 linked to epitope-tagged maltose binding protein (MBP), and immunoblotting with antibodies specific to acetylated H3 and H4. Here we show the specificity and dynamic range of this assay for the histone acetyl-transferases, p300 and PCAF. This assay may be adapted readily for other substrates by simply generating new fusion proteins and for other acetyl-transferases by modifying reaction conditions. PMID:17698235

  1. The radioenzymatic assay of matrix metalloproteinase-1 activity

    International Nuclear Information System (INIS)

    Wu Shiliang; Xu Lan; Zhou Yinghui; Wang Weiping; Shi Ning

    1998-12-01

    The radioenzymatic assay method for tissue collagenase, a metalloproteinase, activity in matrix was established. The matrix collagenase is the most vital catabolic enzyme of collagen in tissue. It mainly acts on type- I, II, III matrix collagen and is also called matrix metalloproteinase-1 (MMPs-1). The assay method for the matrix collagenase was as follows: After type-I collagen was prepared from calf skin and identified with HPLC, it was marked with 3 H-acetic anhydride as the substrate. Then a series of assays were performed in animal experiments and human cases, which showed that the matrix collagenase (MMPs-1) activity assay is feasible and gives reliable results in clinical biochemistry study

  2. Study on quantification of HBs-antibody by immunoradiometric assay

    International Nuclear Information System (INIS)

    Kondo, Yuichi; Itoi, Yoshihiro; Kajiyama, Shizuo

    1989-01-01

    Quantification of HBs-antibody assay was carried out using a commercialized assay kit and standard solutions of HBs-antibody recognised as 1 st reference preparation of hepatitis B immunogloblin by WHO. Standard curve of HBs-antibody was drawn with the function of 3D-spline and the correlation factor was obtained as r = 0.999. Coefficient of intra-assay variance was 3.8 % and that of inter-assay variance was 7.8 %. Dilution tests showed satisfactory results in the range of 2-16 times. Correlation between value of cut-off indices and concentration of HBs-antibody was obtained as the formula of y = 2.599 x-3.894 (r = 0.992) and 2.1 of cut-off index corresponded to about 5 mIU/ml of HBs-antibody concentration. (author)

  3. Dodecylresorufin (C12R) Outperforms Resorufin in Microdroplet Bacterial Assays.

    Science.gov (United States)

    Scheler, Ott; Kaminski, Tomasz S; Ruszczak, Artur; Garstecki, Piotr

    2016-05-11

    This paper proves that dodecylresorufin (C12R) outperforms resorufin (the conventional form of this dye) in droplet microfluidic bacterial assays. Resorufin is a marker dye that is widely used in different fields of microbiology and has increasingly been applied in droplet microfluidic assays and experiments. The main concern associated with resorufin in droplet-based systems is dye leakage into the oil phase and neighboring droplets. The leakage decreases the performance of assays because it causes averaging of the signal between the positive (bacteria-containing) and negative (empty) droplets. Here we show that C12R is a promising alternative to conventional resorufin because it maintains higher sensitivity, specificity, and signal-to-noise ratio over time. These characteristics make C12R a suitable reagent for droplet digital assays and for monitoring of microbial growth in droplets.

  4. A European multicientre study on the comparison of HIV-1 viral loads between VERIS HIV-1 Assay and Roche COBAS® TAQMAN® HIV-1 test, Abbott RealTime HIV-1 Assay, and Siemens VERSANT HIV-1 Assay.

    Science.gov (United States)

    Braun, Patrick; Delgado, Rafael; Drago, Monica; Fanti, Diana; Fleury, Hervé; Hofmann, Jörg; Izopet, Jacques; Kühn, Sebastian; Lombardi, Alessandra; Mancon, Alessandro; Marcos, Mª Angeles; Mileto, Davide; Sauné, Karine; O'Shea, Siobhan; Pérez-Rivilla, Alfredo; Ramble, John; Trimoulet, Pascale; Vila, Jordi; Whittaker, Duncan; Artus, Alain; Rhodes, Daniel

    2017-07-01

    Viral load monitoring is essential for patients under treatment for HIV. Beckman Coulter has developed the VERIS HIV-1 Assay for use on the novel, automated DxN VERIS Molecular Diagnostics System. ¥ OBJECTIVES: Evaluation of the clinical performance of the new quantitative VERIS HIV-1 Assay at multiple EU laboratories. Method comparison with the VERIS HIV-1 Assay was performed with 415 specimens at 5 sites tested with COBAS ® AmpliPrep/COBAS ® TaqMan ® HIV-1 Test, v2.0, 169 specimens at 3 sites tested with RealTime HIV-1 Assay, and 202 specimens from 2 sites tested with VERSANT HIV-1 Assay. Patient monitoring sample results from 4 sites were also compared. Bland-Altman analysis showed the average bias between VERIS HIV-1 Assay and COBAS HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay to be 0.28, 0.39, and 0.61 log 10 cp/mL, respectively. Bias at low end levels below 1000cp/mL showed predicted bias to be DxN VERIS System demonstrated comparable clinical performance to COBAS ® HIV-1 Test, RealTime HIV-1 Assay, and VERSANT HIV-1 Assay. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Discovery of PF-06928215 as a high affinity inhibitor of cGAS enabled by a novel fluorescence polarization assay

    Energy Technology Data Exchange (ETDEWEB)

    Hall, Justin; Brault, Amy; Vincent, Fabien; Weng, Shawn; Wang, Hong; Dumlao, Darren; Aulabaugh, Ann; Aivazian, Dikran; Castro, Dana; Chen, Ming; Culp, Jeffrey; Dower, Ken; Gardner, Joseph; Hawrylik, Steven; Golenbock, Douglas; Hepworth, David; Horn, Mark; Jones, Lyn; Jones, Peter; Latz, Eicke; Li, Jing; Lin, Lih-Ling; Lin, Wen; Lin, David; Lovering, Frank; Niljanskul, Nootaree; Nistler, Ryan; Pierce, Betsy; Plotnikova, Olga; Schmitt, Daniel; Shanker, Suman; Smith, James; Snyder, William; Subashi, Timothy; Trujillo, John; Tyminski, Edyta; Wang, Guoxing; Wong, Jimson; Lefker, Bruce; Dakin, Leslie; Leach, Karen (UMASS, MED); (Pfizer)

    2017-09-21

    Cyclic GMP-AMP synthase (cGAS) initiates the innate immune system in response to cytosolic dsDNA. After binding and activation from dsDNA, cGAS uses ATP and GTP to synthesize 2', 3' -cGAMP (cGAMP), a cyclic dinucleotide second messenger with mixed 2'-5' and 3'-5' phosphodiester bonds. Inappropriate stimulation of cGAS has been implicated in autoimmune disease such as systemic lupus erythematosus, thus inhibition of cGAS may be of therapeutic benefit in some diseases; however, the size and polarity of the cGAS active site makes it a challenging target for the development of conventional substrate-competitive inhibitors. We report here the development of a high affinity (KD = 200 nM) inhibitor from a low affinity fragment hit with supporting biochemical and structural data showing these molecules bind to the cGAS active site. We also report a new high throughput cGAS fluorescence polarization (FP)-based assay to enable the rapid identification and optimization of cGAS inhibitors. This FP assay uses Cy5-labelled cGAMP in combination with a novel high affinity monoclonal antibody that specifically recognizes cGAMP with no cross reactivity to cAMP, cGMP, ATP, or GTP. Given its role in the innate immune response, cGAS is a promising therapeutic target for autoinflammatory disease. Our results demonstrate its druggability, provide a high affinity tool compound, and establish a high throughput assay for the identification of next generation cGAS inhibitors.

  6. Biochemical, hematological and histological parameters induced by cypermetherin toxicity in domestic rabbits.

    Science.gov (United States)

    Dahamna, S; Harzallah, D; Boussahel, S; Belgeit, A; Merghem, M; Bouriche, H

    2010-01-01

    Cypermetherin is widely used in Algeria; this insecticide belongs to the group of pyrethroids classified by the World Health Organization as moderately harmful class II (WHO, 2005). This study was conducted to search the effect of non lethal dose on biochemical parameters, hematological and histological parts of the organs. Male of domestic rabbits Oryctolagus cuniculus (1 kg) received per week and per gavage 1/10 LD50 of Cypermetherin (ARRIVO 25%, active substance of Cypermetherin 250 g/l). blood was collected 72 hours after the treatment. Enzyme activities were assayed in the plasma samples for Glutamate oxalo acetate transaminase (GOT), Glutamate pyruvate transaminase (GPT), Alcaline phosphatase (AlcP), creatinine (Crea) and glucose. Red blood (RBC) cells and white blood cells (WBC) were calculated too. The results indicated a significant increase in transaminases GOT and GPT, and AlcP explain a high energy generating product and dysfunction of the liver. A decline in Crea, Hb, RBC and WBC which is related to the immunity, this is probably due to cell lyses explain the effect of Cypermetherin on erythropoeisis. Histological examination confirmed the biochemical tests by the observation of inflammatory infiltrate and perilobular fibrosis. In conclusion, Cypermetherin with used dose affects biochemical, hematological and histological parameters of the rabbits.

  7. Nutritional and biochemical therapies for neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    George E. Barreto

    2015-04-01

    Full Text Available Neurodegenerative diseases such as Parkinson’s disease, Huntington and Alzheimer’s disease are characterized by neuronal death and loss in different areas of the brain. Downstream signaling mechanisms associated to cellular death/survival are altered, where mitochondrial damage and inflammation, dysfunctional autophagy process, and accumulation of toxins proteins play a central role in the pathogenesis of these diseases. The disabling effects of these diseases on health system are high and greatly affect the health and daily lifestyle of patients. In this context, pharmacological and non-pharmacological therapies, which are used in palliative and preventive treatments, have been widely assessed in human patients, as well as animal and cellular models in the last decades. However, the genetics and epigenetics factors of any disease can cause different paths in its progression. Nutritional and biochemical therapy approaches by activation or manipulation of different transcription factors such as Nrf2, PPARα, CREB and TEFB in animal and cellular models have shown protective effects against neurodegeneration. Some of these therapies include caloric restriction diet, use of glutathione precursors and Mediterranean diet. This work highlights the evidences of different nutritional and biochemical approaches for the treatment of neurodegenerative diseases and how novel research approaches, such as the use of systems biology, will allow a better comprehension of key processes and biological responses involved in these diseases.

  8. Biochemical processes for geothermal brine treatment

    Energy Technology Data Exchange (ETDEWEB)

    Premuzic, E.T.; Lin, M.S.; Bohenek, M.; Joshi-Tope, G.; Zhou, W.; Shelenkova, L.; Wilke, R.

    1998-08-01

    As part of the DOE Geothermal Energy Program, BNL`s Advanced Biochemical Processes for Geothermal Brines (ABPGB) project is aimed at the development of cost-efficient and environmentally acceptable technologies for the disposal of geothermal wastes. Extensive chemical studies of high and low salinity brines and precipitates have indicated that in addition to trace quantities of regulated substances, e.g., toxic metals such as arsenic and mercury, there are significant concentrations of valuable metals, including gold, silver and platinum. Further chemical and physical studies of the silica product have also shown that the produced silica is a valuable material with commercial potential. A combined biochemical and chemical technology is being developed which (1) solubilizes, separates, and removes environmentally regulated constituents in geothermal precipitates and brines, (2) generates an amorphous silica product which may be used as feedstock for the production of revenue generating materials, (3) recover economically valuable trace metals and salts. Geothermal power resources which utilize low salinity brines and use the Stretford process for hydrogen sulfide abatement generate a contaminated sulfur cake. Combined technology converts such sulfur to a commercial grade sulfur, suitable for agricultural use. The R and D activities at BNL are conducted jointly with industrial parties in an effort focused on field applications.

  9. BIOCHEMICAL PROCESSES FOR GEOTHERMAL BRINE TREATMENT

    Energy Technology Data Exchange (ETDEWEB)

    PREMUZIC,E.T.; LIN,M.S.; BOHENEK,M.; JOSHI-TOPE,G.; ZHOU,W.; SHELENKOVA,L.; WILKE,R.

    1998-09-20

    As part of the DOE Geothermal Energy Program, BNL's Advanced Biochemical Processes for Geothermal Brines (ABPGB) project is aimed at the development of cost-efficient and environmentally acceptable technologies for the disposal of geothermal wastes. Extensive chemical studies of high and low salinity brines and precipitates have indicated that in addition to trace quantities of regulated substances, e.g., toxic metals such as arsenic and mercury, there are significant concentrations of valuable metals, including gold, silver and platinum. Further chemical and physical studies of the silica product have also shown that the produced silica is a valuable material with commercial potential. A combined biochemical and chemical technology is being developed which (1) solubilizes, separates, and removes environmentally regulated constituents in geothermal precipitates and brines (2) generates an amorphous silica product which may be used as feedstock for the production of revenue generating materials, (3) recover economically valuable trace metals and salts. Geothermal power resources which utilize low salinity brines and use the Stretford process for hydrogen sulfide abatement generate a contaminated sulfur cake. Combined technology converts such sulfur to a commercial grade sulfur, suitable for agricultural use. The R and D activities at BNL are conducted jointly with industrial parties in an effort focused on field applications.

  10. Biochemical research elucidating metabolic pathways in Pneumocystis*

    Directory of Open Access Journals (Sweden)

    Kaneshiro E.S.

    2010-12-01

    Full Text Available Advances in sequencing the Pneumocystis carinii genome have helped identify potential metabolic pathways operative in the organism. Also, data from characterizing the biochemical and physiological nature of these organisms now allow elucidation of metabolic pathways as well as pose new challenges and questions that require additional experiments. These experiments are being performed despite the difficulty in doing experiments directly on this pathogen that has yet to be subcultured indefinitely and produce mass numbers of cells in vitro. This article reviews biochemical approaches that have provided insights into several Pneumocystis metabolic pathways. It focuses on 1 S-adenosyl-L-methionine (AdoMet; SAM, which is a ubiquitous participant in numerous cellular reactions; 2 sterols: focusing on oxidosqualene cyclase that forms lanosterol in P. carinii; SAM:sterol C-24 methyltransferase that adds methyl groups at the C-24 position of the sterol side chain; and sterol 14α-demethylase that removes a methyl group at the C-14 position of the sterol nucleus; and 3 synthesis of ubiquinone homologs, which play a pivotal role in mitochondrial inner membrane and other cellular membrane electron transport.

  11. [Biochemical principles of early saturnism recognition].

    Science.gov (United States)

    Tsimakuridze, M P; Mansuradze, E A; Zurashvili, D G; Tsimakuridze, M P

    2009-03-01

    The aim of the work is to determine the major sensitive criteria of biochemical indicators that allow timely discovery of negative influence of lead on organism and assist in early diagnosis of primary stages of saturnism. The workers of Georgian typographies, performing technological processes of letterpress printing were observed. Professional groups having contact with lead aerosols (main group of 66 people) and the workers of the same typography not being in touch with the poison (control group of 24 people) were studied. It was distinguished that, protracted professional contact with lead causes moderate increase of lead, coproporphyrin and DALA in daily urine in most cases; it is more clearly evidenced in the professional groups of lead smelters and lino operators and less clearly among typesetter and printers. Upon the checkup of people, having a direct contact with lead, biochemical analysis of urine should be given a preference, especially the determination of quantitative content of lead and coproporphyrin in urine with the aim of revealing the lead carrier, which is one of the first signals for occupational lookout and medical monitoring of the similar contingent.

