Sample records for biochemical chromogenic assays

  1. 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


    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...... kit based on insoluble chromogenic substrates is described here. Two distinct substrate types were produced: Chromogenic Polymer Hydrogel (CPH) substrates (made from purified polysaccharides and proteins) and Insoluble Chromogenic Biomass (ICB) substrates (made from complex biomass materials). Both...

  2. Evaluation of robot automated chromogenic substrate LAL endotoxin assay method for pharmaceutical products testing. (United States)

    Tsuji, K; Martin, P A


    The robot automated chromogenic substrate LAL assay method was evaluated for endotoxin testing using three lots each of 12 pharmaceutical products. As many as 216 assays, including automated standard curve construction and sample preparation, can be performed in a single day of unattended operation. The method is linear (r greater than .99) in the range of 0 to 0.2 EU/ml. The precision of the method determined by assaying a lot of calcium gluconate for four days was 6%, 10%, and 10% for within an assay block, between assay blocks, and between assay days, respectively. Recovery of endotoxin when spiked into products ranged from 81% to 110% and was within the statistical variation (2 sigma limit) of the method. The endotoxin levels detected in a biological raw material by the chromogenic substrate assay method correlated well with that of the gel-clot LAL assay method. The endotoxin content of the majority of the pharmaceutical products tested was well below the sensitivity of both the chromogenic substrate and the gel clot LAL assay methods.

  3. Quantitative digital image analysis of chromogenic assays for high throughput screening of alpha-amylase mutant libraries. (United States)

    Shankar, Manoharan; Priyadharshini, Ramachandran; Gunasekaran, Paramasamy


    An image analysis-based method for high throughput screening of an alpha-amylase mutant library using chromogenic assays was developed. Assays were performed in microplates and high resolution images of the assay plates were read using the Virtual Microplate Reader (VMR) script to quantify the concentration of the chromogen. This method is fast and sensitive in quantifying 0.025-0.3 mg starch/ml as well as 0.05-0.75 mg glucose/ml. It was also an effective screening method for improved alpha-amylase activity with a coefficient of variance of 18%.

  4. Specific, sensitive, precise, and rapid functional chromogenic assay of activated first complement component (C1) in plasma

    DEFF Research Database (Denmark)

    Munkvad, S; Jespersen, J; Sidelmann, Johannes Jakobsen;


    by using an amidolytic rate assay with a chromogenic substrate. We have optimized the assay conditions with respect to incubation time, concentration of antiserum to C1-inh, ionic strength, and pH. Our method determines specifically the concentration in plasma of free activated C1, not complexes...

  5. Analytical assays based on chromogenic and fluorogenic chemosensors for the detection of cyanide

    Directory of Open Access Journals (Sweden)

    Vanderléia Gava Marini


    Full Text Available Cyanide (CN– is an anion well–known for its toxicity, being a chemical agent often related to cases of homicide and suicide. Despite being responsible for the toxicity of many animals and plants, it is used in several industrial activities, with innumerous implications in terms of the environment. Due to its high toxicity, the maximum level of CN– concentration allowed by the World Health Organization in potable water is 1.7 µmol/L. This low concentration limit requires methods of visual detection and quantitative determination which are ever more sensitive, simple, reliable, and economical. Advancements in the field of chromogenic and fluorogenic chemosensors for anionic analytes have led to the development of several methodologies for the detection of CN–. Therefore, this review aims to present the main strategies that have been used in the study of quantitative and naked–eye detection of CN– by means of chromogenic and fluorogenic chemosensors. Aspects related to CN–, such as its reactivity, toxicity, applications, and implications in different domains of knowledge, are presented. Recent work involving the development of chemosensors for CN– based on acid–base reactions, chemodosimeters, chromoreactands, and competition assays is also described. In addition, recent studies that make use of nanotechnology to develop strategies for the detection of CN– are also discussed, as well as the prospects envisioned in this field.

  6. 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


    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...

  7. Chromogenic platform based on recombinant Drosophila melanogaster acetylcholinesterase for visible unidirectional assay of organophosphate and carbamate insecticide residues

    Energy Technology Data Exchange (ETDEWEB)

    Han Zheng [Institute for Agri-food Standards and Testing Technology, Shanghai Academy of Agricultural Sciences, 1018 Jinqi Road, Shanghai 201403 (China); Chi Chensen [School of Life Science and Biotechnology, Bor Luh Food Safety Center, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai 200240 (China); Bai Bing; Liu Gang; Rao Qinxiong [Institute for Agri-food Standards and Testing Technology, Shanghai Academy of Agricultural Sciences, 1018 Jinqi Road, Shanghai 201403 (China); Peng Shaojie [Institute of Shanghai Food and Drug Supervision, 615 Liuzhou Road, Shanghai 200233 (China); Liu Hong [Shanghai Municipal Center for Disease Control and Prevention, 1380 Zhongshan West Road, Shanghai 200336 (China); Zhao Zhihui [Institute for Agri-food Standards and Testing Technology, Shanghai Academy of Agricultural Sciences, 1018 Jinqi Road, Shanghai 201403 (China); Zhang Dabing [School of Life Science and Biotechnology, Bor Luh Food Safety Center, Shanghai Jiao Tong University, 800 Dongchuan Road, Shanghai 200240 (China); Wu Aibo, E-mail: [Institute for Agri-food Standards and Testing Technology, Shanghai Academy of Agricultural Sciences, 1018 Jinqi Road, Shanghai 201403 (China)


    Highlight: Black-Right-Pointing-Pointer A visible chromogenic platform for rapid analysis of OP and CM insecticide residues was developed. Black-Right-Pointing-Pointer The assay has the capabilities of both qualitative measurement and quantitative analysis. Black-Right-Pointing-Pointer The sensitivity, capabilities of resisting interferences and storage stability were desirable. Black-Right-Pointing-Pointer Matrix effects were acceptable and detection performance was satisfactory in real application. - Abstract: In this study we propose a chromogenic platform for rapid analysis of organophosphate (OP) and carbamate (CM) insecticide residues, based on recombinant Drosophila melanogaster acetylcholinesterase (R-DmAChE) as enzyme and indoxyl acetate as substrate. The visible chromogenic strip had the advantages identical to those of commonly used lateral flow assays (LFAs) with utmost simplicity in sample loading and result observation. After optimization, depending on the color intensity (CI) values, the well-established assay has the capabilities of both qualitative measurement via naked eyes and quantitative analysis by colorimetric reader with the desirable IC{sub 50} values against the tested six insecticides (0.06 {mu}g mL{sup -1} of carbofuran, 0.28 {mu}g mL{sup -1} of methomyl, 0.03 {mu}g mL{sup -1} of dichlorvos, 31.6 {mu}g mL{sup -1} of methamidophos, 2.0 {mu}g mL{sup -1} of monocrotophos, 6.3 {mu}g mL{sup -1} of omethoate). Acceptable matrix effects and satisfactory detection performance were confirmed by in-parallel LC-MS/MS analysis in different vegetable varieties at various spiked levels of 10{sup -3} to 10{sup 1} {mu}g g{sup -1}. Overall, the testified suitability and applicability of this novel platform meet the requirements for practical use in food safety management and environmental monitoring, especially in the developing world.

  8. Advancement in biochemical assays in andrology

    Institute of Scientific and Technical Information of China (English)

    Wolf-BernhardSchill; RaftHenkel


    Determination of maikers of sperm function, accessory sex gland secretion and silent male genital tract inflammation is of considerable diagnostic value in the evaluation of male infertility. The introduction of biochemical tests into the analysis of male factor has the advantage that standardized assays with a coefficient of variafion characteristic of clinical chemistry are performed, in contrast to biological test systems with a large variability .Biochemical parameters may be used in clinical practice to evaluate the sperm fertitizing capacity (acrosin, aniline blue,ROS), to characterize male accessory sex gland secretinns (fructose, a-glucosidase, PSA), and to identify men with silent genital tract inflammation (elastase, C'3 complement component, coeruloplasmin, IgA, IgG, ROS). (As/an J Androl 1999 Jun; 1: 45-51)

  9. Patients with deep venous thrombosis and thrombophilia risk factors have a specific prolongation of the lag time in a chromogenic thrombin generation assay

    NARCIS (Netherlands)

    Haas, F.J.L.M.; Kluft, C.; Biesma, D.H.; Schutgens, R.E.G.


    The objective of the present study was to evaluate the influence of thrombophilia risk factors on variables of a chromogenic thrombin generation assay (ETP) in a setting with acute deep venous thrombosis (DVT) and non-DVT patients. In 152 outpatients suspected for DVT, the results of thrombophilia i

  10. Endotoxin Detection in Pharmaceuticals and Medical Devices with Kinetic-QCL, a Kinetic-Quantitative Chromogenic Limulus Amebocyte Lysate Assay. (United States)

    Berzofsky, Ronald N.


    The observation that endotoxin caused gelation in extracts of Limulus amebocytes has been expanded to the development of an in vitro kinetic, quantitative chromogenic LAL assay (Kinetic-QCL) for the detection of endotoxin in aqueous fluids. Within the last 15 years, the use of Limulus amebocyte lysate to detect and control the presence of pyrogenic substances in pharmaceuticals and medical devices has gained wide international acceptance. Both the United States and European Pharmacopoeias contain descriptions of and requirements for the LAL Bacterial Endotoxin Test. Both pharmacopoeias have begun to remove the rabbit pyrogen test requirement in a majority of drug monographs and have substituted endotoxin limits to be determined by LAL. The use of LAL has proved invaluable in controlling the level of endotoxin in finished product. The endotoxin contribution of raw materials and packaging material can be monitored as well. In-process testing at critical production steps can identify additional sources of endotoxin contamination, and depyrogenation processes can be validated by quantitating the degradation of endotoxin challenges. The speed, reproducibility, sensitivity, and economics of the Kinetic-QCL assay, in conjunction with the ppropriate equipment and software, over both the in vivo rabbit pyrogen test and the more traditional LAL gel-clot assay allow a more in-depth approach to the control of endotoxin in pharmaceuticals and medical devices.

  11. Ninhydrin-sodium molybdate chromogenic analytical probe for the assay of amino acids and proteins (United States)

    Anantharaman, Shivakumar; Padmarajaiah, Nagaraja; Al-Tayar, Naef Ghllab Saeed; Shrestha, Ashwinee Kumar


    A sensitive method has been proposed for the quantification of amino acids and proteins using ninhydrin and sodium molybdate as chromogenic substrates in citrate buffer of pH 5.6. A weak molybdate-hydrindantin complex plays the role in the formation of Ruhemann's purple. The linear response for the amino acid, amino acid mixture and Bovine serum albumin is between 0.999 and 66.80 μM, 1.52 and 38 μM and 5 and 100 μg/L, respectively. The molar absorptivity of the individual amino acid by the proposed reaction extends from 0.58 × 104 to 2.86 × 104 M- 1 cm- 1. The linearity equations for the proposed ninhydrin-molybdate for amino acid mixture is Abs = 0.021 × Conc (μM) - 0.002. The applicability of the proposed method has been justified in food and biological samples in conjunction with Kjeldahl method.

  12. Measurement Uncertainty of Chromogenic LAL Assays: Reaction Time and Proportion of Endotoxin and LAL Reagent Affect Release of p-Nitroaniline. (United States)

    Ostronoff, Celina Silva; Lourenço, Felipe Rebello


    Limulus Amebocyte Lysate (LAL) assays are widely used for detection and quantification of bacterial endotoxins in pharmaceuticals and medical devices. However, there are only a few studies on the measurement uncertainty of LAL assays. The aim of this work was to identify and quantify the main sources of measurement uncertainty for end point and kinetic-chromogenic LAL assays. Response surface methodology was used to study how the release of p-nitroaniline (pNA) is affected by reaction time and proportion of endotoxin and LAL reagent in end point and kinetic-chromogenic LAL assays, respectively. Increased release of pNA was observed when reaction time was increased. In addition, if different volumes of sample (or endotoxin standard) and LAL reagent are used, the pNA release rate will be affected. These results may be due to the increased interaction between the bacterial endotoxin and LAL-activated enzyme. Final measurement uncertainties (95% confidence interval) were 90-120% and 90-127% of bacterial endotoxin content for end point and kinetic-chromogenic assays, respectively. These values are reasonable for the scope of the method and allow the application of these measurement uncertainties in routine analysis of pharmaceuticals and medical devices.

  13. Click Chemistry-Mediated Nanosensors for Biochemical Assays. (United States)

    Chen, Yiping; Xianyu, Yunlei; Wu, Jing; Yin, Binfeng; Jiang, Xingyu


    Click chemistry combined with functional nanoparticles have drawn increasing attention in biochemical assays because they are promising in developing biosensors with effective signal transformation/amplification and straightforward signal readout for clinical diagnostic assays. In this review, we focus on the latest advances of biochemical assays based on Cu (I)-catalyzed 1, 3-dipolar cycloaddition of azides and alkynes (CuAAC)-mediated nanosensors, as well as the functionalization of nanoprobes based on click chemistry. Nanoprobes including gold nanoparticles, quantum dots, magnetic nanoparticles and carbon nanomaterials are covered. We discuss the advantages of click chemistry-mediated nanosensors for biochemical assays, and give perspectives on the development of click chemistry-mediated approaches for clinical diagnosis and other biomedical applications.

  14. Click Chemistry-Mediated Nanosensors for Biochemical Assays (United States)

    Chen, Yiping; Xianyu, Yunlei; Wu, Jing; Yin, Binfeng; Jiang, Xingyu


    Click chemistry combined with functional nanoparticles have drawn increasing attention in biochemical assays because they are promising in developing biosensors with effective signal transformation/amplification and straightforward signal readout for clinical diagnostic assays. In this review, we focus on the latest advances of biochemical assays based on Cu (I)-catalyzed 1, 3-dipolar cycloaddition of azides and alkynes (CuAAC)-mediated nanosensors, as well as the functionalization of nanoprobes based on click chemistry. Nanoprobes including gold nanoparticles, quantum dots, magnetic nanoparticles and carbon nanomaterials are covered. We discuss the advantages of click chemistry-mediated nanosensors for biochemical assays, and give perspectives on the development of click chemistry-mediated approaches for clinical diagnosis and other biomedical applications. PMID:27217831

  15. Use of laminar flow patterning for miniaturised biochemical assays

    DEFF Research Database (Denmark)

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


    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...... 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......-streptavidin interactions, DNA-hybridization and DNA-triplex formation. The system is simple, fast, reproducible, flexible, and has small sample requirements....

  16. Comparison of Clot-based, Chromogenic, and Fluorescence Assays for Measurement of Factor VIII Inhibitors in the U.S. Hemophilia Inhibitor Research Study (United States)

    Miller, Connie H.; Rice, Anne S.; Boylan, Brian; Shapiro, Amy D.; Lentz, Steven R.; Wicklund, Brian M.; Kelly, Fiona M.; Soucie, J. Michael


    Summary Background Detection and validation of inhibitors (antibodies) to hemophilia treatment products are important for clinical care, evaluation of product safety, and assessment of population trends. Methods Centralized monitoring for factor VIII (FVIII) inhibitors was conducted for patients in the Hemophilia Inhibitor Research Study using a previously reported modified Nijmegen-Bethesda clotting assay (NBA), a chromogenic Bethesda assay (CBA), and a novel fluorescence immunoassay (FLI). Results NBA and CBA were performed on 1005 specimens and FLI on 272 specimens. CBA was negative on 880/883 specimens (99.7%) with Nijmegen-Bethesda units (NBU)<0.5 and positive on 42/42 specimens (100%) with NBU≥2.0 and 43/80 specimens (53.8%) with NBU 0.5–1.9. Among specimens with positive NBA and negative CBA, 58.1% were FLI-negative, 12.9% had evidence of lupus anticoagulant, and 35.5% had non-time-dependent inhibition. CBA and FLI were positive on 72.4% and 100% of 1.0–1.9 NBU specimens and 43.1% and 50.0% of 0.5–0.9 NBU specimens. FLI detected antibodies in 98.0% of CBA-positive and 81.6% of NBA-positive specimens (P=0.004). Among 21 new inhibitors detected by NBA, 5 (23.8%) with 0.7–1.3 NBU did not react in CBA or FLI. Among previously positive patients with 0.5–1.9 NBU, 7/25 (28%) were not CBA or FLI positive. FLI was positive on 36/169 NBU-negative specimens (21.3%). Conclusions FVIII specificity could not be demonstrated by CBA or FLI for 26% of inhibitors of 0.5–1.9 NBU; such results must be interpreted with caution. Low titer inhibitors detected in clot-based assays should always be repeated, with consideration given to evaluating their reactivity with FVIII using more specific assays. PMID:23601690

  17. Chromogenic smart materials

    Directory of Open Access Journals (Sweden)

    Carl M. Lampert


    Full Text Available Smart materials cover a wide and developing range of technologies. A particular type of smart material, known as chromogenics, can be used for large area glazing in buildings, automobiles, planes, and for certain types of electronic display. These technologies consist of electrically-driven media including electrochromism, suspended particle electrophoresis, polymer dispersed liquid crystals, electrically heated thermotropics, and gaschromics.

  18. Integration of electrochemistry in micro-total analysis systems for biochemical assays: recent developments. (United States)

    Xu, Xiaoli; Zhang, Song; Chen, Hui; Kong, Jilie


    Micro-total analysis systems (microTAS) integrate different analytical operations like sample preparation, separation and detection into a single microfabricated device. With the outstanding advantages of low cost, satisfactory analytical efficiency and flexibility in design, highly integrated and miniaturized devices from the concept of microTAS have gained widespread applications, especially in biochemical assays. Electrochemistry is shown to be quite compatible with microanalytical systems for biochemical assays, because of its attractive merits such as simplicity, rapidity, high sensitivity, reduced power consumption, and sample/reagent economy. This review presents recent developments in the integration of electrochemistry in microdevices for biochemical assays. Ingenious microelectrode design and fabrication methods, and versatility of electrochemical techniques are involved. Practical applications of such integrated microsystem in biochemical assays are focused on in situ analysis, point-of-care testing and portable devices. Electrochemical techniques are apparently suited to microsystems, since easy microfabrication of electrochemical elements and a high degree of integration with multi-analytical functions can be achieved at low cost. Such integrated microsystems will play an increasingly important role for analysis of small volume biochemical samples. Work is in progress toward new microdevice design and applications.

  19. An interlaboratory study on efficient detection of Shiga toxin-producing Escherichia coli O26, O103, O111, O121, O145, and O157 in food using real-time PCR assay and chromogenic agar. (United States)

    Hara-Kudo, Yukiko; Konishi, Noriko; Ohtsuka, Kayoko; Iwabuchi, Kaori; Kikuchi, Rie; Isobe, Junko; Yamazaki, Takumiko; Suzuki, Fumie; Nagai, Yuhki; Yamada, Hiroko; Tanouchi, Atsuko; Mori, Tetsuya; Nakagawa, Hiroshi; Ueda, Yasufumi; Terajima, Jun


    To establish an efficient detection method for Shiga toxin (Stx)-producing Escherichia coli (STEC) O26, O103, O111, O121, O145, and O157 in food, an interlaboratory study using all the serogroups of detection targets was firstly conducted. We employed a series of tests including enrichment, real-time PCR assays, and concentration by immunomagnetic separation, followed by plating onto selective agar media (IMS-plating methods). This study was particularly focused on the efficiencies of real-time PCR assays in detecting stx and O-antigen genes of the six serogroups and of IMS-plating methods onto selective agar media including chromogenic agar. Ground beef and radish sprouts samples were inoculated with the six STEC serogroups either at 4-6CFU/25g (low levels) or at 22-29CFU/25g (high levels). The sensitivity of stx detection in ground beef at both levels of inoculation with all six STEC serogroups was 100%. The sensitivity of stx detection was also 100% in radish sprouts at high levels of inoculation with all six STEC serogroups, and 66.7%-91.7% at low levels of inoculation. The sensitivity of detection of O-antigen genes was 100% in both ground beef and radish sprouts at high inoculation levels, while at low inoculation levels, it was 95.8%-100% in ground beef and 66.7%-91.7% in radish sprouts. The sensitivity of detection with IMS-plating was either the same or lower than those of the real-time PCR assays targeting stx and O-antigen genes. The relationship between the results of IMS-plating methods and Ct values of real-time PCR assays were firstly analyzed in detail. Ct values in most samples that tested negative in the IMS-plating method were higher than the maximum Ct values in samples that tested positive in the IMS-plating method. This study indicates that all six STEC serogroups in food contaminated with more than 29CFU/25g were detected by real-time PCR assays targeting stx and O-antigen genes and IMS-plating onto selective agar media. Therefore, screening

  20. Ultraminiaturized assay for rapid, low cost detection and quantification of clinical and biochemical samples. (United States)

    Parween, Shahila; Nahar, Pradip


    Herein, we report a simple, sensitive, rapid and low-cost ultraminiaturized assay technique for quantitative detection of 1 μl of clinical or biochemical sample on a novel ultraminiaturized assay plate (UAP). UAP is prepared by making tiny cavities on a polypropylene sheet. As UAP cannot immobilize a biomolecule through absorption, we have activated the tiny cavities of UAP by 1-fluoro-2-nitro-4-azidobenzene in a photochemical reaction. Activated UAP (AUAP) can covalently immobilize any biomolecule having an active nucleophilic group such as amino group. Efficacy of AUAP is demonstrated by detecting human IgE, antibody of hepatitis C virus core antigen and oligonucleotides. Quantification is performed by capturing the image of the colored assay solution and digitally quantifying the image by color saturation without using costly NanoDrop spectrophotometer. Image - based detection of human IgE and an oligonucleotide shows an excellent correlation with absorbance - based assay (recorded in a NanoDrop spectrophotometer); it is validated by Pearson's product-moment correlation with correlation coefficient of r = 0.9545088 and r = 0.9947444 respectively. AUAP is further checked by detecting hepatitis C virus Ab where strong correlation of color saturation with absorbance with respect to concentration is observed. Ultraminiaturized assay successfully detects target oligonucleotides by perfectly hybridizing with their respective complementary oligonucleotide probes but not with a random oligonucleotide. Ultraminiaturized assay technique has substantially reduced the requirement of reagents by 100 times and assay timing by 50 times making it a potential alternative to conventional method.

  1. Characterization of the early events in dengue virus cell entry by biochemical assays and single-virus tracking

    NARCIS (Netherlands)

    van der Schaar, Hilde M.; Rust, Michael J.; Waarts, Barry-Lee; van der Ende-Metselaarl, Heidi; Kuhn, Richard J.; Wilschut, Jan; Zhuang, Xiaowei; Smit, Jolanda M.


    In this study, we investigated the cell entry characteristics of dengue virus (DENV) type 2 strain SI on mosquito, BHK-15, and BS-C-1 cells. The concentration of virus particles measured by biochemical assays was found to be substantially higher than the number of infectious particles determined by

  2. A sensitive ferricyanide-mediated biochemical oxygen demand assay for analysis of wastewater treatment plant influents and treated effluents. (United States)

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


    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 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.

  3. Enzyme assays. (United States)

    Reymond, Jean-Louis; Fluxà, Viviana S; Maillard, Noélie


    Enzyme assays are analytical tools to visualize enzyme activities. In recent years a large variety of enzyme assays have been developed to assist the discovery and optimization of industrial enzymes, in particular for "white biotechnology" where selective enzymes are used with great success for economically viable, mild and environmentally benign production processes. The present article highlights the aspects of fluorogenic and chromogenic substrates, sensors, and enzyme fingerprinting, which are our particular areas of interest.

  4. Centrifugal multiplexing fixed-volume dispenser on a plastic lab-on-a-disk for parallel biochemical single-end-point assays. (United States)

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


    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.

  5. Generation and precise control of dynamic biochemical gradients for cellular assays (United States)

    Saka, Yasushi; MacPherson, Murray; Giuraniuc, Claudiu V.


    Spatial gradients of diffusible signalling molecules play crucial roles in controlling diverse cellular behaviour such as cell differentiation, tissue patterning and chemotaxis. In this paper, we report the design and testing of a microfluidic device for diffusion-based gradient generation for cellular assays. A unique channel design of the device eliminates cross-flow between the source and sink channels, thereby stabilizing gradients by passive diffusion. The platform also enables quick and flexible control of chemical concentration that makes highly dynamic gradients in diffusion chambers. A model with the first approximation of diffusion and surface adsorption of molecules recapitulates the experimentally observed gradients. Budding yeast cells cultured in a gradient of a chemical inducer expressed a reporter fluorescence protein in a concentration-dependent manner. This microfluidic platform serves as a versatile prototype applicable to a broad range of biomedical investigations.

  6. Generation and precise control of dynamic biochemical gradients for cellular assays

    CERN Document Server

    Saka, Yasushi; Giuraniuc, Claudiu V


    Spatial gradients of diffusible signalling molecules play crucial roles in controlling diverse cellular behaviour such as cell differentiation, tissue patterning and chemotaxis. Here we present a microfluidic platform for cellular assays that can generate and control diffusion-based gradients dynamically. A unique design of the device eliminates cross-flow between the source and sink channels, thereby stabilising gradients by passive diffusion. The platform also enables quick and flexible control of chemical concentration that makes highly dynamic gradients in diffusion chambers. Budding yeast cells cultured in a gradient of a chemical inducer expressed a reporter fluorescence protein in a concentration-dependent manner. This microfluidic platform serves as a versatile prototype applicable to a broad range of biomedical investigations.

  7. Development and validation of a quick easily used biochemical assay for evaluating the viability of small immobile arthropods. (United States)

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


    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.

  8. Comparative analyses of universal extraction buffers for assay of stress related biochemical and physiological parameters. (United States)

    Han, Chunyu; Chan, Zhulong; Yang, Fan


    Comparative efficiency of three extraction solutions, including the universal sodium phosphate buffer (USPB), the Tris-HCl buffer (UTHB), and the specific buffers, were compared for assays of soluble protein, free proline, superoxide radical (O2∙-), hydrogen peroxide (H2O2), and the antioxidant enzymes such as superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (POD), ascorbate peroxidase (APX), glutathione peroxidase (GPX), and glutathione reductase (GR) in Populus deltoide. Significant differences for protein extraction were detected via sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and two-dimensional electrophoresis (2-DE). Between the two universal extraction buffers, the USPB showed higher efficiency for extraction of soluble protein, CAT, GR, O2∙-, GPX, SOD, and free proline, while the UTHB had higher efficiency for extraction of APX, POD, and H2O2. When compared with the specific buffers, the USPB showed higher extraction efficiency for measurement of soluble protein, CAT, GR, and O2∙-, parallel extraction efficiency for GPX, SOD, free proline, and H2O2, and lower extraction efficiency for APX and POD, whereas the UTHB had higher extraction efficiency for measurement of POD and H2O2. Further comparisons proved that 100 mM USPB buffer showed the highest extraction efficiencies. These results indicated that USPB would be suitable and efficient for extraction of soluble protein, CAT, GR, GPX, SOD, H2O2, O2∙-, and free proline.

  9. 发色底物法在酶促反应初速度内测定α1抗胰蛋白酶的活性%Detection ofα1 antitrypsin activity by chromogenic substrate assay with initial veloci-ty of enzymatic reaction

    Institute of Scientific and Technical Information of China (English)

    张晋超; 赵雄; 吕茂民; 尹惠琼; 王延琳; 章金刚


    Objective To detect the activity of α1 antitrypsin(AAT) with initial velocity of enzymatic reaction in order to detect the activity of samples in the process of separating and purifying plasma protein ,chromogenic substrate assay was optimized.Methods The effect of trypsin concentration and reaction time on enzymatic reaction was acquired by the kinetic monitoring mode of the microplate reader .Initial velocity was calculated to confirm the largest concentration of trypsin which was saturated by substrate .AAT was incubated with trypsin and absorbance produced by enzymatic reaction of remaining trypsin and substrate could reflect the activity of AAT .A standard curve was established with △D fitting with the activity of AAT standard.The activity of related samples was detected and the precision and accuracy of the method were evaluated . Results Trypsin concentration was 0.0625 mg/ml.Within 20 minutes, enzymatic reaction was with initial velocity .The range of the standard curve was 200-1200 IU/ml.Correlation coefficient was more than 0.99.The activity of Cohn Ⅳ, samples of pre-processing and elution were (720.59 ±18.63), (601.84 ±19.18),and (568.09 ±24.83)IU/ml, respec-tively.The relative standard deviation was less than 10%. Sample recovery rate was 90%-110%.Conclusion The optimized chromogenic substrate assay greatly improves accuracy and precision .The method can be used for the detec-tion of AAT activity of samples in laboratories and workshops .%目的:优化发色底物法,使其在酶促反应初速度内测定α1抗胰蛋白酶( AAT)的活性并用于血浆蛋白纯化过程中各样品活性的检测。方法采用酶标仪动态监测模式观察酶浓度和反应时间对酶促反应的影响;计算初速度并确定被底物饱和的最大酶浓度。将AAT与胰蛋白酶孵育,剩余靶酶和底物作用产生的光密度可反映AAT的活性。通过△D与AAT标准品活性进行拟合建立标准曲线,测定相关样品的活

  10. Zn(OTf)2-catalysed indolylation and pyrrolylation of isatins: Efficient synthesis and biochemical assay of 3,3-di(heteroaryl)oxindoles

    Indian Academy of Sciences (India)

    C Praveen; S Narendiran; P Dheenkumar; P T Perumal


    An efficient and cheap synthetic approach to 3,3-di(indolyl)oxindoles and 3,3-di(pyrrolyl) oxindoles has been developed via Zn(OTf)2 catalysed indolylation and pyrrolylation of isatins. A preliminary biochemical assay of the synthesized molecules in rodent models were performed to estimate the serum glutamate oxaloacetate transaminase and malondialdehyde levels.

  11. 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.

  12. Determination of bacterial endotoxin content of diphtheria toxin mutant CRM 197 by kinetic chromogenic assay%动态显色法检测白喉毒素突变体CRM197的细菌内毒素含量

    Institute of Scientific and Technical Information of China (English)

    邓杰; 张珂; 袁涛; 李春阳


    目的 建立检测白喉毒素突变体CRM197细菌内毒素(bacterial endotoxin)含量的动态显色法(kinetic chromogenic assay,KCA).方法 按照《中国药典》三部(2010版)要求,用细菌内毒素检查用水(bacterial endotoxin,BET)稀释细菌内毒素工作标准品制备细菌内毒素标准曲线系列溶液,各浓度均设3个平行孔,分别与动态显色鲎试剂(kinetic chromogenic analysis tachypleus amebocyte lysate,KCA TAL)反应,绘制标准曲线,验证标准曲线的可靠性,确定最佳线性范围、测定范围及最低检测限(limit of quantitation,LOQ),验证方法的精密性和准确性,并进行初步应用.结果 标准曲线的回归方程为:y=-0.263x+2.741,R2=0.997,相关系数的绝对值|r|≥0.980,阴性对照的反应时间大于标准曲线最低点的反应时间,各复孔的变异系数(CV)<10%;内毒素浓度在0.02~2.50EU/ml时,线性关系良好,LOQ为0.02 EU/ml;3个浓度(2.50、0.50、0.02 EU/ml)标准内毒素检测结果的CV均<5%,加入高、中、低3个浓度(2.50、0.50、0.02 EU/ml)标准内毒素的3批供试品检测结果的CV均<10%,回收率在95% ~ 143%之间;采用建立的方法检测10批次CRM197样品的细菌内毒素含量,回收率在77%~118%之间,其中8批样品的内毒素含量合格.结论 动态显色法检测CRM197中内毒素的含量精密性和准确性良好,能快速、定量检测样品中的内毒素含量,抗干扰能力强,可用于CRM197研制过程中的质量控制.