  12. Pulmonary biochemical alterations resulting from ozone exposure

    Energy Technology Data Exchange (ETDEWEB)

    Mustafa, M.G.; Lee, S.D.

    1976-07-01

    Metabolic response of lung tissue to ozone was studied in rats and monkeys after exposure of animals to various levels of ozone (0.1 to 0.8 ppM) for 1 to 30 days. In rats, 0.8 ppM ozone exposure resulted in a 40 to 50 percent augmentation of oxygen utilization in lung homogenate in the presence of an added substrate (e.g., succinate or 2-oxoglutarate). Activities of marker enzymes, viz. mitochondrial succinate-cytochrome c reductase; microsomal NADPH-cytochrome c reductase and cytosolic glucose-6-phosphate dehydrogenase, increased maximally (40 to 70 percent over control) after 3 to 4 days of exposure, and remained elevated throughout the 0.8 ppM ozone exposure for 30 days. In monkeys, the observations were the same except that the magnitude of biochemical changes was relatively smaller. Exposure of animals to lower levels of ozone resulted in proportionately smaller biochemical changes in the lung, and ozone effects were detectable up to the 0.2 ppM level. While 0.1 ppM ozone exposure was ineffective, dietary deficiency of vitamin E, a natural antioxidant, increased the sensitivity of rat lungs to this concentration of ozone. The results suggest that low-level ozone exposures may cause metabolic alterations in the lung, and that dietary supplementation of vitamin E may offer protection against oxidant stress.

  13. Biochemical Manifestation of HIV Lipodystrophy Syndrome.

    Science.gov (United States)

    Ihenetu, Kenneth; Mason, Darius

    2012-01-01

    Highly active anti-retroviral therapy (HAART), including protease inhibitors (PI) have led to dramatic improvements in the quality and quantity of life in patients with acquired immunodeficiency syndrome (AIDS). However, a significant number of AIDS patients on HAART develop characteristic changes in body fat redistribution referred to as lipodystrophy syndrome (LDS). Features of LDS include hypertrophy in the neck fat pad (buffalo hump), increased fat in the abdominal region (protease paunch), gynecomastia and loss of fat in the mid-face and extremities. The aim of this paper is to review the current knowledge regarding this syndrome. This article reviews the published investigations on biochemical manifestation of HIV lipodystrophy syndrome. It is estimated that approximately 64% of patients treated with PI will experience this syndrome. Biochemically, these patients have increased triglycerides (Trig), total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C) and extremely low high-density lipoprotein-cholesterol (HDL-C). It is hoped that awareness of this syndrome would aid in early diagnosis and better patient management, possibly leading to a lower incidence of cardiovascular complications among these patients.

  14. Biochemical markers of neonatal myocardial dysfunction.

    Science.gov (United States)

    Almeida, Cristina Maria; Carrapato, Manuel R G; Pinto, Fernando; Pinto, Mariana; Ferreira, Sofia; Schmitt, Denise; Marinho, Luis

    2011-04-01

    Cardiac ultrasounds (US) are not always available at the bedside. Cardiac Troponin I (cTnI), CK-MB and NT-proBNP may be an alternative or complementary to influence evaluation and treatment. To determine reference ranges of biochemical markers cTnI, CK-MB and NT-proBNP in normal neonates. Cord and blood samples were collected from neonates and the above biochemical markers were determined. Ultrasounds were performed blindly. CK-MB remains constant from cord blood to the first day, declining thereafter to almost half the values (81.5 vs 52.0 U/l); cTnI increases from 0.004 to 0.058 ng/ml by 72 h falling to 0.030 by day 10; NT-proBNP peaks by 24 h (5085.5 pg/ml), subsiding to 3388.5 pg/ml by day 3, falling to 1316.0 pg/ml by day 10. CK-MB, mostly of muscle origin and reflecting labor stress or injury, is not to recommend as a measure of myocardial damage in the neonate. The rise in cTnI may be explained by a degree of myocardial involvement, albeit physiological. The initial rise and subsequent fall of NT-proBNP represents the physiological ventricular overload of transient birth adaptation.

  15. A high throughput biochemical fluorometric method for measuring lipid peroxidation in HDL.

    Directory of Open Access Journals (Sweden)

    Theodoros Kelesidis

    Full Text Available Current cell-based assays for determining the functional properties of high-density lipoproteins (HDL have limitations. We report here the development of a new, robust fluorometric cell-free biochemical assay that measures HDL lipid peroxidation (HDLox based on the oxidation of the fluorochrome Amplex Red. HDLox correlated with previously validated cell-based (r = 0.47, p<0.001 and cell-free assays (r = 0.46, p<0.001. HDLox distinguished dysfunctional HDL in established animal models of atherosclerosis and Human Immunodeficiency Virus (HIV patients. Using an immunoaffinity method for capturing HDL, we demonstrate the utility of this novel assay for measuring HDLox in a high throughput format. Furthermore, HDLox correlated significantly with measures of cardiovascular diseases including carotid intima media thickness (r = 0.35, p<0.01 and subendocardial viability ratio (r = -0.21, p = 0.05 and physiological parameters such as metabolic and anthropometric parameters (p<0.05. In conclusion, we report the development of a new fluorometric method that offers a reproducible and rapid means for determining HDL function/quality that is suitable for high throughput implementation.

  16. Online Fault Diagnosis for Biochemical Process Based on FCM and SVM.

    Science.gov (United States)

    Wang, Xianfang; Du, Haoze; Tan, Jinglu

    2016-12-01

    Fault diagnosis is becoming an important issue in biochemical process, and a novel online fault detection and diagnosis approach is designed by combining fuzzy c-means (FCM) and support vector machine (SVM). The samples are preprocessed via FCM algorithm to enhance the ability of classification firstly. Then, those samples are input to the SVM classifier to realize the biochemical process fault diagnosis. In this study, a glutamic acid fermentation process is chosen as an example to diagnose the fault by this method, the result shows that the diagnosis time is largely shortened, and the accuracy is extremely improved by comparing to a single SVM method.

  17. Reporter Gene Assays in Ecotoxicology.

    Science.gov (United States)

    Elad, Tal; Belkin, Shimshon

    The need for simple and rapid means for evaluating the potential toxic effects of environmental samples has prompted the development of reporter gene assays, based on tester cells (bioreporters) genetically engineered to report on sample toxicity by producing a readily quantifiable signal. Bacteria are especially suitable to serve as bioreporters owing to their fast responses, low cost, convenient preservation, ease of handling, and amenability to genetic manipulations. Various bacterial bioreporters have been introduced for general toxicity and genotoxicity assessment, and the monitoring of endocrine disrupting and dioxin-like compounds has been mostly covered by similarly engineered eukaryotic cells. Some reporter gene assays have been validated, standardized, and accredited, and many others are under constant development. Efforts are aimed at broadening detection spectra, lowering detection thresholds, and combining toxicity identification capabilities with characterization of the toxic effects. Taking advantage of bacterial robustness, attempts are also being made to incorporate bacterial bioreporters into field instrumentation for online continuous monitoring or on-site spot checks. However, key hurdles concerning test validation, cell preservation, and regulatory issues related to the use of genetically modified organisms still remain to be overcome.

  18. Fractionation of sulfur isotopes in the chemical and biochemical oxidation of sulfide to sulfate

    International Nuclear Information System (INIS)

    Maass, I.; Wetzel, K.; Weise, G.; Heyer, J.

    1983-01-01

    The behaviour of sulfur isotopes in the chemical and biochemical oxidation of marcasite (FeS 2 ) to sulfate has been investigated in rest and shaker cultures at 30 0 C. The microbiological oxidation was carried out using a mixed culture of Thiobacillus. The results show a considerably faster formation of sulfate in the biochemical oxidation in comparison with the chemical oxidation. Isotope analyses of the formed sulfates indicate no or only very small isotope fractionations depending on experimental conditions. The highest enrichment of 32 S in the sulfate is 1.7 per mille. In accordance with the results of other authors it is concluded that in both chemical and biochemical weathering of sedimentary sulfides resulting in the formation of sulfates isotope effects are not of importance. (author)

  19. Performance limits and trade-offs in entropy-driven biochemical computers.

    Science.gov (United States)

    Chu, Dominique

    2018-04-14

    It is now widely accepted that biochemical reaction networks can perform computations. Examples are kinetic proof reading, gene regulation, or signalling networks. For many of these systems it was found that their computational performance is limited by a trade-off between the metabolic cost, the speed and the accuracy of the computation. In order to gain insight into the origins of these trade-offs, we consider entropy-driven computers as a model of biochemical computation. Using tools from stochastic thermodynamics, we show that entropy-driven computation is subject to a trade-off between accuracy and metabolic cost, but does not involve time-trade-offs. Time trade-offs appear when it is taken into account that the result of the computation needs to be measured in order to be known. We argue that this measurement process, although usually ignored, is a major contributor to the cost of biochemical computation. Copyright © 2018 Elsevier Ltd. All rights reserved.

  20. Analysis of Bone Mineral Density According to the Biochemical Variable Markers in Adults

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sun Geun [Dept. of Radiology, Woosuk University Hospital, Chonju (Korea, Republic of); Kweon, Dae Cheol; Song, Woon Heung [Shinheung College University, Uijungbu (Korea, Republic of)

    2009-12-15

    To evaluate the bone mineral density (BMD) and biochemical markers. We evaluated the BMD of femoral neck and lumbar spines of 998(male 568, female 430) persons who took a regular health screening in Woosuk University Hospital from September 2007 to March 2008 by dual energy bone mineral densitometry. Results of BMD are different in terms of biochemical markers. Especially aged people showed osteoporotic change progressively. Degree of osteoporosis increases with age. A steep decrease of BMD can be found in postmenopausal women who have low level of female hormone. More persistent effort is needed to find out the factors that can reduce BMD values for prevention of problems by osteoporosis. In essence, research on factors related to other biochemical markers must be studied continuously.

  1. Biochemical changes in low-salt fermentation of solidstate soy sauce

    African Journals Online (AJOL)

    Low-salt solid-state fermentation soy sauce was prepared with defatted soy bean and wheat bran. Biochemical changes during the aging of the soy sauce mash were investigated. Results show that after a 15-day aging period, the contents of total nitrogen, formol titration nitrogen, free amino acids, reducing sugar, total ...

  2. A comparative study on the biochemical effect of ocular and oral ...

    African Journals Online (AJOL)

    STORAGESEVER

    2010-05-24

    May 24, 2010 ... and three months on some biochemical parameters using rabbits as animal model. The results obtained show that the kidney ..... MT is a heavy metal binding protein which is believed to influence the uptake, ... may probably be due to the omission of the portal blood system after ocular uptake of Cd. It is ...

  3. Biochemical changes in low-salt solid-state fermented soy sauce ...

    African Journals Online (AJOL)

    Low-salt solid-state fermentation soy sauce was prepared with defatted soy bean and wheat bran. Biochemical changes during the aging of the soy sauce mash were investigated. Results showed that after a 15-day aging period, the contents of total nitrogen, formol titration nitrogen, free amino acids, reducing sugar, total ...

  4. A comparative study on the biochemical effect of ocular and oral ...

    African Journals Online (AJOL)

    The objective of the present study was to compare the effect of oral and ocular exposure to cadmium for one and three months on some biochemical parameters using rabbits as animal model. The results obtained show that the kidney, femoral muscle, femur and brain of rabbits ocularly exposed to Cd had a significantly ...

  5. A sensitive branched DNA HIV-1 signal amplification viral load assay with single day turnaround.

    Directory of Open Access Journals (Sweden)

    Mark A Baumeister

    Full Text Available Branched DNA (bDNA is a signal amplification technology used in clinical and research laboratories to quantitatively detect nucleic acids. An overnight incubation is a significant drawback of highly sensitive bDNA assays. The VERSANT® HIV-1 RNA 3.0 Assay (bDNA ("Versant Assay" currently used in clinical laboratories was modified to allow shorter target incubation, enabling the viral load assay to be run in a single day. To dramatically reduce the target incubation from 16-18 h to 2.5 h, composition of only the "Lysis Diluent" solution was modified. Nucleic acid probes in the assay were unchanged. Performance of the modified assay (assay in development; not commercially available was evaluated and compared to the Versant Assay. Dilution series replicates (>950 results were used to demonstrate that analytical sensitivity, linearity, accuracy, and precision for the shorter modified assay are comparable to the Versant Assay. HIV RNA-positive clinical specimens (n = 135 showed no significant difference in quantification between the modified assay and the Versant Assay. Equivalent relative quantification of samples of eight genotypes was demonstrated for the two assays. Elevated levels of several potentially interfering endogenous substances had no effect on quantification or specificity of the modified assay. The modified assay with drastically improved turnaround time demonstrates the viability of signal-amplifying technology, such as bDNA, as an alternative to the PCR-based assays dominating viral load monitoring in clinical laboratories. Highly sensitive bDNA assays with a single day turnaround may be ideal for laboratories with especially stringent cost, contamination, or reliability requirements.

  6. Capillary Electrophoresis Analysis of Conventional Splicing Assays

    DEFF Research Database (Denmark)

    de Garibay, Gorka Ruiz; Acedo, Alberto; García-Casado, Zaida

    2014-01-01

    of these assays is often challenging. Here, we explore this issue by conducting splicing assays in 31 BRCA2 genetic variants. All variants were assessed by RT-PCR followed by capillary electrophoresis and direct sequencing. If assays did not produce clear-cut outputs (Class-2 or Class-5 according to analytical...

  7. Multicentre comparison of a diagnostic assay

    DEFF Research Database (Denmark)

    Waters, Patrick; Reindl, Markus; Saiz, Albert

    2016-01-01

    ) assays in neuromyelitis optica spectrum disorders (NMOSD). METHODS: Coded samples from patients with neuromyelitis optica (NMO) or NMOSD (101) and controls (92) were tested at 15 European diagnostic centres using 21 assays including live (n=3) or fixed cell-based assays (n=10), flow cytometry (n=4...

  8. Development of On-Line High Performance Liquid Chromatography (HPLC)-Biochemical Detection Methods as Tools in the Identification of Bioactives

    Science.gov (United States)

    Malherbe, Christiaan J.; de Beer, Dalene; Joubert, Elizabeth

    2012-01-01

    Biochemical detection (BCD) methods are commonly used to screen plant extracts for specific biological activities in batch assays. Traditionally, bioactives in the most active extracts were identified through time-consuming bio-assay guided fractionation until single active compounds could be isolated. Not only are isolation procedures often tedious, but they could also lead to artifact formation. On-line coupling of BCD assays to high performance liquid chromatography (HPLC) is gaining ground as a high resolution screening technique to overcome problems associated with pre-isolation by measuring the effects of compounds post-column directly after separation. To date, several on-line HPLC-BCD assays, applied to whole plant extracts and mixtures, have been published. In this review the focus will fall on enzyme-based, receptor-based and antioxidant assays. PMID:22489144

  9. C8-Linked Pyrrolobenzodiazepine Monomers with Inverted Building Blocks Show Selective Activity against Multidrug Resistant Gram-Positive Bacteria.

    Science.gov (United States)

    Andriollo, Paolo; Hind, Charlotte K; Picconi, Pietro; Nahar, Kazi S; Jamshidi, Shirin; Varsha, Amrit; Clifford, Melanie; Sutton, J Mark; Rahman, Khondaker Miraz

    2018-02-09

    Antimicrobial resistance has become a major global concern. Development of novel antimicrobial agents for the treatment of infections caused by multidrug resistant (MDR) pathogens is an urgent priority. Pyrrolobenzodiazepines (PBDs) are a promising class of antibacterial agents initially discovered and isolated from natural sources. Recently, C8-linked PBD biaryl conjugates have been shown to be active against some MDR Gram-positive strains. To explore the role of building block orientations on antibacterial activity and obtain structure activity relationship (SAR) information, four novel structures were synthesized in which the building blocks of previously reported compounds were inverted, and their antibacterial activity was studied. The compounds showed minimum inhibitory concentrations (MICs) in the range of 0.125-32 μg/mL against MDR Gram-positive strains with a bactericidal mode of action. The results showed that a single inversion of amide bonds reduces the activity while the double inversion restores the activity against MDR pathogens. All inverted compounds did not stabilize DNA and lacked eukaryotic toxicity. The compounds inhibit DNA gyrase in vitro, and the most potent compound was equally active against both wild-type and mutant DNA gyrase in a biochemical assay. The observed activity of the compounds against methicillin resistant S. aureus (MRSA) strains with equivalent gyrase mutations is consistent with gyrase inhibition being the mechanism of action in vivo, although this has not been definitively confirmed in whole cells. This conclusion is supported by a molecular modeling study showing interaction of the compounds with wild-type and mutant gyrases. This study provides important SAR information about this new class of antibacterial agents.

  10. Immunohistochemical and Biochemical Expression Patterns of TTF-1, RAGE, GLUT-1 and SOX2 in HCV-Associated Hepatocellular Carcinomas

    Science.gov (United States)

    Aboushousha, Tarek; Mamdouh, Samah; Hamdy, Hussam; Helal, Noha; Khorshed, Fatma; Safwat, Gehan; Seleem, Mohamed

    2018-01-27

    Objective: To investigate the expression of TTF-1, RAGE, GLUT1 and SOX2 in HCV-associated HCCs and in surrounding non-tumorous liver tissue. Material and Methods: Tissue material from partial hepatectomy cases for HCC along with corresponding serum samples and 30 control serum samples from healthy volunteers were studied. Biopsies were classified into: non-tumor hepatic tissue (36 sections); HCC (33 sections) and liver cell dysplasia (LCD) (15 sections). All cases were positive for HCV. Immunohistochemistry (IHC), gene extraction and quantitative real-time reverse-transcription assays (qRT-PCR) were applied. Results: By IHC, LCD and HCC showed significantly high percentages of positive cases with all markers. SOX2 showed significant increase with higher HCC grades, while RAGE demonstrated an inverse relation and GLUT-1 and TTF-1 lacked any correlation. In nontumorous-HCV tissue, we found significantly high TTF-1, low RAGE and negative SOX2 expression. RAGE, GLUT-1 and SOX2 show non-significant elevation positivity in high grade HCV compared to low grade lesions. TTF-1, RAGE and SOX2 exhibited low expression in cirrhosis compared to fibrosis. Biochemical studies on serum and tissue extracts revealed significant down-regulation of RAGE, GLUT-1 and SOX2 genes, as well as significant up-regulation of the TTF-1 gene in HCC cases compared to controls. All studied genes show significant correlation with HCC grade. In non-tumor tissue, only TTF-1 gene expression had a significant correlation with the fibrosis score. Conclusion: Higher expression of TTF-1, RAGE, GLUT-1 and SOX2 in HCC and dysplasia compared to non-tumor tissues indicates up-regulation of these markers as early events during the development of HCV-associated HCC. Creative Commons Attribution License

  11. Long-term stability of biochemical markers in pediatric serum specimens stored at -80 °C: a CALIPER Substudy.

    Science.gov (United States)

    Brinc, Davor; Chan, Man Khun; Venner, Allison A; Pasic, Maria D; Colantonio, David; Kyriakopolou, Lianna; Adeli, Khosrow

    2012-07-01

    Pediatric serum samples collected from healthy children in the CALIPER (Canadian Laboratory Initiative in Pediatric Reference Interval) project are stored at -80 °C for various periods of time. This study aimed to determine the stability of chemistry, protein, and hormone analytes under these conditions. Serum samples collected from children of 0-18 years of age attending outpatient clinics were pooled into a single pool or into age-group specific pools. Following baseline measurement, each pool was aliquoted and kept frozen at -80 °C until analysis. Samples were analyzed for 57 biochemical markers at monthly intervals over a 10-13 month period and each aliquot was subject to one freeze-thaw cycle before analysis. The analysis was performed on VITROS Chemistry System, COBAS INTEGRA 400 Plus and IMMULITE 2500. Values obtained at monthly intervals were compared to baseline measurements and examined for trends over time. A majority of analytes measured in this study showed no significant time-dependent change relative to baseline or trend over time after up to 13 months of storage. PTH showed up to 27.2% decline after 10 months of storage with most of the decline evident after 2 months. Most analytes showed variability over time, which is thought to reflect assay variability rather than changes in analyte stability. The study shows stability for a majority of analytes stored in serum at -80 °C after up to 13 months of storage. Samples do not require immediate testing for reference interval determination for the selected analytes with possible exception of PTH. Copyright © 2012 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  12. Assaying environmental nickel toxicity using model nematodes

    Science.gov (United States)

    Rudel, David; Douglas, Chandler; Huffnagle, Ian; Besser, John M.; Ingersoll, Christopher G.

    2013-01-01

    Although nickel exposure results in allergic reactions, respiratory conditions, and cancer in humans and rodents, the ramifications of excess nickel in the environment for animal and human health remain largely undescribed. Nickel and other cationic metals travel through waterways and bind to soils and sediments. To evaluate the potential toxic effects of nickel at environmental contaminant levels (8.9-7,600 µg Ni/g dry weight of sediment and 50-800 µg NiCl2/L of water), we conducted assays using two cosmopolitan nematodes, Caenorhabditis elegans and Pristionchus pacificus. We assayed the effects of both sediment-bound and aqueous nickel upon animal growth, developmental survival, lifespan, and fecundity. Uncontaminated sediments were collected from sites in the Midwestern United States and spiked with a range of nickel concentrations. We found that nickel-spiked sediment substantially impairs both survival from larval to adult stages and adult longevity in a concentration-dependent manner. Further, while aqueous nickel showed no adverse effects on either survivorship or longevity, we observed a significant decrease in fecundity, indicating that aqueous nickel could have a negative impact on nematode physiology. Intriguingly, C. elegansand P. pacificus exhibit similar, but not identical, responses to nickel exposure. Moreover, P. pacificus could be tested successfully in sediments inhospitable to C. elegans. Our results add to a growing body of literature documenting the impact of nickel on animal physiology, and suggest that environmental toxicological studies could gain an advantage by widening their repertoire of nematode species.

  13. Assaying environmental nickel toxicity using model nematodes.

    Directory of Open Access Journals (Sweden)

    David Rudel

    Full Text Available Although nickel exposure results in allergic reactions, respiratory conditions, and cancer in humans and rodents, the ramifications of excess nickel in the environment for animal and human health remain largely undescribed. Nickel and other cationic metals travel through waterways and bind to soils and sediments. To evaluate the potential toxic effects of nickel at environmental contaminant levels (8.9-7,600 µg Ni/g dry weight of sediment and 50-800 µg NiCl2/L of water, we conducted assays using two cosmopolitan nematodes, Caenorhabditis elegans and Pristionchus pacificus. We assayed the effects of both sediment-bound and aqueous nickel upon animal growth, developmental survival, lifespan, and fecundity. Uncontaminated sediments were collected from sites in the Midwestern United States and spiked with a range of nickel concentrations. We found that nickel-spiked sediment substantially impairs both survival from larval to adult stages and adult longevity in a concentration-dependent manner. Further, while aqueous nickel showed no adverse effects on either survivorship or longevity, we observed a significant decrease in fecundity, indicating that aqueous nickel could have a negative impact on nematode physiology. Intriguingly, C. elegans and P. pacificus exhibit similar, but not identical, responses to nickel exposure. Moreover, P. pacificus could be tested successfully in sediments inhospitable to C. elegans. Our results add to a growing body of literature documenting the impact of nickel on animal physiology, and suggest that environmental toxicological studies could gain an advantage by widening their repertoire of nematode species.

  14. Assaying environmental nickel toxicity using model nematodes.

    Science.gov (United States)

    Rudel, David; Douglas, Chandler D; Huffnagle, Ian M; Besser, John M; Ingersoll, Christopher G

    2013-01-01

    Although nickel exposure results in allergic reactions, respiratory conditions, and cancer in humans and rodents, the ramifications of excess nickel in the environment for animal and human health remain largely undescribed. Nickel and other cationic metals travel through waterways and bind to soils and sediments. To evaluate the potential toxic effects of nickel at environmental contaminant levels (8.9-7,600 µg Ni/g dry weight of sediment and 50-800 µg NiCl2/L of water), we conducted assays using two cosmopolitan nematodes, Caenorhabditis elegans and Pristionchus pacificus. We assayed the effects of both sediment-bound and aqueous nickel upon animal growth, developmental survival, lifespan, and fecundity. Uncontaminated sediments were collected from sites in the Midwestern United States and spiked with a range of nickel concentrations. We found that nickel-spiked sediment substantially impairs both survival from larval to adult stages and adult longevity in a concentration-dependent manner. Further, while aqueous nickel showed no adverse effects on either survivorship or longevity, we observed a significant decrease in fecundity, indicating that aqueous nickel could have a negative impact on nematode physiology. Intriguingly, C. elegans and P. pacificus exhibit similar, but not identical, responses to nickel exposure. Moreover, P. pacificus could be tested successfully in sediments inhospitable to C. elegans. Our results add to a growing body of literature documenting the impact of nickel on animal physiology, and suggest that environmental toxicological studies could gain an advantage by widening their repertoire of nematode species.

  15. HSimulator: Hybrid Stochastic/Deterministic Simulation of Biochemical Reaction Networks

    Directory of Open Access Journals (Sweden)

    Luca Marchetti

    2017-01-01

    Full Text Available HSimulator is a multithread simulator for mass-action biochemical reaction systems placed in a well-mixed environment. HSimulator provides optimized implementation of a set of widespread state-of-the-art stochastic, deterministic, and hybrid simulation strategies including the first publicly available implementation of the Hybrid Rejection-based Stochastic Simulation Algorithm (HRSSA. HRSSA, the fastest hybrid algorithm to date, allows for an efficient simulation of the models while ensuring the exact simulation of a subset of the reaction network modeling slow reactions. Benchmarks show that HSimulator is often considerably faster than the other considered simulators. The software, running on Java v6.0 or higher, offers a simulation GUI for modeling and visually exploring biological processes and a Javadoc-documented Java library to support the development of custom applications. HSimulator is released under the COSBI Shared Source license agreement (COSBI-SSLA.

  16. Structural and Biochemical Studies of LysM Proteins

    DEFF Research Database (Denmark)

    Wong, Mei Mei Jaslyn Elizabeth

    2017-01-01

    The Lysin Motif (LysM) is a well characterised carbohydrate-binding module that is present, usually as repeated entities, in proteins from all organisms except archaea. The functional significance of the multiplicity of the LysM module was investigated using two homologous NlpC/P60 endopeptidases...... involved in peptidoglycan hydrolysis; the Cell Wall Lytic enzyme associated with cell Separation (CwlS) from Bacillus subtilis, and P60_Tth from Thermus thermopiles. Biochemical studies conducted on purified CwlS showed that multiple LysM modules function cooperatively to bind N-acetylglucosamine (NAG......) polymers, and that the increased affinity was positively correlated to the catalytic activity of the enzyme. The cooperativity of LysM domains was further demonstrated in structural studies on P60_2LysM, a variant of P60_Tth that contains only two LysM domains and lacks the catalytic domain. Ligand...

  17. Biochemical methane potential (BMP) of artichoke waste: the inoculum effect.

    Science.gov (United States)

    Fabbri, Andrea; Serranti, Silvia; Bonifazi, Giuseppe

    2014-03-01

    The aim of this work was to investigate anaerobic digestibility of artichoke waste resulting from industrial transformation. A series of batch anaerobic digestion tests was performed in order to evaluate the biochemical methane potential of the matrix in respect of the process. A comparison of the different performances of the laboratory-scale reactors operating in mesophilic conditions and utilizing three different values of the inoculum/substrate ratio was carried out. The best performance was achieved with an inoculum/substrate ratio of 2. Artichoke-processing byproducts showed a classical organic waste decomposition behaviour: a fast start-up phase, an acclimation stage, and a final stabilization phase. Following this approach, artichoke waste reached chemical oxygen demand removal of about 90% in 40 days. The high methane yield (average 408.62 mL CH4 gvs (-1) voltatile solids), makes artichoke waste a good product to be utilized in anaerobic digestion plants for biogas production.

  18. Archaea Signal Recognition Particle Shows the Way

    Directory of Open Access Journals (Sweden)

    Christian Zwieb

    2010-01-01

    Full Text Available Archaea SRP is composed of an SRP RNA molecule and two bound proteins named SRP19 and SRP54. Regulated by the binding and hydrolysis of guanosine triphosphates, the RNA-bound SRP54 protein transiently associates not only with the hydrophobic signal sequence as it emerges from the ribosomal exit tunnel, but also interacts with the membrane-associated SRP receptor (FtsY. Comparative analyses of the archaea genomes and their SRP component sequences, combined with structural and biochemical data, support a prominent role of the SRP RNA in the assembly and function of the archaea SRP. The 5e motif, which in eukaryotes binds a 72 kilodalton protein, is preserved in most archaea SRP RNAs despite the lack of an archaea SRP72 homolog. The primary function of the 5e region may be to serve as a hinge, strategically positioned between the small and large SRP domain, allowing the elongated SRP to bind simultaneously to distant ribosomal sites. SRP19, required in eukaryotes for initiating SRP assembly, appears to play a subordinate role in the archaea SRP or may be defunct. The N-terminal A region and a novel C-terminal R region of the archaea SRP receptor (FtsY are strikingly diverse or absent even among the members of a taxonomic subgroup.