  13. Hormonal and biochemical serum assay in relation to the estrous cycle and follicular growth in Arabian mare

    Institute of Scientific and Technical Information of China (English)

    Amal M Abo-El maaty; K H El-Shahat


    Objective:The current study is an endeavor for profound understanding hormonal and biochemical constituents in Arabian mare serum during the estrous cycle.Methods:Ten Arab brood mares of previous foaling records were scanned with ultrasound each other day, and blood samples were collected with each ultrasound examination. The diameter of follicles and corpus luteum was measured. The follicles were classified into small, medium, dominant, 1st and 2nd largest according to their diameter. Hormonal concentrations of thyroxin, progesterone, estradiol and testosterone were assayed. Superoxide dismutase (SOD), malondialdehyde (MDA), nitric oxide (NO) and total antioxidants concentrations (TAC) were measured. Besides, the levels of total proteins, albumin, globulin, glucose, total cholesterol, and triglycerides were assayed. Results:The data revealed that multiple follicles underwent progressive enlargement (≤30 mm). Only the largest (dominant) follicle reached a maximal diameter of (35.82±1.57) mm during estrous phase. During the luteal phase, the corpus luteum reached a maximum diameter of (31.78±1.4) mm. The serum progesterone levels in mare were significantly (P<0.05) higher in luteal phase than those recorded at time of estrus. The reverse was true for estradiol. However, the levels of testosterone and thyroxin did not significantly change during estrous cycle in mare. No significant difference was observed in the serum levels of total protein, albumin and globulin in both phases of estrous cycle. Similar finding was observed in SOD concentrations. In contrast, the concentrations of glucose, cholesterol and triglyceride tended to be significantly (P<0.05) higher at estrous stage than those recorded at luteal one. The same finding was observed in NO levels. On other hand, TAC significantly increased (P<0.05) in mare serum obtained from luteal phase than those obtained at estrous one. A reverse was true for MDA levels.Conlcusions:The steroid hormones and metabolic

  14. Exudative epidermitis in pigs caused by toxigenic Staphylococcus chromogenes

    DEFF Research Database (Denmark)

    Andresen, Lars Ole; Ahrens, Peter; Daugaard, Lise


    Staphylococcus chromogenes is closely related to Staphylococcus hyicus, which is recognised as the causative agent of exudative epidermitis (EE) in pigs. S. chromogenes is part of the normal skin flora of pigs, cattle and poultry and has so far been considered non-pathogenic to pigs. A strain of S...

  15. Revisiting catechol derivatives as robust chromogenic hydrogen donors working in alkaline media for peroxidase mimetics. (United States)

    Drozd, Marcin; Pietrzak, Mariusz; Pytlos, Jakub; Malinowska, Elżbieta


    Colloidal noble metal-based nanoparticles are able to catalyze oxidation of chromogenic substrates by H2O2, similarly to peroxidases, even in basic media. However, lack of robust chromogens, which work in high pH impedes their real applications. Herein we demonstrate the applicability of selected catechol derivatives: bromopyrogallol red (BPR) and pyrogallol (PG) as chromogenic substrates for peroxidase-like activity assays, which are capable of working over wide range of pH, covering also basic values. Hyperbranched polyglycidol-stabilized gold nanoparticles (HBPG@AuNPs) were used as model enzyme mimetics. Efficiency of several methods of improving stability of substrates in alkaline media by means of selective suppression of their autoxidation by molecular oxygen was evaluated. In a framework of presented studies the impact of borate anion, applied as complexing agent for PG and BPR, on their stability and reactivity towards oxidation mediated by catalytic AuNPs was investigated. The key role of high concentration of hydrogen peroxide in elimination of non-catalytic oxidation of PG and improvement of optical properties of BPR in alkaline media containing borate was underlined. Described methods of peroxidase-like activity characterization with the use of BPR and PG can become universal tools for characterization of nanozymes, which gain various applications, among others, they are used as catalytic labels in bioassays and biosensors.

  16. Comparative Study of a Novel Biochemical Assay, the Rapidec Carba NP Test, for Detecting Carbapenemase-Producing Enterobacteriaceae. (United States)

    Lifshitz, Ziv; Adler, Amos; Carmeli, Yehuda


    The novel biochemical test, the Rapidec Carba NP (RCNP), was evaluated using carbapenemase- and non-carbapenemase-producing Enterobacteriaceae isolates. The RCNP test was compared with the Carba NP test (CNP) and the modified Hodge test. Compared to the CNP test, the RCNP test had identical sensitivity (96%) and lower specificity (93% versus 100%). The medium used to culture the isolates significantly affected test sensitivity and specificity. The RCNP test was quicker and easier to perform than the other tests.

  17. 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


    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...

  18. Selective chromogenic detection of thiol-containing biomolecules using carbonaceous nanospheres loaded with silver nanoparticles as carrier. (United States)

    Hu, Bo; Zhao, Yang; Zhu, Hai-Zhou; Yu, Shu-Hong


    Thiol-containing biomolecules show strong affinity with noble metal nanostructures and could not only stably protect them but also control the self-assembly process of these special nanostructures. A highly selective and sensitive chromogenic detection method has been designed for the low and high molecular weight thiol-containing biomolecules, including cysteine, glutathione, dithiothreitol, and bovine serum albumin, using a new type of carbonaceous nanospheres loaded with silver nanoparticles (Ag NPs) as carrier. This strategy relies upon the place-exchange process between the reporter dyes on the surface of Ag NPs and the thiol groups of thiol-containing biomolecules. The concentration of biomolecules can be determined by monitoring with the fluorescence intensity of reporter dyes dispersed in solution. This new chromogenic assay method could selectively detect these biomolecules in the presence of various other amino acids and monosaccharides and even sensitively detect the thiol-containing biomolecules with different molecular weight, even including proteins.

  19. The effect of multiple analysers on the biochemical diagnosis of myocardial infarction using a contemporary troponin-I assay. (United States)

    Pethick, James; Patel, Prashanth; Davies, Timothy; Thompson, John; Nallagonda, Madhavi; Beech, Alison; Collinson, Paul; Lee, Virginia; Gupta, Pankaj


    Background The measurement of cardiac troponin is central for the diagnosis of myocardial infarction (MI). It is recommended that a coefficient of variation of ≤10% is achieved at the diagnostic threshold and significant change between serial measurements reported. Many modern laboratories use multiple analysers linked by automation where samples are randomly assigned to an analyser. It is therefore important to consider the combined effect of all analysers on the analytical performance of troponin measurement. Method The performance of a contemporary troponin-I (cTn-I) assay run on three analysers, linked by an automated track, was undertaken across a range of cTn-I concentrations. The data for the three analysers were aggregated to obtain the combined analytical coefficient of variation (CVA) and reference change values (RCVs). Results The CVA improved with increasing concentration and calculated RCVs ranged from 67.2% (±13 ng/L) to 32% (±160 ng/L) between cTn-I values 20 ng/L and 500 ng/L. Although there were significant differences in cTn-I measurement between analysers around the diagnostic threshold ( P diagnosis of MI. We also show that the RCV varies according to baseline cTn-I values and that reporting a single RCV across the analytical range of cTn-I may not be appropriate.

  20. Anaerobic digestion of solid waste in RAS: Effect of reactor type on the biochemical acidogenic potential (BAP) and assessment of the biochemical methane potential (BMP) by a batch assay

    DEFF Research Database (Denmark)

    Suhr, Karin Isabel; Letelier-Gordo, Carlos Octavio; Lund, Ivar


    additional 14 and 20 days) in continuously stirred tank reactors. Generally, the VFA yield increased with time and no effect of the reactor type used was found within the time frame of the experiment. At 10 days HT or 10 days HRT the VFA yield reached 222.3 ± 30.5 and 203.4 ± 11.2 mg VFA g-1 TVS0 (total...... volatile solids at day 0) in batch and fed-batch reactor, respectively. For the fedbatch reactor, increasing HRT from 5 to 10 days gained no significant additional VFA yield. Prolonging the batch reactor experiment to 20 days increased VFA production further (273.9 ± 1.6 mg VFA g-1 TVS0, n=2). After 10...... for the design of an acidogenic continuously stirred reactor tank in a RAS single-sludge denitrification set-up. The biochemical methane potential of the sludge was estimated to 318 ± 29 g CH4 g-1 TVS0 by a batch assay and represented a higher utility of the solid waste when comparing the methane yield...

  1. Fast, Continuous, and High-Throughput (Bio)Chemical Activity Assay for N-Acyl-l-Homoserine Lactone Quorum-Quenching Enzymes (United States)

    Last, Daniel; Krüger, Georg H. E.; Dörr, Mark


    ABSTRACT Quorum sensing, the bacterial cell-cell communication by small molecules, controls important processes such as infection and biofilm formation. Therefore, it is a promising target with several therapeutic and technical applications besides its significant ecological relevance. Enzymes inactivating N-acyl-l-homoserine lactones, the most common class of communication molecules among Gram-negative proteobacteria, mainly belong to the groups of quorum-quenching lactonases or quorum-quenching acylases. However, identification, characterization, and optimization of these valuable biocatalysts are based on a very limited number of fundamentally different methods with their respective strengths and weaknesses. Here, a (bio)chemical activity assay is described, which perfectly complements the other methods in this field. It enables continuous and high-throughput activity measurements of purified and unpurified quorum-quenching enzymes within several minutes. For this, the reaction products released by quorum-quenching lactonases and quorum-quenching acylases are converted either by a secondary enzyme or by autohydrolysis to l-homoserine. In turn, l-homoserine is detected by the previously described calcein assay, which is sensitive to α-amino acids with free N and C termini. Besides its establishment, the method was applied to the characterization of three previously undescribed quorum-quenching lactonases and variants thereof and to the identification of quorum-quenching acylase-expressing Escherichia coli clones in an artificial library. Furthermore, this study indicates that porcine aminoacylase 1 is not active toward N-acyl-l-homoserine lactones as published previously but instead converts the autohydrolysis product N-acyl-l-homoserine. IMPORTANCE In this study, a novel method is presented for the identification, characterization, and optimization of quorum-quenching enzymes that are active toward N-acyl-l-homoserine lactones. These are the most common

  2. Chromogenic switchable glazing: Towards the development of the smart window

    Energy Technology Data Exchange (ETDEWEB)

    Lampert, C.M.


    The science and technology of chromogenic materials for switchable glazings in building applications is discussed. These glazings can be used for dynamic control of solar and visible energy. Currently many researchers and engineers are involved with the development of products in this field. A summary of activities in Japan, Europe, Australia, USA and Canada is made. The activities of the International Energy Agency are included. Both non-electrically activated and electrically activated glazings are discussed. Technologies covered in the first category are photochromics, and thermochromics and thermotropics. A discussion of electrically activated chromogenic glazings includes dispersed liquid crystals, dispersed particles and electrochromics. A selection of device structures and performance characteristics are compared. A discussion of transparent conductors is presented. Technical issues concerning large-area development of smart windows are discussed.

  3. Chromogenic in situ hybridization: a multicenter study comparing silver in situ hybridization with FISH. (United States)

    Bartlett, J M S; Campbell, Fiona M; Ibrahim, Merdol; Wencyk, Peter; Ellis, Ian; Kay, Elaine; Connolly, Yvonne; O'Grady, Anthony; Di Palma, Silvana; Starczynski, Jane; Morgan, John M; Jasani, Bharat; Miller, Keith


    Our purposes were to perform a robust assessment of a new HER2 chromogenic in situ hybridization test and report on concordance of silver in situ hybridization (SISH) data with fluorescence in situ hybridization (FISH) data and on intraobserver and interlaboratory scoring consistency. HER2 results were scored from 45 breast cancers in 7 laboratories using the Ventana (Tucson, AZ) INFORM HER-2 SISH assay and in 1 central laboratory using a standard FISH assay. Overall, 94.8% of cases were successfully analyzed by SISH across the 6 participating laboratories that reported data. Concordance for diagnosis of HER2 amplification by SISH compared with FISH was high (96.0% overall). Intraobserver variability (8.0%) and intersite variability (12.66%) of absolute HER2/chromosome 17 ratios appear to be tightly controlled across all 6 participating laboratories. The Ventana INFORM HER-2 SISH assay is robust and reproducible, shows good concordance with a standard FISH assay, and complies with requirements in national guidelines for performance of diagnostic tests.

  4. A selective spectrophotometric method for determination of rosoxacin antibiotic using sodium nitroprusside as a chromogenic reagent (United States)

    Askal, Hassan F.; Refaat, Ibrahim H.; Darwish, Ibrahim A.; Marzouq, Mostafa A.


    A selective spectrophotometric method for the determination of rosoxacin (ROS), a 4-quinolone antimicrobial agent, has been developed and validated. The method was based on the reaction of ROS with alkaline sodium nitroprusside (SNP) reagent at room temperature forming a red colored chromogen measured at 455 nm. The conditions affecting the reaction (SNP concentration, pH, color-developing time, temperature, diluting solvent and chromogen stability time) were optimized. Under the optimum conditions, good linear relationship ( r = 0.9987) was obtained between the absorbance and the concentration of ROS in the range of 20-50 μg ml -1. The assay limits of detection and quantitation were 2.5 and 8.4 μg ml -1, respectively. The method was successfully applied to the analysis of bulk drug and laboratory-prepared tablets; the mean percentage recoveries were 100.1 ± 0.33 and 101.24 ± 1.28%, respectively. The results were compared favourably with those obtained by the reported method; no significant difference in the accuracy and precision as revealed by the accepted values of t- and F-tests, respectively. The robustness and ruggedness of the method was checked and satisfactory results were obtained. The proposed method was found to be highly selective for ROS among the fluoroquinolone antibiotics. The reaction mechanism was proposed and it proceeded in two steps; the formation of nitroferrocyanide by the action of sodium hydroxide alkalinity on SNP and the subsequent formation of the colored nitrosyl-ROS derivative by the attack at position 6 of ROS.

  5. Identification of microorganisms grown on chromogenic media by MALDI-TOF MS. (United States)

    Lüthje, Petra; Pranada, Arthur B; Carruthers-Lay, Duncan; Desjardins, Marc; Gaillot, Olivier; Wareham, David; Ciesielczuk, Holly; Özenci, Volkan


    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and chromogenic media are widely used in clinical microbiology laboratories to facilitate the rapid selection and identification of pathogens. The aim of this study was to evaluate whether usage of chromogenic media limits the diagnostic performance of MALDI-TOF MS for microbial identification. A total of 386 microorganisms collected and analyzed at five laboratories were included. Isolates were cultured on relevant chromogenic media and non-selective agar plates in parallel and identified using the Bruker MALDI-TOF MS. Among the tested isolates, no misidentification was recorded and there was no medium-related difference in the identification level. However, score values were overall slightly but significantly lower for isolates grown on chromogenic media. In conclusion, the use of chromogenic culture media tested here had no relevant impact on MALDI-TOF MS performance for diagnostic purposes.

  6. Comparison of Fluorescence In Situ Hybridization and Chromogenic In Situ Hybridization for Low and High Throughput HER2 Genetic Testing

    DEFF Research Database (Denmark)

    Poulsen, Tim S; Espersen, Maiken Lise Marcker; Kofoed, Vibeke;


    results show that the differences between the HER2 genetic assays do not have an effect on the analytic performance and the CISH technology is superior to high throughput HER2 genetic testing due to scanning speed, while the IQ-FISH may still be a choice for fast low throughput HER2 genetic testing.......The purpose was to evaluate and compare 5 different HER2 genetic assays with different characteristics that could affect the performance to analyze the human epidermal growth factor 2 (HER2) gene copy number under low and high throughput conditions. The study included 108 tissue samples from breast...... cancer patients with HER2 immunohistochemistry (IHC) results scored as 0/1+, 2+, and 3+. HER2 genetic status was analysed using chromogenic in situ hybridization (CISH) and fluorescence in situ hybridization (FISH). Scoring results were documented through digital image analysis. The cancer region...

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


    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, w

  8. Colorimetric protein assay techniques. (United States)

    Sapan, C V; Lundblad, R L; Price, N C


    There has been an increase in the number of colorimetric assay techniques for the determination of protein concentration over the past 20 years. This has resulted in a perceived increase in sensitivity and accuracy with the advent of new techniques. The present review considers these advances with emphasis on the potential use of such technologies in the assay of biopharmaceuticals. The techniques reviewed include Coomassie Blue G-250 dye binding (the Bradford assay), the Lowry assay, the bicinchoninic acid assay and the biuret assay. It is shown that each assay has advantages and disadvantages relative to sensitivity, ease of performance, acceptance in the literature, accuracy and reproducibility/coefficient of variation/laboratory-to-laboratory variation. A comparison of the use of several assays with the same sample population is presented. It is suggested that the most critical issue in the use of a chromogenic protein assay for the characterization of a biopharmaceutical is the selection of a standard for the calibration of the assay; it is crucial that the standard be representative of the sample. If it is not possible to match the standard with the sample from the perspective of protein composition, then it is preferable to use an assay that is not sensitive to the composition of the protein such as a micro-Kjeldahl technique, quantitative amino acid analysis or the biuret assay. In a complex mixture it might be inappropriate to focus on a general method of protein determination and much more informative to use specific methods relating to the protein(s) of particular interest, using either specific assays or antibody-based methods. The key point is that whatever method is adopted as the 'gold standard' for a given protein, this method needs to be used routinely for calibration.

  9. Study of endotoxin release by Salmonella thyphi by antibiotics in Chromogenic Limulus Amebocyte Lyssate Method

    Directory of Open Access Journals (Sweden)

    Shagari G


    Full Text Available It seems, rapid destruction of gram negative bacteria by antibiotics contribute to the clinical deterioration of some patients with lethal endotoxemia. In this research we evaluated LPS (lipopolysaccharide release during antibiotic killing of salmonella typhi (Ty2-5536. The organism was incubated in the presence of Chloramphenicol, Ampicillin and Co-trimoxazole at concentrations that killed >99.9% of the organisms as determined by quantitative culture techniques. After incubation the antibiotic-bacterial cultures were centrifuged and the supernatants were filtered and collected for "in vivo" and "in vitro" studies. Injection of 1 ml/kg of filtrates in rabbits raised normal temperature of the animals by 1.2°C that it showes the presence of LPS in the filtrates. Quantitative chromogenic limulus amebocyte lyssate (L.A.L assay was used to determine the amount of LPS in the filtrates. The amount of LPS was 86.67±2.53 Pg/ml for chloramphenicol, 113.33±8.07 Pg/ml for Ampicillin and 134.18±11.59 Pg/ml for Co-trimoxazole. According to our investigation chloramphenicol is the best antibiotic against S.typhi because it decrease the induced-pathological effects of LPS in gram negative infection

  10. Biochemical studies of mouse brain tubulin: colchicine binding (DEAE-cellulose filter) assay and subunits (. cap alpha. and. beta. ) biosynthesis and degradation (in newborn brain)

    Energy Technology Data Exchange (ETDEWEB)

    Tse, Cek-Fyne


    A DEAE-cellulose filter assay, measuring (/sup 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 ..cap alpha.. and ..beta.. subunits of two day mouse brain 100,000g supernatant were studied after intracerebral injection of (/sup 3/H)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)

  11. The role of chromogenic bacteria in external tooth discoloration in children

    Directory of Open Access Journals (Sweden)

    Zuhal Kırzıoğlu


    Full Text Available Tooth discoloration can cause social and psychological problems and lead to loss of self-confidence in children. Concerns of parents and children on appearance and esthetics increase the importance of dental esthetic treatments in children. External tooth discoloration in children can be seen in both the primary and the permanent teeth. Poor oral hygiene, use of iron preparations and chromogenic bacteria are among the etiology of external tooth discoloration. Chromogenic bacteria can cause black/brown, green, orange or blue discolorations on the tooth surfaces of children. For clinicians, most interesting external tooth discoloration in children has been the black discoloration, which is more frequently observed compared to the other types. Most of the studies also have focused on the chromogenic bacteria associated with black discolorations, and the data concerning other types of discoloration are limited. Our knowledge on the existing chromogenic bacterial mechanisms is increasing by the developing microbiological diagnostic procedures, and new species associated with discoloration are becoming recognized. Still, studies dealing with the mechanisms of the chromogenic bacteria leading to discoloration are insufficient in the accessible sources. The objective of this article is to emphasize the role of the chromogenic bacteria in external tooth discoloration in children, to review precautions that can be taken to prevent discoloration, and current treatment methods, and to contribute to the current knowledge of the clinicians on this subject.

  12. 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. (United States)

    Zhu, Ming-Zhi; Wu, Wei; Jiao, Li-Li; Yang, Ping-Fang; Guo, Ming-Quan


    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.

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

    KAUST Repository

    Breuer, Sebastian


    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.

  14. 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


    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.

  15. [Evaluation of chromogenic medium Uriselect4 in urine culture]. (United States)

    Ferjani, Asma; Marzouk, Manel; Idriss, Nadia; Sammoud, Sammoud; Hannachi, Naila; Boukadida, Janel


    We evaluated the performance and the cost of chromogenic medium Uriselect4 agar with regard to the standard medium for the detection and identification of urinary tract pathogens. A total of 503 clinical urine specimens containing leucocytes greater or equal to 104/mL were analysed prospectively, in parallel by two different persons on blood agar (GS) and Uriselect4 according to the manufacturers' instructions. Of the 503 urine specimens tested, 210 gave a positive culture on Uriselect4 versus 181 on GS. The majority of bacterial species grew on both media; enterobacteria grew on Uriselect4 better than GS. The identification of Escherichia coli (E. coli), Proteus mirabilis (P. mirabilis), KES group and Enterococcus faecalis (E. faecalis) did not require the use of galleries Api and has a gain of 24  h. Positive pure cultures on Uriselect4 corresponding to negative cultures of GS were noted in 17 ases. Conversely, in seven cases a positive pure culture on GS was noted while the corresponding Uriselect4 cultures were negative. The cost of identification on GS (including the cost of galleries Api), was about two times higher than Uriselect4. Uriselect4 medium isolates the most frequent urinary tract pathogens and identify them so almost immediately, with a lower cost.

  16. Recent Advance in Chemiluminescence Assay and Its Biochemical Applications%化学发光在生化分析中的应用研究进展

    Institute of Scientific and Technical Information of China (English)

    刘萌; 王子月; 张春阳


    化学发光分析是利用化学发光反应的发光现象,对化学发光物质由激发态跃迁回基态时发出的光信号进行测量的一种分析方法。化学发光分析具有无需外来光源、灵敏度高、操作方便、分析快速以及易于实现自动化等优点,可与其它分析技术联用,在临床检验、药物分析和环境监测等领域具有广泛应用。近年来,纳米材料、生物芯片及微流控技术的引入促进了化学发光分析技术的发展。本文综述了化学发光分析与高效液相色谱、毛细管电泳、量子点、微流控芯片和微阵列、以及滚环扩增、等温指数扩增和两级等温扩增联用技术的发展,介绍了化学发光分析技术在DNA、生物小分子、生物酶、蛋白质和金属离子检测中的应用研究进展,并展望了其发展趋势。%Chemiluminescence ( CL) assay is able to measure the optical signal emitted from the CL reagents as a result of the transition from the excited state back to the ground state. CL assay has significant advantages such as no external light source, high sensitivity, convenient operation, rapid analysis and easy automation, and has wide applications in clinical test, drug analysis and environmental monitoring. Recently, the introduction of nanomaterials, biochip and microfluidic techniques promote the development of CL assay. In this review, we summarize the recent progress in CL assay with the integration of high performance liquid chromatography, capillary electrophoresis, quantum dots, microfluidic chips, microarrays, rolling circle amplification, isothermal exponential amplification, and two-stage isothermal amplification for the detection of DNA, small biological molecules, enzymes, proteins and metal ions. We also give a summary of its future directions and highlight its potential applications.

  17. Covalently deposited dyes: a new chromogen paradigm that facilitates analysis of multiple biomarkers in situ. (United States)

    Day, William A; Lefever, Mark R; Ochs, Robert L; Pedata, Anne; Behman, Lauren J; Ashworth-Sharpe, Julia; Johnson, Donald D; May, Eric J; Grille, James G; Roberts, Esteban A; Kosmeder, Jerry W; Morrison, Larry E


    Multiplexed analysis of multiple biomarkers in a tissue sample requires use of reporter dyes with specific spectral properties that enable discrimination of signals. Conventional chromogens with broad absorbance spectra, widely used in immunohistochemistry (IHC), offer limited utility for multiplexed detection. Many dyes with narrow absorbance spectra, eg rhodamines, fluoresceins, and cyanines, potentially useful for multiplexed detection are well-characterized; however, generation of a chromogenic reagent useful for IHC analysis has not been demonstrated. Studies reported herein demonstrate utility of tyramine-chemistry for synthesis of a wide variety of new chromogenic dye conjugates useful for multiplexed in situ analysis using conventional light microscopes. The dyes, useful individually or in blends to generate new colors, provide signal sensitivity and dynamic range similar to conventional DAB chromogen, while enabling analysis of co-localized biomarkers. It is anticipated that this new paradigm will enable generation of a wide variety of new chromogens, useful for both research and clinical biomarker analysis that will benefit clinicians and patients.

  18. Specific and global coagulation assays in the diagnosis of discrepant mild hemophilia A. (United States)

    Bowyer, Annette E; Van Veen, Joost J; Goodeve, Anne C; Kitchen, Steve; Makris, Michael


    The activity of the factor VIII coagulation protein can be measured by three methods: a one or two-stage clotting assay and a chromogenic assay. The factor VIII activity of most individuals with mild hemophilia A is the same regardless of which method is employed. However, approximately 30% of patients show marked discrepancies in factor VIII activity measured with the different methods. The objective of this study was to investigate the incidence of assay discrepancy in our center, assess the impact of alternative reagents on factor VIII activity assays and determine the usefulness of global assays of hemostasis in mild hemophilia A. Factor VIII activity was measured in 84 individuals with mild hemophilia A using different reagents. Assay discrepancy was defined as a two-fold or greater difference between the results of the one-stage and two-stage clotting assays. Rotational thromboelastometry and calibrated automated thrombography were performed. Assay discrepancy was observed in 31% of individuals; 12% with lower activity in the two-stage assay and 19% with lower activity in the one-stage assay. The phenotype could not always be predicted from the individual's genotype. Chromogenic assays were shown to be a suitable alternative to the two-stage clotting assay. Thromboelastometry was found to have poor sensitivity in hemophilia. Calibrated automated thrombography supported the results obtained by the two-stage and chromogenic assays. The current international guidelines do not define the type of assay to be used in the diagnosis of mild hemophilia A and some patients could be misclassified as normal. In our study, 4% of patients would not have been diagnosed on the basis of the one-stage factor VIII assay. Laboratories should use both one stage and chromogenic (or two-stage) assays in the diagnosis of patients with possible hemophilia A.

  19. Selective chromogenic and fluorogenic peptide substrates for the assay of cysteine peptidases in complex mixtures (United States)

    Cysteine peptidases are important in many biological processes. In this study, we describe the design, synthesis and use of selective peptide substrates for cysteine peptidases of the C1 papain family. The structure of the proposed substrates can be expressed by the general formula Glp-Xaa-Ala-Y, wh...

  20. High-throughput analysis of endogenous fruit glycosyl hydrolases using a novel chromogenic hydrogel substrate assay

    DEFF Research Database (Denmark)

    Schückel, Julia; Kracun, Stjepan Kresimir; Lausen, Thomas Frederik;


    an important role in fruit development and ripening processes by modulating the plant cell wall. Knowledge about these enzymes is important for research in fruit development and also important for industry regarding postharvest properties. Although advances in genetic control and cell wall biochemistry have...

  1. Chromogenic agar medium for detection and isolation of Escherichia coli serogroups O26, O45, O103, O111, O121, and O145 from fresh beef and cattle feces. (United States)

    Kalchayanand, Norasak; Arthur, Terrance M; Bosilevac, Joseph M; Wells, James E; Wheeler, Tommy L


    Non-O157 Shiga toxin-producing Escherichia coli (STEC) strains are clinically important foodborne pathogens. Unlike E. coli O157:H7, these foodborne pathogens have no unique biochemical characteristics to readily distinguish them from other E. coli strains growing on plating media. In this study, a chromogenic agar medium was developed in order to differentiate among non-O157 STEC strains of serogroups O26, O45, O103, O111, O121, and O145 on a single agar medium. The ability of this chromogenic agar medium to select and distinguish among these pathogens is based on a combination of utilization of carbohydrates, b -galactosidase activity, and resistance to selective agents. The agar medium in combination with immunomagnetic separation was evaluated and successfully allowed for the detection and isolation of these six serogroups from artificially contaminated fresh beef. The agar medium in combination with immunomagnetic separation also allowed successful detection and isolation of naturally occurring non-O157 STEC strains present in cattle feces. Thirty-five strains of the top six non-O157 STEC serogroups were isolated from 1,897 fecal samples collected from 271 feedlot cattle. This chromogenic agar medium could help significantly in routine screening for the top six non-O157 STEC serogroups from beef cattle and other food.

  2. Equivalent intraperitoneal doses of ibuprofen supplemented in drinking water or in diet: a behavioral and biochemical assay using antinociceptive and thromboxane inhibitory dose-response curves in mice. (United States)

    Salama, Raghda A M; El Gayar, Nesreen H; Georgy, Sonia S; Hamza, May


    Background. Ibuprofen is used chronically in different animal models of inflammation by administration in drinking water or in diet due to its short half-life. Though this practice has been used for years, ibuprofen doses were never assayed against parenteral dose-response curves. This study aims at identifying the equivalent intraperitoneal (i.p.) doses of ibuprofen, when it is administered in drinking water or in diet. Methods. Bioassays were performed using formalin test and incisional pain model for antinociceptive efficacy and serum TXB2 for eicosanoid inhibitory activity. The dose-response curve of i.p. administered ibuprofen was constructed for each test using 50, 75, 100 and 200 mg/kg body weight (b.w.). The dose-response curves were constructed of phase 2a of the formalin test (the most sensitive phase to COX inhibitory agents), the area under the 'change in mechanical threshold'-time curve in the incisional pain model and serum TXB2 levels. The assayed ibuprofen concentrations administered in drinking water were 0.2, 0.35, 0.6 mg/ml and those administered in diet were 82, 263, 375 mg/kg diet. Results. The 3 concentrations applied in drinking water lay between 73.6 and 85.5 mg/kg b.w., i.p., in case of the formalin test; between 58.9 and 77.8 mg/kg b.w., i.p., in case of the incisional pain model; and between 71.8 and 125.8 mg/kg b.w., i.p., in case of serum TXB2 levels. The 3 concentrations administered in diet lay between 67.6 and 83.8 mg/kg b.w., i.p., in case of the formalin test; between 52.7 and 68.6 mg/kg b.w., i.p., in case of the incisional pain model; and between 63.6 and 92.5 mg/kg b.w., i.p., in case of serum TXB2 levels. Discussion. The increment in pharmacological effects of different doses of continuously administered ibuprofen in drinking water or diet do not parallel those of i.p. administered ibuprofen. It is therefore difficult to assume the equivalent parenteral daily doses based on mathematical calculations.