  19. THE INVESTIGATION OF BIOCHEMICAL CONTENT OF Elaeagnus angustifolia

    OpenAIRE

    yıldırım, Işıl

    2018-01-01

    AbstractStudies about herbal products are increasing every day due to their rich biochemical content. Elaeagnus angustofolia is one of the best known plant species to have a strong biochemical substance spectrum. This work was performed to identify the some biochemical content of Eleagnus angustofolia. In this study, vitamins A, E, and C, total sugar content, inverted sugar content, cellulose content, amount of total protein, and fatty acid properties were studied. Our investigations revealed...

  20. BioNessie - a grid enabled biochemical networks simulation environment

    OpenAIRE

    Liu, X.; Jiang, J.; Ajayi, O.; Gu, X.; Gilbert, D.; Sinnott, R.O.

    2008-01-01

    The simulation of biochemical networks provides insight and understanding about the underlying biochemical processes and pathways used by cells and organisms. BioNessie is a biochemical network simulator which has been developed at the University of Glasgow. This paper describes the simulator and focuses in particular on how it has been extended to benefit from a wide variety of high performance compute resources across the UK through Grid technologies to support larger scale simulations.

  1. Biochemical analysis of CTLA-4 immunoreactive material from human blood

    Directory of Open Access Journals (Sweden)

    Dennert Kate

    2009-09-01

    Full Text Available Abstract Background CTLA-4 was initially described as a membrane-bound molecule that inhibited lymphocyte activation by interacting with B7.1 and B7.2 molecules on antigen presenting cells. Alternative splicing of mRNA encoding the CTLA-4 receptor leads to the production of a molecule (sCTLA-4 that lacks a membrane anchor and is therefore secreted into the extracellular space. Despite studies finding that people with autoimmune disease more frequently express high levels of sCTLA-4 in their blood than apparently healthy people, the significance of these findings is unclear. Methods Molecules isolated from blood using CTLA-4 specific antibodies were analyzed with ligand binding assays, mass spectroscopy, and biochemical fractionation in an effort to increase our understanding of CTLA-4 immunoreactive material. Results Mass spectroscopy analysis of the molecules recognized by multiple CTLA-4-specific antibodies failed to identify any CTLA-4 protein. Even though these molecules bind to the CTLA-4 receptors B7.1 and B7.2, they also exhibit properties common to immunoglobulins. Conclusion We have identified molecules in blood that are recognized by CTLA-4 specific antibodies but also exhibit properties of immunoglobulins. Our data indicates that what has been called sCTLA-4 is not a direct product of the CTLA-4 gene, and that the CTLA-4 protein is not part of this molecule. These results may explain why the relationship of sCTLA-4 to immune system activity has been difficult to elucidate.

  2. Biochemical characterization of the GM2 gangliosidosis B1 variant

    Directory of Open Access Journals (Sweden)

    Tutor J.C.

    2004-01-01

    Full Text Available The deficiency of the A isoenzyme of ß-hexosaminidase (Hex produced by different mutations of the gene that codes for the alpha subunit (Tay-Sachs disease has two variants with enzymological differences: the B variant consists of the absence of Hex A isoenzyme and the B1 variant produces an inactive Hex A isoenzyme for the hydrolysis of the GM2 ganglioside and synthetic substrates with negative charge. In contrast to the early childhood form of the B variant, the B1 variant appears at a later clinical stage (3 to 7 years of age with neurodegenerative symptoms leading to the death of the patient in the second decade of life. The most frequent mutation responsible for the GM2 gangliosidosis B1 variant is R178H, which has a widespread geographic and ethnic distribution. The highest incidence has been described in Portugal, which has been suggested as the point of origin of this mutation. Biochemical characterization of this lysosomal disease is carried out using negatively charged synthetic alpha subunit-specific sulfated substrates, since Hex A isoenzyme heat-inactivation assays are not applicable. However, the determination of the apparent activation energy of Hex using the neutral substrate 3,3'-dichlorophenolsulfonphthaleinyl N-acetyl-ß-D-glucosaminide, may offer a valid alternative. The presence of an alpha subunit in the alphaß heterodimer Hex A means that its activation energy (41.8 kJ/mol is significantly lower than that of the ßß homodimer Hex B (75.1 kJ/mol; however, as mutation inactivates the alpha subunit, the Hex A of the B1 variant presents an activation energy that is similar to that of the Hex B isoenzyme.

  3. Non destructive assay (NDA) techniques

    International Nuclear Information System (INIS)

    Mafra Guidicini, Olga; Llacer, Carlos D.; Rojo, Marcelo

    2001-01-01

    In the IAEA Safeguards System the basic verification method used is nuclear material accountancy, with containment and surveillance as important complementary measures. If nuclear material accountancy is to be effective, IAEA inspectors have to make independent measurements to verify declared material quantities. Most of the equipment available to the inspectors is designed to measure gamma rays and/or neutrons emitted by various nuclear materials. Equipment is also available to measure the gross weight of an item containing nuclear material. These types of measurement techniques are generally grouped under the title of nondestructive assay (NDA). The paper describes the NDA techniques and instruments used to verify the total amount of nuclear material held at a nuclear facility. (author)

  4. Micronutrient influence in Nannochloropsis sp. growth and biochemical profile

    Directory of Open Access Journals (Sweden)

    Catarina Rosado Correia

    2014-06-01

    Full Text Available Microalgae are a heterogeneous group, which includes various and distinct taxa. However, few genera are actively used in aquaculture, due to the lack of specific requirements, like adequate size, shape and non-toxic. Nannochloropsis sp. is one of the main microalgae used in aquaculture, due to its nutritional profile and for being a fast-growing microalgae. Despite its characteristics, large-scale culture of Nannochloropsis sp. is in constant improvement, in order to accomplish the best productivity, combined with an adequate biochemical profile. In an attempt to achieve this, an experimental set-up was tested with microalgae grown in four different culture media: (1 NutriBloom [NB] (commercial medium used at Necton’s facilities, (2 NutriBloom without cobalt [NB w/Co], (3 Simplex [S] (no addition of iron or any micronutrient and (4 Sea Mineral Solution [SMS]. Nannochloropsis’s contents of protein, carbohydrates, total lipid and PUFA’s were controlled at logarithmic and stationary phases of growth, using classical techniques, as mentioned in Lowry (1951, Dubois (1956, Bligh and Dyer (1959 and Lepage & Roy (1986. Higher productivity, dry weight and number of cells were achieved when cultured in NB and NB w/Co. In the logarithmic phase, NB w/Co presented the higher protein content, SMS showed the higher percentage of lipid and polyunsaturated fatty acids (PUFAS. In the stationary phase, S medium showed higher lipid percentage; nevertheless, NB medium presented the higher content of protein and PUFAs. In both phases, NB medium had the higher sugar content. Differences between micronutrient concentrations explain the verified variations in the microalgae’s biochemical profile, particularly iron, copper, cobalt and molybdenium variations.

  5. Biochemical Hypermedia: Glucose as a Central Molecule in Metabolism

    Directory of Open Access Journals (Sweden)

    J.K. Sugai

    2008-05-01

    Full Text Available The technologies of information, together with education resources, have been pointed out as a solution to the improvement of teaching approach, but they still claim for programs to fulfill the demands of didactic materials. So, a biochemical software was developed aiming to contribute for the better understanding of the glycolysis. It was prepared with the help of concept maps, ISIS Draw, ADOBE Photoshop and FLASH MX Program. The introduction screen shows a teacher in a theater presenting glucose as a central molecule in the metabolism of animals, plants and many microorganisms. She invites for a better knowledge of glucose through a view of its discovery and its metabolism. A step by step animation process shows the interaction of glucose in aerobic conditions with the enzymes of the glycolytic pathways and its products. An explanation text of each enzyme catalytic process is provided by links. A static pathway is always available through a link. The fates of pyruvate yielding lactic acid and ethanol under anaerobic conditions are shown as well. The overall reactions of gluconeogenesis and the functional significance of this pathway are presented. The experimental treatment involved the presentation of this hypermedia for Nutrition undergraduate students (UFSC as a tool for better comprehension of the theme. The students revealed that it was extremely effective in promoting the understanding of the enzymatic mechanisms involved in glycolysis. This suggests that there is a significant added value in employing the software as an instructional effort to enhance student’s abilities to understand biochemical pathways.

  6. Biochemical Measurements of the Human Stress Response

    Science.gov (United States)

    1984-03-01

    8217 biochemical stress response. Dr Thomas Longridge was the project scientist and Dr Joe De Maio was the task scientist. This research was conducted by the...under condition A described in next paragraph. Second, DOPAC, MHPG, HVA, 5-HIAA, and VMA were determined by a modified method of Joseph, Kadam , and...n tt Ln 0 0 vJ 0 (N ’lw %.D w 0 0) to CI la, co 0 (r) Ch 𔃺I I 0 𔃺 0 0 0 (’) v 01-4 V~ 4 Hn qqJ Ln NOD & OD N r- ai r, m 0 ~ $4 c4 H 0 OI H r) (n t

  7. Prions: the danger of biochemical weapons

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    Eric Almeida Xavier

    2014-09-01

    Full Text Available The knowledge of biotechnology increases the risk of using biochemical weapons for mass destruction. Prions are unprecedented infectious pathogens that cause a group of fatal neurodegenerative diseases by a novel mechanism. They are transmissible particles that are devoid of nucleic acid. Due to their singular characteristics, Prions emerge as potential danger since they can be used in the development of such weapons. Prions cause fatal infectious diseases, and to date there is no therapeutic or prophylactic approach against these diseases. Furthermore, Prions are resistant to food-preparation treatments such as high heat and can find their way from the digestive system into the nervous system; recombinant Prions are infectious either bound to soil particles or in aerosols. Therefore, lethal Prions can be developed by malicious researchers who could use it to attack political enemies since such weapons cause diseases that could be above suspicion.

  8. Biochemical composition of marine monogenean parasite eggs.

    Science.gov (United States)

    Brazenor, Alexander K; Francis, David S; Hutson, Kate S; Carton, Alexander G

    2017-12-01

    This study on the eggs of the tropical monogenean Neobenedenia girellae presents the first detailed quantitative biochemical information of a marine parasite species' eggs. Moisture and protein composed the majority of the contents of freshly laid eggs (79.12±0.82 and 11.51±0.49% respectively) followed by lipid (2.50±0.15%). Lipids were composed of approximately equal amounts of saturated, monounsaturated, and polyunsaturated fatty acids and the predominant lipid class was triacylglycerol (33.82±1.20%). This study represents a fundamental step towards a better understanding of the early life biology of this important species of parasite. Copyright © 2017. Published by Elsevier B.V.

  9. Biochemical Basis of Sestrin Physiological Activities

    Energy Technology Data Exchange (ETDEWEB)

    Ho, Allison; Cho, Chun-Seok; Namkoong, Sim; Cho, Uhn-Soo; Lee, Jun Hee (Michigan)

    2016-05-10

    Excessive accumulation of reactive oxygen species (ROS) and chronic activation of mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) are well-characterized promoters of aging and age-associated degenerative pathologies. Sestrins, a family of highly conserved stress-inducible proteins, are important negative regulators of both ROS and mTORC1 signaling pathways; however, the mechanistic basis of how Sestrins suppress these pathways remains elusive. In the past couple of years, breakthrough discoveries about Sestrin signaling and its molecular nature have markedly increased our biochemical understanding of Sestrin function. These discoveries have also uncovered new potential therapeutic strategies that may eventually enable us to attenuate aging and age-associated diseases.

  10. Highly valuable microalgae: biochemical and topological aspects.

    Science.gov (United States)

    Pignolet, Olivier; Jubeau, Sébastien; Vaca-Garcia, Carlos; Michaud, Philippe

    2013-08-01

    The past decade has seen a surge in the interest in microalgae culture for biodiesel production and other applications as renewable biofuels as an alternative to petroleum transport fuels. The development of new technologies for the culture of these photosynthetic microorganisms and improved knowledge of their biochemical composition has spurred innovation in the field of high-value biomolecules. These developments are only economically viable if all the microalgae fractions are valorized in a biorefinery strategy. Achieving this objective requires an understanding of microalgae content and the cellular localization of the main biomolecular families in order to develop efficient harvest and sequential recovery technologies. This review summarizes the state of the art in microalgae compositions and topologies using some examples of the main industrially farmed microalgae.

  11. Physico-Biochemical aspects of Shock

    Directory of Open Access Journals (Sweden)

    Kulkarni M.D.

    2009-08-01

    Full Text Available Shock is a clinical condition characterized by decreased blood flow to vital organs due to imbalance between size of vascular bed and effective circulating blood volume and the inability of body tissues to metabolize nutrients normally. The decreased blood flow to vital organs like kidney, liver, spleen, brain etc. is caused by pulling and stagnation blood elsewhere in circulation. The defective blood flow to tissues implies incomplete oxygen supply to the cells, tissues and organs consequent effect being interference with metabolism. It is very essential to study physicobiochemical aspect of shock because after knowing the pathways involved in various physio -pathological processes, we can undertake effective treatment and there by shock can be satisfactorily treated and prevented. Following are the various physico-biochemical processes stated by various workers to study the shock. [Vet. World 2009; 2(4.000: 161-162

  12. Hemoglobin Variants: Biochemical Properties and Clinical Correlates

    Science.gov (United States)

    Thom, Christopher S.; Dickson, Claire F.; Gell, David A.; Weiss, Mitchell J.

    2013-01-01

    Diseases affecting hemoglobin synthesis and function are extremely common worldwide. More than 1000 naturally occurring human hemoglobin variants with single amino acid substitutions throughout the molecule have been discovered, mainly through their clinical and/or laboratory manifestations. These variants alter hemoglobin structure and biochemical properties with physiological effects ranging from insignificant to severe. Studies of these mutations in patients and in the laboratory have produced a wealth of information on hemoglobin biochemistry and biology with significant implications for hematology practice. More generally, landmark studies of hemoglobin performed over the past 60 years have established important paradigms for the disciplines of structural biology, genetics, biochemistry, and medicine. Here we review the major classes of hemoglobin variants, emphasizing general concepts and illustrative examples. PMID:23388674

  13. Detection of Streptococcus pyogenes using rapid visual molecular assay.

    Science.gov (United States)

    Zhao, Xiangna; He, Xiaoming; Li, Huan; Zhao, Jiangtao; Huang, Simo; Liu, Wei; Wei, Xiao; Ding, Yiwei; Wang, Zhaoyan; Zou, Dayang; Wang, Xuesong; Dong, Derong; Yang, Zhan; Yan, Xiabei; Huang, Liuyu; Du, Shuangkui; Yuan, Jing

    2015-09-01

    Streptococcus pyogenes is an increasingly important pathogen in many parts of the world. Rapid and accurate detection of S. pyogenes aids in the control of the infection. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed and validated for the specific detection of S. pyogenes. The assay incorporates two methods: a chromogenic analysis using a calcein/Mn(2+) complex and real-time turbidity monitoring to assess the reaction. Both methods detected the target DNA within 60 min under 64°C isothermal conditions. The assay used specifically designed primers to target spy1258, and correctly identified 111 strains of S. pyogenes and 32 non-S. pyogenes strains, including other species of the genus Streptococcus. Tests using reference strains showed that the LAMP assay was highly specific. The sensitivity of the assay, with a detection limit of 1.49 pg DNA, was 10-fold greater than that of PCR. The LAMP assay established in this study is simple, fast and sensitive, and does not rely upon any special equipment; thus, it could be employed in clinical diagnosis. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  14. Determination of Nitrate Reductase Assay Depending on the Microbial Growth

    International Nuclear Information System (INIS)

    El-Kabbany, H.M.