  3. Equivalent intraperitoneal doses of ibuprofen supplemented in drinking water or in diet: a behavioral and biochemical assay using antinociceptive and thromboxane inhibitory dose–response curves in mice

    Directory of Open Access Journals (Sweden)

    Raghda A.M. Salama


    Full Text Available Background. Ibuprofen is used chronically in different animal models of inflammation by administration in drinking water or in diet due to its short half-life. Though this practice has been used for years, ibuprofen doses were never assayed against parenteral dose–response curves. This study aims at identifying the equivalent intraperitoneal (i.p. doses of ibuprofen, when it is administered in drinking water or in diet. Methods. Bioassays were performed using formalin test and incisional pain model for antinociceptive efficacy and serum TXB2 for eicosanoid inhibitory activity. The dose–response curve of i.p. administered ibuprofen was constructed for each test using 50, 75, 100 and 200 mg/kg body weight (b.w.. The dose–response curves were constructed of phase 2a of the formalin test (the most sensitive phase to COX inhibitory agents, the area under the ‘change in mechanical threshold’-time curve in the incisional pain model and serum TXB2 levels. The assayed ibuprofen concentrations administered in drinking water were 0.2, 0.35, 0.6 mg/ml and those administered in diet were 82, 263, 375 mg/kg diet. Results. The 3 concentrations applied in drinking water lay between 73.6 and 85.5 mg/kg b.w., i.p., in case of the formalin test; between 58.9 and 77.8 mg/kg b.w., i.p., in case of the incisional pain model; and between 71.8 and 125.8 mg/kg b.w., i.p., in case of serum TXB2 levels. The 3 concentrations administered in diet lay between 67.6 and 83.8 mg/kg b.w., i.p., in case of the formalin test; between 52.7 and 68.6 mg/kg b.w., i.p., in case of the incisional pain model; and between 63.6 and 92.5 mg/kg b.w., i.p., in case of serum TXB2 levels. Discussion. The increment in pharmacological effects of different doses of continuously administered ibuprofen in drinking water or diet do not parallel those of i.p. administered ibuprofen. It is therefore difficult to assume the equivalent parenteral daily doses based on mathematical calculations.

  4. Novel Chromogenic Chemosensors for Fluoride Anion Based on 8-Hydroxyquinoline Azo Derivatives

    Institute of Scientific and Technical Information of China (English)

    CHENG,Yun-Fei; LIU,Zhi-Qiang; SHI,Mei; ZHAO,Qiang; LI,Fu-You; YI,Tao; HUANG,Chun-Hui


    A series of 8-hydroxyquinoline azo derivatives with diverse conjugated structures were synthesized and studied to chromogenically detect anions. All the dyes allowed selective detection for fluoride anion in CH3CN via instant deprotonation of the compounds, which was affirmed by UV-Vis absorption and 1H NMR spectra. The chromogenically responding ability increases as the substituent changes from phenyl to naphthyl or anthryl. This result is likely to be related to the enhancement of intramolecular charge transfer (ICT) induced by extension of conjugated structure.

  5. Determination of renal clearances of amylase/creatinine with chromogenic and enzymatic methods. (United States)

    Hohenwallner, W; Wimmer, E; Sommer, R


    Urinary amylase was estimated by chromogenic (amylochrome Roche) as well as enzymatic methods (SKI and Beckman: substrate starch and substrate maltotetraose respectively). Random and timed urines (24 hour collections) were analysed. Clearances of amylase gave different results dependent upon the amylase-test used and the glomerular filtration rate. Correlation between chromogenic and enzymatic methods (starch as substrate) was poor. The ratio of amylase and creatinine clearance was used to test different methods. Reference values for this ratio for the amylochrome method (N = 106) were 2.85 +/- 0.99% and for the Beckman-DS method (N = 60) 2.82 +/- 0.87%.

  6. Chromogenic detection of Sarin by discolouring decomplexation of a metal coordination complex. (United States)

    Ordronneau, Lucie; Carella, Alexandre; Pohanka, Miroslav; Simonato, Jean-Pierre


    An innovative chromogenic sensing concept based on decomplexation of a tris-(bipyridine)iron(II) coordination complex has been developed for the detection of organophosphorus nerve agents. It was evaluated both on a simulant and real Sarin in vapour and liquid phases.

  7. Establishment of a serum iron assay system on Dimension RxL Max automatic biochemical analyzer%Dimension RxL MAX全自动生化分析仪血清铁测定系统的建立

    Institute of Scientific and Technical Information of China (English)

    陈敏; 易浔飞; 廖剑


    Objective; To evaluate methodologically the serum iron assay system establised on Demension RxL Max automatic biochemical analyzer. Methods: The blank reagent cartridge was filled with commercial reagents and the assay parameters were programmed on the open channel. The precision and linearity of the method were evaluated using the guidelines of NCCLS EP5A and EP6A, respectively. The test results of this method were compared with those of commercial method using serum samples of patients. Results; The linear range is 2-1000 μg/dL, the total CVs for high value and low value samples are 5. 55% and 7. 49%, respectively. The average recovery is 104% , and the regression equation is,y = 0. 9691x-3. 6437 (r = 0. 9890) . Conclusion: The method is sensitive, accurate, and feasible for serum ion level monitoring.%在Dimension RxL Max全自动生化分析仪的开放通道上用空白试剂盒装载商品化亚铁嗪检测试剂,建立了血清铁的测定体系,并对所建方法进行了方法学评价.结果表明, 该方法的高、低值样本总变异系数分别为5.55%、7.49%,测定线性范围为2~1000μg/dL,平均回收率为104%.实验证明,在该仪器开放通道中运用亚铁嗪法进行血清铁测定是可行的.

  8. Anionic chromogenic chemosensors highly selective for fluoride or cyanide based on 4-(4-Nitrobenzylideneamine)phenol

    Energy Technology Data Exchange (ETDEWEB)

    Nicoleti, Celso R.; Marini, Vanderleia G.; Zimmermann, Lizandra M.; Machado, Vanderlei G., E-mail: [Departamento de Quimica, Universidade Federal de Santa Catarina (UFSC), Florianopolis, SC (Brazil)


    4-(4-Nitrobenzylideneamine)phenol was used in two strategies allowing the highly selective detection of F{sup -} and CN{sup -}. Firstly, the compound in acetonitrile acts as a chromogenic chemosensor based on the idea that more basic anions cause its deprotonation (colorless solution), generating a colored solution containing phenolate. The discrimination of CN{sup -} over F{sup -} was obtained by adding 1.4% water to acetonitrile: water preferentially solvates F{sup -}, leaving the CN{sup -} free to deprotonate the compound. Another strategy involved an assay comprised of the competition between phenolate dye and the analyte for calyx[4]pyrrole in acetonitrile, a receptor highly selective for F{sup -}. Phenolate and calyx[4]pyrrole form a hydrogen-bonded complex, which changes the color of the medium. On the addition of various anions, only F{sup -} was able to restore the original color corresponding to phenolate in solution due to the fact that the anion dislodges phenolate from the complexation site. (author)

  9. Comparison of three chromogenic media and evaluation of two molecular-based identification systems for the detection of Enterobacter sakazakii from environmental samples from infant formulae factories. (United States)

    Derzelle, Sylviane; Dilasser, Françoise; Maladen, Véronique; Soudrie, Nicole; Leclercq, Alexandre; Lombard, Bertrand; Lafarge, Veŕonique


    Enterobacter sakazakii is an occasional contaminant of powdered infant formula that can cause rare but severe foodborne infections in infants. To determine optimal methods for the detection and identification of E. sakazakii, 38 naturally contaminated samples from infant formulae factories were analyzed by two PCR-based methods and by a method (TS 22964/RM 210) developed by the International Organization for Standardization and the International Dairy Federation (ISO-IDF) using three different commercial chromogenic agars. The ISO-IDF method includes two enrichment steps, plating of the second enrichment broth on E. sakazakii isolation agar (a chromogenic selective agar), picking of five typical colonies for transfer onto tryptone soy agar, and subsequent confirmation of yellow-pigmented colonies by biochemical characterization. Twenty-two of the 38 samples were positive by the culture method. E. sakazakii isolation agar (ESIA; AES Laboratoires), COMPASS agar (Biokar Diagnostics), and Druggan-Forsythe-Iversen agar (Oxoid) compared favorably with violet red bile glucose agar (VRBG, a selective medium for Enterobacteriaceae), with positive predictive values of 86.96, 88, and 74.07%, respectively, in contrast to 47.83% for VRBG. One additional positive sample was detected using the nonpatented real-time PCR method evaluated, and those results were in 97.3% concordance with the ISO-IDF results. Some discrepancies between the results of the DuPont Qualicon BAX system and those of the ISO-IDF method could be explained by heterogeneity of contamination and sampling. Thus, both PCR-based systems were suitable for detecting and specifically identifying E. sakazakii within 1 to 2 days, and COMPASS agar and ESIA could be used interchangeably as a first-step medium to isolate presumptive E. sakazakii colonies.

  10. Synthesis of Chiral Phenolic 1,1?-Binaphthocrown Ethers and Some Proton-Ionisable Chromogenic Derivatives


    Köszegi, Eva; Grün, Alajos; Bitter, Istvan


    Abstract Synthesis of mono-and bis(1,1?-bi-2-naphthocrown)ethers containing bis(2,6-methylene)anisyl subunit in the crown ring were developed. These chiral macrocycles are suitable precursors to introduce chromogenic function, as exemplified by two novel crowned azophenol chromoionophores. Their coloration process induced by various achiral and chiral amines was studied by UV-vis spectrophotometry. (Koszegi, Eva) (Grun, Alajos) ibit...

  11. Chromogenic and fluorogenic detection and discrimination of nerve agents Tabun and Vx. (United States)

    Kumar, Vinod; Rana, Hemlata


    Our approach uses squaraine (SQ) as the molecular-receptor as well as an indicator for the chromogenic and fluorogenic detection and discrimination of nerve agents Tabun and Vx. To mimic a real-life scenario, the protocols were implemented in spiked water and soil samples, on surfaces, and in the gas phase. The lower detection limit will be useful to protect human health and national security.

  12. Automatic Digital Analysis of Chromogenic Media for Vancomycin-Resistant-Enterococcus Screens Using Copan WASPLab. (United States)

    Faron, Matthew L; Buchan, Blake W; Coon, Christopher; Liebregts, Theo; van Bree, Anita; Jansz, Arjan R; Soucy, Genevieve; Korver, John; Ledeboer, Nathan A


    Vancomycin-resistant enterococci (VRE) are an important cause of health care-acquired infections (HAIs). Studies have shown that active surveillance of high-risk patients for VRE colonization can aid in reducing HAIs; however, these screens generate a significant cost to the laboratory and health care system. Digital imaging capable of differentiating negative and "nonnegative" chromogenic agar can reduce the labor cost of these screens and potentially improve patient care. In this study, we evaluated the performance of the WASPLab Chromogenic Detection Module (CDM) (Copan, Brescia, Italy) software to analyze VRE chromogenic agar and compared the results to technologist plate reading. Specimens collected at 3 laboratories were cultured using the WASPLab CDM and plated to each site's standard-of-care chromogenic media, which included Colorex VRE (BioMed Diagnostics, White City, OR) or Oxoid VRE (Oxoid, Basingstoke, United Kingdom). Digital images were scored using the CDM software after 24 or 40 h of growth, and all manual reading was performed using digital images on a high-definition (HD) monitor. In total, 104,730 specimens were enrolled and automation agreed with manual analysis for 90.1% of all specimens tested, with sensitivity and specificity of 100% and 89.5%, respectively. Automation results were discordant for 10,348 specimens, and all discordant images were reviewed by a laboratory supervisor or director. After a second review, 499 specimens were identified as representing missed positive cultures falsely called negative by the technologist, 1,616 were identified as containing borderline color results (negative result but with no package insert color visible), and 8,234 specimens were identified as containing colorimetric pigmentation due to residual matrix from the specimen or yeast (Candida). Overall, the CDM was accurate at identifying negative VRE plates, which comprised 84% (87,973) of the specimens in this study.

  13. 酮糖的显色试验研究%Research on Chromogenic Test of Ketose

    Institute of Scientific and Technical Information of China (English)

    曹会兰; 张秀芹


    采用盐酸—间苯二酚溶液对酮糖的显色反应,探讨了显色反应机理与糖的结构关系,将正交试验法应用于显色条件的选择。通过将果糖、葡萄糖、蔗糖、乳糖与盐酸—间苯二酚试剂共热煮沸,在不同浓度、不同时间下观察实验现象,得出酮糖在浓度为1%左右、时间为2~5 min之间,反应最明显,而醛糖最不明显。可以用该试剂迅速地把酮糖和醛糖区别开来,测定结果令人满意。%Relationship between the mechanism of chromogenic reaction and the structure of carbohydrate was discussed and the orthogonal testes were applied to choose the chromogenic condition through the chromogenic reaction of hydrochloric acid -resor-cinol solution and ketoses .The ketoses and aldoses were quickly distinguished by observing the co -heating experiment of different carbohydrates at different concentration and time .The result was that the reaction phenomenon of the ketoses ( about 1%, t=2~5 min) were obvious , but aldoses were not .

  14. Detection of Extracellular Enzyme Activity in Penicillium using Chromogenic Media. (United States)

    Yoon, Ji Hwan; Hong, Seung Beom; Ko, Seung Ju; Kim, Seong Hwan


    A total of 106 Penicillium species were tested to examine their ability of degrading cellobiose, pectin and xylan. The activity of β-glucosidase was generally strong in all the Penicillium species tested. P. citrinum, P. charlesii, P. manginii and P. aurantiacum showed the higher ability of producing β-glucosidase than other tested species. Pectinase activity was detected in 24 Penicillium species. P. paracanescens, P. sizovae, P. sartoryi, P. chrysogenum, and P. claviforme showed strong pectinase activity. In xylanase assay, 84 Penicillium species showed activity. Strong xylanase activity was detected from P. megasporum, P. sartoryi, P. chrysogenum, P. glandicola, P. discolor, and P. coprophilum. Overall, most of the Penicillium species tested showed strong β-glucosidase activity. The degree of pectinase and xylanase activity varied depending on Penicillium species.

  15. Novel method based on chromogenic media for discrimination and selective enumeration of lactic acid bacteria in fermented milk products. (United States)

    Galat, Anna; Dufresne, Jérôme; Combrisson, Jérôme; Thépaut, Jérôme; Boumghar-Bourtchai, Leyla; Boyer, Mickaël; Fourmestraux, Candice


    Microbial analyses of fermented milk products require selective methods to discriminate between close species simultaneously present in high amounts. A culture-based method combining novel chromogenic agar media and appropriate incubation conditions was developed to enumerate lactic acid bacteria (LAB) strains in fermented milk. M1 agar, containing two chromogenic substrates, allowed selective enumeration of Lactobacillus rhamnosus, two strains of Lactobacillus paracasei subsp. paracasei and Streptococcus salivarius subsp. thermophilus based on differential β-galactosidase and β-glucosidase activities. Depending on the presence of some or all of the above strains, M1 agar was supplemented with L-rhamnose or vancomycin and incubations were carried out at 37 °C or 44 °C to increase selectivity. A second agar medium, M2, containing one chromogenic substrates was used to selectively enumerate β-galactosidase producing Lactobacillus delbrueckii subsp. bulgaricus at 47 °C. By contrast with the usual culture media, the chromogenic method allowed unambiguous enumeration of each species, including discrimination between the two L. paracasei, up to 10(9) CFU/g of fermented milk. In addition, the relevance of the method was approved by enumerating reference ATCC strains in pure cultures and fermented milk product. The method could also be used for enumerations on non-Danone commercial fermented milk products containing strains different from those used in this study, showing versatility of the method. To our knowledge, this is the first description of a chromogenic culture method applied to selective enumeration of LAB.

  16. Evolution and characterization of a new reversibly photoswitching chromogenic protein, Dathail. (United States)

    Langan, Patricia S; Close, Devin W; Coates, Leighton; Rocha, Reginaldo C; Ghosh, Koushik; Kiss, Csaba; Waldo, Geoff; Freyer, James; Kovalevsky, Andrey; Bradbury, Andrew R M


    We report the engineering of a new reversibly switching chromogenic protein, Dathail. Dathail was evolved from the extremely thermostable fluorescent proteins thermal green protein (TGP) and eCGP123 using directed evolution and ratiometric sorting. Dathail has two spectrally distinct chromogenic states with low quantum yields, corresponding to absorbance in a ground state with a maximum at 389nm, and a photo-induced metastable state with a maximum at 497nm. In contrast to all previously described photoswitchable proteins, both spectral states of Dathail are non-fluorescent. The photo-induced chromogenic state of Dathail has a lifetime of ~50min at 293K and pH7.5 as measured by UV-Vis spectrophotometry, returning to the ground state through thermal relaxation. X-ray crystallography provided structural insights supporting a change in conformation and coordination in the chromophore pocket as being responsible for Dathail's photoswitching. Neutron crystallography, carried out for the first time on a protein from the green fluorescent protein family, showed a distribution of hydrogen atoms revealing protonation of the chromophore 4-hydroxybenzyl group in the ground state. The neutron structure also supports the hypothesis that the photo-induced proton transfer from the chromophore occurs through water-mediated proton relay into the bulk solvent. Beyond its spectroscopic curiosity, Dathail has several characteristics that are improvements for applications, including low background fluorescence, large spectral separation, rapid switching time, and the ability to switch many times. Therefore, Dathail is likely to be extremely useful in the quickly developing fields of imaging and biosensors, including photochromic Förster resonance energy transfer, high-resolution microscopy, and live tracking within the cell.

  17. Evaluation of biochemical and serological methods to identify and clustering yeast cells of oral Candida species by CHROMagar test, SDS-PAGE and ELISA

    Directory of Open Access Journals (Sweden)

    J. A. de O. Rodrigues

    Full Text Available The purpose of this work was to evaluate biochemical and serological methods to characterize and identify Candida species from the oral cavity. The strains used were five Candida species previously identified: C. albicans, C. guilliermondii, C. parapsilosis, C. krusei, C. tropicalis, and Kluyveromyces marxianus, as a negative control. The analyses were conducted through the SDS-PAGE associated with statistical analysis using software, chromogenic medium, and CHROMagar Candida (CA, as a differential medium for the isolation and presumptive identification of clinically important yeasts and an enzyme-linked immunoabsorbent assay (ELISA, using antisera produced against antigens from two C. albicans strains. This method enabled the screening of the three Candida species: C. albicans, C. tropicalis, and C. Krusei, with 100% of specificity. The ELISA using purified immunoglobulin G showed a high level of cross-reaction against protein extracts of Candida species. The SDS-PAGE method allowed the clustering of species-specific isolates using the Simple Matching coefficient, S SM = 1.0. The protein profile analysis by SDS-PAGE increases what is known about the taxonomic relationships among oral yeasts. This methodology showed good reproducibility and allows collection of useful information for numerical analysis on information relevant to clinical application, and epidemiological and systematical studies.

  18. Synthesis and characterization of chromogenic fluoran compounds containing 4-ketoquinazolinone moieties

    Directory of Open Access Journals (Sweden)



    Full Text Available Chromogenic fluoran compounds containing 4-ketoquinazolinone were synthesized by reacting 2-(4-diethylamino-2-hydroxybenzoylbenzoic acid with various substituted 4-ketoquinazolinones in the presence of sulfuric acid. The 4-ketoquinazolinones were obtained by reacting various substituted benzoxazin-4-ones with 4-aminophenol or 2-nitro-p-anisidine. All the synthesized derivatives were identified by conventional methods, such as melting points, elemental analysis, IR, 1H-NMR, and UV-visible spectroscopy in organic solvent and 95 % acetic acid. All the fluoran compounds develop colour on contact with acidic or electron-accepting compounds.

  19. Using selective chromogenic plates to optimize isolation of group B Streptococcus in pregnant women

    Directory of Open Access Journals (Sweden)

    Romano Mattei


    Full Text Available Group B Streptococcus (GBS remains the leading cause of severe bacterial infections (sepsis, meningitis, pneumonia in neonates. We compared the detection of GBS from recto-vaginal swabs on blood agar and two chromogenic media and evaluated their antibiotic susceptibility. A total of 1351 swabs were taken from pregnant women at 35-37 weeks of gestation. Following enrichment in Todd Hewitt broth + nalidixic acid and colistin, the samples were plated on Columbia CNA agar (CNA, chromID Strepto B agar (STRB and Granada Agar (GRAN, respectively. GBS were found in 22.4% of recto-vaginal swabs from pregnant women. Sensitivity, specificity, positive and negative predictive values of GBS detection were 88%, 88%, 81% and 96% for CNA, 99%, 97%, 90% and 99% for STRB and 94%, 99%, 98% e 99% for GRAN; Cohen’s k index concordances for CNA, STREB and GRAN were 0.68, 0.92 and 0.96, respectively. All isolates were susceptible to penicillin, whereas resistances of erythromycin and clindamycin were 40% and 42%, respectively. To conclude, selective broth enrichment combined with chromogenic plates is recommended for GBS screening in pregnant women.

  20. A selective chromogenic agar that distinguishes Bacillus anthracis from Bacillus cereus and Bacillus thuringiensis. (United States)

    Juergensmeyer, Margaret A; Gingras, Bruce A; Restaino, Lawrence; Frampton, Elon W


    A selective and differential plating medium, R & F anthracis chromogenic agar (ACA), has been developed for isolating and identifying presumptive colonies of Bacillus anthracis. ACA contains the chromogenic substrate 5-bromo-4-chloro-3-indoxyl-choline phosphate that upon hydrolysis yields teal (blue green) colonies indicating the presence of phosphatidylcholine-specific phospholipase C (PC-PLC) activity. Among seven Bacillus species tested on ACA, only members of the Bacillus cereus group (B. anthracis, B. cereus, and B. thuringiensis) produced teal colonies (PC-PLC positive) having cream rings. Examination of colony morphology in 18 pure culture strains of B. anthracis (15 ATCC strains plus AMES-1-RIID, ANR-1, and AMED-RIID), with one exception, required 48 h at 35 to 37 degrees C for significant color production, whereas only 24 h was required for B. cereus and B. thuringiensis. This differential rate of PC-PLC synthesis in B. anthracis (due to the truncated plcR gene and PlcR regulator in B. anthracis) allowed for the rapid differentiation on ACA of presumptive colonies of B. anthracis from B. cereus and B. thuringiensis in both pure and mixed cultures. Effective recovery of B. anthracis from a variety of matrices having both high (soil and sewage) and low microbial backgrounds (cloth, paper, and blood) spiked with B. anthracis ANR-1 spores suggests the probable utility of ACA plating for B. anthracis recovery in a diversity of applications.

  1. A multi writable thiophene-based selective and reversible chromogenic fluoride probe with dual -NH functionality (United States)

    Vishwakarma, Siddharth; Kumar, Ajit; Pandey, Abha; Upadhyay, K. K.


    A chromogenic fluoride probe bearing bis imine groups having dual -NH functionality (BSB) has been designed, synthesised and structurally characterized by its single crystal X-ray diffraction studies. The BSB could visually and spectroscopically recognise F- with high selectivity over other anions by exhibiting intense chromogenic response (from colourless to red) for F- in acetonitrile solution. The UV-visible titration and 1H NMR titration experiments indicated that the observed changes occur via a combined process including hydrogen bonding and deprotonation between the BSB and F-. Moreover theoretical calculations at the Density Functional Theory (DFT) level shed further light upon probe design strategy and the nature of interactions between BSB and F-. The limit of detection and binding constant of BSB towards F- were found to be 6.9 × 10- 7 M and 1.42 ± 0.069 × 108 M- 2 respectively. Finally, by using F- and H+ as chemical inputs and the absorbance as output, a INHIBIT logic gate was constructed, which exhibits "Multi-write" ability without obvious degradation in its optical output.

  2. Performance of chromogenic media for Candida in rapid presumptive identification of Candida species from clinical materials

    Directory of Open Access Journals (Sweden)

    M V Pravin Charles


    Full Text Available Background: In perspective of the worldwide increase in a number of immunocompromised patients, the need for identification of Candida species has become a major concern. The development of chromogenic differential media, introduced recently, facilitate rapid speciation. However, it can be employed for routine mycology workup only after an exhaustive evaluation of its benefit and cost effectiveness. This study was undertaken to evaluate the benefit and cost effectiveness of chromogenic media for speciation of Candida clinical isolates. Materials and Methods: Sputum samples of 382 patients were screened for the presence of Candida spp. by Gram stain and culture on sabouraud dextrose agar. Candida species were identified using Gram stain morphology, germ tube formation, cornmeal agar with Tween-80, sugar fermentation tests and morphology on HiCrome Candida differential agar. All the Candida isolates were inoculated on HiCrome Candida agar (HiMedia, Mumbai, India. Results: The sensitivity and specificity of HiCrome agar for identification of Candida albicans were 90% and 96.42%, respectively whereas sensitivity and specificity of carbohydrate fermentation test were 86.67% and 74.07%, respectively. Sensitivity and specificity values of HiCrome agar for detection of C. albicans, Candida parapsilosis and Candida glabrata were above 90%. Conclusions: We found HiCrome agar has high sensitivity and specificity comparable to that of the conventional method. In addition, use of this differential media could significantly cut down the turnaround time as well as cost of sample processing.

  3. Spectral Imaging of Multi-Color Chromogenic Dyes in Pathological Specimens

    Directory of Open Access Journals (Sweden)

    Merryn V. E. Macville


    Full Text Available We have investigated the use of spectral imaging for multi‐color analysis of permanent cytochemical dyes and enzyme precipitates on cytopathological specimens. Spectral imaging is based on Fourier‐transform spectroscopy and digital imaging. A pixel‐by‐pixel spectrum‐based color classification is presented of single‐, double‐, and triple‐color in situ hybridization for centromeric probes in T24 bladder cancer cells, and immunocytochemical staining of nuclear antigens Ki‐67 and TP53 in paraffin‐embedded cervical brush material (AgarCyto. The results demonstrate that spectral imaging unambiguously identifies three chromogenic dyes in a single bright‐field microscopic specimen. Serial microscopic fields from the same specimen can be analyzed using a spectral reference library. We conclude that spectral imaging of multi‐color chromogenic dyes is a reliable and robust method for pixel color recognition and classification. Our data further indicate that the use of spectral imaging (a may increase the number of parameters studied simultaneously in pathological diagnosis, (b may provide quantitative data (such as positive labeling indices more accurately, and (c may solve segmentation problems currently faced in automated screening of cell‐ and tissue specimens. Figures on‐3/macville.htm.

  4. Assay discrepancy in mild haemophilia A: entire population study in a National Haemophilia Centre. (United States)

    Poulsen, A L; Pedersen, L H; Hvas, A-M; Poulsen, L H; Thykjaer, H; Ingerslev, J


    Assay discrepancy in mild haemophilia, here defined by a significantly higher factor VIII (FVIII):C response by the one-stage procoagulant assay as compared with a two-stage enzymatic method, has repeatedly been reported in literature. The purpose of this study was to determine the overall prevalence of this phenomenon amongst mild haemophilia families from a population of 2.95 million inhabitants in the Western Danish region. Information was collected retrospectively through a thorough search of archives of the National Haemophilia Centre in Aarhus. We identified 109 patients with mild haemophilia A amongst whom 92 were eligible to enter the study. These represent a total of 53 unrelated families. Our data illustrate that this assay discrepancy pattern is found quite frequently amongst our mild haemophilia A families. While the ratio of FVIII:C chromogenic/FVIII:C clot values was quite consistent amongst patients belonging to same family pattern, ratios in the entire cohort of families ranged from 0.18 to 1.00. Selecting a cut-off level for the FVIII:C chromogenic/FVIII:C clot ratios at 0.7, 0.6 and 0.5, respectively, we found that 38 (72%), 27 (51%) and 19 (36%) of families, respectively, displayed this assay discrepancy. In 10 patients, the FVIII:C chromogenic level was inside the category of moderate haemophilia at >0.01-area.

  5. Influence of amylase assay technique on renal clearance of amylase-creatinine ratio. (United States)

    Levitt, M D; Johnson, S G; Ellis, C J; Engel, R R


    The influence of amylase assay technique on the renal amylase/creatinine clearance measurement was determined by analysis of serum and urine specimens obtained from 10 normal subjects. CAm/CCr averaged 2.19 +/- 0.18% with a saccharogenic technique, 1.52 +/- 0.2% with an iodometric technique, and 0.80 +/- 0.08% with a chromogenic technique. Each of these values differed significantly (P less than 0.05) from the other two. Recovery studies were carried out by adding partially purified human salivary or pancreatic amylase to human newborn serum or urine (which contain minimal endogenous amylase). Equal amylase activity was recovered from serum and urine by the saccharogenic technique whereas recovery from urine was less than 50% of that from serum using the iodometric and chromogenic techniques. The accuracy of the chromogenic technique is markedly improved by the addition of albumin to the urine assay system. Although it appears that only the saccharogenic method provides an accurate estimate of CAm/CCr, each assay technique distinguished the elevated CAm/CCr of patients with pancreatitis from the normal range established for that technique. Accurate clinical interpretation of CAm/CCr measurment requires knowledge of the amylase assay technique used.

  6. MMP-9 and MMP-2 activities in stomach and breast tumours, as measured by a novel MMP activity assay using modified urokinase as a substrate

    NARCIS (Netherlands)

    Hanemaaijer, R.; Visser, H.; Duffy, J.; Verspaget, H.W.; Verheijen, J.H.; Maguire, T.


    Matrix metalloproteinases (MMPs) play an important role in many pathological processes. However, MMP activities are difficult to determine since no simple specific and/or chromogenic substrates exist. Therefore, we have developed a novel MMP activity assay using a modified urokinase as a substrate.

  7. Biochemical filter with sigmoidal response: increasing the complexity of biomolecular logic. (United States)

    Privman, Vladimir; Halámek, Jan; Arugula, Mary A; Melnikov, Dmitriy; Bocharova, Vera; Katz, Evgeny


    The first realization of a designed, rather than natural, biochemical filter process is reported and analyzed as a promising network component for increasing the complexity of biomolecular logic systems. Key challenge in biochemical logic research has been achieving scalability for complex network designs. Various logic gates have been realized, but a "toolbox" of analog elements for interconnectivity and signal processing has remained elusive. Filters are important as network elements that allow control of noise in signal transmission and conversion. We report a versatile biochemical filtering mechanism designed to have sigmoidal response in combination with signal-conversion process. Horseradish peroxidase-catalyzed oxidation of chromogenic electron donor by H(2)O(2) was altered by adding ascorbate, allowing to selectively suppress the output signal, modifying the response from convex to sigmoidal. A kinetic model was developed for evaluation of the quality of filtering. The results offer improved capabilities for design of scalable biomolecular information processing systems.

  8. Biochemical Filter with Sigmoidal Response: Increasing the Complexity of Biomolecular Logic

    CERN Document Server

    Privman, Vladimir; Arugula, Mary A; Melnikov, Dmitriy; Bocharova, Vera; Katz, Evgeny


    The first realization of a designed, rather than natural, biochemical filter process is reported and analyzed as a promising network component for increasing the complexity of biomolecular logic systems. Key challenge in biochemical logic research has been achieving scalability for complex network designs. Various logic gates have been realized, but a "toolbox" of analog elements for interconnectivity and signal processing has remained elusive. Filters are important as network elements that allow control of noise in signal transmission and conversion. We report a versatile biochemical filtering mechanism designed to have sigmoidal response in combination with signal-conversion process. Horseradish peroxidase-catalyzed oxidation of chromogenic electron donor by hydrogen peroxide, was altered by adding ascorbate, allowing to selectively suppress the output signal, modifying the response from convex to sigmoidal. A kinetic model was developed for evaluation of the quality of filtering. The results offer improved...

  9. Standardization of reagents and methods used in cytological and histological practice with emphasis on dyes, stains and chromogenic reagents

    DEFF Research Database (Denmark)

    Lyon, H O; De Leenheer, A P; Horobin, R W;


    The need for the standardization of reagents and methods used in the histology laboratory is demonstrated. After definitions of dyes, stains, and chromogenic reagents, existing standards and standards organizations are discussed. This is followed by practical instructions on how to standardize dy...

  10. Prospective Two-Center Comparison of Three Chromogenic Agars for Methicillin-Resistant Staphylococcus aureus Screening in Hospitalized Patients

    NARCIS (Netherlands)

    Dodémont, Magali; Verhulst, Carlo; Nonhoff, Claire; Nagant, Carole; Denis, Olivier; Kluytmans, Jan


    Three chromogenic media, chromID MRSA SMART (SMART), chromID MRSA first generation (chromID), and Brilliance MRSA (OX2), were evaluated for methicillin-resistant Staphylococcus aureus (MRSA) screening using 1,220 samples. The sensitivity at 24 h was significantly better with the SMART agar (66.4%) t

  11. The correlation between dual-color chromogenic in situ hybridization and fluorescence in situ hybridization in assessing HER2 gene amplification in breast cancer

    DEFF Research Database (Denmark)

    Pedersen, Marianne; Rasmussen, Birgitte Bruun


    and the generated chromogenic signals are also stable. This study presents a dual color CISH for simultaneous detection of the HER2 gene and chromosome 17. The CISH method performs a chromogenic detection "on top" of the Food and Drug Administration (FDA)-approved HER2 FISH pharmDx method, where the fluorochrome......-labeled probes are detected using enzyme-labeled antibodies and visualized by chromogenic enzymatic reactions. The HER2 status (amplified/not amplified and HER2 ratios) was evaluated by the CISH method and compared with results obtained by the FDA-approved FISH method. Of the 72 successfully investigated...

  12. [Contribution of the chromogenic medium CHROMagar(®)Candida in mycological diagnosis of yeasts]. (United States)

    Ouanes, A; Kouais, A; Marouen, S; Sahnoun, M; Jemli, B; Gargouri, S


    The incidence of invasive candidiasis has increased dramatically over the last decades due to a larger number of patients at risk. The diagnosis remains difficult as the clinical presentation is not specific and the biological diagnosis usually takes several days to become positive. We propose in this work through a prospective study to evaluate the contribution of a chromogenic medium CHROMagar(®) (Becton-Dickinson) in the mycological diagnosis of Candida. We selected 680 samples from patients hospitalized in the intensive care unit for epidemiological surveillance over a period of 11 weeks. We treated samples by culture on Sabouraud and on CHROMagar(®). The species identification was performed by chlamydosporulation test and carbohydrate assimilation tests. We found that the CHROMagar(®)Candida evaluated in our work was a valuable tool in the primary culture in differentiating the most frequently isolated yeast species and in better detection of mixed cultures.

  13. Naked-eye detection of biologically important anions by a new chromogenic azo-azomethine sensor. (United States)

    Rezaeian, Khatereh; Khanmohammadi, Hamid


    A new chromogenic azo-azomethine sensor, containing active phenolic sites, has been designed and synthesized via condensation reaction of N,N,N',N'-tetrakis(2-aminoethyl)-2,2-dimethyl propane-1,3-diamine with 1-(3-formyl-4-hydroxyphenylazo)-4-nitrobenzene. The anion recognition ability of the synthesized receptor was evaluated using UV-Vis spectroscopy and (1)H NMR technique. The anion recognition studies exhibited that the receptor acts as a sensor for biologically important anions such as F(-), AcO(-) and H2PO4(-) over other anions. The binding stoichiometry between sensor and anions was found to be 1:2. (1)H NMR experiment revealed that sensor recognizes anions via H-bonds and subsequent deprotonation to elicit a vivid color change. Interestingly, the sensory system not only let for the naked eye detection without any spectroscopic instrumentation but also helped to discriminate between anions.

  14. A novel method for the spectrophotometric determination of cefradine by using sodium nitroprusside as chromogenic reagent

    Institute of Scientific and Technical Information of China (English)


    A novel method is developed for the determination of cefradine by using sodium nitroprusside as chromogenic reagent. The experiment indicates that a russety product is formed by the reaction of cefradine with sodium nin'oprusside in basic solution, and the maximum absorption wavelength (λmax) of russety product is 505 nm. And the sensitization of tetradecyl benzyl dimethyl ammonium chloride for the reaction of cefradine with sodium nitroprusside is remarkable. The apparent molar absorption coefficient (ε505)is 2.81 x 103 L/mol cm. The linear equation is A = 0.0657 + 0.00804C (μg/mL) in the range of 1.50-55.0 μg/mL of cefradine with a correlation coefficient r = 0.9992, and the detection limit is 1.38 μg/mL. This method has been applied to determine cefradine in capsule and tablet samples.

  15. Application of high refractive index and/or chromogenic layers to control solar and thermal radiations (United States)

    Suzuki, Motofumi; Nishiura, Kensuke; Masunaka, Shoma; Muroi, Naoto; Namura, Kyoko


    In this presentation, we demonstrate that high refractive index materials such as β-FeSi2 and/or chromogenic materials such as VO2 are the key to control solar and thermal radiations. β-FeSi2 is known as an eco-friendly semiconductor and for sputtered polycrystalline β-FeSi2 thin films, we recently found that λ 0.3 in IR region, while n is higher than 5. On the other hand, another interesting optical property of β-FeSi2 is that both n and k are considerably high in visible to NIR region ( λ designed multilayers consisting of β-FeSi2/SiO2/β-FeSi2/W, where the upper β-FeSi2 layer absorbs VIS and NIR (λ <= 1.0 μm) and the bottom β-FeSi2 layer/W absorbs IR (1.0 <= λ <=2.0 μm). The optimized multilayers absorb more than 90% of solar energy and the eminence at 450 °C is lower than 10%. The perfect absorbers with high refractive index layers are useful for applications to solar selective absorbers for solar thermal power generation and spectrally selective thermal emitters for thermophotovoltaic power generation, IR heaters, radiation cooling. Replacing one of β-FeSi2 layers with a chromogenic material allows active control of solar and thermal radiation. In the presentation, we also demonstrate the active perfect absorbers including a VO2 layer in NIR region.

  16. A new generation of versatile chromogenic substrates for high-throughput analysis of biomass-degrading enzymes

    DEFF Research Database (Denmark)

    Kracun, Stjepan Kresimir; Schückel, Julia; Westereng, Bjørge;


    Background: Enzymes that degrade or modify polysaccharides are widespread in pro- and eukaryotes and have multiple biological roles and biotechnological applications. Recent advances in genome and secretome sequencing, together with associated bioinformatic tools, have enabled large numbers...... of carbohydrate-acting enzymes to be putatively identified. However, there is a paucity of methods for rapidly screening the biochemical activities of these enzymes, and this is a serious bottleneck in the development of enzyme-reliant bio-refining processes. Results: We have developed a new generation of multi...... carbohydrate-acting enzymes, and the assays have the potential to be incorporated into fully or semi-automated robotic enzyme screening systems...

  17. Magnetic bead-based enzyme-chromogenic substrate system for ultrasensitive colorimetric immunoassay accompanying cascade reaction for enzymatic formation of squaric acid-iron(III) chelate. (United States)

    Lai, Wenqiang; Tang, Dianping; Zhuang, Junyang; Chen, Guonan; Yang, Huanghao


    This work reports on a simple and feasible colorimetric immunoassay with signal amplification for sensitive determination of prostate-specific antigen (PSA, used as a model) at an ultralow concentration by using a new enzyme-chromogenic substrate system. We discovered that glucose oxidase (GOx), the enzyme broadly used in enzyme-linked immunosorbent assay (ELISA), has the ability to stimulate in situ formation of squaric acid (SQA)-iron(III) chelate. GOx-catalyzed oxidization of glucose leads to the formation of gluconic acid and hydrogen peroxide (H2O2). The latter can catalytically oxidize iron(II) to iron(III), which can rapidly (immunoassay protocol with GOx-labeled anti-PSA detection antibody can be designed for the detection of target PSA on capture antibody-functionalized magnetic immunosensing probe, monitored by recording the color or absorbance (λ = 468 nm) of the generated SQA-iron(III) chelate. The absorbance intensity shows to be dependent on the concentration of target PSA. A linear dependence between the absorbance and target PSA concentration is obtained under optimal conditions in the range from 1.0 pg mL(-1) to 30 ng mL(-1) with a detection limit (LOD) of 0.5 pg mL(-1) (0.5 ppt) estimated at the 3Sblank level. The sensitivity displays to be 3-5 orders of magnitude better than those of most commercialized human PSA ELISA kits. In addition, the developed colorimetric immunoassay was validated by assaying 12 human serum samples, receiving in good accordance with those obtained by the commercialized PSA ELISA kit. Importantly, the SQA-based immunosensing system can be further extended for the detection of other low-abundance proteins or biomarkers by controlling the target antibody.

  18. Biochemical Abnormalities in Batten's Syndrome

    DEFF Research Database (Denmark)

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


    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 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...... in whole blood. In normal human beings a connection was found between the erythrocyte selenium content and GSHPx activity assayed by cumene hydroperoxide as a peroxide donor....

  19. Biochemical markers of bone turnover

    Energy Technology Data Exchange (ETDEWEB)

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


    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.

  20. Chromogenic medium for direct susceptibility testing of Candida spp. isolated from urine. (United States)

    de Vasconcelos, Antônio Alexandre; Menezes, Everardo Albuquerque; Cunha, Francisco Afrânio


    Currently, there has been an increased frequency of fungal infections. Candida albicans and other Candida spp. have been proven to be major causes for urinary tract infection. Increased resistance to antifungals indicates the need to develop strategies in order to prevent the spread of resistance. Chromogenic medium have been proven to be useful in the detection of yeasts in clinical specimens containing mixed cultures of Candida. The aim of this study was to compare the results of antifungal susceptibility testing with fluconazole and amphotericin B on strains of Candida spp. isolated from urine, conducted on a Mueller-Hinton Agar with Glucose and Methylene Blue (MHAGMB) medium and on a Hicrome Candida® Agar with 2% Glucose (HCAG) medium. We used 40 samples of Candida spp. isolated from urine samples from inpatients and outpatients. The results showed that both media presented high rates of agreement, above 94%. The use of the HCAG medium decreases the release time of the results by 24-48 h, which may be decisive for initiating the correct drug treatment.

  1. Primary isolation of Candida species from urine specimens using chromogenic medium. (United States)

    Okulicz, J F; Rivard, R G; Conger, N G; Nguyen, M X; Hospenthal, D R


    CHROMagar Candida (CaC) is a chromogenic medium that can be used to detect Candida species, including Candida albicans, Candida krusei, Candida tropicalis, and perhaps Candida glabrata. We evaluated the utility of CaC to detect candiduria in high-risk patients and the potential usefulness of this information in directing initial antifungal therapy in those later identified with candidaemia. CaC was compared in parallel to standard laboratory methods (SM) for the detection of Candida from urine collected from high-risk units and wards. Of 893 samples, Candida was recovered by CaC from 104 compared with 35 using SM. No isolates detected by SM were undetected by CaC. More than one Candida species were recovered by CaC in 19 of the 104 (18.3%); only two mixed cultures were detected by SM. The identification was more rapid with CaC. Five of 69 patients with candiduria detected by CaC developed candidaemia on or after the date of urine culture. SM recovered fungus in only two of these patients. CaC can be used as primary media for the detection of Candida species from urine specimens. Primary isolation by CaC may enable clinicians to make earlier, directed selection of antifungal agents and potentially reduce patient morbidity and mortality.

  2. Non-Peptide-based Small-Molecule Probe for Fluorogenic and Chromogenic Detection of Chymotrypsin. (United States)

    Wu, Lei; Yang, Shu-Hou; Xiong, Hao; Yang, Jia-Qian; Guo, Jun; Yang, Wen-Chao; Yang, Guang-Fu


    We report herein a non-peptide-based small molecule probe for fluorogenic and chromogenic detection of chymotrypsin, and the primary application. This probe was rationally designed by mimicking the peptide substrate and optimized by adjusting the recognization group. The refined probe 2 exhibits good specificity toward chymotrypsin, producing about 25-fold higher enhancement in both the fluorescence intensity and absorbance upon the catalysis by chymotrypsin. Compared with the most widely used peptide substrate (AMC-FPAA-Suc) of chymotrypsin, probe 2 shows about 5-fold higher binding affinity, and comparable catalytical efficiency against chymotrypsin. Furthermore, it was successfully applied for the inhibitor characterization. To the best of our knowledge, probe 2 is the first non-peptide-based small-molecule probe for chymotrypsin, with the advantages of simple structure and high sensitivity compared to the widely used peptide-based substrates. This small-molecule probe is expected to be a useful molecular tool for drug discovery and chymotrypsin-related diseases diagnosis.

  3. Chromogene properties of the betalains before gamma photons; Propiedades cromogenas de las betalainas ante fotones gamma

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez N, S.; Quintero M, C. L. [Universidad Autonoma de Zacatecas, Unidad Academica de Ciencias Quimicas, Programa de Quimica en Alimentos, Km. 0.5 Carretera a Guadalajara Ejido La Escondida, Zacatecas (Mexico); Vega C, H. R., E-mail: [Universidad Autonoma de Zacatecas, Unidad Academica de Ciencias Nucleares, Calle Cipres No. 10, Fracc. La Penuela, 98068 Zacatecas (Mexico)


    The coloration changes of four natural extracts in function of the absorbed dose produced by a gamma rays source of {sup 137}Cs have been studied. The natural extracts were obtained of tuna varieties that contain betalains that are natural pigments of some plants as the beet of where is derived its name. These also are found in abundant form in the fruits of some species of opuntia genus (tunas). The extracts were obtained by maceration, starting from beet and three tuna varieties that were stabilized to a p H of 5.5. The extracts were exposed to the gamma rays of a {sup 137}Cs source and the change in the coloration was observed by means of an ultra violet/visible spectrophotometer through of the absorption of the samples to photons of wave longitude 535 nm. The absorption was measured, to different time intervals. The relation between the absorbed dose in D{sub w} water and the chromogene properties of the pigment was established, with the intention of using it as possible dosemeter. (Author)

  4. Transition metal cations extraction by ester and ketone derivatives of chromogenic azocalix[4]arenes. (United States)

    Ak, Metin; Taban, Deniz; Deligöz, Hasalettin


    The molecule of azocalix[n]arene is a macrocyclic used effectively in the complexation of the heavy metal pollutants (like silver and mercury). In this work, our main aim is to prepare new chromogenic azocalix[n]arene molecules to elaborate an extractant with high extractant selectivity for metal ions able to detect this type of pollutant. The solvent extraction properties of four acetyls, four methyl ketones and four benzoyls derivatives from azocalix[4]arenes which were prepared by linking 4-ethyl, 4-n-butyl, 4-acetamid anilin and 2-aminothiazol to calix[4]arene through a diazo-coupling reaction, the alkaline earth (Sr2+) and the transition (Ag+, Hg2+, Co2+, Ni2+, Cu2+, Zn2+, Cd2+, Cr3+) metal cations have been determined by extraction studies with metal picrates. Both ketones are better extractants than esters, and show a strong preference for Ag+, while Cu2+ and Cr3+ are the most extracted cation with the esters. Both acetyl and benzoyl esters are good carriers for Ag+ and Hg2+.

  5. extended-spectrum-lactamases (ESBLs) chromogenic medium in Escherichia coli and Klebsiella pneumoniae strains carried%ChromID ESBLs显色培养基的临床应用评价

    Institute of Scientific and Technical Information of China (English)

    乔宁; 喻华; 姜伟; 黄湘宁; 张凯


    目的 评价超广谱β-内酰胺酶(ESBLs)显色培养基对患者标本中携带ESBLs大肠埃希菌、肺炎克雷伯菌等细菌的快速筛选作用.方法 特患者的肛拭子标本接种ESBLs显色培养基,对平板上生长的细菌做鉴定和双纸片增效确认实验.结果 显色平板为紫红色的菌株中大肠埃希菌为98.3%,且100%为ESBLs确证阳性:显色平板为绿色的菌株中仅73.3%为肺炎克雷伯菌,90.1%为ESBLs确证阳性.119株中有24株无色细菌和5株棕色细菌,其中2株ESBIs确证为阴性.结论 ChromID ESBLs显色培养基能快速检测筛选ESBLs阳性的大肠埃希菌、肺炎克雷伯菌等肠杆菌科细菌,对控制院内感染、指导临床合理使用抗菌药物有积极意义,但是鉴定到种需加以生化反应,ESBLs株必须附加试验进行确定.%To evaluate the role of rapid screening of extended-spectrum β-laetamases (ESBLs) chromogenic medium in Escherichia coli and Klebsiella pneumoniae strains earned ESBLs. Methods Hie samples of archos swab from patients were inoculated in ESBLs chromogenic medium. The growing bacteria on the plate were identified and confirmed using double-disk synergy test. Results On the chromogenic plate, 98. 3% of Escherichia coli strains were shown purple color and 100% of ESBLs strains were confirmed as positive; 73. 3% of Klebsiella pneumoniae strains were shown green color and 90. 1% of ESBLs strains were confirmed as positive. Twenty-four bacteria were colorless and Eve strains were shown brown in the 119 strains, and two ESBLs strains were confirmed as negative. Conclusion As a s rapid screening method to analyze the ESBLs of Enterobacteri-aceae bacteria, such as Escherichia coli and Klebsiella pneumonia, ChromID ESBLe chromogenic medium can provide positive effects on control of nosocomial infections and direction of clinical rational use of antibiotics. However, it needs biochemical reactions in identification of species, and ESBLs strains must to

  6. β-Glucuronidase-coupled assays of glucuronoyl esterases. (United States)

    Fraňová, Lucia; Puchart, Vladimír; Biely, Peter


    Glucuronoyl esterases (GEs) are microbial enzymes with potential to cleave the ester bonds between lignin alcohols and xylan-bound 4-O-methyl-d-glucuronic acid in plant cell walls. This activity renders GEs attractive research targets for biotechnological applications. One of the factors impeding the progress in GE research is the lack of suitable substrates. In this work, we report a facile preparation of methyl esters of chromogenic 4-nitrophenyl and 5-bromo-4-chloro-3-indolyl β-D-glucuronides for qualitative and quantitative GE assay coupled with β-glucuronidase as the auxiliary enzyme. The indolyl derivative affording a blue indigo-type product is suitable for rapid and sensitive assay of GE in commercial preparations as well as for high throughput screening of microorganisms and genomic and metagenomic libraries.

  7. Development of a Novel Chromogenic Medium for Improved Campylobacter Detection from Poultry Samples. (United States)

    Teramura, Hajime; Iwasaki, Mihoko; Ogihara, Hirokazu


    The presence of expanded-spectrum β-lactamase (ESBL)-producing Escherichia coli is a common problem in the isolation of Campylobacter from poultry samples using conventional cefoperazone-based selective media. A novel chromogenic medium (CM-HT), based on modified charcoal cefoperazone deoxycholate agar (mCCDA), has been developed as a solution for improved Campylobacter detection from poultry samples. Although the basic components of CM-HT are the same as mCCDA, CM-HT uses both granular charcoal and sodium cefoxitin to enhance viewability and inhibit ESBL-producing bacteria. All tested Campylobacter jejuni (n = 31) and Campylobacter coli (n = 6) strains grew and formed purple-colored colonies on CM-HT. In contrast, the growth of all other tested microorganisms, including ESBL-producing E. coli strains, was suppressed by this medium. Additionally, 84 poultry samples were examined for the presence of Campylobacter using the ISO 10272-1 method (enrichment with Bolton broth) and the NIHSJ-02 method (enrichment with Preston broth) with mCCDA and CM-HT media for the isolation. The numbers of samples from which Camplylobacter was detected on CM-HT using Preston and Bolton broth were 22 and 18, whereas the numbers on mCCDA were 22 and 13, respectively. Only Campylobacter was detected on CM-HT using both enrichment broths; however, there were 5 and 19 samples from which ESBL-producing E. coli was detected on mCCDA using Preston and Bolton broth, respectively. Thus, there was a significant difference between CM-HT and mCCDA in selectivity for ESBL-producing E. coli regardless of which enrichment broth was used. The results obtained demonstrated that CM-HT is a possible solution for the improved isolation of Campylobacter from poultry samples.

  8. Detection of Escherichia coli colonies on confluent plates of chromogenic media used in membrane filtration. (United States)

    Maheux, Andrée F; Dion-Dupont, Vanessa; Bisson, Marc-Antoine; Bouchard, Sébastien; Rodriguez, Manuel J


    MI agar (MI), Chromocult® Coliform agar ES (Chromocult), and DC with BCIG agar (DC) are chromogenic membrane filtration culture-based methods used to assess microbiological water quality. In this study, their ability to detect Escherichia coli colonies on confluent growth plates was determined by testing water samples containing increasing concentrations of a non-E. coli growing bacterium, Citrobacter youngae. Then, their ability to inhibit the growth of non-coliform bacteria was determined by simultaneously testing 603 well water samples. Results were compared with those obtained with mFC and Colilert® methods. Results showed that the E. coli count was difficult to determine on mFC, Chromocult and DC when non-E. coli colonies reached levels of 10(4)colony forming units (CFU)/100 mL. However, the E. coli count did not interfere with Colilert until non-E. coli colonies reached concentrations of 10(7)CFU/100 mL. No inhibition was observed with MI as E. coli colonies could be easily detected in the presence of at least 10(7)CFU/100 mL of C. youngae. Using well water samples, confluent growth plates were observed for 144, 177, and 185 of the 603 well water samples tested with the MI, Chromocult and DC methods, respectively. Among these confluent growth plates, E. coli colonies were not detected for 10, 20, and 31 water samples. However, they were detected by the mFC and/or Colilert methods. Thus, among the three methods tested, the MI method presented the lowest grow rate of atypic colonies and was the only one that presents no interference in the E. coli count.

  9. 2株H5N1亚型禽流感病毒人工感染鸭的11项血液生化指标测定%Eleven Blood Biochemical Indexes Assay of Ducks Experimentally Infected with Two Avian Influenza Viruses(H5N1)

    Institute of Scientific and Technical Information of China (English)

    叶远兰; 周泉鹤; 李玉谷


    鸭肌肉接种禽流感病毒A/duck/Guangdong/185/2004(H5N1)和A/duck/Guangdong/221/2004(H5N1)后,血清中的丙氨酸氨基转移酶、天门冬氨酸氨基转移酶、碱性磷酸酶、总蛋白、肌酐、总胆固醇、钙和磷明显升高;葡萄糖和甘油三酯明显降低;白蛋白表现为先下降后升高的变化趋势.作者认为,检测这些血液生化指标对于诊断禽流感具有一定的参考价值.%Ducks intramuscularly inoculated with avian influenza virus strains, A/duck/Guangdong/185/2004 (H5N1) and A/duck/Guangdong/221/2004(H5N1), showed that the serum alanine aminotransferase(glutamic-pyruvic transminase), aspartate aminotransferase(glutamic-oxaloacetic transminase), alkaline phosphatase, total protein, creatinine, cholesterol, calcium and phosphorus were significantly increased; while glucose and triglycerides were obviously decreased; albumin was decreased at first and increased later. We considered that assaying these blood biochemical indexes were useful for diagnosing avian influenza.


    Korobova, A G; Frolova, L N; Kliasova, G A


    The detection of enterobacteria with production of beta-lactamases of extended spectrum in selective chromogenic agar was analyzed The results ofdetection of beta-lactamases of extended spectrum was compared with "double disc" technique. The smears from mucous membrane of guttur and rectum from patients were analyzed in parallel on solid growth agar (Endo or Mac Conkey) and on selective agar CHROMagartm ESBL (CHROMagar France). The production of beta-lactamases of extended spectrum was confirmed using "double discs" technique. To exclude hyper-production of ampC beta-lactamases E-test was applied containing cefotetan and cefotetan with cloxacillin. The sampling consisted of 1552 samples from patients. The study permitted to isolate 1243 strains of enterobacteria on agar Endo or Mac Conkey and 409 strains of enterobacteria on selective agar CHROMagartm ESBL (Escherichia coli n = 226, Klebsiella pneumoniae n = 105, enterobacter spp. n = 35, Citrobacter spp. n = 21, others n = 22). The application of "double discs" technique confirmed production of beta-lactamases of extended spectrum in 386 (94%) out of 409 strains isolated on agar CHROMagartm ESBL. In 23 (6%) of strains no confirmation was established and hyper-production of ampC of beta-lactamases was established 15 out of total. Additionally, 8 were sensitive to cephalosporin of third generation. All enterobacteria isolated on agar Endo or Mac Conkey also were tested by "double discs" technique. Overall, 394 strains of enterobacteria with production of beta-lactamases of extended spectrum were obtained. On all agars (agar Endo or Mac Conkey and CHROMagartm ESBL)--263 (67%) strains; only on CHROMagartm ESBL--123 (31%) and only on agar Endo or Mac Conkey--8 (2%) (p agar CHROMagartm ESBL made up to 98% and specificity--97%. The resolution about detection of enterobacteria producing beta-lactamases of extended spectrum were submitted to clinic in 18-24 hours after arrival ofsamplesfrom patients in laboratory. The CHR

  11. A new chromogenic agent for iron(III): Synthesis, structure and spectroscopic studies

    Indian Academy of Sciences (India)

    Chandrama Basu; Santanu Chowdhury; Helen Stoeckli-Evans; Soma Mukherjee


    A heterocyclic hydrazone ligand, diacetyl monoxime-2-pyridyl hydrazone, HL, 1, was investigated as a new chromogenic agent for selective detection of Fe3+. The ligand 1, undergoes 1 : 2 complexation with Fe3+ and Ni2+ to form complexes [FeIII(HL)2]Cl3, 1a and [NiII(HL)2]Cl2, 1b respectively. The iron(III) complex 1a gives a characteristic absorption peak at 487 nm with distinct reddish-pink colouration. The change in colour can easily be distinguished from other metal complexes by the naked eye. No obvious interference was observed in presence of other metal ions (Na+, K+, Ca2+, Mg2+, Al3+, Mn2+, Ni2+, Cu2+, Zn2+, Co2+, Pb2+, Hg2+, Cd2+). The bands appearing in the UV region (200-340 nm) are characteristics of the ligand, HL, 1. In the complexes [FeIII(HL)2]Cl3, 1a, and [NiII(HL)2]Cl2, 1b, these ligand centered bands are accompanied by multiple bands extending into the visible region (350-500 nm). The association constants (ass, UV-Vis) were found to be (6.4865 ± 0.004) × 105 for the complex 1a and (1.1960 ± 0.002) × 105 for the complex 1b at 298 K determined by the UV-Vis spectroscopy. On excitation at 285 nm, the ligand HL, 1 strongly emits at 364 nm due to an intraligand1( - *) transition. The complexes are luminescent (ex 285 nm, em 365 nm) with /0 0.75 for 1a and 0.81 for 1b. In both the cases, the 1 : 2 binding is confirmed by Job’s method. Molecular structure of the complex 1b has been determined by single crystal X-ray diffraction studies. Here, two crystallographically distinct but metrically very similar molecules making an enantiomeric pair constitute the asymmetric unit in which both metal atoms are tris chelated in meridional geometry.

  12. ESBL Detection: Comparison of a Commercially Available Chromogenic Test for Third Generation Cephalosporine Resistance and Automated Susceptibility Testing in Enterobactericeae (United States)

    El-Jade, Mohamed Ramadan; Parcina, Marijo; Schmithausen, Ricarda Maria; Stein, Christoph; Meilaender, Alina; Hoerauf, Achim; Molitor, Ernst


    Rapid detection and reporting of third generation cephalosporine resistance (3GC-R) and of extended spectrum betalactamases in Enterobacteriaceae (ESBL-E) is a diagnostic and therapeutic priority to avoid inefficacy of the initial antibiotic regimen. In this study we evaluated a commercially available chromogenic screen for 3GC-R as a predictive and/or confirmatory test for ESBL and AmpC activity in clinical and veterinary Enterobacteriaceae isolates. The test was highly reliable in the prediction of cefotaxime and cefpodoxime resistance, but there was no correlation with ceftazidime and piperacillin/tazobactam minimal inhibitory concentrations. All human and porcine ESBL-E tested were detected with exception of one genetically positive but phenotypically negative isolate. By contrast, AmpC detection rates lay below 30%. Notably, exclusion of piperacillin/tazobactam resistant, 3GC susceptible K1+ Klebsiella isolates increased the sensitivity and specificity of the test for ESBL detection. Our data further imply that in regions with low prevalence of AmpC and K1 positive E. coli strains chromogenic testing for 3GC-R can substitute for more time consuming ESBL confirmative testing in E. coli isolates tested positive by Phoenix or VITEK2 ESBL screen. We, therefore, suggest a diagnostic algorithm that distinguishes 3GC-R screening from primary culture and species-dependent confirmatory ESBL testing by βLACTATM and discuss the implications of MIC distribution results on the choice of antibiotic regimen. PMID:27494134

  13. ESBL Detection: Comparison of a Commercially Available Chromogenic Test for Third Generation Cephalosporine Resistance and Automated Susceptibility Testing in Enterobactericeae. (United States)

    El-Jade, Mohamed Ramadan; Parcina, Marijo; Schmithausen, Ricarda Maria; Stein, Christoph; Meilaender, Alina; Hoerauf, Achim; Molitor, Ernst; Bekeredjian-Ding, Isabelle


    Rapid detection and reporting of third generation cephalosporine resistance (3GC-R) and of extended spectrum betalactamases in Enterobacteriaceae (ESBL-E) is a diagnostic and therapeutic priority to avoid inefficacy of the initial antibiotic regimen. In this study we evaluated a commercially available chromogenic screen for 3GC-R as a predictive and/or confirmatory test for ESBL and AmpC activity in clinical and veterinary Enterobacteriaceae isolates. The test was highly reliable in the prediction of cefotaxime and cefpodoxime resistance, but there was no correlation with ceftazidime and piperacillin/tazobactam minimal inhibitory concentrations. All human and porcine ESBL-E tested were detected with exception of one genetically positive but phenotypically negative isolate. By contrast, AmpC detection rates lay below 30%. Notably, exclusion of piperacillin/tazobactam resistant, 3GC susceptible K1+ Klebsiella isolates increased the sensitivity and specificity of the test for ESBL detection. Our data further imply that in regions with low prevalence of AmpC and K1 positive E. coli strains chromogenic testing for 3GC-R can substitute for more time consuming ESBL confirmative testing in E. coli isolates tested positive by Phoenix or VITEK2 ESBL screen. We, therefore, suggest a diagnostic algorithm that distinguishes 3GC-R screening from primary culture and species-dependent confirmatory ESBL testing by βLACTATM and discuss the implications of MIC distribution results on the choice of antibiotic regimen.