    2012-01-01

    A rapid micro-dilution assay for determination of the antimicrobial susceptibility of different bacterial isolates was developed. This assay is based on the ability of the most of viable organisms to reduce nitrates. The MIC or MBC could be determined by nitrate reductase (NR) only after 30 to 90 min of incubation depending on the behaviour of microbial growth. Bacterial viability is detected by a positive nitrite reduction rather than visible turbidity. The nitrate reduction assay was compared with standard micro-assay using 250 isolates of different taxa against 10 antibiotics belonging to different classes. An excellent agreement of 82.5 % was found between the two methods and only 17.5 % of 1794 trials showed difference in the determined MIC by tow-dilution interval above or below the MIC determined by the turbidimetric method under the same test conditions. However, the nitrate reduction assay was more rapid and sensitive in detecting viable bacteria and so, established an accurate estimate of the minimal inhibitory concentration (MIC) or the minimal bacterial concentration (MBC). The nitrate reduction assay offers the additional advantage that it could be used to determine the MBC without having to subculture the broth. 232 cases of resistance were detected by NR and 4 different media were tested for susceptibility test. The bacterial isolates were exposed to ultra violet (UV) light for different period

  15. Biochemical Characterization of the Lactobacillus reuteri Glycoside Hydrolase Family 70 GTFB Type of 4,6-α-Glucanotransferase Enzymes That Synthesize Soluble Dietary Starch Fibers.

    Science.gov (United States)

    Bai, Yuxiang; van der Kaaij, Rachel Maria; Leemhuis, Hans; Pijning, Tjaard; van Leeuwen, Sander Sebastiaan; Jin, Zhengyu; Dijkhuizen, Lubbert

    2015-10-01

    4,6-α-Glucanotransferase (4,6-α-GTase) enzymes, such as GTFB and GTFW of Lactobacillus reuteri strains, constitute a new reaction specificity in glycoside hydrolase family 70 (GH70) and are novel enzymes that convert starch or starch hydrolysates into isomalto/maltopolysaccharides (IMMPs). These IMMPs still have linear chains with some α1→4 linkages but mostly (relatively long) linear chains with α1→6 linkages and are soluble dietary starch fibers. 4,6-α-GTase enzymes and their products have significant potential for industrial applications. Here we report that an N-terminal truncation (amino acids 1 to 733) strongly enhances the soluble expression level of fully active GTFB-ΔN (approximately 75-fold compared to full-length wild type GTFB) in Escherichia coli. In addition, quantitative assays based on amylose V as the substrate are described; these assays allow accurate determination of both hydrolysis (minor) activity (glucose release, reducing power) and total activity (iodine staining) and calculation of the transferase (major) activity of these 4,6-α-GTase enzymes. The data show that GTFB-ΔN is clearly less hydrolytic than GTFW, which is also supported by nuclear magnetic resonance (NMR) analysis of their final products. From these assays, the biochemical properties of GTFB-ΔN were characterized in detail, including determination of kinetic parameters and acceptor substrate specificity. The GTFB enzyme displayed high conversion yields at relatively high substrate concentrations, a promising feature for industrial application. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  16. Dynamic analysis of biochemical network using complex network method

    Directory of Open Access Journals (Sweden)

    Wang Shuqiang

    2015-01-01

    Full Text Available In this study, the stochastic biochemical reaction model is proposed based on the law of mass action and complex network theory. The dynamics of biochemical reaction system is presented as a set of non-linear differential equations and analyzed at the molecular-scale. Given the initial state and the evolution rules of the biochemical reaction system, the system can achieve homeostasis. Compared with random graph, the biochemical reaction network has larger information capacity and is more efficient in information transmission. This is consistent with theory of evolution.

  17. Making transuranic assay measurements using modern controllers

    International Nuclear Information System (INIS)

    Kuckertz, T.H.; Caldwell, J.T.; Medvick, P.A.; Kunz, W.E.; Hastings, R.D.

    1987-01-01

    This paper describes methodology and computer-controlled instrumentation developed at the Los Alamos National Laboratory that accurately performs nondestructive assays of large containers bearing transuranic wastes and nonradioactive matrix materials. These assay systems can measure fissile isotopes with 1-mg sensitivity and spontaneous neutron-emitting isotopes at a 10-mg sensitivity. The assays are performed by neutron interrogation, detection, and counting in a custom assay chamber. An International Business Machines Personal Computer (IBM-PC) is used to control the CAMAC-based instrumentation system that acquires the assay data. 6 refs., 7 figs

  18. Pattern Selection by Dynamical Biochemical Signals

    Science.gov (United States)

    Palau-Ortin, David; Formosa-Jordan, Pau; Sancho, José M.; Ibañes, Marta

    2015-01-01

    The development of multicellular organisms involves cells to decide their fate upon the action of biochemical signals. This decision is often spatiotemporally coordinated such that a spatial pattern arises. The dynamics that drive pattern formation usually involve genetic nonlinear interactions and positive feedback loops. These complex dynamics may enable multiple stable patterns for the same conditions. Under these circumstances, pattern formation in a developing tissue involves a selection process: why is a certain pattern formed and not another stable one? Herein we computationally address this issue in the context of the Notch signaling pathway. We characterize a dynamical mechanism for developmental selection of a specific pattern through spatiotemporal changes of the control parameters of the dynamics, in contrast to commonly studied situations in which initial conditions and noise determine which pattern is selected among multiple stable ones. This mechanism can be understood as a path along the parameter space driven by a sequence of biochemical signals. We characterize the selection process for three different scenarios of this dynamical mechanism that can take place during development: the signal either 1) acts in all the cells at the same time, 2) acts only within a cluster of cells, or 3) propagates along the tissue. We found that key elements for pattern selection are the destabilization of the initial pattern, the subsequent exploration of other patterns determined by the spatiotemporal symmetry of the parameter changes, and the speeds of the path compared to the timescales of the pattern formation process itself. Each scenario enables the selection of different types of patterns and creates these elements in distinct ways, resulting in different features. Our approach extends the concept of selection involved in cellular decision-making, usually applied to cell-autonomous decisions, to systems that collectively make decisions through cell

  19. Biochemically enhanced methane production from coal

    Science.gov (United States)

    Opara, Aleksandra

    For many years, biogas was connected mostly with the organic matter decomposition in shallow sediments (e.g., wetlands, landfill gas, etc.). Recently, it has been realized that biogenic methane production is ongoing in many hydrocarbon reservoirs. This research examined microbial methane and carbon dioxide generation from coal. As original contributions methane production from various coal materials was examined in classical and electro-biochemical bench-scale reactors using unique, developed facultative microbial consortia that generate methane under anaerobic conditions. Facultative methanogenic populations are important as all known methanogens are strict anaerobes and their application outside laboratory would be problematic. Additional testing examined the influence of environmental conditions, such as pH, salinity, and nutrient amendments on methane and carbon dioxide generation. In 44-day ex-situ bench-scale batch bioreactor tests, up to 300,000 and 250,000 ppm methane was generated from bituminous coal and bituminous coal waste respectively, a significant improvement over 20-40 ppm methane generated from control samples. Chemical degradation of complex hydrocarbons using environmentally benign reagents, prior to microbial biodegradation and methanogenesis, resulted in dissolution of up to 5% bituminous coal and bituminous coal waste and up to 25% lignite in samples tested. Research results confirm that coal waste may be a significant underutilized resource that could be converted to useful fuel. Rapid acidification of lignite samples resulted in low pH (below 4.0), regardless of chemical pretreatment applied, and did not generate significant methane amounts. These results confirmed the importance of monitoring and adjusting in situ and ex situ environmental conditions during methane production. A patented Electro-Biochemical Reactor technology was used to supply electrons and electron acceptor environments, but appeared to influence methane generation in a

  20. Robust simplifications of multiscale biochemical networks

    Directory of Open Access Journals (Sweden)

    Zinovyev Andrei

    2008-10-01

    Full Text Available Abstract Background Cellular processes such as metabolism, decision making in development and differentiation, signalling, etc., can be modeled as large networks of biochemical reactions. In order to understand the functioning of these systems, there is a strong need for general model reduction techniques allowing to simplify models without loosing their main properties. In systems biology we also need to compare models or to couple them as parts of larger models. In these situations reduction to a common level of complexity is needed. Results We propose a systematic treatment of model reduction of multiscale biochemical networks. First, we consider linear kinetic models, which appear as "pseudo-monomolecular" subsystems of multiscale nonlinear reaction networks. For such linear models, we propose a reduction algorithm which is based on a generalized theory of the limiting step that we have developed in 1. Second, for non-linear systems we develop an algorithm based on dominant solutions of quasi-stationarity equations. For oscillating systems, quasi-stationarity and averaging are combined to eliminate time scales much faster and much slower than the period of the oscillations. In all cases, we obtain robust simplifications and also identify the critical parameters of the model. The methods are demonstrated for simple examples and for a more complex model of NF-κB pathway. Conclusion Our approach allows critical parameter identification and produces hierarchies of models. Hierarchical modeling is important in "middle-out" approaches when there is need to zoom in and out several levels of complexity. Critical parameter identification is an important issue in systems biology with potential applications to biological control and therapeutics. Our approach also deals naturally with the presence of multiple time scales, which is a general property of systems biology models.

  1. Revealing the ability of a novel polysaccharide bioflocculant in bioremediation of heavy metals sensed in a Vibrio bioluminescence reporter assay.

    Science.gov (United States)

    Sajayan, Arya; Seghal Kiran, G; Priyadharshini, S; Poulose, Navya; Selvin, Joseph

    2017-09-01

    A bioflocculant-producing bacterial strain, designated MSI021, was isolated from the marine sponge Dendrilla nigra and demonstrated 94% flocculation activity in a kaolin clay suspension. MSI021 was identified as Bacillus cereus based on phylogenetic affiliation and biochemical characteristics. The purified extra-cellular bioflocculant was chemically elucidated as a polysaccharide molecule. The polysaccharide bioflocculant was stable under both acidic and alkaline conditions (pH 2.0-10.0) and temperatures up to 100 °C. The purified bioflocculant efficiently nucleated the formation of silver nanoparticles which showed broad spectrum antibacterial activity. The ability of the bioflocculant to remediate heavy metal toxicity was evaluated by measuring the inhibition of bioluminescence expression in Vibrio harveyi. Enrichment of heavy metals such as zinc, mercury and copper at concentrations of 1, 2 and 3 mM in culture media showed significant reduction of bioluminescence in Vibrio, whereas media enriched with heavy metals and bioflocculant showed dose dependent improvement in the expression of bioluminescence. The assay results demonstrated that the polysaccharide bioflocculant effectively mitigates heavy metal toxicity, thereby improving the expression of bioluminescence in Vibrio. This bioluminescence reporter assay can be developed into a high-throughput format to monitor and evaluate of heavy metal toxicity. The findings of this study revealed that a novel polysaccharide bioflocculant produced by a marine B. cereus demonstrated strong flocculating performance and was effective in nucleating the formation antibacterial silver nanoparticles and removing heavy metals. These results suggest that the MSI021 polysaccharide bioflocculant can be used to develop greener waste water treatment systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Biochemical and morphological changes in endothelial cells in response to hypoxic interstitial edema

    Directory of Open Access Journals (Sweden)

    Miserocchi Giuseppe

    2006-01-01

    Full Text Available Abstract Background A correlation between interstial pulmonary matrix disorganization and lung cellular response was recently documented in cardiogenic interstitial edema as changes in the signal-cellular transduction platforms (lipid microdomains: caveoale and lipid rafts. These findings led to hypothesize a specific "sensing" function by lung cells resulting from a perturbation in cell-matrix interaction. We reason that the cell-matrix interaction may differ between the cardiogenic and the hypoxic type of lung edema due to the observed difference in the sequential degradation of matrix proteoglycans (PGs family. In cardiogenic edema a major fragmentation of high molecular weight PGs of the interfibrillar matrix was found, while in hypoxia the fragmentation process mostly involved the PGs of the basement membrane controlling microvascular permeability. Based on these considerations, we aim to describe potential differences in the lung cellular response to the two types of edema. Methods We analysed the composition of plasma membrane and of lipid microdomains in lung tissue samples from anesthetized rabbits exposed to mild hypoxia (12 % O2 for 3–5 h causing interstitial lung edema. Lipid analysis was performed by chromatographic techniques, while protein analysis by electrophoresis and Western blotting. Lipid peroxidation was assessed on total plasma membranes by a colorimetric assay (Bioxytech LPO-586, OxisResearch. Plasma membrane fluidity was also assessed by fluorescence. Lipid microdomains were isolated by discontinuous sucrose gradient. We also performed a morphometric analysis on lung cell shape on TEM images from lung tissue specimen. Results After hypoxia, phospholipids content in plasma membranes remained unchanged while the cholesterol/phospholipids ratio increased significantly by about 9% causing a decrease in membrane fluidity. No significant increase in lipid peroxidation was detected. Analysis of lipid microdomains showed a

  3. Pi overlapping ring systems contained in a homogeneous assay: a novel homogeneous assay for antigens

    Science.gov (United States)

    Kidwell, David A.

    1993-05-01

    A novel immunoassay, Pi overlapping ring systems contained in a homogeneous assay (PORSCHA), is described. This assay relies upon the change in fluorescent spectral properties that pyrene and its derivatives show with varying concentration. Because antibodies and other biomolecules can bind two molecules simultaneously, they can change the local concentration of the molecules that they bind. This concentration change may be detected spectrally as a change in the fluorescence emission wavelength of an appropriately labeled biomolecule. Several tests of PORSCHA have been performed which demonstrate this principle. For example: with streptavidin as the binding biomolecule and a biotin labeled pyrene derivative, the production of the excimer emitting at 470 nm is observed. Without the streptavidin present, only the monomer emitting at 378 and 390 nm is observed. The ratio of monomer to excimer provides the concentration of unlabeled biotin in the sample. Approximately 1 ng/mL of biotin may be detected with this system using a 50 (mu) l sample (2 X 10-16 moles biotin). The principles behind PORSCHA, the results with the streptavidin/biotin system are discussed and extensions of the PORSCHA concept to antibodies as the binding partner and DNA in homogeneous assays are suggested.