  14. Detection of Streptococcus pyogenes using rapid visual molecular assay. (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


    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.

  15. Digital magnetic tagging for multiplexed suspension-based biochemical assays (United States)

    Mitrelias, T.; Trypiniotis, T.; Palfreyman, J. J.; Hong, B.; Vyas, K.; Hayward, T. J.; Llandro, J.; Kopper, K. P.; Bland, J. A. C.; Robertson, P. A.; Barnes, C. H. W.


    Microarrays and suspension (or bead)-based technologies have attracted significant interest for their broad applications in high throughput molecular biology. However, the throughput of microarrays will always be limited by the array density and the slow diffusion of molecules to their binding sites. Suspension-based technologies, in which all the reactions take place directly on the surface of microcarriers functionalized with molecular probes, could offer true multiplexing due to the possibility of extending their detection capability by a straightforward expansion of the size of the chemical library of probes. To fully exploit their potential, the microcarriers must be tagged, but the number of distinct codes available from spectrometric/graphical/physical encoding methods is currently fairly limited. A digital magnetic tagging method based on magnetic microtags, which have been anisotropy engineered to provide stable magnetization directions which correspond to digital codes, is reported. The tags can be suspended in solution and functionalized with a variety of biological molecular probes. Magnetic tagging offers several benefits compared to the traditional optical encoding techniques currently employed. It offers minimal background signals, potential for a large number of distinct codes, miniaturization of devices, and the ability to write a code in situ. Experimental data showing the reading of individual magnetic microbars from samples comprising 50×20 μm2 Ni elements, as well as micromagnetic simulations that show the feasibility of stray field detection, are presented. The stray fields of the magnetic microbars spanning a range of 60 mOe were detected by a microfabricated fluxgate sensor scanned in a raster fashion over the sample that was placed about 70 μm away. Free floating tags have also been fabricated for use in microfluidic systems. A magnetic lab-on-a-chip device could be used for tagging biomolecular probes for applications in genome sequencing, immunoassays, clinical diagnostics, drug discovery, and general pathogen detection and screening.

  16. Modularity of Biochemical Filtering for Inducing Sigmoid Response in Both Inputs in an Enzymatic AND Gate

    CERN Document Server

    Bakshi, Saira; Halamek, Jan; Privman, Vladimir; Katz, Evgeny


    We report the first systematic study of designed two-input biochemical systems as information processing gates with favorable noise-transmission properties accomplished by modifying the gate's response from convex shape to sigmoid in both inputs. This is realized by an added chemical "filter" process which recycles some of the output back into one of the inputs. We study a system involving the biocatalytic function of the enzyme horseradish peroxidase, functioning as an AND gate. We consider modularity properties, such as the use of three different input chromogens that, when oxidized yield signal-detection outputs for various ranges of the primary input, hydrogen peroxide. We also examine possible uses of different filter-effect chemicals (reducing agents) to induce the sigmoid-response. A modeling approach is developed and applied to our data, allowing us to describe the enzymatic kinetics in the framework of a formulation suitable for evaluating the noise-handling properties of the studied systems as logic...

  17. A highly sensitive and selective turn-on fluorogenic and chromogenic sensor based on BODIPY-functionalized magnetic nanoparticles for detecting lead in living cells. (United States)

    Son, Hyunjong; Lee, Hye Young; Lim, Jung Mi; Kang, Dongmin; Han, Won Seok; Lee, Shim Sung; Jung, Jong Hwa


    A new fluoro-chromogenic chemosensor based on BODIPY-functionalized Fe(3)O(4)@SiO(2) core/shell nanoparticles 1 has been prepared. Chemosensor 1 exhibits a high affinity and selectivity for Pb(2+) over competing metal ions tested. Moreover, confocal microscopy, and flow cytometry experiments established that 1 can be used for detecting Pb(2+) levels within living cell.

  18. Determination of HER-2 status on FNAC material from breast carcinomas using in situ hybridization with dual chromogen visualization with silver enhancement (dual SISH

    Directory of Open Access Journals (Sweden)

    Beraki Elsa


    Full Text Available During the last years, HER-2 status kits and protocols for chromogen visualization of hybridization signals have come on the market. The first generation using chromogen visualization used single color probes. The second generation, now emerging on the market, uses dual chromogen visualization. The aim of this study has been to test a new dual color chromogen kit (Ventana INFORM HER2 Dual Colour ISH Roche ® and compare the results with our in-house method(s. The material consisted primarily of cytological material from invasive breast carcinomas in 49 women. Dual SISH was done on all 49 cytological and histological specimens. The histological specimens were treated according to the manufacturer′s recommendations. The procedure was modified in several steps in order to adapt it to the cytological material. Hybridization failed in two cytological specimens. Dual SISH showed concordant results on cytological and histological material as to amplified/not amplified. The included cases had the same HER-2 expression in the invasive and the in situ components on histology. Four IDC showed HER-2 amplification (8.5%. Polysomy was found in two cases. All dual SISH results except for one concurred with the results of the in-house method(s (1/47=2.1%. The dual SISH is suitable for cytological examination of HER-2 status. The protocol must be optimized for cytological material.

  19. Synthesis and application of resorufin β-D-glucuronide, a low-cost chromogenic substrate for detecting Escherichia coli in drinking water. (United States)

    Magro, Germinal; Bain, Robert E S; Woodall, Claire A; Matthews, Robert L; Gundry, Stephen W; Davis, Anthony P


    The development of low-cost tests for Escherichia coli is hampered by the expense and limited choice of enzyme substrates. Most chromogenic substrates are required in costly amounts, while fluorogenic substrates require an additional apparatus (e.g., an ultraviolet lamp) to be detected. Herein, we propose an alternative chromogenic substrate, resorufin β-d-glucuronide (REG), which is exceptionally sensitive and may be employed in very small amounts. We show that REG can be produced similarly to other simple glucuronides and should therefore be no more expensive. The compound is used by both healthy and injured E. coli, resulting in a pronounced color change from orange to a bright pink. Because the released dye (resorufin) has a high extinction coefficient, substantially lower amounts are needed than for commercially available substrates. The potential of this substrate is demonstrated by a presence/absence test requiring just 0.1 mg of REG/100 mL of water sample, one hundredth of the quantity needed for common chromogenic substrates, with an estimated bulk cost of ≤0.1 U.S. cents/test. REG shows promise as a chromogenic substrate for E. coli detection and should be considered in the development of new water tests, especially for low-income settings.

  20. Prospective evaluation of the chromogenic medium CandiSelect 4 for differentiation and presumptive identification of non-Candida albicans Candida species

    NARCIS (Netherlands)

    Zhao, Liang; de Hoog, G Sybren; Cornelissen, Akke; Lyu, Qian; Mou, Lili; Liu, Taohua; Cao, Yu; Vatanshenassan, Mansoureh; Kang, Yingqian


    Rapid identification of pathogenic yeasts is a crucial step in timely and appropriate antifungal therapy. For diagnostics in the clinical laboratory, simplified alternatives to barcoding are needed. CandiSelect 4 (CS4) medium, a chromogenic medium for isolation of clinical yeasts, allows routine rec

  1. Biochemical Neuroscience Laboratory (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...

  2. Comparison of two rapid biochemical tests and four chromogenic selective media for detection of carbapenemase-producing Gram-negative bacteria. (United States)

    Hinić, Vladimira; Amrein, Ivo; Stammler, Sabrina; Heckendorn, Judith; Meinel, Dominik; Frei, Reno; Egli, Adrian


    We evaluated RAPIDEC® CARBA NP, Neo-Rapid CARB, chromID® CARBA SMART (CARB/OXA), Brilliance™ CRE/ESBL, ChromArt CRE and BBL™ CHROMagar™ CPE for the detection of carbapenemase-producing bacteria. The analytical sensitivity of RAPIDEC® CARBA NP was better than that of Neo-Rapid CARB. A combination of carbapenemase and ESBL screening plates could be advantageous.

  3. Measures of Biochemical Sociology (United States)

    Snell, Joel; Marsh, Mitchell


    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…

  4. Biochemical Education in Brazil. (United States)

    Vella, F.


    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)

  5. Changes in the chromogene properties of the betalaine; Cambios en las propiedades cromogenas de las betalainas inducidos por radiacion gamma

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez N, S.; Pinedo S, A.; Amador V, P.; Chacon R, A.; Arcos P, A.; Vega C, H.R. [Unidad Academica de Ciencias Nucleares, C. Cipres 10, Fracc. La Penuela, 98068, Zacatecas, Zac. (Mexico)


    The changes of coloration of four natural extracts were determined in function of the absorbed dose taken place by a source of rays gamma of Cs-137. The used natural extracts contain betalaine that are natural pigments of some plants as the beet root, of there their name. They are also in abundant form in the fruits (tunas) of some species of the Opuntia gender. The extracts were obtained by maceration, starting from beet root and three tuna varieties, and they were stabilized to pH 5.5 the change of coloration you determines in a spectrophotometer ultraviolet/visible by means of the absorbance from the samples to photons of 475, 535 and 600 nm of wave longitude. The absorbance was measured, to different intervals of time. The relationship settled down between the absorbed dose and the chromogene properties of the pigment, with the intention of using it as possible dosemeter. (Author)

  6. A comparison of standard cultural methods for the detection of foodborne Salmonella species including three new chromogenic plating media. (United States)

    Schönenbrücher, Vanessa; Mallinson, Edward T; Bülte, Michael


    In this study the draft of the horizontal method for the detection of Salmonella species from human food and animal feed (ISO 6579:2002) was compared to the European gold standard (DIN EN 12824:1998), including the three new chromogenic plating media AES Salmonella Agar Plate (ASAP), Oxoid Salmonella Chromogen Media (OSCM) and Miller-Mallinson agar (MM). First the growth and appearance of 36 bacterial type strains (Salmonella and other 21 species) on ASAP, OSCM and MM were compared to those on the three traditional agars Brilliant Green Agar according to Edel and Kampelmacher (BGA), Xylose Lysine Deoxycholate Agar (XLD) and Xylose Lysine Tergitol 4 Agar (XLT4). Only on MM agar, did all of 36 tested type strains produce typical colonies, especially strains of S. Senftenberg, Salmonella arizonae, S. Dublin and S. Derby. Artificial inoculation experiments using raw pork ground meat (n=92) were subsequently conducted. A shortened incubation time of 24 h in RVS broth yielded a Salmonella species recovery of 100% from spiked meat samples. Finally, 286 naturally contaminated raw porcine and bovine minced meat samples and raw poultry meat samples were investigated. Forty-three strains from a total of 39 Salmonella-positive samples were found. S. Typhimurium (n=21), with DT 104 L, DT 012 and RDNC being the most prevalent subtypes isolated. D-tartrate-positive S. Paratyphi B (n=2) and S. Saint-Paul (n=3) were also recovered. They were cultured from poultry meat and were multi-resistant against antibiotics including nalidixic acid. Rappaport Vassiliadis broth with soypeptone (RVS) yielded the highest recovery of Salmonella spp. (97,4%) compared to Tetrathionate broth with Novobiocin according to Muller and Kauffman (MKTTn, 94,9%) and Selenite Cystine broth (SC, 38,5%). However, no significant difference was obtained by comparing the ISO 6579:2002 draft to the gold standard.

  7. Fluorogenic and chromogenic probe for rapid detection of a nerve agent simulant DCP. (United States)

    Wu, Wei-hui; Dong, Jun-jun; Wang, Xin; Li, Jian; Sui, Shao-hui; Chen, Gao-yun; Liu, Ji-wei; Zhang, Ming


    A fluorogenic and visual probe was devised to detect diethyl chlorophosphate (DCP), a nerve agent simulant. The probe, N-(rhodamine B)-lactam-2-aminoethanol (RB-AE), undergoes oxazoline formation following phosphorylation in the presence of DCP, which gives rapid and clear fluorescence and color change in the assay solutions.

  8. Multiplexing oscillatory biochemical signals. (United States)

    de Ronde, Wiet; ten Wolde, Pieter Rein


    In recent years it has been increasingly recognized that biochemical signals are not necessarily constant in time and that the temporal dynamics of a signal can be the information carrier. Moreover, it is now well established that the protein signaling network of living cells has a bow-tie structure and that components are often shared between different signaling pathways. Here we show by mathematical modeling that living cells can multiplex a constant and an oscillatory signal: they can transmit these two signals simultaneously through a common signaling pathway, and yet respond to them specifically and reliably. We find that information transmission is reduced not only by noise arising from the intrinsic stochasticity of biochemical reactions, but also by crosstalk between the different channels. Yet, under biologically relevant conditions more than 2 bits of information can be transmitted per channel, even when the two signals are transmitted simultaneously. These observations suggest that oscillatory signals are ideal for multiplexing signals.

  9. 自主研制志贺氏菌显色培养基的应用效果评价%Evaluation of applications for Shigella chromogenic medium based on independent innovation

    Institute of Scientific and Technical Information of China (English)

    胡杨峰; 何艳玲; 刘沛


    Objective To explore the application effect of the shigella chromogenic medium based on independent innovation. Methods A total of 30 standard strains were used to test the chromogenic medium, and SO food samples on sale were detected by shigella chromogenic medium based on independent innovation, R&F shigella chromogenic medium, XLD and MacConkey agar. Results Shigella spp. Could be effectively detected by Shigella chromogenic medium based on independent innovation, the detection rate of food samples was 10% by innovative products and R&F Shigella chromogenic medium. Conclusion The specialty,sensitivity and detection rate of innovative shigella chromogenic medium and R&F shigella chromogenic medium are consistency.%目的 探讨自主研制的志贺氏菌显色培养基的应用效果.方法 采用30株标准菌株及50份自然样品检测试验,比对自主研制的志贺氏菌显色培养基、梅里埃(R&F)志贺氏菌显色培养基、木糖-赖氨酸-去氧胆酸盐培养基(XLD培养基)和麦康凯琼脂培养基的选择性.结果 自主研制的志贺氏菌显色培养基能有效分离志贺氏菌属菌株,50份自然样品的检出率与R&F志贺氏菌显色培养基相同,均达到10%.结论 自主研制的志贺氏菌显色培养基与R&F志贺氏菌显色培养基在特异性及检出率达到一致.

  10. Double-staining chromogenic in situ hybridization as a useful alternative to split-signal fluorescence in situ hybridization in lymphoma diagnostics

    DEFF Research Database (Denmark)

    van Rijk, A.; Svenstroup-Poulsen, T.; Jones, M.;


    , their detection is an important adjunct for increasing the reliability of the diagnosis. Recently, split-signal fluorescence hi situ hybridization has become available as a robust method to detect chromosomal breaks in paraffin-embedded formalin-fixed tissues. A bright field approach would bring this technology...... within the reach of every pathology laboratory. Design and Methods Our study was initiated to determine the consistency between chromogenic in situ hybridization and fluorescence in situ hybridization, both using split-signal probes developed for the detection of chromosomal breaks. Five hundred...... after split-signal fluorescence in situ hybridization staining. Conclusions We conclude that double-staining chromogenic in situ hybridization is equally reliable as fluorescence in situ hybridization in detecting chromosomal breaks in lymphoid tissue. Although differences in morphology, hematoxylin...

  11. 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;


    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...

  12. High concentration of human lactoferrin in milk of rhLf-transgenic cows relieves signs of bovine experimental Staphylococcus chromogenes intramammary infection. (United States)

    Simojoki, Heli; Hyvönen, Paula; Orro, Toomas; Pyörälä, Satu


    Six transgenic cows producing recombinant human lactoferrin (rhLf) in their milk and five normal cows at the same lactation stage were experimentally infected with Staphylococcus chromogenes to study the effect of a high concentration of lactoferrin in milk. Coagulase-negative staphylococci such as S. chromogenes have become very common as agents causing mild or subclinical mastitis. All transgenic cows became infected but showed no clinical signs, unlike the control cows, which developed mild clinical mastitis. Transgenic cows eliminated bacteria faster from the quarters than did the controls. Local clinical signs were milder, and the inflammatory reaction assessed by NAGase activity in the milk and by the concentration of milk amyloid A was lower in the transgenic cows. The mild response probably reflected the rapid elimination of bacteria. The milk concentration of rhLf remained constant throughout the study period, but the total concentration of bovine lactoferrin in the milk peaked in both groups at 46h post-challenge. Three cows, all in the control group, exhibited systemic acute phase response as increased concentrations of serum amyloid A in the blood circulation. Transgenic cows with a high concentration of human lactoferrin in their milk seemed to be protected from clinical disease and from prolonged inflammatory reaction, but not from experimental intramammary infection induced by S. chromogenes.

  13. Usefulness of Chromogenic CromoCen® AGN agar medium for the identification of the genus Aeromonas: Assessment of faecal samples. (United States)

    Aguilera-Arreola, M G; Portillo-Muñoz, M I; Rodríguez-Martínez, C; Castro-Escarpulli, G


    Selective screening media for the detection and identification of Aeromonas strains are needed to guide primary isolation procedures in the clinical laboratory. This study compared the selective CromoCen® AGN chromogenic agar medium for the detection and identification of Aeromonas strains that were isolated from various samples against the conventional selective agar media that are commonly used for the isolation of this organism in food, environmental and clinical samples. The Miles and Misra and ecometric methods were used to evaluate the microbiological performance of CromoCen® AGN chromogenic agar medium, which was shown to be satisfactory. A total of 14 reference Aeromonas strains, 44 wild strains and 106 clinical stool specimens were examined using both non-chromogenic selective agars that are commonly used for Aeromonas isolation and CromoCen® AGN agar. The latter exhibited 94.73% sensitivity and 100% specificity for the various samples. On CromoCen® AGN agar medium, Aeromonas formed colonies with light green, greenish and salmon pigments with or without a surrounding wide transparent zone (halo) of 2-3mm in diameter around the entire border. This medium is recommended for the isolation and potential identification of the Aeromonas genus.

  14. Detection of c-myc amplification in formalin-fixed paraffin-embedded tumor tissue by chromogenic in situ hybridization (CISH). (United States)

    Todorović-Raković, Nataša


    In situ hybridization (ISH) allows evaluation of genetic abnormalities, such as changes in chromosome number, chromosome translocations or gene amplifications, by hybridization of tagged DNA (or RNA) probes with complementary DNA (or RNA) sequences in interphase nuclei of target tissue. However, chromogenic in situ hybridization (CISH) is also applicable to formalin-fixed, paraffin-embedded (FFPE) tissues, besides metaphase chromosome spreads. CISH is similar to fluorescent in situ hybridization (FISH) regarding pretreatments and hybridization protocols but differs in the way of visualization. Indeed, CISH signal detection is similar to that used in immunohistochemistry, making use of a peroxidase-based chromogenic reaction instead of fluorescent dyes. In particular, tagged DNA probes are indirectly detected using an enzyme-conjugated antibody targeting the tags. The enzymatic reaction of the chromogenic substrate leads to the formation of strong permanent brown signals that can be visualized by bright-field microscopy at 40 × magnification. The advantage of CISH is that it allows the simultaneous observation of gene amplification and tissue morphology and the slides can be stored for a long time.

  15. Assays for Determination of Protein Concentration. (United States)

    Olson, Bradley J S C


    Biochemical analysis of proteins relies on accurate quantification of protein concentration. Detailed in this appendix are some commonly used methods for protein analysis, e.g., Lowry, Bradford, bicinchoninic acid (BCA), UV spectroscopic, and 3-(4-carboxybenzoyl)quinoline-2-carboxaldehyde (CBQCA) assays. The primary focus of this report is assay selection, emphasizing sample and buffer compatibility. The fundamentals of generating protein assay standard curves and of data processing are considered, as are high-throughput adaptations of the more commonly used protein assays. Also included is a rapid, inexpensive, and reliable BCA assay of total protein in SDS-PAGE sample buffer that is used for equal loading of SDS-PAGE gels. © 2016 by John Wiley & Sons, Inc.

  16. Study of tolerance of enterobacteria to chlorine-based biocides in experimental models using chromogenic indicator tests

    Directory of Open Access Journals (Sweden)

    N.R. Efimochkina


    Full Text Available The species-specific composition of microbial contaminants of vegetable raw materials and equipment used in the production of biotechnological products and beverages fermentation are studied. 85 enterobacteria strains was isolated and investigated, 46 strains of the genera Enterobacter, Pantoea, Citrobacter, Serratia, Escherichia, Cronobacter was identified to the species level; the most frequently detected bacteria of the genera Enterobacter and Pantoea (about 50 %. For the first time developed and tested chromogenic in vitro model based that allows to quantify the degree of inhibition of gram-negative microflora under the influence of antimicrobial agents depending on the concentrations of biocides and density of bacterial populations. A comparative analysis of the tolerance of Enterobacteriaceae strains from different biotopes was conducted. Sensitivity to the treatment of chlorine-containing biocides in 26 strains of enterobacteria from plant material and 9 strains of Escherichia coli from the intestine of male rats of Wistar line was tested. Enterobacteria from vegetable raw materials and swabs were more resistant to antimicrobial action of chlorine, than the representatives of the populations of the normal intestinal microbiota. It is established that the active chlorine concentration of 50–100 mg/dm 3 , the most commonly used in the processing of vegetable raw materials, is not effective for Enterobacteriaceae, if the density of the microbial population is 10 5–7 cells/cm 3 and above. At an initial level of contamination with Enterobacteriaceae not more than 10 3 cells/cm 3 processing solutions with a concentration of active chlorine of 75 to 100 mg/dm 3 can provide effective disinfection of raw materials, equipment, or inventory. Experimental chromogenic in vitro model proposed to assess the impact of chlorine-based biocides on the degree of the enterobacteria inhibition, can be used to justify the selection and doses of

  17. Spectrophotometric Determination of Various Drugs Using Chloranilic Acid as Chromogenic Reagent - II

    Directory of Open Access Journals (Sweden)

    V. Annapurna


    Full Text Available Simple, accurate and reproducible visible spectrophotometric method for the assay of drugs such as pyrilamine and fluvoxamine as maleates was established based on the formation of charge-transfer complex and redox reaction between the corresponding drug and substituted p-benzoquinone (chloranilic acid. The optical characteristics such as Beer’s law limits, molar absorptivity and Sandell’s sensitivity are reported. Regression analysis using the method of least squares was made to evaluate the slope (b, intercept (a and correlation coefficient (r and standard error of estimation (Se for each drug.

  18. Biochemical Hypermedia: Galactose Metabolism.

    Directory of Open Access Journals (Sweden)

    J.K. Sugai


    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.

  19. Chromogenic behaviors of the Humboldt squid (Dosidicus gigas) studied in situ with an animal-borne video package. (United States)

    Rosen, Hannah; Gilly, William; Bell, Lauren; Abernathy, Kyler; Marshall, Greg


    Dosidicus gigas (Humboldt or jumbo flying squid) is an economically and ecologically influential species, yet little is known about its natural behaviors because of difficulties in studying this active predator in its oceanic environment. By using an animal-borne video package, National Geographic's Crittercam, we were able to observe natural behaviors in free-swimming D. gigas in the Gulf of California with a focus on color-generating (chromogenic) behaviors. We documented two dynamic displays without artificial lighting at depths of up to 70 m. One dynamic pattern, termed 'flashing' is characterized by a global oscillation (2-4 Hz) of body color between white and red. Flashing was almost always observed when other squid were visible in the video frame, and this behavior presumably represents intraspecific signaling. Amplitude and frequency of flashing can be modulated, and the phase relationship with another squid can also be rapidly altered. Another dynamic display termed 'flickering' was observed whenever flashing was not occurring. This behavior is characterized by irregular wave-like activity in neighboring patches of chromatophores, and the resulting patterns mimic reflections of down-welled light in the water column, suggesting that this behavior may provide a dynamic type of camouflage. Rapid and global pauses in flickering, often before a flashing episode, indicate that flickering is under inhibitory neural control. Although flashing and flickering have not been described in other squid, functional similarities are evident with other species.

  20. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Directory of Open Access Journals (Sweden)

    F.E. Rosa


    Full Text Available Human epidermal growth factor receptor 2 (HER2 has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC. HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH in moderate immunoexpression (IHC 2+ cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH, which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

  1. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Directory of Open Access Journals (Sweden)

    F.E. Rosa

    Full Text Available Human epidermal growth factor receptor 2 (HER2 has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC. HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH in moderate immunoexpression (IHC 2+ cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH, which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

  2. Targeting topoisomerase IIa in endometrial adenocarcinoma: a combined chromogenic in situ hybridization and immunohistochemistry study based on tissue microarrays. (United States)

    Tsiambas, E; Alexopoulou, D; Lambropoulou, S; Gerontopoulos, K; Karakitsos, P; Karameris, A


    Topoisomerase IIa is a nucleic enzyme that affects the topological structure of DNA and also is a target for chemotherapy (ie, anthracyclines). In this study, we coevaluated its protein expression with chromosome 17 and gene status. Using tissue microarrays, 40 cases of sporadic, primary endometrial adenocarcinomas, 5 cases of atypical hyperplasia, and 5 cases of benign hyperplasia were obtained and reembedded into two paraffin blocks with a core diameter of 1 mm. Immunohistochemistry combined with chromogenic in situ hybridization was performed in 2 and 5 microm sections, respectively. Finally using a semiautomated Image Analysis System, we evaluated the levels of Nuclear labeling index of topoisomerase IIa expression. Statistical analysis was performed by SPSS version 11.0 software. The results indicate that chromosome 17 instability (aneuploidy in 7/40 cases) and Topo IIa gene deregulation (amplification in 3/40 and deletion in 1/40 cases) are significant genetic events correlated with biologic behavior in endometrial adenocarcinoma. Because protein overexpression was observed in a significant proportion of the tumors (18/40), detection of the specific gene deregulation mechanism is a crucial process for application of targeted chemotherapies, which are characterized by different levels of cardiotoxicity and other serious effects.

  3. Presumptive identification of Candida species other than C. albicans, C. krusei, and C. tropicalis with the chromogenic medium CHROMagar Candida

    Directory of Open Access Journals (Sweden)

    Horvath Lynn L


    Full Text Available Abstract Background CHROMagar Candida (CaC is increasingly being reported as a medium used to differentiate Candida albicans from non-albicans Candida (NAC species. Rapid identification of NAC can assist the clinician in selecting appropriate antifungal therapy. CaC is a differential chromogenic medium designed to identify C. albicans, C. krusei, and C. tropicalis based on colony color and morphology. Some reports have proposed that CaC can also reliably identify C. dubliniensis and C. glabrata. Methods We evaluated the usefulness of CaC in the identification of C. dubliniensis, C. famata, C. firmetaria, C. glabrata, C. guilliermondii, C. inconspicua, C. kefyr, C. lipolytica, C. lusitaniae, C. norvegensis, C. parapsilosis, and C. rugosa. Results Most NAC produced colonies that were shades of pink, lavender, or ivory. Several isolates of C. firmetaria and all C. inconspicua produced colonies difficult to differentiate from C. krusei. Most C. rugosa isolates produced unique colonies with morphology like C. krusei except in a light blue-green color. C. glabrata isolates produced small dark violet colonies that could be differentiated from the pink and lavender colors produced by other species. All seventeen isolates of C. dubliniensis produced green colonies similar to those produced by C. albicans. Conclusion C. glabrata and C. rugosa appear distinguishable from other species using CaC. Some NAC, including C. firmetaria and C. inconspicua, could be confused with C. krusei using this medium.

  4. Comparison of two chromogenic media for the detection of vancomycin-resistant enterococcal carriage by nursing home residents. (United States)

    Gouliouris, Theodore; Blane, Beth; Brodrick, Hayley J; Raven, Kathy E; Ambridge, Kirsty E; Kidney, Angela D; Hadjirin, Nazreen F; Török, M Estée; Limmathurotsakul, Direk; Peacock, Sharon J


    We compared ChromID VRE and Brilliance VRE media for the detection of vancomycin-resistant enterococci (VRE). Using a panel of 28 enterococcal isolates, 10 vanA Enterococcus faecium and three vanA Enterococcus faecalis isolates grew as per manufacturers' instructions whilst growth of two vanC and eight vancomycin-susceptible enterococci was inhibited on both media. Important differences were noted in the selectivity and chromogenic properties of the two media for vanA Enterococcus raffinosus and vanB E. faecium. The two media were further evaluated using 295 stool samples from nursing home residents, 34 of which grew VRE (11.5%). ChromID and Brilliance had comparable sensitivity, which was increased markedly by prolonging incubation to 48 hours (from 29% to 82%, and from 41% to 85%, respectively) and by a pre-enrichment step (to 97% and 100%, respectively). Brilliance VRE agar had higher selectivity at 48 hours, and after pre-enrichment.

  5. Hemin-block copolymer micelle as an artificial peroxidase and its applications in chromogenic detection and biocatalysis. (United States)

    Qu, Rui; Shen, Liangliang; Chai, Zhihua; Jing, Chen; Zhang, Yufeng; An, Yingli; Shi, Linqi


    Following an inspiration from the fine structure of natural peroxidases, such as horseradish peroxidase (HRP), an artificial peroxidase was constructed through the self-assembly of diblock copolymers and hemin, which formed a functional micelle with peroxidase-like activity. The pyridine moiety in block copolymer poly(ethylene glycol)-block-poly(4-vinylpyridine) (PEG-b-P4VP) can coordinate with hemin, and thus hemin is present in a five-coordinate complex with an open site for binding substrates, which mimics the microenvironment of heme in natural peroxidases. The amphiphilic core-shell structure of the micelle and the coordination interaction of the polymer to the hemin inhibit the formation of hemin μ-oxo dimers, and thereby enhance the stability of hemin in the water phase. Hemin-micelles exhibited excellent catalytic performance in the oxidation of phenolic and azo compounds by H2O2. In comparison with natural peroxidases, hemin-micelles have higher catalytic activity and better stability over wide temperature and pH ranges. Hemin-micelles can be used as a detection system for H2O2 with chromogenic substrates, and they anticipate the possibility of constructing new biocatalysts tailored to specific functions.