  4. A functional assay to measure postsynaptic gamma-aminobutyric acidB responses in cultured spinal cord neurons: Heterologous regulation of the same K+ channel

    Energy Technology Data Exchange (ETDEWEB)

    Kamatchi, G.L.; Ticku, M.K. (Univ. of Texas Health Science Center, San Antonio (USA))

    1991-02-01

    The stimulation of postsynaptic gamma-aminobutyric acid (GABA)B receptors leads to slow inhibitory postsynaptic potentials due to the influx of K(+)-ions. This was studied biochemically, in vitro in mammalian cultured spinal cord neurons by using 86Rb as a substitute for K+. (-)-Baclofen, a GABAB receptor agonist, produced a concentration-dependent increase in the 86Rb-influx. This effect was stereospecific and blocked by GABAB receptor antagonists like CGP 35 348 (3-aminopropyl-diethoxymethyl-phosphonic acid) and phaclofen. Apart from the GABAB receptors, both adenosine via adenosine1 receptors and 5-hydroxytryptamine (5-HT) via 5-HT1 alpha agonists also increased the 86Rb-influx. These agonists failed to show any additivity between them when they were combined in their maximal concentration. In addition, their effect was antagonized specifically by their respective antagonists without influencing the others. These findings suggest the presence of GABAB, adenosine1 and 5-HT1 alpha receptors in the cultured spinal cord neurons, which exhibit a heterologous regulation of the same K(+)-channel. The effect of these agonists were antagonized by phorbol 12,13-didecanoate, an activator of protein kinase C, and pretreatment with pertussis toxin. This suggests that these agonists by acting on their own receptors converge on the same K(+)-channel through the Gi/Go proteins. In summary, we have developed a biochemical functional assay for studying and characterizing GABAB synaptic pharmacology in vitro, using spinal cord neurons.

  5. Different clinical and biochemical phenotypes resulting from the same substitution at the same glycine residue in different chains of the type I collagen molecule

    Energy Technology Data Exchange (ETDEWEB)

    Paepe, A. De; Nuytinck, L. [Univ. of Gent, Brussels (Belgium); Spotila, L. [Jefferson Institute of Molecular Medicine, Philadelphia, PA (United States)] [and others

    1994-09-01

    Mutations in type I collagen produce osteogenesis imperfecta (OI; brittle bone disease), ranging in severity from lethal to very mild. This phenotypic variation is largely determined by the position and nature of the mutation. Because type I collagen consists of two {alpha}1 chains and one {alpha}2 chain, {alpha}1(I) mutations are generally regarded to have more serious consequences than {alpha}2(I) mutations. We have characterized a point mutation causing substitution of serine for glycine at position 661 of the {alpha}1(I) chain in a child with severe OI. This is precisely the same substitution that had been detected in the {alpha}2(I) chain in a woman with post-menopausal osteoporosis. She and two of her sons were heterozygous and the third son was homozygous as a result of uniparental disomy. Biochemical collagen profiles were studied in each of the patients and compared with a control. Medium and cell-layer collagens were overmodified in all patients. Overmodification was pronounced in the patient with the {alpha}1(I) mutation, but mild in the patients with the {alpha}2(I) mutation, being slightly less evident in the heterozygotes than in the homozygote. Thermal stability assays showed that the melting temperature of the mutant {alpha}1(I) chains was reduced by 3{degrees}C, whereas the melting curves in the patients with the {alpha}2(I) mutation were not significantly different from the control. These results show that the type of {alpha}-chain harboring the mutation influences the fundamental biochemical behavior of type I collagen molecules and strikingly emphasizes the predominant role of {alpha}1(I) chains compared with {alpha}2(I) in this respect.

  6. Biofertilizing efficiency of Sargassum polycystum extract on growth and biochemical composition of Vigna radiata and Vigna mungo

    Directory of Open Access Journals (Sweden)

    Bharath B

    2018-01-01

    Full Text Available Objective: To evaluate the effect of marine brown alga Sargassum polycystum extract on growth and biochemical parameters of Vigna radiata and Vigna mungo. Methods: Different concentrations of algal extracts (0.5%, 1.0%, 2.0%, 3.0%, 4.0%, and 5.0% were prepared and applied to the crops at every 10-day intervals under natural conditions. After 30 d, the plants were harvested to evaluate the growth and biochemical parameters. Results: Seaweed liquid fertilizers treated seedlings showed maximum growth in 3.0% concentration when compared to the untreated seedlings. Similarly, biochemical parameters such as photosynthetic pigments, protein, reducing sugar, total sugar and amino acids exhibited increases in 3.0% concentration seaweed extract. Decreases in growth and biochemical parameters were noticed in concentrations higher than 3.0%. Conclusions: Presence of micronutrients and growth regulating substances in the liquid extract help healthier and faster productivity of the crop.

  7. Application of the microbiological method DEFT/APC and DNA comet assay to detect ionizing radiation processing of minimally processed vegetables

    International Nuclear Information System (INIS)

    Araujo, Michel Mozeika

    2008-01-01

    Marketing of minimally processed vegetables (MPV) are gaining impetus due to its convenience, freshness and apparent healthy. However, minimal processing does not reduce pathogenic microorganisms to safe levels. Food irradiation is used to extend the shelf life and inactivation of food-borne pathogens, Its combination with minimal processing could improve the safety and quality of MPV. Two different food irradiation detection methods, a biological, the DEFT/APC, and another biochemical, the DNA Comet Assay were applied to MPV in order to test its applicability to detect irradiation treatment. DEFT/APC is a microbiological screening method based on the use of the direct epi fluorescent filter technique (DEFT) and the aerobic plate count (APC). DNA Comet Assay detects DNA damage due to ionizing radiation. Samples of lettuce, chard, watercress, dandelion, kale, chicory, spinach, cabbage from retail market were irradiated O.5 kGy and 1.0 kGy using a 60 Co facility. Irradiation treatment guaranteed at least 2 log cycle reduction for aerobic and psychotropic microorganisms. In general, with increasing radiation doses, DEFT counts remained similar independent of irradiation processing while APC counts decreased gradually. The difference of the two counts gradually increased with dose increment in all samples. It could be suggested that a DEFT/APC difference over 2.0 log would be a criteria to judge if a MPV was treated by irradiation. DNA Comet Assay allowed distinguishing non-irradiated samples from irradiated ones, which showed different types of comets owing to DNA fragmentation. Both DEFT/APC method and DNA Comet Assay would be satisfactorily used as a screening method for indicating irradiation processing. (author)

  8. Development of a new oxygen consumption rate assay in cultures of Acanthamoeba (Protozoa: Lobosea) and its application to evaluate viability and amoebicidal activity in vitro.

    Science.gov (United States)

    Heredero-Bermejo, I; Criado-Fornelio, A; Soliveri, J; Díaz-Martín, J A; Matilla-Fuentes, J; Sánchez-Arias, J A; Copa-Patiño, J L; Pérez-Serrano, J

    2015-08-01

    A new fluorometric method has been developed for measuring the oxygen consumption rate (OCR) of Acanthamoeba cultures in microplates and for screening molecules with amoebicidal activity against this microorganism. The use of a biofunctional matrix (containing an oxygen-sensitive fluorogenic probe) attached to the microplate wells allowed continuous measurement of OCR in the medium, hence assessment of amoebic growth. The new OCR method applied to cell viability yielded a linear relationship and monitoring was much quicker than with indirect viability assays previously used. In addition, two drugs were tested in a cytotoxicity assay monitored by the new OCR viability test. With this procedure, the standard amoebicidal drug chlorhexidine digluconate showed an IC50 of 3.53 + 1.3 mg/l against Acanthamoeba polyphaga and 3.19 + 1.2 mg/l against Acanthamoeba castellanii, whereas a cationic dendrimer [G1Si(NMe3+)4] showed an IC50 of 6.42 + 1.3 mg/l against A. polyphaga. These data agree with previous studies conducted in our laboratory. Therefore, the new OCR method has proven powerful and quick for amoebicidal drug screening and is likely to be applied in biochemical studies concerning protozoa respiration and metabolism. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Biochemical and proteomic analyses of the physiological response induced by individual housing in gilts provide new potential stress markers.

    Science.gov (United States)

    Marco-Ramell, Anna; Arroyo, Laura; Peña, Raquel; Pato, Raquel; Saco, Yolanda; Fraile, Lorenzo; Bendixen, Emøke; Bassols, Anna

    2016-11-25

    The objective assessment of animal stress and welfare requires proper laboratory biomarkers. In this work, we have analyzed the changes in serum composition in gilts after switching their housing, from pen to individual stalls, which is generally accepted to cause animal discomfort. Blood and saliva samples were collected a day before and up to four days after changing the housing system. Biochemical analyses showed adaptive changes in lipid and protein metabolism after the housing switch, whereas cortisol and muscular markers showed a large variability between animals. 2D-DIGE and iTRAQ proteomic approaches revealed variations in serum protein composition after changing housing and diet of gilts. Both techniques showed alterations in two main homeostatic mechanisms: the innate immune and redox systems. The acute phase proteins haptoglobin, apolipoprotein A-I and α1-antichymotrypsin 3, and the antioxidant enzyme peroxiredoxin 2 were found differentially expressed by 2D-DIGE. Other proteins related to the innate immune system, including lactotransferrin, protegrin 3 and galectin 1 were also identified by iTRAQ, as well as oxidative stress enzymes such as peroxiredoxin 2 and glutathione peroxidase 3. Proteomics also revealed the decrease of apolipoproteins, and the presence of intracellular proteins in serum, which may indicate physical injury to tissues. Housing of gilts in individual stalls and diet change increase lipid and protein catabolism, oxidative stress, activate the innate immune system and cause a certain degree of tissue damage. We propose that valuable assays for stress assessment in gilts may be based on a score composed by a combination of salivary cortisol, lipid metabolites, innate immunity and oxidative stress markers and intracellular proteins.

  10. Expert system for transuranic waste assay

    International Nuclear Information System (INIS)

    Zoolalian, M.L.; Gibbs, A.; Kuhns, J.D.

    1989-01-01

    Transuranic wastes are generated at the Savannah River Site (SRS) as a result of routine production of nuclear materials. These wastes contain Pu-238 and Pu-239 and are placed into lined 55-gallon waste drums. The drums are placed on monitored storage pads pending shipment to the Waste Isolation Pilot Plant in New Mexico. A passive-active neutron (PAN) assay system is used to determine the mass of the radioactive material within the waste drums. Assay results are used to classify the wastes as either low-level or transuranic (TRU). During assays, the PAN assay system communicates with an IBM-AT computer. A Fortran computer program, called NEUT, controls and performs all data analyses. Unassisted, the NEUT program cannot adequately interpret assay results. To eliminate this limitation, an expert system shell was used to write a new algorithm, called the Transuranic Expert System (TRUX), to drive the NEUT program and add decision making capabilities for analysis of the assay results. The TRUX knowledge base was formulated by consulting with human experts in the field of neutron assay, by direct experimentation on the PAN assay system, and by observing operations on a daily basis. TRUX, with its improved ability to interpret assay results, has eliminated the need for close supervision by a human expert, allowing skilled technicians to operate the PAN assay system. 4 refs., 1 fig., 4 tabs

  11. POTENTIALITIES AND RISKS OF BIOCHEMICAL EDUCATIONAL RESEARCH VIA INTERNET

    Directory of Open Access Journals (Sweden)

    M.C. Gomes

    2004-05-01

    Full Text Available The internet is more and more present in worldwide education. There are innumerable projects thatstimulate its usage for aiding the teaching-learning process. This new trend modies the reality of thewhole school. However, many copies of identical information are found in several sites: graphs, gures,texts, links and even errors. The aim of this work is to analyse the biochemical issues photosynthesisand cellular respiration available in web pages, taking into account contents quality, trustworthinessand eectiveness. Firstly, 6th secondary level students were inquired by a questionnaire on their use ofinternet resources. More than 80 percent of them were regular users. The result conrms the alreadyknown potential of internet in education. Fourteen sites were analysed regarding to contents, presenceof bibliographical references, authorship, titles of responsibles and adequacy to the target public. Inrelation to contents, presence of conceptual errors, illustrations and other stimulant elements wereanalysed. None of the sites presented biblioghraphic references; less than half divulged responsiblenames and their graduation. The great majority do not mention intended public target. In general,the sites did not contained grave conceptual errors, except for two that mention estoma insteadestroma, cilica/acilica phosphorylation (for cyclic and acyclic, grama (for grana and place oxygenas essential for anaerobic respiration. However, one of these sites was the only one that mentionedenzymes and regulation steps of cellular respiration. Half of the sites presented identical texts andgures. None used livened up resources. The majority showed contents as virtual versions of printedbooks. Illustrations did not present necessary legends. None of the analysed sites, thus, was consideredexcellent. Trustworthiness is also questionable as much of the contents arte clearly copies of otherpages, recurrent even on the errors. Our data strengthen the need for

  12. Simulation methods with extended stability for stiff biochemical Kinetics

    Directory of Open Access Journals (Sweden)

    Rué Pau

    2010-08-01

    Full Text Available Abstract Background With increasing computer power, simulating the dynamics of complex systems in chemistry and biology is becoming increasingly routine. The modelling of individual reactions in (biochemical systems involves a large number of random events that can be simulated by the stochastic simulation algorithm (SSA. The key quantity is the step size, or waiting time, τ, whose value inversely depends on the size of the propensities of the different channel reactions and which needs to be re-evaluated after every firing event. Such a discrete event simulation may be extremely expensive, in particular for stiff systems where τ can be very short due to the fast kinetics of some of the channel reactions. Several alternative methods have been put forward to increase the integration step size. The so-called τ-leap approach takes a larger step size by allowing all the reactions to fire, from a Poisson or Binomial distribution, within that step. Although the expected value for the different species in the reactive system is maintained with respect to more precise methods, the variance at steady state can suffer from large errors as τ grows. Results In this paper we extend Poisson τ-leap methods to a general class of Runge-Kutta (RK τ-leap methods. We show that with the proper selection of the coefficients, the variance of the extended τ-leap can be well-behaved, leading to significantly larger step sizes. Conclusions The benefit of adapting the extended method to the use of RK frameworks is clear in terms of speed of calculation, as the number of evaluations of the Poisson distribution is still one set per time step, as in the original τ-leap method. The approach paves the way to explore new multiscale methods to simulate (biochemical systems.

  13. Biochemical markers of bone turnover in benign paroxysmal positional vertigo

    Science.gov (United States)

    Lee, Sun Bin; Lee, Chang Ho; Kim, Young Ju; Kim, Hyoung-Mi

    2017-01-01

    Objective Several studies have suggested a possible relationship between recurrent benign paroxysmal positional vertigo (BPPV) and altered calcium homeostasis in the endolymph of the inner ear. The present study aimed to evaluate the association between Ca2+ and vitamin D status and BPPV occurrence as well as the status of bone biochemical markers in osteoporotic patients who were diagnosed with idiopathic BPPV. Methods The study included total 132 patients who were referred to our clinic between August 2008 and October 2013. Based on the bone mineral density (BMD) results, the subjects were divided into three groups: normal BMD (n = 34), osteopenia (n = 40) and osteoporosis (n = 58). The biochemical markers of bone turnover including serum Carboxy-terminal telopeptide of type I collagen (s-CTX), osteocalcin, alkaline phosphatase (ALP) and urinary free deoxypyridinoline (u-DPD), were analyzed, along with the serum Ca2+ and vitamin D levels. Results The mean serum calcium, phosphate and creatinine clearance levels were within the standard laboratory reference range. The incidence of vitamin D deficiency was 11.8% (4/34) in the normal BMD group, 15% (6/40) in the osteopenia group and 43.1% (25/58) in the osteoporosis group. There was a positive correlation between the 25(OH)D and BMD results in the patients with BPPV. Among the bone turnover markers, the osteocalcin and u-DPD levels were significantly elevated in the osteoporotic patients with BPPV. Multiple logistic regression analyses showed that osteoporosis and vitamin D deficiency were associated with BPPV. Conclusion Our findings suggest that the prevalence of BPPV in osteoporotic patients is associated with vitamin D deficiency and high bone turnover rates at systemic level, which could disturb local Ca2+ homeostasis in the inner ear. PMID:28467451

  14. Evaluation of Liver Biochemical Parameters in Manganese Miners

    Directory of Open Access Journals (Sweden)

    S. Mohammad Hossein Razavian

    2014-06-01

    Full Text Available Background: Manganese (Mn related jobs may cause manganism especially in miners. Side effects include neural and pathological disorders. In spite of liver is the main organ that filters Mn (99% but few studies has performed about Mn toxicity in liver so no specific biochemical indicator is available (Gunnar. In this study, the relation between blood, urine and saliva Mn level and its hepatotoxic effects is evaluated. Materials and Methods: Blood, urine, saliva of 50 accidently selected miners collected in acid washed tubes for an experience study. Samples were used to evaluation not only biochemical parameters by pars azmoon kits but also Mn concentration by mass spectroscopy. Results: Manganese concentration in all miners in addition to blood AST, ALT, ALP increased significantly (p<0.001 related to controls. Miners with 10-15 years background had higher blood total, direct & indirect billirubin and ALP levels compared to others. Mn concentration in serum declined but in urine and saliva had no changes by working in mine. AST & ALT increased significantly in miners with 300 μg/L serum Mn concentration. Mn concentration in various samples and serum AST & ALT level were higher in native miners than non-native but in both not related to background. Conclusion: Significantly higher levels of billirubin, AST & ALT in miners compared to controls revealed Mn hepatotoxic effects in them. Also significant ALP increasing showed cholestasis in miners that supported by AST, ALT level. Significant billirubin, AST, ALT, ALP in miners with 10-15 years background revealed the importance of this period in miners liver check up. Higher Mn levels in different sources of native miners can be due to more environmental contact. Higher AST, ALT and lower ALP level in native miners indicate more hepatastoxic and less cholestasis and therefore arthrosclerosis and parkinson risk in these workers.

  15. Radiometric assays for glycerol, glucose, and glycogen

    International Nuclear Information System (INIS)

    Bradley, D.C.; Kaslow, H.R.

    1989-01-01

    We have developed radiometric assays for small quantities of glycerol, glucose and glycogen, based on a technique described by Thorner and Paulus for the measurement of glycerokinase activity. In the glycerol assay, glycerol is phosphorylated with [32P]ATP and glycerokinase, residual [32P]ATP is hydrolyzed by heating in acid, and free [32P]phosphate is removed by precipitation with ammonium molybdate and triethylamine. Standard dose-response curves were linear from 50 to 3000 pmol glycerol with less than 3% SD in triplicate measurements. Of the substances tested for interference, only dihydroxyacetone gave a slight false positive signal at high concentration. When used to measure glycerol concentrations in serum and in media from incubated adipose tissue, the radiometric glycerol assay correlated well with a commonly used spectrophotometric assay. The radiometric glucose assay is similar to the glycerol assay, except that glucokinase is used instead of glycerokinase. Dose response was linear from 5 to 3000 pmol glucose with less than 3% SD in triplicate measurements. Glucosamine and N-acetylglucosamine gave false positive signals when equimolar to glucose. When glucose concentrations in serum were measured, the radiometric glucose assay agreed well with hexokinase/glucose-6-phosphate dehydrogenase (H/GDH)-based and glucose oxidase/H2O2-based glucose assays. The radiometric method for glycogen measurement incorporates previously described isolation and digestion techniques, followed by the radiometric assay of free glucose. When used to measure glycogen in mouse epididymal fat pads, the radiometric glycogen assay correlated well with the H/GDH-based glycogen assay. All three radiometric assays offer several practical advantages over spectral assays

  16. Harmonization of radiobiological assays: why and how?

    International Nuclear Information System (INIS)

    Prasanna, Pataje G.

    2014-01-01

    The International Atomic Energy Agency has made available a technical manual for cytogenetic biodosimetry assays (dicentric chromosome aberration (DCA) and cytokinesis-block micronucleus (CBMN) assays) used for radiation dose assessment in radiation accidents. The International Standardization Organization, which develops standards and guidelines, also provides an avenue for laboratory accreditation, has developed guidelines and recommendations for performing cytogenetic biodosimetry assays. Harmonization of DCA and CBMN assays, has improved their accuracy. Double-blinded inter-laboratory comparison studies involving several networks have further validated DCA and CBMN assays and improved the confidence in their potential use for radiation dose assessment in mass casualties. This kind of international harmonization is lacking for pre-clinical radiobiology assays. The widely used pre-clinical assays that are relatively important to set stage for clinical trials include clonogenic assays, flow-cytometry assays, apoptotic assays, and tumor regression and growth delay assays. However, significant inter-laboratory variations occur with respect to data among laboratories. This raises concerns on the reliability and reproducibility of preclinical data that drives further development and translation. Lack of reproducibility may stem from a variety of factors such as poor scientist training, less than optimal experimental design, inadequate description of methodology, and impulse to publish only the positive data etc. Availability of technical manuals, standard operating procedures, accreditation avenues for laboratories performing such assays, inter-laboratory comparisons, and use of standardized protocols are necessary to enhance reliability and reproducibility. Thus, it is important that radiobiological assays are harmonized for laboratory protocols to ensure successful translation of pre-clinical research on radiation effect modulators to help design clinic trials with

  17. Automated amperometric plutonium assay system

    International Nuclear Information System (INIS)

    Burt, M.C.

    1985-01-01

    The amperometric titration for plutonium assay has been used in the nuclear industry for over twenty years and has been in routine use at the Hanford Engineering Development Laboratory since 1976 for the analysis of plutonium oxide and mixed oxide fuel material for the Fast Flux Test Facility. It has proven itself to be an accurate and reliable method. The method may be used as a direct end point titration or an excess of titrant may be added and a back titration performed to aid in determination of the end point. Due to the slowness of the PuVI-FeII reaction it is difficult to recognize when the end point is being approached and is very time consuming if the current is allowed to decay to the residual value after each titrant addition. For this reason the back titration in which the rapid FeII-CrVI reaction occurs is used by most laboratories. The back titration is performed by the addition of excess ferrous solution followed by two measured aliquots of standard dichromate with measurement of cell current after each addition

  18. Qualification of standard membrane-feeding assay with Plasmodium falciparum malaria and potential improvements for future assays.

    Directory of Open Access Journals (Sweden)

    Kazutoyo Miura

    Full Text Available Vaccines that interrupt malaria transmission are of increasing interest and a robust functional assay to measure this activity would promote their development by providing a biologically relevant means of evaluating potential vaccine candidates. Therefore, we aimed to qualify the standard membrane-feeding assay (SMFA. The assay measures the transmission-blocking activity of antibodies by feeding cultured P. falciparum gametocytes to Anopheles mosquitoes in the presence of the test antibodies and measuring subsequent mosquito infection. The International Conference on Harmonisation (ICH Harmonised Tripartite Guideline Q2(R1 details characteristics considered in assay validation. Of these characteristics, we decided to qualify the SMFA for Precision, Linearity, Range and Specificity. The transmission-blocking 4B7 monoclonal antibody was tested over 6 feeding experiments at several concentrations to determine four suitable concentrations that were tested in triplicate in the qualification experiments (3 additional feeds to evaluate Precision, Linearity and Range. For Specificity, 4B7 was tested in the presence of normal mouse IgG. We determined intra- and inter-assay variability of % inhibition of mean oocyst intensity at each concentration of 4B7 (lower concentrations showed higher variability. We also showed that % inhibition was dependent on 4B7 concentration and the activity is specific to 4B7. Since obtaining empirical data is time-consuming, we generated a model using data from all 9 feeds and simulated the effects of different parameters on final readouts to improve the assay procedure and analytical methods for future studies. For example, we estimated the effect of number of mosquitoes dissected on variability of % inhibition, and simulated the relationship between % inhibition in oocyst intensity and % inhibition of prevalence of infected mosquitos at different mean oocysts in the control. SMFA is one of the few biological assays used in

  19. Biochemical diagnosis of pheochromocytoma: which test is best?

    NARCIS (Netherlands)

    Lenders, J.W.M.; Pacak, K.; Walther, M.M.; Linehan, W.M.; Mannelli, M.; Friberg, P.; Keiser, H.R.; Goldstein, D.S.; Eisenhofer, G.

    2002-01-01

    CONTEXT: Diagnosis of pheochromocytoma depends on biochemical evidence of catecholamine production by the tumor. However, the best test to establish the diagnosis has not been determined. OBJECTIVE: To determine the biochemical test or combination of tests that provides the best method for diagnosis

  20. Biochemical changes occurring during fermentation of camel milk by ...

    African Journals Online (AJOL)

    Biochemical changes occurring during fermentation of camel milk by selected bacterial starter cultures. ... Abstract. The biochemical changes in amino acids, water soluble vitamins, soluble sugars and organic acids occurring during fermentation (at 43°C for 6 h) of camel milk inoculated with Streptococcus thermophilus 37, ...

  1. Biochemical markers of mineral bone disorder in South African ...

    African Journals Online (AJOL)

    Associations between log transformed PTH and other biochemical parameters were assessed by multiple linear regression analyses following significant associa- tions obtained from univariate regression analyses. A lo- gistic regression model was used to evaluate the effect of other biochemical parameters on the odds of ...

  2. Developments in commercially produced microbials at Biochem Products

    Science.gov (United States)

    John Lublinkhof; Douglas H. Ross

    1985-01-01

    Biochem Products is part of a large industrial and scientific family - the Solvay Group. Solvay, headquartered in Brussels, Belgium is a multinational company with 46,000 employees worldwide. In the U.S., our working partners include a large polymer manufacturer, a peroxygen producer and a leading poultry and animal health products company. Biochem Products is a...

  3. Exploring basic biochemical constituents in the body tissues of ...

    African Journals Online (AJOL)

    Feeding regime did not influence susceptibility to mass loss during export. Animal age influenced the biochemical composition and export performance of abalone. Keywords: abalone; aquaculture; feeds; Haliotis midae; live export; mass loss; tissue biochemical constituents. African Journal of Marine Science 2010, 32(1): ...

  4. Biochemical evaluation of phenylketonuria (PKU: from diagnosis to treatment

    Directory of Open Access Journals (Sweden)

    Leticia Belmont-Martínez

    2014-07-01

    Besides periodical Phe and Tyr testing, biochemical follow-up includes the measurement of necessary elements that guarantee normal physical and intellectual development such as selenium, zinc, B12 vitamin, folates, iron and long chain fatty acids. Clinical context is as important as biochemical status so periodic evaluation of nutritional, medical, social and psychological aspects should be included.

  5. Reference Ranges for Some Biochemical Parameters in Adult ...

    African Journals Online (AJOL)

    PURPOSE: To establish the reference ranges of some biochemical parameters for adult Kenyan population. METHODS: In a prospective involving 1100 healthy blood donors (age: 18-55 yr) in Kenyatta National Hospital, Kenya reference ranges of some biochemical analytes were constructed by using the parametric ...

  6. An improved solution of first order kinetics for biochemical oxygen ...

    African Journals Online (AJOL)

    This paper evaluated selected Biochemical Oxygen Demand first order kinetics methods. Domesticinstitutional wastewaters were collected twice in a month for three months from the Obafemi Awolowo University, Ile-Ife waste stabilization ponds. Biochemical Oxygen Demand concentrations at different days were determined ...

  7. Biochemical characterization of three Aspergillus niger β-galactosidases

    Directory of Open Access Journals (Sweden)

    Dandan Niu

    2017-05-01

    Conclusions: Three new β-galactosidases belonging to glycosyl hydrolase family 35 from A. niger F0215 were cloned and biochemically characterized. In addition to the known LacA, A. niger has at least three β-galactosidase family members with remarkably different biochemical properties.

  8. Haematological and blood biochemical indices of West African ...

    African Journals Online (AJOL)

    Haematological and blood biochemical indices of West African dwarf goats vaccinated against Pestes des petit ruminants (PPR) ... blood biochemical indices of forty randomly selected West African dwarf (WAD) goats were studied. Packed cell volume ... neutrophil/lymphocyte ratio and white blood cells (WBC) than females.

  9. Differential response of biochemical parameters to EMS and MMS ...

    African Journals Online (AJOL)

    B.B.D. Khalandar

    2015-06-10

    Jun 10, 2015 ... Abstract Aim: To study the effect of alkylating agents such as EMS and MMS on chromosomes and biochemical parameters in induced diabetic mouse. Methods: Chromosome preparations from bone marrow was made using the method of Evans et al. (1964) and biochemical estimations from the liver was ...

  10. Local biochemical and morphological differences in human Achilles tendinopathy

    DEFF Research Database (Denmark)

    Pingel, Jessica; Fredberg, U.; Qvortrup, Klaus

    2012-01-01

    The incidence of Achilles tendinopathy is high and underlying etiology as well as biochemical and morphological pathology associated with the disease is largely unknown. The aim of the present study was to describe biochemical and morphological differences in chronic Achilles tendinopathy....... The expressions of growth factors, inflammatory mediators and tendon morphology were determined in both chronically diseased and healthy tendon parts....

  11. Determination of fecal contamination origin in reclaimed water open-air ponds using biochemical fingerprinting of enterococci and fecal coliforms.

    Science.gov (United States)

    Casanovas-Massana, Arnau; Blanch, Anicet R

    2013-05-01

    Low levels of fecal indicator bacteria (FIB) were recently detected in two reclaimed water open-air ponds used to irrigate a golf course located in Northeastern Spain. The aim of this study was to evaluate the feasibility of a biochemical fingerprinting method to track the origin of fecal contamination in water with low FIB levels, as in the aforementioned ponds. We also aimed to determine whether FIB presence was due to regrowth of the reclaimed water populations or to a contribution of fecal matter whose source was in the golf facility. Three hundred and fifty enterococcal strains and 308 fecal coliform strains were isolated from the ponds and reclamation plant, and they were biochemically phenotyped. In addition, the inactivation of several microbial fecal pollution indicators (fecal coliforms, total bifidobacteria, sorbitol-fermenting bifidobacteria, somatic bacteriophages, and bacteriophages infecting Bacteroides thetaiotaomicron) was studied using a mesocosm in situ in order to obtain information about their decay rate. Although FIB concentration was low, the biochemical fingerprinting provided evidence that the origin of the fecal contamination in the ponds was not related to the reclaimed water. Biochemical fingerprinting thus proved to be a successful approach, since other microbial source-tracking methods perform poorly when dealing with low fecal load matrices. Furthermore, the mesocosm assays indicated that none of the microbial fecal indicators was able to regrow in the ponds. Finally, the study highlights the fact that reclaimed water may be recontaminated in open-air reservoirs, and therefore, its microbial quality should be monitored throughout its use.