  6. Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

    Energy Technology Data Exchange (ETDEWEB)

    Rosa, F.E.; Santos, R.M. [Departamento de Patologia, Hospital das Clínicas, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil); Rogatto, S.R. [Departamento de Urologia, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil); Hospital A.C. Camargo, CIPE, São Paulo, SP (Brazil); Domingues, M.A.C. [Departamento de Patologia, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil)


    Human epidermal growth factor receptor 2 (HER2) has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC). HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH) in moderate immunoexpression (IHC 2+) cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH), which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH) and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

  7. Ex vivo effects of low-dose rivaroxaban on specific coagulation assays and coagulation factor activities in patients under real life conditions. (United States)

    Mani, Helen; Hesse, Christian; Stratmann, Gertrud; Lindhoff-Last, Edelgard


    Global coagulation assays display variable effects at different concentrations of rivaroxaban. The aim of this study is to quantify the ex vivo effects of low-dose rivaroxaban on thrombophilia screening assays and coagulation factor activities based on the administration time, and to show how to mask possible interferences. Plasma samples from 40 patients receiving rivaroxaban 10 mg daily were investigated to measure activities of clotting factor II, V, VII, VIII, IX, XI, XII and XIII; protein C- and protein S-levels; lupus anticoagulants; anticardiolipin IgG and IgM; D-dimer, heparin-platelet factor 4 (HPF4) antibodies and screening tests for von Willebrand disease (VWD). Two hours after rivaroxaban administration, the activities of clotting factors were significantly decreased to different extents, except for factor XIII. Dilution of plasma samples resulted in neutralisation of these interferences. The chromogenic protein C activity assay was not affected by rivaroxaban. Depending on the timing of tablet intake in relation to blood sampling protein S activity was measured falsely high when a clotting assay was used. False-positive results for lupus anticoagulants were observed depending on the assay system used and the administration time of rivaroxaban. ELISA-based assays such as anticardiolipin IgG and IgM, D-dimer, HPF4-antibodies and the turbidimetric assays for VWD were not affected by rivaroxaban. Specific haemostasis clotting tests should be performed directly prior to rivaroxaban intake. Assay optimisation in the presence of rivaroxaban can be achieved by plasma dilution. Immunologic assays are not influenced by rivaroxaban, while chromogenic assays can be used, when they do not depend on factor Xa.

  8. Rotating disk sorbent extraction for pre-concentration of chromogenic organic compounds and direct determination by solid phase spectrophotometry. (United States)

    Richter, Pablo; Cañas, Alejandro; Muñoz, Carlos; Leiva, Claudio; Ahumada, Inés


    A novel and very simple microextraction approach for pre-concentration and direct solid phase spectrophotometric measurement has been developed for the determination of chromogenic analytes. The model analyte to assess this approach was the chromophore malachite green (MG). The analyte was extracted from water samples onto a small rotating disk made of Teflon containing a sorbent phase of polydimethylsiloxane (PDMS) on one of its surfaces. We refer to the extraction procedure as rotating disk sorptive extraction (RDSE). After extraction, the sorbent phase with the concentrated analyte was separated from the Teflon disk and used directly for MG determination by solid phase spectrophotometry at 624 nm, without the necessity of a desorption step. Chemical and extraction variables such as concentration of sodium sulfate, pH, disk rotational velocity, extraction time, and temperature were studied in order to establish the best conditions for extraction. Under optimum conditions, the extraction of MG was carried out in 18 min and 90 min, for sample volumes of 100mL or 1000 mL, respectively. The detection limit, based on three times the standard deviation of the blank phase (3σ(b)), was 1.4 μg L⁻¹ and the repeatability, expressed as relative standard deviation (RSD), for 20 μg L⁻¹ MG was 8.1%. This study also applied the method to real samples, obtaining quantitative recovery (mean recovery of 99.3%). The PDMS phases could be reused after desorbing the MG into methanol for 3h. Replacement of the PDMS film onto the disk is very easy and low cost.

  9. Measuring dabigatran with the dilute Russell viper venom confirm assay in an anticoagulation clinic population. (United States)

    McGlasson, David L; Fritsma, George A


    The dabigatran dose-response is predictable; however, it is necessary to measure plasma levels in a variety of clinical conditions. We evaluated a novel dabigatran measure - the 'dilute Russell viper venom confirm (DRVVC) assay' - against current developmental assays and a reference method. We measured plasma dabigatran and compared results from the Stago Sta-Clot DRVVC assay, Stago Ecarin Chromogenic Assay, Biophen Hemoclot Thrombin Inhibitor, and liquid chromatography tandem mass spectrometry. We obtained dabigatran calibrators and controls from Biophen, and performed the coagulation assays using a Stago STA-R Evolution coagulometer. Liquid chromatography tandem mass spectrometry method specimens were performed on an AB Sciex instrument at LabCorp. We enrolled 97 anticoagulation clinic patients (mean age 76 years) who were taking 150 mg dabigatran twice daily. All had creatinine clearances above 30 ml/min; patients were not excluded for concurrent medications or health issues. Citrated blood specimens were processed immediately, and stored at -70°C. We did not correlate collection time with medication time. We employed descriptive statistics, analysis of variance, and the Bland-Altman difference plot to assess the data. The range for all assays was 11.6-917 ng/ml. Analysis of variance generated a P value of 0.1 and Bland-Altman differences were all below 4.0% compared with DRVVC. The DRVVC measures dabigatran with validity comparable to other methods.

  10. Evaluation of three selective chromogenic media, CHROMagar ESBL, CHROMagar CTX-M and CHROMagar KPC, for the detection of Klebsiella pneumoniae producing OXA-48 carbapenemase. (United States)

    Hornsey, Michael; Phee, Lynette; Woodford, Neil; Turton, Jane; Meunier, Daniele; Thomas, Claire; Wareham, David W


    Three selective chromogenic culture media (CHROMagars ESBL, CTX-M and KPC) were evaluated for their ability to support the growth of nine Klebsiella pneumoniae isolates producing OXA-48 carbapenemase in combination with other β-lactamases. CHROMagar ESBL and CHROMagar KPC were the most sensitive media, supporting growth of all isolates with a detection limit as low as  10(6) CFU/ml. Both CHROMagar ESBL and CHROMagar KPC may be useful for enhanced isolation of K pneumoniae producing OXA-48-like carbapenemases.

  11. Detection of Listeria monocytogenes by direct colony hybridization on hydrophobic grid-membrane filters by using a chromogen-labeled DNA probe. (United States)

    Peterkin, P I; Idziak, E S; Sharpe, A N


    A DNA probe specific for Listeria monocytogenes was isolated from a beta-hemolytic recombinant clone of an L. monocytogenes gene bank. It was labeled with horseradish peroxidase and used in a direct colony hybridization method on hydrophobic grid-membrane filters for the detection of the organism. Following color development of the chromogen, a commercial counter (HGMF Interpreter) was able to detect and count the organisms electronically. The method gave a positive reaction with 70 L. monocytogenes strains, while showing a negative reaction with 10 strains of other Listeria spp. and with 20 organisms of other genera.

  12. Ouroboros - Playing A Biochemical

    Directory of Open Access Journals (Sweden)

    D. T. Rodrigues


    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

  13. Enzyme and biochemical producing fungi

    DEFF Research Database (Denmark)

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


    factories for sustainable production of important molecules. For developing fungi into efficient cell factories, the project includes identification of important factors that control the flux through the pathways using metabolic flux analysis and metabolic engineering of biochemical pathways....

  14. Mathematical Models of Biochemical Oscillations


    Conrad, Emery David


    The goal of this paper is to explain the mathematics involved in modeling biochemical oscillations. We first discuss several important biochemical concepts fundamental to the construction of descriptive mathematical models. We review the basic theory of differential equations and stability analysis as it relates to two-variable models exhibiting oscillatory behavior. The importance of the Hopf Bifurcation will be discussed in detail for the central role it plays in limit cycle behavior and...

  15. Biochemical and behavioral effects of long-term citalopram administration and discontinuation in rats Role of serotonin synthesis

    NARCIS (Netherlands)

    Bosker, Fokko J.; Tanke, Marit A. C.; Jongsma, Minke E.; Cremers, Thomas I. F. H.; Jagtman, Evelien; Pietersen, Charmaine Y.; van der Hart, Marieke G. C.; Gladkevich, Anatoliy V.; Kema, Ido P.; Westerink, Ben H. C.; Korf, Jakob; den Boer, Johan A.


    We have investigated effects of continuous SSRI administration and abrupt discontinuation on biochemical and behavioral indices of rat brain serotonin function and attempted to identify underlying mechanisms Biochemistry of serotonin was assessed with brain tissue assays and microdialysis behavior w

  16. A rapid kinetic chromogenic method for quantification of bacterial endotoxins in lyophilized reagents for labeling with {sup 99m}Tc radiopharmaceuticals

    Energy Technology Data Exchange (ETDEWEB)

    Fukumori, Neuza T.O.; Campos, Domingos G.; Silva, Laercio; Fernandes, Adriana V.; Mengatti, Jair; Silva, Constancia P.G.; Matsuda, Margareth M.N. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)


    A rapid quantitative kinetic chromogenic test in an automated Portable Test System (PTS) has been developed for determination of bacterial endotoxins in water, in-process and end-products using the Limulus amebocyte lysate (LAL). The aim of this work was to validate the method for lyophilized reagents for labeling with {sup 99m}Tc radiopharmaceuticals with no interfering factors. Experiments were performed in three consecutive batches of the lyophilized reagents Methylenediphosphonic Acid (MDP) and Pyrophosphate (PYRO) produced at IPEN-CNEN/ SP using the PTS from Endosafe, Inc.{sup TM}, Charleston, SC. The Maximum Valid Dilution (MVD) was calculated to establish the extent of dilution to avoid interfering test conditions (MVD=500). Better results were obtained above 1:20 dilution factor for MDP and 1:100 for PYRO. The parameters of coefficient correlation (R) -0.980, RPPC between 50 - 200% and coefficient variation (CV) of the samples less than 25% were satisfied and the endotoxin concentration was lower than the lowest concentration of the standard curve (0.05 EU mL{sup -1}), therefore less than the established limit in pharmacopoeias. The PTS is a rapid, simple and accurate technique using the quantitative kinetic chromogenic method for bacterial endotoxin determination. For this reason, it is very practical in the radiopharmaceutical area and it trends to be the method of choice for the pyrogen test. For MDP and PYRO, the validation was successfully performed. (author)

  17. Application of 2-Trichloromethylbenzimidazole in Analytical Chemistry: A Highly Selective Chromogenic Reagent for Thin-Layer Chromatography and Some Other Analytical Uses

    Directory of Open Access Journals (Sweden)

    Leszek Konopski


    Full Text Available 2-Trichloromethylbenzimidazole (TCMB was used as a chromogenic reagent in organic or inorganic analysis, mainly in thin-layer chromatography (TLC. In reactions of TCMB with some heteroaromatic nitrogen containing compounds, such as azines, azoles and benzazoles, a formation of high colored products occurred. For azines, the chromogenic reaction was highly regioselective, since the both adjacent α-positions versus the nitrogen atom(s must not be substituted. A TLC method of detection was developed. Thirty azines, azoles, and benzazoles were detected at the detection limit 10 ng to 1 μg. This method was also applied for detection of heteroaromatic pesticides, and the attempts to construct active and passive dosimeters for nicotine were made. In a prechromatographic reaction of aromatic o-diamines with methyl trichloroacetimidate, TCMB or its derivatives were formed in situ. Followed by TLC and visualization in pyridine vapors, this procedure was applied for detection of o-phenylenediamine derivatives. The reaction product of TCMB and pyridine (LI Complex was identified and fully characterized. Two different reaction mechanisms: with electron deficient basic heteroaromatic compounds, like pyridine, and with more acidic compounds, for example, pyrrole, were discussed. In aqueous solutions, the LI Complex may be also used as a new indicator for complexometric, adsorption and acid-base titration of inorganic compounds.

  18. Performance of the chromID Salmonella Elite chromogenic agar in comparison with CHROMagar™ Salmonella, Oxoid™ Brilliance™ Salmonella and Hektoen agars for the isolation of Salmonella from stool specimens. (United States)

    Martiny, Delphine; Dediste, Anne; Anglade, Claire; Vlaes, Linda; Moens, Catherine; Mohamed, Souad; Vandenberg, Olivier


    chromID™ Salmonella Elite is compared with 3 culture media commonly used for Salmonella isolation from stool specimens. As results were equivalent to other chromogenic media (100% sensitivity, 98% specificity), only financial arguments should guide the choice for a medium with respect to another.

  19. BEST: Biochemical Engineering Simulation Technology

    Energy Technology Data Exchange (ETDEWEB)


    The idea of developing a process simulator that can describe biochemical engineering (a relatively new technology area) was formulated at the National Renewable Energy Laboratory (NREL) during the late 1980s. The initial plan was to build a consortium of industrial and U.S. Department of Energy (DOE) partners to enhance a commercial simulator with biochemical unit operations. DOE supported this effort; however, before the consortium was established, the process simulator industry changed considerably. Work on the first phase of implementing various fermentation reactors into the chemical process simulator, ASPEN/SP-BEST, is complete. This report will focus on those developments. Simulation Sciences, Inc. (SimSci) no longer supports ASPEN/SP, and Aspen Technology, Inc. (AspenTech) has developed an add-on to its ASPEN PLUS (also called BioProcess Simulator [BPS]). This report will also explain the similarities and differences between BEST and BPS. ASPEN, developed by the Massachusetts Institute of Technology for DOE in the late 1970s, is still the state-of-the-art chemical process simulator. It was selected as the only simulator with the potential to be easily expanded into the biochemical area. ASPEN/SP, commercially sold by SimSci, was selected for the BEST work. SimSci completed work on batch, fed-batch, and continuous fermentation reactors in 1993, just as it announced it would no longer commercially support the complete ASPEN/SP product. BEST was left without a basic support program. Luckily, during this same time frame, AspenTech was developing a biochemical simulator with its version of ASPEN (ASPEN PLUS), which incorporates most BEST concepts. The future of BEST will involve developing physical property data and models appropriate to biochemical systems that are necessary for good biochemical process design.

  20. Evaluation of chromogenic medium and direct latex agglutination test for detection of group B streptococcus in vaginal specimens from pregnant women in Lebanon and Kuwait. (United States)

    Ghaddar, Nahed; Alfouzan, Wadha; Anastasiadis, Elie; Al Jiser, Tamima; Itani, Saad Eddine; Dernaika, Racha; Eid, Toufic; Ghaddar, Ali; Charafeddine, Adib; Dhar, Rita; El Hajj, Hiba


    This study was undertaken to evaluate chromogenic medium and a direct latex agglutination test (DLA) for detection of Group B Streptococcus (GBS) in the vaginal specimens of pregnant women, and to ascertain the prevalence of GBS in this population in Kuwait and Lebanon. Vaginal swabs, collected from women at 35-37 weeks of gestation, were cultured on 5 % sheep blood agar (SBA), colistin nalidixic acid agar (CNA), Strept B Select chromogenic agar (SBS) as well as Lim enrichment broth in 168 cases in Lebanon while only SBA was used for 1391 samples in Kuwait. In addition, vaginal samples from 102 GBS-positive and 20 GBS-negative women near the time of delivery were collected in Kuwait for evaluation of the DLA test. During the study period, the prevalence of GBS colonization was determined to be 20.7 % (288/1391) in Kuwait while 18.4 % (31) of 168 pregnant women in Lebanon had vaginal cultures positive for GBS. By direct plating of vaginal swabs on the three media used, the isolation rates of GBS were 51.6, 64.5 and 77.4 % on SBA, CNA and SBS, respectively, which increased to 90.35, 93.1 and 96.8 %, respectively, following subculture in Lim broth after 18 h of incubation. The sensitivity of the DLA test was found to be dependent on the density of GBS colonization, resulting in 100 % sensitivity and 100 % specificity for heavy (>10(2) c.f.u. per swab) and moderately heavy (50-100 c.f.u. per swab) growth of GBS. However, for vaginal specimens yielding <50 c.f.u. per swab, the sensitivity, specificity, positive and negative predictive values of the DLA test were 100, 55.5, 63.6 and 100 %, respectively. In conclusion, a chromogenic agar, such as SBS, and a DLA test can be used for rapid detection of GBS in pregnant women. The DLA test, in particular, could prove to be a useful tool for immediate detection of GBS in women near delivery so that intrapartum antibiotic prophylaxis can be initiated.

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

    DEFF Research Database (Denmark)

    Tørring, Niels


    Background Screening for fetal trisomy 21 in the first trimester includes analysis of the serological markers pregnancy-associated plasma protein A (PAPP-A) and free beta human choriogonadotropin (free βhCG). With the recent launch of the PAPP-A free βhCG and assays on the Roche Cobas and Elecsys...... platforms, we investigated their clinical and analytical performance in samples from gestaional weeks 8+0 to 14+0. Methods. We conducted a multicenter study based on serum samples from 5397 pregnancies including 107 samples from cases of verified fetal trisomy 21 at 8 to 14 weeks of gestation. A technical...... 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....

  2. Thin-Film Transistor-Based Biosensors for Determining Stoichiometry of Biochemical Reactions (United States)

    Wang, Yi-Wen; Chen, Ting-Yang; Yang, Tsung-Han; Chang, Cheng-Chung; Yang, Tsung-Lin; Lo, Yu-Hwa


    The enzyme kinetic in a biochemical reaction is critical to scientific research and drug discovery but can hardly be determined experimentally from enzyme assays. In this work, a charge-current transducer (a transistor) is proposed to evaluate the status of biochemical reaction by monitoring the electrical charge changes. Using the malate-aspartate shuttle as an example, a thin-film transistor (TFT)-based biosensor with an extended gold pad is demonstrated to detect the biochemical reaction between NADH and NAD+. The drain current change indicates the status of chemical equilibrium and stoichiometry. PMID:28033412

  3. Microbead agglutination based assays

    KAUST Repository

    Kodzius, Rimantas


    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.

  4. Absolute nuclear material assay (United States)

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


    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.

  5. Utilization of the chromogenic agar MRSA ID in the surveillance of methicillin-resistant Staphylococcus aureus in a low endemic unit for MRSA

    Directory of Open Access Journals (Sweden)

    Laura Squarzon


    Full Text Available Methicillin-resistant Staphylococcus aureus (MRSA is one of the most common pathogen responsible for nosocomial infections.Approximately 20% of patients undergoing surgical operations acquires at least one nosocomial infection. Nasal carriers of S. aureus have been identified as one of the populations at major risk to develop this type of infection, particularly, after operations, dialysis, transplants and admission in intensive care unit. Our study was conducted on patients who came from a Neurosurgery Intensive Care Unit, where the incidence of MRSA is very low (8% and where it is very important to identify MRSA carriers in order to adopt measures to avoid transmission.The use of chromogenic agar MRSA ID allowed us to identify green colonies of MRSA within 24 hours and to make a timely and careful diagnosis.

  6. [Detection of TDH-producing Vibrio parahaemolyticus O3:K6 from naturally contaminated shellfish using an immunomagnetic separation method and chromogenic agar medium]. (United States)

    Hara-Kudo, Y; Sugiyama, K; Nishina, T; Saitoh, A; Nakagawa, H; Ichihara, T; Konuma, H; Hasegawa, J; Kumagai, S


    We attempted to isolate TDH-producing Vibrio parahaemolyticus O3:K6 from shellfish. Asari samples were incubated with TSB supplemented with 2% (w/v) NaCl for 6 h, and then the 6-h cultures were incubated with salt polymyxin broth for 18 h. After the two-step enrichment, a 1 ml portion of the culture was treated with magnetic beads coated with K6 antibody for immunoconcentration of V. parahaemolyticus O3:K6. The immunoconcentrated and untreated cultures were plated onto a chromogenic agar and TCBS agar media for isolation of V. parahaemolyticus. TDH-producing V. parahaemolyticus O3:K6 was isolated from 3 out of 66 lots (4.5%) of naturally contaminated Asari. Six of 4,265 colonies suspected as V. parahaemolyticus (0.14%) were TDH-producing V. parahaemolyticus O3:K6.

  7. Prospective evaluation of the chromogenic medium CandiSelect 4 for differentiation and presumptive identification of non-Candida albicans Candida species. (United States)

    Zhao, Liang; de Hoog, G Sybren; Cornelissen, Akke; Lyu, Qian; Mou, Lili; Liu, Taohua; Cao, Yu; Vatanshenassan, Mansoureh; Kang, Yingqian


    Rapid identification of pathogenic yeasts is a crucial step in timely and appropriate antifungal therapy. For diagnostics in the clinical laboratory, simplified alternatives to barcoding are needed. CandiSelect 4 (CS4) medium, a chromogenic medium for isolation of clinical yeasts, allows routine recognition of Candida albicans and presumptive identification of Candida tropicalis, Candida glabrata, and Candida krusei. We evaluated an extension of this method with 46 non-Candida albicans Candida (NCAC) and 7 Malassezia species. The medium supported growth of all species tested and a wide diversity of cultural types were observed. Colony colours were in violet, turquoise (including green and blue), or white tinges. Eight NCAC species produced violet pigmentation similar to that of C. albicans. Most NCAC species, including C. glabrata and C. tropicalis were distributed in the turquoise group. Malassezia species were invariably blue.

  8. Changes in the chromogen properties of the betalaine induced by gamma radiation; Cambios en las propiedades cromogenas de las betalainas inducidos por radiacion gamma

    Energy Technology Data Exchange (ETDEWEB)

    Rodriguez N, S.; Pinedo S, A.; Amador V, P.; Chacon R, A.; Arcos P, A.; Vega C, H.R. [Unidad Academica de Ciencias Nucleares, C. Cipres 10, Fracc. La Penuela, 98068 Zacatecas (Mexico)


    The changes of coloration of four natural extracts in function of the absorbed dose produced by a gamma rays source of {sup 137} Cs were determined. The natural extracts used contain betalaines that are natural pigments of some plants as the beet about that their name. They are also in abundant form in the fruits (tunas) of some species of the Opuntia genus. The extracts were obtained by maceration, starting from beet and three tuna varieties, and they were stabilized to pH 5.5 The change of coloration it was determined in a visible ultraviolet spectrophotometer by means of the absorbance from the samples to photons of 475, 535 and 600 nm of wavelength. The absorbance to different intervals of time was measured. The relationship between the absorbed dose and the chromogene properties of the pigment, with the intention of using it as possible dosemeter was settled down. (Author)

  9. Associative learning in biochemical networks


    Ghandi, Nikhil; Ashkenasy, Gonen; Tannenbaum, Emmanuel


    We develop a simple, chemostat-based model illustrating how a process analogous to associative learning can occur in a biochemical network. Associative learning is a form of learning whereby a system "learns" to associate two stimuli with one another. In our model, two types of replicating molecules, denoted A and B, are present in some initial concentration in the chemostat. Molecules A and B are stimulated to replicate by some growth factors, denoted GA and GB, respectively. It is also assu...

  10. Simultaneous Detection of Antigen-Specific IgG- and IgM-Secreting Cells with a B Cell Fluorospot Assay

    Directory of Open Access Journals (Sweden)

    M. Anthony Moody


    Full Text Available The traditional enzyme-linked immunospot (ELISpot assay is the gold standard for the enumeration of antigen-specific B cells. Since B cell availability from biological samples is often limited, either because of sample size/volume or the need of performing multiple analyses on the same sample, the implementation of ELISpot assay formats that allow the simultaneous detection of multiple antibody types is desirable. While dual-color ELISpot assays have been described, technical complexities have so far prevented their wide utilization as well as further expansion of their multicolor capability. An attractive solution is to replace the chromogenic reaction of the traditional ELISpot assay with a fluorescent detection system (fluorospot assay. Fluorospot assays using fluorophore-conjugated secondary antibodies in conjunction with fluorescence enhancers, FITC/anti-FITC and biotin/avidin amplification systems and dedicated equipment for spot detection have been developed to enumerate T-cells secreting two or three specific cytokines and, more recently, IgG and IgA antibody-secreting cells (ASCs. We hereby report a method for a multiplex B cell fluorospot assay that utilizes quantum-dot nanocrystals as reporters without further amplification systems or need of dedicated equipment. With this method we simultaneously enumerated HIV-1 gp41 envelope glycoprotein-specific IgG and IgM antibody-secreting cells with sensitivity comparable to that of the traditional ELISpot assay.

  11. Biochemical adaptation to ocean acidification. (United States)

    Stillman, Jonathon H; Paganini, Adam W


    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. Transgenic Animal Mutation Assays

    Institute of Scientific and Technical Information of China (English)

    Tao Chen; Ph.D.D.A.B.T.


    @@ The novel transgenic mouse and rat mutation assays have provided a tool for analyzing in vivo mutation in any tissue, thus permitting the direct comparison of cancer incidence with mutant frequency.

  13. Tube-Forming Assays. (United States)

    Brown, Ryan M; Meah, Christopher J; Heath, Victoria L; Styles, Iain B; Bicknell, Roy


    Angiogenesis involves the generation of new blood vessels from the existing vasculature and is dependent on many growth factors and signaling events. In vivo angiogenesis is dynamic and complex, meaning assays are commonly utilized to explore specific targets for research into this area. Tube-forming assays offer an excellent overview of the molecular processes in angiogenesis. The Matrigel tube forming assay is a simple-to-implement but powerful tool for identifying biomolecules involved in angiogenesis. A detailed experimental protocol on the implementation of the assay is described in conjunction with an in-depth review of methods that can be applied to the analysis of the tube formation. In addition, an ImageJ plug-in is presented which allows automatic quantification of tube images reducing analysis times while removing user bias and subjectivity.

  14. Cell viability assays: introduction. (United States)

    Stoddart, Martin J


    The measurement of cell viability plays a fundamental role in all forms of cell culture. Sometimes it is the main purpose of the experiment, such as in toxicity assays. Alternatively, cell viability can be used to -correlate cell behaviour to cell number, providing a more accurate picture of, for example, anabolic -activity. There are wide arrays of cell viability methods which range from the most routine trypan blue dye exclusion assay to highly complex analysis of individual cells, such as using RAMAN microscopy. The cost, speed, and complexity of equipment required will all play a role in determining the assay used. This chapter aims to provide an overview of many of the assays available today.

  15. Near infrared probes for biochemical, cellular, and whole animal analysis of disease processes (United States)

    Kovar, Joy; Boveia, Vince; Chen, Huaxian; Peng, Xinzhan; Skopp, Rose; Little, Garrick; Draney, Dan; Olive, D. M.


    The study of disease processes requires a number of tools for detection of proteins and biomarkers in cell and animal based assays. Near infrared (NIR) technologies offer the advantage of high signal without interference from background producing factors such as tissues, blood, or plastics. NIR fluorescence quenching biochemical assays employing a novel NIR quencher are homogeneous and sensitive. NIR-based immunocytochemical assays offer a means of quantitatively evaluating cell signaling pathways. The technology can be extended to the development of targeted molecular imaging agents for disease analysis in animal models. We describe here model assays for each of these categories. A fluorescence quenching caspase-3 assay was developed employing a novel, broadly applicable quencher dye suitable for use with both visible and NIR dye chemistries. An NIR cell based assay is described for assessment of phosphorylation of p53 in response to a cellular stimulus. Finally, we describe the development and application of a targeted NIR optical imaging agent for monitoring tumor growth in whole animals. The NIR biochemical and cell based assays are robust with Z' factors greater than 0.7. The use of an IRDye (R)800CW-labeled cyclic RGD peptide is presented as a model for development and application of targeted imaging agents. NIR technologies are compatible with the complete spectrum of assay needs for disease analysis and therapeutic development.

  16. New Rapid Spore Assay (United States)

    Kminek, Gerhard; Conley, Catharine


    The presentation will detail approved Planetary Protection specifications for the Rapid Spore Assay for spacecraft components and subsystems. Outlined will be the research and studies on which the specifications were based. The research, funded by ESA and NASA/JPL, was conducted over a period of two years and was followed by limited cleanroom studies to assess the feasibility of this assay during spacecraft assembly.

  17. Biochemical structure of Calendula officinalis. (United States)

    Korakhashvili, A; Kacharava, T; Kiknavelidze, N


    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.

  18. Biochemical aspects of Huntington's chorea. (United States)

    Caraceni, T; Calderini, G; Consolazione, A; Riva, E; Algeri, S; Girotti, F; Spreafico, R; Branciforti, A; Dall'olio, A; Morselli, P L


    Fifteen patients affected by Huntington's chorea were divided into two groups, 'slow' and 'fast', according to IQ scores on the Wechsler-Bellevue scale, and scores on some motor performance tests. A possible correlation was looked for between some biochemical data (cerebrospinal fluid (CSF), homovanillic acid (HVA), and 5-hydroxyindolacetic acid (5HIAA) levels, plasma dopamine-beta-hydroxylase (DBH), dopamine (DA) uptake by platelets), and clinical data (duration of illness, severity of symptoms, age of patients, IQ scores, 'slow' and 'fast' groups). The CSF, HVA, and 5HIAA levels were found to be significantly lowered in comparison with normal controls. DBH activity and DA uptake by platelets did not differ significantly from normal subjects. Treatment with haloperidol in all patients and with dipropylacetic acid in three patients did not appear to modify the CSF, HVA, and 5HIAA concentrations, the plasma DBH activity, or the DA uptake. There were no significant differences in the CSF, HVA, and 5HIAA contents between the two groups of patients, and there was no correlation between biochemical data and clinical features. PMID:143508

  19. Biochemical abnormalities in Pearson syndrome. (United States)

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


    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.

  20. Vector Encoding in Biochemical Networks (United States)

    Potter, Garrett; Sun, Bo

    Encoding of environmental cues via biochemical signaling pathways is of vital importance in the transmission of information for cells in a network. The current literature assumes a single cell state is used to encode information, however, recent research suggests the optimal strategy utilizes a vector of cell states sampled at various time points. To elucidate the optimal sampling strategy for vector encoding, we take an information theoretic approach and determine the mutual information of the calcium signaling dynamics obtained from fibroblast cells perturbed with different concentrations of ATP. Specifically, we analyze the sampling strategies under the cases of fixed and non-fixed vector dimension as well as the efficiency of these strategies. Our results show that sampling with greater frequency is optimal in the case of non-fixed vector dimension but that, in general, a lower sampling frequency is best from both a fixed vector dimension and efficiency standpoint. Further, we find the use of a simple modified Ornstein-Uhlenbeck process as a model qualitatively captures many of our experimental results suggesting that sampling in biochemical networks is based on a few basic components.

  1. Mycobacterium pseudoshottsii sp. nov., a slowly growing chromogenic species isolated from Chesapeake Bay striped bass (Morone saxatilis) (United States)

    Rhodes, M.W.; Kator, H.; McNabb, A.; Deshayes, C.; Reyrat, J.-M.; Brown-Elliott, B. A.; Wallace, R.; Trott, K.A.; Parker, J.M.; Lifland, B.; Osterhout, G.; Kaattari, I.; Reece, K.; Vogelbein, W.; Ottinger, C.A.


    A group of slowly growing photochromogenic mycobacteria was isolated from Chesapeake Bay striped bass (Morone saxatilis) during an epizootic of mycobacteriosis. Growth characteristics, acid-fastness and 16S rRNA gene sequencing results were consistent with those of the genus Mycobacterium. Biochemical reactions, growth characteristics and mycolic acid profiles (HPLC) resembled those of Mycobacterium shottsii, a non-pigmented mycobacterium also isolated during the same epizootic. Sequencing of the 16S rRNA genes, the gene encoding the exported repeated protein (erp) and the gene encoding the 65 kDa heat-shock protein (hsp65) and restriction enzyme analysis of the hsp65 gene demonstrated that this group of isolates is unique. Insertion sequences associated with Mycobacterium ulcerans, IS2404 and IS2606, were detected by PCR. These isolates could be differentiated from other slowly growing pigmented mycobacteria by their inability to grow at 37 ??C, production of niacin and urease, absence of nitrate reductase, negative Tween 80 hydrolysis and resistance to isoniazid (1 ??g ml-1), p-nitrobenzoic acid, thiacetazone and thiophene-2-carboxylic hydrazide. On the basis of this polyphasic study, it is proposed that these isolates represent a novel species, Mycobacterium pseudoshottsii sp. nov. The type strain, L15T, has been deposited in the American Type Culture Collection as ATCC BAA-883T and the National Collection of Type Cultures (UK) as NCTC 13318T. ?? 2005 IUMS.