  12. A short 18 items food frequency questionnaire biochemically validated to estimate zinc status in humans.

    Science.gov (United States)

    Trame, Sarah; Wessels, Inga; Haase, Hajo; Rink, Lothar

    2018-02-21

    Inadequate dietary zinc intake is wide-spread in the world's population. Despite the clinical significance of zinc deficiency there is no established method or biomarker to reliably evaluate the zinc status. The aim of our study was to develop a biochemically validated questionnaire as a clinically useful tool that can predict the risk of an individual being zinc deficient. From 71 subjects aged 18-55 years blood and urine samples were collected. Zinc concentrations in serum and urine were determined by atomic absorption spectrometry. A food frequency questionnaire (FFQ) including 38 items was filled out representing the consumption during the last 6 months obtaining nutrient diet scores. Latter were calculated by multiplication of the particular frequency of consumption, the nutrient intake of the respective portion size and the extent of the consumed quantity. Results from the FFQ were compared with nutrient intake information gathered in 24-h dietary recalls. A hemogram was performed and cytokine concentrations were obtained using Enzyme-linked Immunosorbent Assay. Reducing the items of the primary FFQ from 38 to 18 did not result in a significant variance between both calculated scores. Zinc diet scores showed highly significant correlation with serum zinc (r = 0.37; p < 0.01) and urine zinc concentrations (r = 0.34; p < 0.01). Serum zinc concentrations and zinc diet scores showed a significant positive correlation with animal protein intake (r = 0.37; p < 0.01/r = 0.54; p < 0.0001). Higher zinc diet scores were found in omnivores compared to vegetarians (213.5 vs. 111.9; p < 0.0001). The 18 items FFQ seems to be a sufficient tool to provide a good estimation of the zinc status. Moreover, shortening of the questionnaire to 18 items without a loss of predictive efficiency enables a facilitated and resource-saving routine use. A validation of the questionnaire in other cohorts could enable the progression towards clinical

  13. A novel nucleic acid analogue shows strong angiogenic activity

    Energy Technology Data Exchange (ETDEWEB)

    Tsukamoto, Ikuko, E-mail: tukamoto@med.kagawa-u.ac.jp [Department of Pharmaco-Bio-Informatics, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki, Kita, Kagawa 761-0793 (Japan); Sakakibara, Norikazu; Maruyama, Tokumi [Kagawa School of Pharmaceutical Sciences, Tokushima Bunri University, 1314-1 Shido, Sanuki, Kagawa 769-2193 (Japan); Igarashi, Junsuke; Kosaka, Hiroaki [Department of Cardiovascular Physiology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki, Kita, Kagawa 761-0793 (Japan); Kubota, Yasuo [Department of Dermatology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki, Kita, Kagawa 761-0793 (Japan); Tokuda, Masaaki [Department of Cell Physiology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki, Kita, Kagawa 761-0793 (Japan); Ashino, Hiromi [The Tokyo Metropolitan Institute of Medical Science, 1-6 Kamikitazawa2-chome, Setagaya-ku, Tokyo 156-8506 (Japan); Hattori, Kenichi; Tanaka, Shinji; Kawata, Mitsuhiro [Teikoku Seiyaku Co., Ltd., Sanbonmatsu, Higashikagawa, Kagawa 769-2695 (Japan); Konishi, Ryoji [Department of Pharmaco-Bio-Informatics, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe, Miki, Kita, Kagawa 761-0793 (Japan)

    2010-09-03

    Research highlights: {yields} A novel nucleic acid analogue (2Cl-C.OXT-A, m.w. 284) showed angiogenic potency. {yields} It stimulated the tube formation, proliferation and migration of HUVEC in vitro. {yields} 2Cl-C.OXT-A induced the activation of ERK1/2 and MEK in HUVEC. {yields} Angiogenic potency in vivo was confirmed in CAM assay and rabbit cornea assay. {yields} A synthesized small angiogenic agent would have great clinical therapeutic value. -- Abstract: A novel nucleic acid analogue (2Cl-C.OXT-A) significantly stimulated tube formation of human umbilical endothelial cells (HUVEC). Its maximum potency at 100 {mu}M was stronger than that of vascular endothelial growth factor (VEGF), a positive control. At this concentration, 2Cl-C.OXT-A moderately stimulated proliferation as well as migration of HUVEC. To gain mechanistic insights how 2Cl-C.OXT-A promotes angiogenic responses in HUVEC, we performed immunoblot analyses using phospho-specific antibodies as probes. 2Cl-C.OXT-A induced robust phosphorylation/activation of MAP kinase ERK1/2 and an upstream MAP kinase kinase MEK. Conversely, a MEK inhibitor PD98059 abolished ERK1/2 activation and tube formation both enhanced by 2Cl-C.OXT-A. In contrast, MAP kinase responses elicited by 2Cl-C.OXT-A were not inhibited by SU5416, a specific inhibitor of VEGF receptor tyrosine kinase. Collectively these results suggest that 2Cl-C.OXT-A-induces angiogenic responses in HUVEC mediated by a MAP kinase cascade comprising MEK and ERK1/2, but independently of VEGF receptor tyrosine kinase. In vivo assay using chicken chorioallantoic membrane (CAM) and rabbit cornea also suggested the angiogenic potency of 2Cl-C.OXT-A.

  14. Performance evaluation of new automated hepatitis B viral markers in the clinical laboratory: two quantitative hepatitis B surface antigen assays and an HBV core-related antigen assay.

    Science.gov (United States)

    Park, Yongjung; Hong, Duck Jin; Shin, Saeam; Cho, Yonggeun; Kim, Hyon-Suk

    2012-05-01

    We evaluated quantitative hepatitis B surface antigen (qHBsAg) assays and a hepatitis B virus (HBV) core-related antigen (HBcrAg) assay. A total of 529 serum samples from patients with hepatitis B were tested. HBsAg levels were determined by using the Elecsys (Roche Diagnostics, Indianapolis, IN) and Architect (Abbott Laboratories, Abbott Park, IL) qHBsAg assays. HBcrAg was measured by using Lumipulse HBcrAg assay (Fujirebio, Tokyo, Japan). Serum aminotransferases and HBV DNA were respectively quantified by using the Hitachi 7600 analyzer (Hitachi High-Technologies, Tokyo, Japan) and the Cobas AmpliPrep/Cobas TaqMan test (Roche). Precision of the qHBsAg and HBcrAg assays was assessed, and linearity of the qHBsAg assays was verified. All assays showed good precision performance with coefficients of variation between 4.5% and 5.3% except for some levels. Both qHBsAg assays showed linearity from 0.1 to 12,000.0 IU/mL and correlated well (r = 0.9934). HBsAg levels correlated with HBV DNA (r = 0.3373) and with HBcrAg (r = 0.5164), and HBcrAg also correlated with HBV DNA (r = 0.5198; P < .0001). This observation could provide impetus for further research to elucidate the clinical usefulness of the qHBsAg and HBcrAg assays.

  15. First Evaluation of the Biologically Active Substances and Antioxidant Potential of Regrowth Velvet Antler by means of Multiple Biochemical Assays

    Directory of Open Access Journals (Sweden)

    Yujiao Tang

    2015-01-01

    Full Text Available We investigated the biologically active substances contained in RVA (regrowth velvet antler by comparing the composition of biologically active substances and antioxidant potential of different antler segments. RVA was subjected to extraction using DW (distilled water. RVA was divided into 3 segments: T-RVA (top RVA, M-RVA (middle RVA, and B-RVA (base RVA. The T-RVA section possessed the greatest amounts of uronic acid (36.251 mg/g, sulfated GAGs (sulfated glycosaminoglycans (555.76 mg/g, sialic acid (111.276 mg/g, uridine (0.957 mg/g, uracil (1.084 mg/g, and hypoxanthine (1.2631 mg/g. In addition, the T-RVA section possessed the strongest antioxidant capacity as determined by DPPH, H2O2 (hydrogen peroxide, hydroxyl, and ABTS (2,2′-azinobis-3-ethylbenzthiazoline-6-sulphonate radical scavenging activity as well as FRAP (ferric reducing antioxidant power and ORAC (oxygen radical absorbance capacity. The values of those were 53.44, 23.09, 34.12, 60.31, and 35.81 TE/μM at 1 mg/mL and 113.57 TE/μM at 20 μg/mL. These results indicate that the T-RVA section possesses the greatest amount of biologically active substances and highest antioxidant potential. This is the first report on the biologically active substances and antioxidant potential of RVA.

  16. High-throughput screening of carbohydrate-degrading enzymes using novel insoluble chromogenic substrate assay kits

    DEFF Research Database (Denmark)

    Schückel, Julia; Kracun, Stjepan Kresimir; Willats, William George Tycho

    2016-01-01

    Carbohydrates active enzymes (CAZymes) have multiple roles in vivo and are widely used for industrial processing in the biofuel, textile, detergent, paper and food industries. A deeper understanding of CAZymes is important from both fundamental biology and industrial standpoints. Vast numbers...... of CAZymes exist in nature (especially in microorganisms) and hundreds of thousands have been cataloged and described in the carbohydrate active enzyme database (CAZy). However, the rate of discovery of putative enzymes has outstripped our ability to biochemically characterize their activities. One reason...... for this is that advances in genome and transcriptome sequencing, together with associated bioinformatics tools allow for rapid identification of candidate CAZymes, but technology for determining an enzyme's biochemical characteristics has advanced more slowly. To address this technology gap, a novel high-throughput assay...

  17. Assay development status report for total cyanide

    Energy Technology Data Exchange (ETDEWEB)

    Simpson, B.C. [Westinghouse Hanford Co., Richland, WA (United States); Jones, T.E.; Pool, K.H. [Pacific Northwest Lab., Richland, WA (United States)

    1993-02-01

    A validated cyanide assay that is applicable to a variety of tank waste matrices is necessary to resolve certain waste tank safety issues and for purposes of overall waste characterization. The target for this effort is an assay with an applicable range of greater than 1,000 ppM (0.10 wt%) total cyanide and a confidence level greater than 80%. Figure 1 illustrates the operating regime of the proposed cyanide assay method. The Assay Development Status Report for Total Cyanide will summarize the past experience with cyanide analyses on-tank waste matrices and will rate the status of the analytical methods used to assay total cyanide (CN{sup {minus}} ion) in the tank waste matrices as acceptable or unacceptable. This paper will also briefly describe the current efforts for improving analytical resolution of the assays and the attempts at speciation.

  18. Cell-based lipid flippase assay employing fluorescent lipid derivatives

    DEFF Research Database (Denmark)

    Jensen, Maria Stumph; Costa, Sara; Günther-Pomorski, Thomas

    2016-01-01

    , studies of individual P4-ATPase family members from fungi, plants, and animals show that P4-ATPases differ in their substrate specificities and mediate transport of a broader range of lipid substrates. Here, we describe an assay based on fluorescent lipid derivatives to monitor and characterize lipid...

  19. Sandwich assay for tacrolimus using 2 antitacrolimus antibodies.

    Science.gov (United States)

    Wei, Tie Q; Zheng, Yi F; Dubowy, Michael; Sharma, Manoj

    2014-04-01

    Although detection of natural haptens by antihapten antibodies in sandwich assay format has the theoretical advantages of high analytical specificity and sensitivity, this type of assay has not been reported because of the seemingly insurmountable task of avoiding steric hindrance between the 2 bindings. This is especially true for ring-structured hydrophobic haptens. The macrolide drug tacrolimus (FK506, Prograf®, 804 Da) is such a hapten. Here we show the detection of tacrolimus using 2 antitacrolimus monoclonal antibodies in a sandwich assay. Both antibodies were developed by use of an intact tacrolimus molecule covalently linked to a carrier protein but via 2 different positions separated by 10 carbon atoms. Epitope analysis based on drug analog binding was used to show no overlap between the binding sites of the 2 antibodies, indicating the 10-carbon separation resulted in 2 distinct epitopes. The distinct epitopes suggested that the drug might be approachable by the antibodies from 2 separate directions, which predicted simultaneous binding as in sandwich formation. This prediction was confirmed in sandwich ELISA and affinity column-mediated immunoassay formats. The assay demonstrated good imprecision and significantly lower metabolite cross-reactivity than competitive assay counterparts. Comparison with liquid chromatography-tandem mass spectrometry (LC-MS/MS) using 55 whole-blood samples from transplant patients with tacrolimus concentrations ranging from 0.9 to 29.5 ng/mL showed a linear regression: sandwich = 0.99 × LC-MS/MS + 0.10 ng/mL, r = 0.991, Sy|x = 1.08 ng/mL. This work demonstrates that a highly specific sandwich assay using 2 antihapten antibodies is feasible for the measurement of a hapten drug.

  20. Biochemical and mutational analysis of glutamine synthetase type III from the rumen anaerobe Ruminococcus albus 8.

    Science.gov (United States)

    Amaya, Kensey R; Kocherginskaya, Svetlana A; Mackie, Roderick I; Cann, Isaac K O

    2005-11-01

    Two different genes encoding glutamine synthetase type I (GSI) and GSIII were identified in the genome sequence of R. albus 8. The identity of the GSIII protein was confirmed by the presence of its associated conserved motifs. The glnN gene, encoding the GSIII, was cloned and expressed in Escherichia coli BL21 cells. The recombinant protein was purified and subjected to biochemical and physical analyses. Subunit organization suggested a protein present in solution as both monomers and oligomers. Kinetic studies using the forward and the gamma-glutamyl transferase (gamma-GT) assays were carried out. Mutations that changed conserved glutamic acid residues to alanine in the four GSIII motifs resulted in drastic decreases in GS activity using both assays, except for an E380A mutation, which rather resulted in an increase in activity in the forward assay compared to the wild-type protein. Reduced GSIII activity was also exhibited by mutating, individually, two lysines (K308 and K318) located in the putative nucleotide-binding site to alanine. Most importantly, the presence of mRNA transcripts of the glnN gene in R. albus 8 cells grown under ammonia limiting conditions, whereas little or no transcript was detected in cells grown under ammonia sufficient conditions, suggested an important role for the GSIII in the nitrogen metabolism of R. albus 8. Furthermore, the mutational studies on the conserved GSIII motifs demonstrated, for the first time, their importance in the structure and/or function of a GSIII protein.