  2. Against vaccine assay secrecy. (United States)

    Herder, Matthew; Hatchette, Todd F; Halperin, Scott A; Langley, Joanne M


    Increasing the transparency of the evidence base behind health interventions such as pharmaceuticals, biologics, and medical devices, has become a major point of critique, conflict, and policy focus in recent years. Yet the lack of publicly available information regarding the immunogenicity assays upon which many important, widely used vaccines are based has received no attention to date. In this paper we draw attention to this critical public health problem by reporting on our efforts to secure vaccine assay information in respect of 10 vaccines through Canada's access to information law. We argue, under Canadian law, that the public health interest in having access to the methods for these laboratory procedures should override claims by vaccine manufacturers and regulators that this information is proprietary; and, we call upon several actors to take steps to ensure greater transparency with respect to vaccine assays, including regulators, private firms, researchers, research institutions, research funders, and journal editors.

  3. Against vaccine assay secrecy (United States)

    Herder, Matthew; Hatchette, Todd F; Halperin, Scott A; Langley, Joanne M


    Increasing the transparency of the evidence base behind health interventions such as pharmaceuticals, biologics, and medical devices, has become a major point of critique, conflict, and policy focus in recent years. Yet the lack of publicly available information regarding the immunogenicity assays upon which many important, widely used vaccines are based has received no attention to date. In this paper we draw attention to this critical public health problem by reporting on our efforts to secure vaccine assay information in respect of 10 vaccines through Canada's access to information law. We argue, under Canadian law, that the public health interest in having access to the methods for these laboratory procedures should override claims by vaccine manufacturers and regulators that this information is proprietary; and, we call upon several actors to take steps to ensure greater transparency with respect to vaccine assays, including regulators, private firms, researchers, research institutions, research funders, and journal editors. PMID:25826194

  4. Rover waste assay system

    Energy Technology Data Exchange (ETDEWEB)

    Akers, D.W.; Stoots, C.M.; Kraft, N.C.; Marts, D.J. [Idaho National Engineering Lab., Idaho Falls, ID (United States)


    The Rover Waste Assay System (RWAS) is a nondestructive assay system designed for the rapid assay of highly-enriched {sup 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 {sup 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.

  5. Assays for calcitonin receptors

    Energy Technology Data Exchange (ETDEWEB)

    Teitelbaum, A.P.; Nissenson, R.A.; Arnaud, C.D.


    The assays for calcitonin receptors described focus on their use in the study of the well-established target organs for calcitonin, bone and kidney. The radioligand used in virtually all calcitonin binding studies is /sup 125/I-labelled salmon calcitonin. The lack of methionine residues in this peptide permits the use of chloramine-T for the iodination reaction. Binding assays are described for intact bone, skeletal plasma membranes, renal plasma membranes, and primary kidney cell cultures of rats. Studies on calcitonin metabolism in laboratory animals and regulation of calcitonin receptors are reviewed.

  6. CTL ELISPOT assay. (United States)

    Ranieri, Elena; Popescu, Iulia; Gigante, Margherita


    Enzyme-linked immune absorbent spot (Elispot) is a quantitative method for measuring relevant parameters of T cell activation. The sensitivity of Elispot allows the detection of low-frequency antigen-specific T cells that secrete cytokines and effector molecules, such as granzyme B and perforin. Cytotoxic T cell (CTL) studies have taken advantage with this high-throughput technology by providing insights into quantity and immune kinetics. Accuracy, sensitivity, reproducibility, and robustness of Elispot resulted in a wide range of applications in research as well as in diagnostic field. Actually, CTL monitoring by Elispot is a gold standard for the evaluation of antigen-specific T cell immunity in clinical trials and vaccine candidates where the ability to detect rare antigen-specific T cells is of relevance for immune diagnostic. The most utilized Elispot assay is the interferon-gamma (IFN-γ) test, a marker for CD8(+) CTL activation, but Elispot can also be used to distinguish different subsets of activated T cells by using other cytokines such as T-helper (Th) 1-type cells (characterized by the production of IFN-γ, IL-2, IL-6, IL-12, IL-21, and TNF-α), Th2 (producing cytokines like IL-4, IL-5, IL-10, and IL-13), and Th17 (IL-17) cells. The reliability of Elispot-generated data, by the evaluation of T cell frequency recognizing individual antigen/peptide, is the core of this method currently applied widely to investigate specific immune responses in cancer, infections, allergies, and autoimmune diseases. The Elispot assay is competing with other methods measuring single-cell cytokine production, e.g., intracellular cytokine by FACS or Miltenyi cytokine secretion assay. Other types of lymphocyte frequency and function assays include limiting dilution assay (LDA), cytotoxic T cell assay (CTL), and tetramer staining. Compared with respect to sensitivity the Elispot assay is outranking other methods to define frequency of antigen-specific lymphocytes. The method

  7. Microfluidic assay without blocking for rapid HIV screening and confirmation. (United States)

    Song, Lusheng; Zhang, Yi; Wang, Wenjun; Ma, Liying; Liu, Yong; Hao, Yanlin; Shao, Yiming; Zhang, Wei; Jiang, Xingyu


    The essential step for HIV spreading limitation is the screening tests. However, there are multiple disadvantages in current screening assays which need further confirmation test. Herein we developed a rapid HIV assay combining screening and confirmation test by using the microfluidic network assay. Meanwhile, the assay is accelerated by bypassing the step of blocking. We call this method as microfluidic assay without blocking (MAWB). Both the limit of detection and reagent incubation time of MAWB are determined by screening of one model protein pair: ovalbumin and its antibody. The assay time is accelerated about 25% while the limit of detection (LOD) is well kept. Formatting the method in for both HIV screening (testing 8 HIV-related samples) and confirmation (assaying 6 kinds of HIV antibodies of each sample) within 30 min was successful. Fast HIV screening and confirmation of 20 plasma samples were also demonstrated by this method. MAWB improved the assay speed while keeping the LOD of conventional ELISA. Meanwhile, both the accuracy and throughput of MAWB were well improved, which made it an excellent candidate for a quick HIV test for both screening and confirmation. Methods like this one will find wide applications in clinical diagnosis and biochemical analysis based on the interactions between pairs of molecules.

  8. Nonlinear Biochemical Signal Processing via Noise Propagation


    Kim, Kyung Hyuk; Qian, Hong; Sauro, Herbert M.


    Single-cell studies often show significant phenotypic variability due to the stochastic nature of intra-cellular biochemical reactions. When the numbers of molecules, e.g., transcription factors and regulatory enzymes, are in low abundance, fluctuations in biochemical activities become significant and such "noise" can propagate through regulatory cascades in terms of biochemical reaction networks. Here we develop an intuitive, yet fully quantitative method for analyzing how noise affects cell...

  9. A UK NEQAS ISH multicenter ring study using the Ventana HER2 dual-color ISH assay.

    LENUS (Irish Health Repository)

    Bartlett, J M S


    We performed a multicenter assessment of a new HER2 dual-color chromogenic in situ hybridization (CISH) test and herein report on concordance of CISH data with fluorescence in situ hybridization (FISH) data and intraobserver and interlaboratory scoring consistency. HER2 results were evaluated using duplicate cores from 30 breast cancers in 5 laboratories using the Ventana HER2 dual-color ISH assay (Ventana Medical Systems, Cambridgeshire, England) and in 1 central laboratory using a standard FISH assay. Overall 93.3% of cases were successfully analyzed by CISH across the 5 participating laboratories. There was excellent concordance (98.0% overall) for diagnosis of HER2 amplification by CISH compared with FISH. Intraobserver variability (7.7%) and intersite variability (9.1%) of absolute HER2\\/chromosome enumeration probe 17 ratios were tightly controlled across all participating laboratories. The Ventana HER2 dual-color ISH assay is robust and reproducible, shows good concordance with a standard FISH assay, and complies with requirements in national and international guidelines for performance of ISH-based diagnostic tests.

  10. A UK NEQAS ISH multicenter ring study using the Ventana HER2 dual-color ISH assay. (United States)

    Bartlett, J M S; Campbell, Fiona M; Ibrahim, Merdol; O'Grady, Anthony; Kay, Elaine; Faulkes, Catherine; Collins, Nadine; Starczynski, Jane; Morgan, John M; Jasani, Bharat; Miller, Keith


    We performed a multicenter assessment of a new HER2 dual-color chromogenic in situ hybridization (CISH) test and herein report on concordance of CISH data with fluorescence in situ hybridization (FISH) data and intraobserver and interlaboratory scoring consistency. HER2 results were evaluated using duplicate cores from 30 breast cancers in 5 laboratories using the Ventana HER2 dual-color ISH assay (Ventana Medical Systems, Cambridgeshire, England) and in 1 central laboratory using a standard FISH assay. Overall 93.3% of cases were successfully analyzed by CISH across the 5 participating laboratories. There was excellent concordance (98.0% overall) for diagnosis of HER2 amplification by CISH compared with FISH. Intraobserver variability (7.7%) and intersite variability (9.1%) of absolute HER2/chromosome enumeration probe 17 ratios were tightly controlled across all participating laboratories. The Ventana HER2 dual-color ISH assay is robust and reproducible, shows good concordance with a standard FISH assay, and complies with requirements in national and international guidelines for performance of ISH-based diagnostic tests.

  11. Determination of rivaroxaban in patient’s plasma samples by anti-Xa chromogenic test associated to High Performance Liquid Chromatography tandem Mass Spectrometry (HPLC-MS/MS) (United States)

    Derogis, Priscilla Bento Matos; Sanches, Livia Rentas; de Aranda, Valdir Fernandes; Colombini, Marjorie Paris; Mangueira, Cristóvão Luis Pitangueira; Katz, Marcelo; Faulhaber, Adriana Caschera Leme; Mendes, Claudio Ernesto Albers; Ferreira, Carlos Eduardo dos Santos; França, Carolina Nunes; Guerra, João Carlos de Campos


    Rivaroxaban is an oral direct factor Xa inhibitor, therapeutically indicated in the treatment of thromboembolic diseases. As other new oral anticoagulants, routine monitoring of rivaroxaban is not necessary, but important in some clinical circumstances. In our study a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was validated to measure rivaroxaban plasmatic concentration. Our method used a simple sample preparation, protein precipitation, and a fast chromatographic run. It was developed a precise and accurate method, with a linear range from 2 to 500 ng/mL, and a lower limit of quantification of 4 pg on column. The new method was compared to a reference method (anti-factor Xa activity) and both presented a good correlation (r = 0.98, p < 0.001). In addition, we validated hemolytic, icteric or lipemic plasma samples for rivaroxaban measurement by HPLC-MS/MS without interferences. The chromogenic and HPLC-MS/MS methods were highly correlated and should be used as clinical tools for drug monitoring. The method was applied successfully in a group of 49 real-life patients, which allowed an accurate determination of rivaroxaban in peak and trough levels. PMID:28170419

  12. A Selective Chromogenic Plate, YECA, for the Detection of Pathogenic Yersinia enterocolitica: Specificity, Sensitivity, and Capacity to Detect Pathogenic Y. enterocolitica from Pig Tonsils

    Directory of Open Access Journals (Sweden)

    M. Denis


    Full Text Available A new selective chromogenic plate, YECA, was tested for its specificity, sensitivity, and accuracy to detect pathogenic Y. enterocolitica from pig tonsils. We tested a panel of 26 bacterial strains on YECA and compared it to PCA, CIN, and YeCM media. Detection of pathogenic Y. enterocolitica was carried out on 50 pig tonsils collected in one slaughter house. Enrichment was done in PSB and ITC broths. Streaking on YECA and CIN was done in direct, after 24H incubation of ITC, after 48H incubation of PSB and ITC. All the plates were incubated at 30∘C during 24 hours. Presence of typical colonies on CIN and YECA was checked, and isolates were biotyped. Pathogenic Y. enterocolitica strains showed an important growth on YECA with small and red fuchsia colonies while biotype 1A exhibited very few violet colonies. Enrichment in ITC during 48H gave the best performance for detecting positive samples in pathogenic Y. enterocolitica, and YECA could detect directly pathogenic Y. enterocolitica strains (2, 3, and 4. Use of YECA in combination with ITC generates a time-saver by giving a positive test in 72H.

  13. Comparison and Recovery of Escherichia coli and Thermotolerant Coliforms in Water with a Chromogenic Medium Incubated at 41 and 44.5°C (United States)

    Alonso, Jose L.; Soriano, Adela; Carbajo, Oscar; Amoros, Inmaculada; Garelick, Hemda


    This study compared the performance of a commercial chromogenic medium, CHROMagarECC (CECC), and CECC supplemented with sodium pyruvate (CECCP) with the membrane filtration lauryl sulfate-based medium (mLSA) for enumeration of Escherichia coli and non-E. coli thermotolerant coliforms (KEC). To establish that we could recover the maximum KEC and E. coli population, we compared two incubation temperature regimens, 41 and 44.5°C. Statistical analysis by the Fisher test of data did not demonstrate any statistically significant differences (P = 0.05) in the enumeration of E. coli for the different media (CECC and CECCP) and incubation temperatures. Variance analysis of data performed on KEC counts showed significant differences (P = 0.01) between KEC counts at 41 and 44.5°C on both CECC and CECCP. Analysis of variance demonstrated statistically significant differences (P = 0.05) in the enumeration of total thermotolerant coliforms (TTCs) on CECC and CECCP compared with mLSA. Target colonies were confirmed to be E. coli at a rate of 91.5% and KEC of likely fecal origin at a rate of 77.4% when using CECCP incubated at 41°C. The results of this study showed that CECCP agar incubated at 41°C is efficient for the simultaneous enumeration of E. coli and KEC from river and marine waters. PMID:10427079

  14. Assessment of commercial chromogenic solid media for the detection of non-O157 Shiga toxin-producing Escherichia coli (STEC). (United States)

    Zelyas, Nathan; Poon, Alan; Patterson-Fortin, Laura; Johnson, Roger P; Lee, Winki; Chui, Linda


    Detection of Shiga toxin-producing Escherichia coli (STEC) has evolved significantly since the introduction of sorbitol-MacConkey agar. This study compares four chromogenic media (CHROMagar™ STEC, Rainbow® O157 agar, CHROMagar™ O157, and Colorex® O157) in their identification of non-O157 STEC. When 161 non-O157 STEC were directly inoculated onto each medium, detection rates on CHROMagar™ STEC, Rainbow® O157 agar, CHROMagar™ O157 and Colorex® O157 were 90%, 70%, 3.7% and 6.8%, respectively. Tellurite minimal inhibitory concentrations (MICs) correlated with growth on CHROMagar™ STEC as 20 of 22 isolates with poor or no growth had MICs ≤1μg/mL. Stool spiking experiments revealed that CHROMagar™ STEC had the highest recovery of the six most common non-O157 STEC, ranging from 30% (in mucoid stool) to 98% (in watery stool). When using clinical stool samples, CHROMagar™ STEC had a sensitivity, specificity, positive predictive value, and negative predictive value of 84.6%, 87%, 13.9%, and 99.6%, respectively.

  15. Prevalence and molecular characterization of Enterobacteriaceae producing NDM-1 carbapenemase at a military hospital in Pakistan and evaluation of two chromogenic media. (United States)

    Day, Kathryn M; Ali, Shamshad; Mirza, Irfan Ali; Sidjabat, Hanna E; Silvey, Anna; Lanyon, Clare V; Cummings, Stephen P; Abbasi, Shahid Ahmed; Raza, Muhammad W; Paterson, David L; Perry, John D


    The aim of this study was to assess the frequency and genotypic diversity of carbapenemase-producing Enterobacteriaceae (CPE) in stool samples from patients attending a military hospital in Pakistan. Further aims included the identification of factors that might predispose to faecal carriage and evaluation of 2 chromogenic culture media: Brilliance CRE and chromID CARBA. Of 175 patients, 32 (18.3%) had faecal carriage of CPE and all produced NDM-1 carbapenemase. All of these 32 patients were detected using chromID CARBA compared with 20 patients (62.5%) detected using Brilliance CRE (P = 0.0015). Duration of hospitalization and treatment with co-amoxyclav were statistically associated with a higher likelihood of carriage of CPE (P ≤ 0.05). The majority of NDM-1-producing Enterobacteriaceae co-produced CTX-M-1 group extended spectrum β-lactamase, and one third produced armA-type methylase. NDM-1 carbapenemase was most commonly found amongst commensal types of Escherichia coli, especially phylogenetic group B1.

  16. Comparison of Six Chromogenic Agar Media for the Isolation of a Broad Variety of Non-O157 Shigatoxin-Producing Escherichia coli (STEC) Serogroups. (United States)

    Verhaegen, Bavo; De Reu, Koen; Heyndrickx, Marc; De Zutter, Lieven


    The isolation of non-O157 STEC from food samples has proved to be challenging. The selection of a suitable selective isolation agar remains problematic. The purpose of this study was to qualitatively and quantitatively evaluate six chromogenic agar media for the isolation of STEC: Tryptone Bile X-glucuronide agar (TBX), Rainbow® Agar O157 (RB), Rapid E. coli O157:H7 (RE), Modified MacConkey Agar (mMac), CHROMagarTM STEC (Chr ST) and chromIDTM EHEC (Chr ID). During this study, 45 E. coli strains were used, including 39 STEC strains belonging to 16 different O serogroups and 6 non-STEC E. coli. All E. coli strains were able to grow on TBX and RB, whereas one STEC strain was unable to grow on Chr ID and a number of other STEC strains did not grow on mMac, CHROMagar STEC and Rapid E. coli O157:H7. However, only the latter three agars were selective enough to completely inhibit the growth of the non-STEC E. coli. Our conclusion was that paired use of a more selective agar such as CHROMagar STEC together with a less selective agar like TBX or Chr ID might be the best solution for isolating non-O157 STEC from food.

  17. Recidiva del color dentario por té, café y vino: In vitro Dental bleaching regression caused by chromogenic beverages: In vitro

    Directory of Open Access Journals (Sweden)

    M Arévalo Pineda


    Full Text Available Este estudio, in vitro, determinó si los dientes con clareamiento presentan mayor cambio de color en el tiempo que los no tratados, al someterse a tinción con bebidas cromógenas, café, té y vino. Se utilizaron 45 incisivos sanos de bovino conservados en suero a 37ºC. Cada espécimen se dividió en dos mitades, una sometida a clareamiento con peróxido de hidrógeno al 35% y otra control. Se midió color con Espectrofotómetro Vita EasyShade. Se dividieron los especímenes al azar en grupos de 15 y fueron sumergidos en café, té y vino, durante 10 minutos, 20 veces, registrando color después de cada inmersión. Los datos fueron analizados con ANOVA y Test de Tukey, con 95% de intervalo de confianza. Los resultados mostraron que, el clareamiento modifica significativamente (p=0.05 el color en los tres grupos (GC=85.8 a 95.1; GT=87.4 a 97.3 y GV=90.8 a 99.3, la recidiva de color se observa a lo largo de las 20 inmersiones, siendo significativa la diferencia de valores ΔE iniciales (GC=18.89; GT=22.97; GV=56.46 y finales (GC=5.56; GT=5.38; GV=12.49. El grupo tratado presenta mayor descenso de unidades ΔE a lo largo de las inmersiones, por lo que es el más teñido (GCcontrol=20.98-5.01; GTcontrol=17.11-3.66; GVcontrol=54.62-11.49. Las tres bebidas cromógenas causan recidiva de color en los dientes clareados, siendo el vino el que causa mayor tinción. Se concluyó que las piezas tratadas, sometidas a los tres tipos de cromógenos, tienen mayor cambio de color que las que no lo son, pero finalmente no se oscurecen más que las no tratadas.This in vitro study established if teeth treated with dental bleaching have a higher change of color over time than those that aren’t treated, when subjected to three chromogenic beverages (coffee, tea and red wine. 45 healthy bovine incisors were used, maintained in 0.9% sodium chloride, at 37ºC. Every specimen was divided into 2 half; one half was subjected to dental bleaching with 35% hydrogen

  18. Naked eye and spectrophotometric detection of chromogenic insecticide in aquaculture using amine functionalized gold nanoparticles in the presence of major interferents (United States)

    Loganathan, C.; John, S. Abraham


    Detection of a chromogenic insecticide, malachite green (MG) using 3,5-diamino-1,2,4-triazole capped gold nanoparticles (DAT-AuNPs) by both naked eye and spectrophotometry was described in this paper. The DAT-AuNPs were prepared by wet chemical method and show absorption maximum at 518 nm. The zeta potential of DAT-AuNPs was found to be - 39.9 mV, suggesting that one of the amine groups of DAT adsorbed on the surface of AuNPs and the other amine group stabilizes the AuNPs from aggregation. The wine red color DAT-AuNPs changes to violet while adding 25 μM MG whereas the absorption band at 518 nm was increased and shifted towards longer wavelength. However, addition of 70 μM MG leads to the aggregation of DAT-AuNPs. This is due to strong electrostatic interaction between ammonium ion of MG and the free amine group of DAT. Based on the color change and shift in SPR band, 25 and 5 μM MG can be easily detected by naked eye and spectrophotometry. The DAT-AuNPs show high selectivity towards MG even in the presence of 5000-fold higher concentrations of common interferents. The practical application was successfully demonstrated by determining MG in fish farm water.

  19. Correlation and comparison of immunohistochemistry for HER2/neu, using the antibody SP3 and chromogenic in situ hybridization in breast carcinomas samples

    Directory of Open Access Journals (Sweden)

    Franciele F. Wolf


    Full Text Available ABSTRACT Introduction: Advances in the field of molecular biology have provided the differentiation of molecular subtypes of breast tumors, providing better prognosis and important tools for the treatment of patients with breast cancer. Among these subtypes, the changes in the human epidermal growth factor receptor 2 gene (HER2/neu, increase its copy number and generating HER2 protein amplification. Studies show that patients with breast cancer HER2/neu amplified tend to relapse earlier and have shorter survival time, the monoclonal antibody Trastuzumab is the therapy indicated. The eligibility of patients for therapy is initially made by the immunohistochemistry (IHC technique, which evaluates the expression level of the HER2 protein. After this evaluation, the cases with equivocal diagnosis (score 2+, are referred to a more accurate technique, the chromogenic in situ hybridization (CISH. Objective: To analyze the sensitivity and specificity of the antibody SP3, and determine their level of agreement with the CISH technique. Material and methods: Retrospective study in the database of the anatomy-pathology laboratory, in CISH tests reports for HER2/neu. Conclusion: The results revealed that clone SP3 showed 100% specificity and 92% sensitivity. IHC reveals variability in its results; however, it is known that the technique is an important tool in the daily routine of laboratories, contributing to the initial screening of patients with breast cancer, which later showed satisfactory results when compared with the CISH technique.

  20. Proto-oncogene HER-2 in normal, dysplastic and tumorous feline mammary glands: an immunohistochemical and chromogenic in situ hybridization study

    Directory of Open Access Journals (Sweden)

    Martín de las Mulas Juana


    Full Text Available Abstract Background Feline mammary carcinoma has been proposed as a natural model of highly aggressive, hormone-independent human breast cancer. To further explore the utility of the model by adding new similarities between the two diseases, we have analyzed the oncogene HER-2 status at both the protein and the gene levels. Methods Formalin-fixed, paraffin-embedded tissue samples from 30 invasive carcinomas, 7 benign lesions and two normal mammary glands were analyzed. Tumour features with prognostic value were recorded. The expression of protein HER-2 was analyzed by immunohistochemistry and the number of gene copies by means of DNA chromogenic in situ hybridization. Results Immunohistochemical HER-2 protein overexpression was found in 40% of feline mammary carcinomas, a percentage higher to that observed in human breast carcinoma. As in women, feline tumours with HER-2 protein overexpression had pathological features of high malignancy. However, amplification of HER-2 was detected in 16% of carcinomas with protein overexpression, a percentage much lower than that observed in their human counterpart. Conclusion Feline mammary carcinoma would be a suitable natural model of that subset of human breast carcinomas with HER-2 protein overexpression without gene amplification.

  1. Naked eye and spectrophotometric detection of chromogenic insecticide in aquaculture using amine functionalized gold nanoparticles in the presence of major interferents. (United States)

    Loganathan, C; John, S Abraham


    Detection of a chromogenic insecticide, malachite green (MG) using 3,5-diamino-1,2,4-triazole capped gold nanoparticles (DAT-AuNPs) by both naked eye and spectrophotometry was described in this paper. The DAT-AuNPs were prepared by wet chemical method and show absorption maximum at 518nm. The zeta potential of DAT-AuNPs was found to be -39.9mV, suggesting that one of the amine groups of DAT adsorbed on the surface of AuNPs and the other amine group stabilizes the AuNPs from aggregation. The wine red color DAT-AuNPs changes to violet while adding 25μM MG whereas the absorption band at 518nm was increased and shifted towards longer wavelength. However, addition of 70μM MG leads to the aggregation of DAT-AuNPs. This is due to strong electrostatic interaction between ammonium ion of MG and the free amine group of DAT. Based on the color change and shift in SPR band, 25 and 5μM MG can be easily detected by naked eye and spectrophotometry. The DAT-AuNPs show high selectivity towards MG even in the presence of 5000-fold higher concentrations of common interferents. The practical application was successfully demonstrated by determining MG in fish farm water.

  2. Biochemical nature of Russell Bodies. (United States)

    Mossuto, Maria Francesca; Ami, Diletta; Anelli, Tiziana; Fagioli, Claudio; Doglia, Silvia Maria; Sitia, Roberto


    Professional secretory cells produce and release abundant proteins. Particularly in case of mutations and/or insufficient chaperoning, these can aggregate and become toxic within or amongst cells. Immunoglobulins (Ig) are no exception. In the extracellular space, certain Ig-L chains form fibrils causing systemic amyloidosis. On the other hand, Ig variants lacking the first constant domain condense in dilated cisternae of the early secretory compartment, called Russell Bodies (RB), frequently observed in plasma cell dyscrasias, autoimmune diseases and chronic infections. RB biogenesis can be recapitulated in lymphoid and non-lymphoid cells by expressing mutant Ig-μ, providing powerful models to investigate the pathophysiology of endoplasmic reticulum storage disorders. Here we analyze the aggregation propensity and the biochemical features of the intra- and extra-cellular Ig deposits in human cells, revealing β-aggregated features for RB.

  3. Instrument for assaying radiation

    Energy Technology Data Exchange (ETDEWEB)

    Coleman, Jody Rustyn; Farfan, Eduardo B.


    An instrument for assaying radiation includes a flat panel detector having a first side opposed to a second side. A collimated aperture covers at least a portion of the first side of the flat panel detector. At least one of a display screen or a radiation shield may cover at least a portion of the second side of the flat panel detector.

  4. New oligosaccharyltransferase assay method. (United States)

    Kohda, Daisuke; Yamada, Masaki; Igura, Mayumi; Kamishikiryo, Jun; Maenaka, Katsumi


    We developed a new in vitro assay for oligosaccharyltransferase (OST), which catalyzes the transfer of preassembled oligosaccharides on lipid carriers onto asparagine residues in polypeptide chains. The asparagine residues reside in the sequon, Asn-X-Thr/Ser, where X can be any amino acid residue except Pro. We demonstrate the potency of our assay using the OST from yeast. In our method, polyacrylamide gel electrophoresis is used to separate the glycopeptide products from the peptide substrates. The substrate peptide is fluorescently labeled and the formation of glycopeptides is analyzed by fluorescence gel imaging. Two in vitro OST assay methods are now widely used, but both the methods depend on previous knowledge of the oligosaccharide moiety: One method uses lectin binding as the separation mechanism and the other method uses biosynthetically or chemoenzymatically synthesized lipid-linked oligosaccharides as donors. N-linked protein glycosylation is found in all three domains of life, but little is known about the N-glycosylation in Archaea. Thus, our new assay, which does not require a priori knowledge of the oligosaccharides, will be useful in such cases. Indeed, we have detected the OST activity in the membrane fraction from a hyperthermophilic archaeon, Pyrococcus furiosus.

  5. Hyaluronic Acid Assays

    DEFF Research Database (Denmark)

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


    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...

  6. Development of an enzyme-linked immunosorbent assay for detection of chicken osteocalcin and its use in evaluation of perch effects on bone remodeling in caged White Leghorns (United States)

    Osteocalcin (OC) is a sensitive biochemical marker for evaluating bone turnover in mammals. The role of avian OC is less clear because of a need for a chicken assay. Our objectives were to develop an assay using indirect competitive ELISA for detecting chicken serum OC and use the assay to examine t...

  7. Associative learning in biochemical networks. (United States)

    Gandhi, Nikhil; Ashkenasy, Gonen; Tannenbaum, Emmanuel


    It has been recently suggested that there are likely generic features characterizing the emergence of systems constructed from the self-organization of self-replicating agents acting under one or more selection pressures. Therefore, structures and behaviors at one length scale may be used to infer analogous structures and behaviors at other length scales. Motivated by this suggestion, we seek to characterize various "animate" behaviors in biochemical networks, and the influence that these behaviors have on genomic evolution. Specifically, in this paper, we develop a simple, chemostat-based model illustrating how a process analogous to associative learning can occur in a biochemical network. Associative learning is a form of learning whereby a system "learns" to associate two stimuli with one another. Associative learning, also known as conditioning, is believed to be a powerful learning process at work in the brain (associative learning is essentially "learning by analogy"). In our model, two types of replicating molecules, denoted as A and B, are present in some initial concentration in the chemostat. Molecules A and B are stimulated to replicate by some growth factors, denoted as G(A) and G(B), respectively. It is also assumed that A and B can covalently link, and that the conjugated molecule can be stimulated by either the G(A) or G(B) growth factors (and can be degraded). We show that, if the chemostat is stimulated by both growth factors for a certain time, followed by a time gap during which the chemostat is not stimulated at all, and if the chemostat is then stimulated again by only one of the growth factors, then there will be a transient increase in the number of molecules activated by the other growth factor. Therefore, the chemostat bears the imprint of earlier, simultaneous stimulation with both growth factors, which is indicative of associative learning. It is interesting to note that the dynamics of our model is consistent with certain aspects of

  8. Kinetic Tetrazolium Microtiter Assay (United States)

    Pierson, Duane L.; Stowe, Raymond; Koenig, David


    Kinetic tetrazolium microtiter assay (KTMA) involves use of tetrazolium salts and Triton X-100 (or equivalent), nontoxic, in vitro color developer solubilizing colored metabolite formazan without injuring or killing metabolizing cells. Provides for continuous measurement of metabolism and makes possible to determine rate of action of antimicrobial agent in real time as well as determines effective inhibitory concentrations. Used to monitor growth after addition of stimulatory compounds. Provides for kinetic determination of efficacy of biocide, greatly increasing reliability and precision of results. Also used to determine relative effectiveness of antimicrobial agent as function of time. Capability of generating results on day of test extremely important in treatment of water and waste, disinfection of hospital rooms, and in pharmaceutical, agricultural, and food-processing industries. Assay also used in many aspects of cell biology.

  9. The corneal pocket assay. (United States)

    Ziche, Marina; Morbidelli, Lucia


    The cornea in most species is physiologically avascular, and thus this assay allows the measurement of newly formed vessels. The continuous monitoring of neovascular growth in the same animal allows the evaluation of drugs acting as suppressors or stimulators of angiogenesis. Under anesthesia a micropocket is produced in the cornea thickness and the angiogenesis stimulus (tumor tissue, cell suspension, growth factor) is placed into the pocket in order to induce vascular outgrowth from the limbal capillaries. Neovascular development and progression can be modified by the presence of locally released or applied inhibitory factors or by systemic treatments. In this chapter the experimental details of the avascular cornea assay, the technical challenges, and advantages and disadvantages in different species are discussed. Protocols for local drug treatment and tissue sampling for histology and pharmacokinetic profile are reported.

  10. B cell helper assays. (United States)

    Abrignani, Sergio; Tonti, Elena; Casorati, Giulia; Dellabona, Paolo


    Activation, proliferation and differentiation of naïve B lymphocytes into memory B cells and plasma cells requires engagement of the B cell receptor (BCR) coupled to T-cell help (1, 2). T cells deliver help in cognate fashion when they are activated upon recognition of specific MHC-peptide complexes presented by B cells. T cells can also deliver help in a non-cognate or bystander fashion, when they do not find specific MHC-peptide complexes on B cells and are activated by alternative mechanisms. T-cell dependent activation of B cells can be studied in vitro by experimental models called "B cell helper assays" that are based on the co-culture of B cells with activated T cells. These assays allow to decipher the molecular bases for productive T-dependent B cell responses. We show here examples of B cell helper assays in vitro, which can be reproduced with any subset of T lymphocytes that displays the appropriate helper signals.

  11. Biochemical genetic markers in sugarcane. (United States)

    Glaszmann, J C; Fautret, A; Noyer, J L; Feldmann, P; Lanaud, C


    Isozyme variation was used to identify biochemical markers of potential utility in sugarcane genetics and breeding. Electrophoretic polymorphism was surveyed for nine enzymes among 39 wild and noble sugarcane clones, belonging to the species most closely related to modern varieties. Up to 114 distinct bands showing presence versus absence type of variation were revealed and used for qualitative characterization of the materials. Multivariate analysis of the data isolated the Erianthus clone sampled and separated the Saccharum spontaneum clones from the S. robustum and S. officinarum clones; the latter two were not differentiated from one another. The analysis of self-progenies of a 2n=112 S. spontaneum and of a commercial variety showed examples of mono- and polyfactorial segregations. Within the progeny of the variety, co-segregation of two isozymes frequent in S. spontaneum led to them being assigned to a single chromosome initially contributed by a S. spontaneum donor. This illustrates how combined survey of ancestral species and segregation analysis in modern breeding materials should permit using the lack of interspecific cross-over to establish linkage groups in a sugarcane genome.


    Directory of Open Access Journals (Sweden)



    Full Text Available Diabetic nephropathy is a clinical syndrome characterized by the following- Persistent albuminuria (>300mg/d or >200μg/min, that is confirmed on at least 2 occasions 3-6 months apart diabetic, progressive decline in the Glomerular Filtration Rate (GFR, elevated arterial blood pressure. The earliest biochemical criteria for the diagnosis of diabetic nephropathy is the presence of micro-albumin in the urine, which if left untreated will eventually lead to End-Stage Renal Disease (ESRD. Micro-albuminuria refers to the excretion of albumin in the urine at a rate that exceeds normal limits. The current study was conducted to establish the prevalence of micro-albuminuria in a sequential sample of diabetic patients attending hospital and OPD Clinic to determine its relationship with known and putative risk factors to identify micro- and normo-albuminuric patients in their sample for subsequent comparison in different age, sex, weight and creatinine clearance of the micro- and normo-albuminuric patients. This cross-sectional analytical study was conducted in one hundred patients at Saraswathi Institute of Medical Sciences, Anwarpur, Hapur, U. P. Patients having diabetes mellitus in different age group ranging from 30 to 70 years were selected. Data was analysed by SPSS software. Micro-albuminuria was observed in 35% in patients with type 2 diabetes mellitus. It was observed that 65% patients were free from any type of albuminuria. Also micro-albuminuria was present in 10% of the patients less than 50 yrs. of age, while 15% of the patients more than 50 yrs. of age were having micro-albuminuria. There was a statistically significant correlation of micro-albuminuria with duration of diabetes. Incidence of micro-albuminuria increases with age as well as increased duration of diabetes mellitus. Our study shows that only 5% patients developed macro-albuminuria. Glycosylated haemoglobin and fasting plasma glucose was significantly raised among all these

  13. Use of substitute Nonidet P-40 nonionic detergents in intracellular tubulin polymerization assays for screening of microtubule targeting agents. (United States)

    Sinha, Saptarshi; Field, Jessica J; Miller, John H


    Shell Chemical Company Nonidet P-40 has been used for decades in many biochemical assays as a nonionic, nondenaturing detergent; however, Shell no longer produces this product. Four commercially available substitutes were investigated and their activities titrated in an intracellular tubulin polymerization assay. Although claimed by the supply companies to be identical to the Shell Nonidet P-40, all four substitutes were about 10-fold more potent and needed to be diluted accordingly. As microtubule targeting drugs are a major class of anticancer agent, and many researchers use the intracellular tubulin polymerization assay, this information is important to help troubleshoot assay development with the new substitutes. As the Shell Nonidet P-40 has been used in many biochemical buffers, these results will be of general interest to the biochemical, cell, and molecular research community.

  14. Radon assay for SNO+

    Energy Technology Data Exchange (ETDEWEB)

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


    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+.


    DEFF Research Database (Denmark)


    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......, when operatively connected, one or more chambers (21) comprising the chemical entities (41), the inlet(s) (5) and outlet(s) (6) and chambers (21) being in fluid connection. The device further comprise means for providing differing chemical conditions in each chamber (21)....

  16. Growth cone collapse assay. (United States)

    Cook, Geoffrey M W; Jareonsettasin, Prem; Keynes, Roger J


    The growth cone collapse assay has proved invaluable in detecting and purifying axonal repellents. Glycoproteins/proteins present in detergent extracts of biological tissues are incorporated into liposomes, added to growth cones in culture and changes in morphology are then assessed. Alternatively purified or recombinant molecules in aqueous solution may be added directly to the cultures. In both cases after a defined period of time (up to 1 h), the cultures are fixed and then assessed by inverted phase contrast microscopy for the percentage of growth cones showing a collapsed profile with loss of flattened morphology, filopodia, and lamellipodia.

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

    Directory of Open Access Journals (Sweden)

    U. U. Ukoha


    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.

  18. RAS - Screens & Assays - Drug Discovery (United States)

    The RAS Drug Discovery group aims to develop assays that will reveal aspects of RAS biology upon which cancer cells depend. Successful assay formats are made available for high-throughput screening programs to yield potentially effective drug compounds.

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

    NARCIS (Netherlands)

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


    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 ena

  20. Photonic Crystal Surfaces as a General Purpose Platform for Label-Free and Fluorescent Assays


    Cunningham, Brian T.


    Photonic crystal surfaces can be designed to provide a wide range of functions that are used to perform biochemical and cell-based assays. Detection of the optical resonant reflections from photonic crystal surfaces enables high sensitivity label-free biosensing, while the enhanced electromagnetic fields that occur at resonant wavelengths can be used to enhance the detection sensitivity of any surface-based fluorescence assay. Fabrication of photonic crystals from inexpensive plastic material...

  1. Evaluating biochemical methane production from brewer's spent yeast. (United States)

    Sosa-Hernández, Ornella; Parameswaran, Prathap; Alemán-Nava, Gibrán Sidney; Torres, César I; Parra-Saldívar, Roberto


    Anaerobic digestion treatment of brewer's spent yeast (SY) is a viable option for bioenergy capture. The biochemical methane potential (BMP) assay was performed with three different samples (SY1, SY2, and SY3) and SY1 dilutions (75, 50, and 25 % on a v/v basis). Gompertz-equation parameters denoted slow degradability of SY1 with methane production rates of 14.59-4.63 mL/day and lag phases of 10.72-19.7 days. Performance and kinetic parameters were obtained with the Gompertz equation and the first-order hydrolysis model with SY2 and SY3 diluted 25 % and SY1 50 %. A SY2 25 % gave a 17 % of TCOD conversion to methane as well as shorter lag phase (methane production. Methane capture and biogas composition were dependent upon the SY source, and co-digestion (or dilution) can be advantageous.

  2. A Program on Biochemical and Biomedical Engineering. (United States)

    San, Ka-Yiu; McIntire, Larry V.


    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)

  3. In vitro reparative dentin: a biochemical and morphological study

    Directory of Open Access Journals (Sweden)

    G. Teti


    Full Text Available In this study, starting from human dental pulp cells cultured in vitro, we simulated reparative dentinogenesis using a medium supplemented with different odontogenic inductors. The differentiation of dental pulp cells in odontoblast-like cells was evaluated by means of staining, and ultramorphological, biochemical and biomolecular methods. Alizarin red staining showed mineral deposition while transmission electron microscopy revealed a  synthesis of extracellular matrix fibers during the differentiation process. Biochemical assays demonstrated that the differentiated phenotype expressed odontoblast markers, such as Dentin Matrix Protein 1 (DMP1 and Dentin Sialoprotein (DSP, as well as type I collagen. Quantitative data regarding the mRNA expression of DMP1, DSP and type I  collagen were obtained by Real Time PCR. Immunofluorescence data demonstrated the various localizations of DSP and DMP1 during odontoblast differentiation. Based on our results, we obtained odontoblast-like cells which simulated the reparative dentin processes in order to better investigate the mechanism of odontoblast differentiation, and dentin extracellular matrix deposition and mineralization. 

  4. Biochemical activity and gene analysis of inherited protein C and antithrombin deficiency in two Chinese pedigrees

    Institute of Scientific and Technical Information of China (English)

    周荣富; 傅启华; 王文斌; 谢爽; 胡翊群; 王学锋; 王振义; 王鸿利


    Background We identified the gene mutations in two Chinese pedigree of type Ⅰ hereditary protein C deficiency and type Ⅰ hereditary antithrombin deficiency.Methods The plasma level of protein C activity (PC∶ A), protein C antigen (PC∶ Ag) , protein S activity, antithrombin activity (AT∶ A) and antithrombin antigen (AT∶ Ag) of propositi and two family members were detected using ELISA and chromogenic assay, respectively. All exons and intron-exon boundaries of protein C gene and antithrombin gene were analyzed by direct sequencing of the corresponding amplified PCR products in DNA from the propositus. Results The plasma PC∶ A and PC∶ Ag of propositus 1 was 26% and 1.43 mg/dl, respectively. The PC∶ Ag and PC∶ A of his father were normal. The decreased PC∶ A level was seen in his mother and 4 of his maternal pedigree. PS∶ A and AT∶ A were all normal in pedigree 1 members. A C5498T heterozygous mutation in exon 3 of protein C gene, resulting in the substitution of Arg for Trp at the 15th amino acid, was identified in propositus 1 and 8 of his relatives. The plasma AT∶ A and AT∶ Ag of propositus 2 was 48.6% and 10.4 mg/dl, respectively. The reduced AT∶ A and AT∶ Ag levels were found in his father and 5 of paternal pedigree. PC∶ A, PC∶ Ag and PS∶ A were all in normal range. A heterozygous 13387-9G deletion in exon 6 of antithrombin gene was identified in propositus 2. This mutation introduced a frameshift and a premature stop at codon 426 and existed in 6 members of pedigree 2.Conclusion The C5498T heterozygous mutation in exon 3 of protein C gene, first reported in China, leads to type I hereditary protein C deficiency. The 13387-9G deletion, a novel mutation, can cause antithrombin deficiency and thrombosis.

  5. 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.


    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

  6. Bacterial assays for recombinagens. (United States)

    Hoffmann, G R


    Two principal strategies have been used for studying recombinagenic effects of chemicals and radiation in bacteria: (1) measurement of homologous recombination involving defined alleles in a partially diploid strain, and (2) measurement of the formation and loss of genetic duplications in the bacterial chromosome. In the former category, most methods involve one allele in the bacterial chromosome and another in a plasmid, but it is also possible to detect recombination between two chromosomal alleles or between two extrachromosomal alleles. This review summarizes methods that use each of these approaches for detecting recombination and tabulates data on agents that have been found to be recombinagenic in bacteria. The assays are discussed with respect to their effectiveness in testing for recombinagens and their potential for elucidating mechanisms underlying recombinagenic effects.

  7. A facile low-cost enzymatic paper-based assay for the determination of urine creatinine. (United States)

    Talalak, Kwanrutai; Noiphung, Julaluk; Songjaroen, Temsiri; Chailapakul, Orawon; Laiwattanapaisal, Wanida


    Creatinine is one of many markers used to investigate kidney function. This paper describes a low-cost enzymatic paper-based analytical device (enz-PAD) for determining urine creatinine. The disposable dead volumes of creatinine enzyme reagents from an automatic analyser cassette were utilised. Whatman No. 3 paper was cut into long rectangular shapes (4×40 mm(2)) on which the enzyme reagents, R1 and R2, were adsorbed in two consecutive regions. The assay was performed by immersing test strips into urine samples contained in microwells to allow creatinine in the sample to react with immobilised active ingredients and, then, traverse via capillary action to the detection area where chromogen products accumulated. The method is based on hydrogen peroxide (H2O2) formation via creatinine conversion using creatininase, creatinase, and sarcosine oxidase. The liberated H2O2 reacts with 4-aminophenazone and 2,4,6-triiodo-3-hydroxybenzoic acid to form quinoneimine with a pink-red colour at the detection zone. The linear range of the creatinine assay was 2.5-25 mg dL(-1) (r(2)=0.983), and the detection limit was 2.0 mg dL(-1). The colorimetric enz-PAD for the creatinine assay was highly correlated with a conventional alkaline picrate method when real urine samples were evaluated (r(2)=0.977; n=40). This simple and nearly zero-cost paper-based device provides a novel alternative method for screening urinary creatinine and will be highly beneficial for developing countries.

  8. Diagnostic difficulties of factor XI deficiencies: interferences' assay or real deficit? (United States)

    Gaymard, Alexandre; Nougier, Christophe


    Madam P, 77 years old, consulted in the hemostasis department after a coagulation anomaly was discovered during her preoperative test for a total hip prosthesis. After confirmation of a persistent and increased aPTT, additional tests were performed and showed the presence of antiphospholipid antibodies. Factor VIII level could be corrected after the plasma dilution to 1/40(th). But successive dilutions were not enough to obtain a correct factor IX (FIX) and factor XI (FXI) level. FIX level was obtained by chromogenic method in order to avoid the interferences caused by the antibodies. Finally, despite the change of reagents and dilutions up to 1/160(th), the FXI level couldn't be determined. Despite these results and those of the thrombin generation assay, the surgery was successfully done without specific treatment thanks to the absence of hemorrhagic history. This observation highlights the diagnostic and monitoring difficulties for uncommon clotting factor deficit. The development of interference free test could increase the support for these patients.

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

    DEFF Research Database (Denmark)

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


    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 optim...

  10. HER2/neu Expression and Gene Alterations in Pancreatic Ductal Adenocarcinoma: A Comparative mmunohistochemistry and Chromogenic in Situ Hybridization Study Based on Tissue Microarrays and Computerized Image Analysis

    Directory of Open Access Journals (Sweden)

    Evangelos Tsiambas


    Full Text Available Context: HER2/neu overexpression is observed in many cancers including pancreatic ductal adenocarcinoma. Although immunohistochemistry remains the basic method for evaluating HER2/neu protein expression, significant information regarding gene status cannot be assessed. Design: Using tissue microarray technology, fifty histologically confirmed pancreatic ductal adenocarcinomas were cored twice and re-embedded in one paraffin block. Immunohistochemistry (clone TAB 250 and chromogenic (HER2/neu amplification Spot Light kit in situ hybridization protocols were performed. The immunostained slides were evaluated by conventional eye microscopy and digital image analysis. The chi square test and the kappa statistic were applied by running the SPSS package. Main outcome measures :The levels of staining intensity were estimated by the performance of a semi automated image analysis system. Results :HER2/neu gene amplification was detected in 8/50 cases (16%. Chromosome 17 aneuploidy was detected in 19 cases (38%. Significant improvement in interobserver agreement (kappa=0.76 vs. 0.94 was achieved correlating the immunohistochemical results obtained by conventional eye and digital microscopy, especially in the cases of overexpression (2+, 3+. Finally, 29 (58%, 11 (22%, 6 (12% and 4 (8% cases were characterized as 0, 1+, 2+ and 3+, respectively. HER2/neu protein expression was significantly associated with grade (P=0.019, but not with stage (P=0.466. in addition, chromosome 17 and gene status were not correlated with stage and grade.. Conclusion :Our results indicate that a subset of pancreatic ductal adenocarcinomas is characterized by HER2/neu gene amplification. In contrast to breast cancer, protein overexpression does not predict this specific gene deregulation mechanism. This event may reflect the different biological role of the molecule in those two solid tumours, affecting the response to novel targeted agents, such as monoclonal anti-HER2/neu

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

    Directory of Open Access Journals (Sweden)

    Moisi Daniella


    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

  12. The various assays for measuring activity states of factor VIIa in plasma and therapeutic products: Diagnostic value and analytical usefulness in various pathophysiological states. (United States)

    Amiral, Jean; Dunois, Claire; Amiral, Cédric; Seghatchian, Jerard


    The key coagulation factor FVII, and its activated form FVIIa, present a major interest for their role at the initiation phase of blood coagulation, and because they can activate all blood coagulation cascade, through the extrinsic, but also the intrinsic pathway. Blood activation initiated through FVII is first presented, as it is understood nowadays. Measurement of FVII and FVIIa were of main interest for epidemiological studies, but FVIIa contribution to assay results was only deduced. The introduction of specific FVIIa assays, functional or immunoassays, allowed measuring directly FVIIa without any interference of non-activated FVII, or other coagulation factors or their activated forms. The various methods available, and their characteristics are presented, with a special focus on two assays developed by our group for FVIIa (a clotting one and a chromogenic one). The FVIIa clotting assay shows evident superiority for measuring its activity in plasma, in pathophysiological conditions. The normal range is products, or for following recoveries in treated patients, including hemophiliacs with inhibitors, patients with severe bleeding risk (liver diseases, surgery, trauma, …), and lastly for measurement of its activity in therapeutic products.

  13. Herbicide resistance screening assay. (United States)

    Peterson, Joan M


    Herbicide resistance screening is a method that can be used not only to determine presence of the enzyme, phosphinothricin acetyltransferase, encoded by either the Bar or the Pat gene in transgenic maize, but also to assess the inheritance ratio of those genes in a segregating population. Herbicide screening can also be used to study linkage of a transgene of interest that was cotransformed with the herbicide resistance marker gene. By combining the herbicide screen assay with a PCR-based screen of leaf tissue DNA for the presence of both the Bar or the Pat gene marker and a cotransformed transgene of interest from the same seedling tissue and maintaining that seedling identity, the researcher can identify linkage or the possible breakdown in linkage of the marker gene and the transgene of interest. Further, the occurrence of "DNA silencing" can be evaluated if an individual seedling that was susceptible to the applied herbicide nonetheless gave PCR data that indicated presence of the gene responsible for herbicide resistance. Similarly, "DNA silencing" of the gene of interest may be investigated if the seeds can be screened and scored for that phenotypic trait in a nondestructive manner prior to planting.

  14. 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: [Universidad de Guanajuato, Departamento de Ciencias Medicas, DCS, 20 de Enero No. 929, Col. Obregon, 37000 Leon, Guanajuato (Mexico)


    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)

  15. 40 CFR 158.2010 - Biochemical pesticides data requirements. (United States)


    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false Biochemical pesticides data...) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2010 Biochemical pesticides... required to support registration of biochemical pesticides. Sections 158.2080 through 158.2084 identify...

  16. Nonlinear biochemical signal processing via noise propagation. (United States)

    Kim, Kyung Hyuk; Qian, Hong; Sauro, Herbert M


    Single-cell studies often show significant phenotypic variability due to the stochastic nature of intra-cellular biochemical reactions. When the numbers of molecules, e.g., transcription factors and regulatory enzymes, are in low abundance, fluctuations in biochemical activities become significant and such "noise" can propagate through regulatory cascades in terms of biochemical reaction networks. Here we develop an intuitive, yet fully quantitative method for analyzing how noise affects cellular phenotypes based on identifying a system's nonlinearities and noise propagations. We observe that such noise can simultaneously enhance sensitivities in one behavioral region while reducing sensitivities in another. Employing this novel phenomenon we designed three biochemical signal processing modules: (a) A gene regulatory network that acts as a concentration detector with both enhanced amplitude and sensitivity. (b) A non-cooperative positive feedback system, with a graded dose-response in the deterministic case, that serves as a bistable switch due to noise-induced ultra-sensitivity. (c) A noise-induced linear amplifier for gene regulation that requires no feedback. The methods developed in the present work allow one to understand and engineer nonlinear biochemical signal processors based on fluctuation-induced phenotypes.

  17. eQuilibrator--the biochemical thermodynamics calculator. (United States)

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


    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 ( 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.

  18. Advances in Biochemical Indices of Zooplankton Production. (United States)

    Yebra, L; Kobari, T; Sastri, A R; Gusmão, F; Hernández-León, S


    Several new approaches for measuring zooplankton growth and production rates have been developed since the publication of the ICES (International Council for the Exploration of the Sea) Zooplankton Methodology Manual (Harris et al., 2000). In this review, we summarize the advances in biochemical methods made in recent years. Our approach explores the rationale behind each method, the design of calibration experiments, the advantages and limitations of each method and their suitability as proxies for in situ rates of zooplankton community growth and production. We also provide detailed protocols for the existing methods and information relevant to scientists wanting to apply, calibrate or develop these biochemical indices for zooplankton production.

  19. 过氧化氢酶活性测定的新方法%A novel method for assaying catalase activity

    Institute of Scientific and Technical Information of China (English)

    刘砚韬; 王振伟; 张伶俐


    OBJECTIVE To establish a novel method for determination of catalase activity.METHODS Two-step method was adopted in this research for determination of catalase activity.Firstly,the catalase catalyzed the breakdown of H2O2.Secondly,the peroxidase reacted with residual H2O2 and oxygen donor-TOOS which was oxygenated to a blue quinoneimine dye,then absorbance at 555 nm was determined to calculate the activity.The influence of factors in the reaction,such as chromogenic reagent,substrate concentration,reaction time,enzyme concentration,and sensitivity and stability of chromogenic reagent were compared with other methods.RESULTS Optimal reaction system was composed of pH 7.5,30 ℃ and 16.2 mmol·L-1 of H2O2 as well as TOOS as the chromogenic reagent.CONCLUSION This was a simple,quick and high sensitive method to determine catalase activity and was especially appropriate for assaying catalase activity in some enzymes used in diagnostic kit.%目的 建立一种简单、灵敏和快速检测过氧化氢酶(CAT)活性的方法.方法 采用两步法检测CAT活性.第一步是CAT催化H2O2分解,第二步是剩余的H2O2在CAT作用下,氧供体(TOOS)被氧化成蓝色醌亚胺物质,测定在555 nm处的吸光度,计算CAT活性.对反应的影响因素,如显色剂、底物浓度、反应时间、酶浓度、灵敏度和稳定性等,与其它CAT活性测定方法进行比较分析.结果 从pH7.5、30℃、16.2 mmol·L-1作为最佳反应条件,TOOS作显色试剂.结论 所建方法用于测定CAT活性,具有简单、快速和灵敏度高等特点,尤其适用于诊断试剂盒中CAT活性的检测.

  20. Biochemical investigation and biological evaluation of the methanolic extract of the leaves of Nyctanthes arbortristis in vitro

    Institute of Scientific and Technical Information of China (English)

    Repon Kumer Saha; Srijan Acharya; Syed Sohidul Haque Shovon; Apurba Sarker Apu; Priyanka Roy


    Objective: Nyctanthes arbortristis is a common plant in Bangladesh. The objective of our research was to biochemical and biological analysis of the methanolic extract of the dried leaves of Nyctanthesarbortristis found in Bangladesh. Methods: We investigated the presence of polyphenols, flavanoids and other types of compounds by thin layer chromatography, infrared spectroscopy, and UV spectroscopy analysis. We performed antioxidant assay by colorimetric methods. We investigated antibacterial assay by disk diffusion method. Cell surface receptor binding assay was performed by hemagglutination inhibition assay and hemolysis assay. Results: Methanolic extract of the leaves of Nyctanthes arbortristis contains flavanoids and other biologically active compounds. The extract showed antioxidant, peroxide scavenging and total reducing activity. The extract also showed antibacterial activities against several strains of bacteria. It also showed hemaglutination inhibition activities and hydrogen peroxide induced hemolysis inhibition activity in human blood cells. Conclusions: Therefore, Nyctanthes arbortristis may be considered as a plant of various health benefits.

  1. Practical assay issues with the PERT/PBRT assay: a highly sensitive reverse transcriptase assay. (United States)

    Chang, A; Dusing, S


    Product safety testing for retroviruses can be achieved by a panel of screening assays, including electron microscopy, viral gene specific PCRs, virus propagation, and detection of reverse transciptase activity. The application of PCR-based reverse transcriptase assays (PERT) that are approximately a million-fold more sensitive than conventional nucleotide incorporation assays in the testing of biologicals is described. Use of PERT assays can be applied to three areas: (i) screening for adventitious retrovirus contamination; (ii) detecting and quantifying endogenous viral particle load and (iii) monitoring levels of infectious retrovirus generation in cell lines that contain endogenous retroviruses.

  2. High frequency lateral flow affinity assay using superparamagnetic nanoparticles (United States)

    Lago-Cachón, D.; Rivas, M.; Martínez-García, J. C.; Oliveira-Rodríguez, M.; Blanco-López, M. C.; García, J. A.


    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.

  3. 23种致癌芳香胺的显色法测定%Chromogenic method determination of 23 kinds of carcinogenic aromatic amines

    Institute of Scientific and Technical Information of China (English)

    叶曦雯; 李引; 牛增元; 张清智; 高永刚


    In order to reduce the dependence on large-scale chromatographs, improve the detection speed and cut the cost of testing, a chromogenic determination method for 23 kinds of carcinogenic aromatic amines through reacting with guaiacol was developed. According to the structural features of aromatic amines and mechanism of diazo-coupling reaction, all terms of diazo-coupling reaction process were studied and optimized effectively. The optimal conditions were obtained as follows; at room temperature, 23 kinds of carcinogenic aromatic amines reacted with sodium nitrite and diazo salt was formed at a pH value of 2.56, and then added ammonium sulfamate, removing the excessive sodium nitrite, guaiacol was added to produce coupling /developing reaction with the diazo salt; the best developing pH ranged from 10 to 11, and developing reaction could be completed quickly, and the resulting azo compound remained stable within 180 min. Linearity was excellent in the concentration range of 1 - 50 mg/L. This method could be effectively applied to textiles in qualitative screening test of 23 kinds of carcinogenic aromatic amines.%为了减少禁用芳香胺检测过程对大型色谱仪的依赖,提高检测速度,降低检测成本,建立了23种致癌芳香胺的邻甲氧基苯酚显色测定方法.根据芳香胺的结构特征,引入重氮化-偶合反应机制,有效考察优化了23种芳香胺重氮化反应和偶合反应过程中的各项条件,得到最佳的反应条件:在室温下,23种芳香胺和亚硝酸钠在pH值为2.56时反应生成重氮盐,加入氨基磺酸铵除去多余的亚硝酸钠后,加入邻甲氧基苯酚与重氮盐偶联显色,显色的最佳pH值范围为10~11,显色反应可以迅速完成,生成的偶氮化合物可稳定180 min以上.方法的线性范围为1~ 50 mg/L.该方法操作简单易行,可有效地应用于纺织品中23种致癌芳香胺的同时定性筛选检测.

  4. 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.

  5. Biochemical Applications in the Analytical Chemistry Lab (United States)

    Strong, Cynthia; Ruttencutter, Jeffrey


    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…

  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)


    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. Biochemical and Anatomical Characteristics of Dolphin Muscles. (United States)


    hexosamines. These data suggest a biochemical difference in dolphin and rat tendon, which may be of relevance to the unique myofascial design of the...glycosaminoglycan (GAG) chains . The GAG chain is a polymer of repeating disaccharides, each disaccharide containing a hexosamine and either a

  8. Biochemical Thermodynamics under near Physiological Conditions (United States)

    Mendez, Eduardo


    The recommendations for nomenclature and tables in Biochemical Thermodynamics approved by IUBMB and IUPAC in 1994 can be easily introduced after the chemical thermodynamic formalism. Substitution of the usual standard thermodynamic properties by the transformed ones in the thermodynamic equations, and the use of appropriate thermodynamic tables…

  9. Reactions of 1-methyl-2-phenylindole with malondialdehyde and 4-hydroxyalkenals. Analytical applications to a colorimetric assay of lipid peroxidation. (United States)

    Gérard-Monnier, D; Erdelmeier, I; Régnard, K; Moze-Henry, N; Yadan, J C; Chaudière, J


    Under acidic and mild-temperature conditions, 1-methyl-2-phenylindole was found to react with malondialdehyde (MDA) and 4-hydroxyalkenals to yield a stable chromophore with intense maximal absorbance at 586 nm. The use of methanesulfonic acid results in optimal yields of chromophore produced from MDA as well as from 4-hydroxynonenal. By contrast, the use of hydrochloric acid results in an optimal yield of chromophore produced from MDA and a negligible reaction of 4-hydroxynonenal. Taking advantage of such chromogenic reactions, we developed a new colorimetric assay of lipid peroxidation. Using a methanesulfonic acid-based medium, MDA and 4-hydroxyalkenals can be measured at the 586 nm wavelength. However, the presence of endogenous inhibitors of the reaction with 4-hydroxyalkenals is common, and this means that the latter may be underestimated in some biological samples. The assay performed in a hydrochloric acid-based medium enables the specific measurement of MDA in the presence of 4-hydroxyalkenals. Upon hydrolysis of Schiff bases in hydrochloric acid (pH 1.5), either assay can be used to specifically measure the amount of total MDA in biological samples because 4-hydroxyalkenals undergo an irreversible cyclization reaction under the hydrochloric acid-based conditions of hydrolysis. The two assays were applied to the determination of the amount of MDA alone and of MDA and 4-hydroxyalkenals in an in vitro model of lipid peroxidation. This methodology was also used to clarify complex patterns of tissue-specific MDA production in vivo, following hydrolysis of Schiff bases, in rodents treated with doxorubicin.


    Directory of Open Access Journals (Sweden)

    Mark Dysinger


    Full Text Available As biomarkers grow in relevance for both the design and support of therapeutics and the clinical trials associated with them, there is an ever increasing need for accurate quantitation of these biochemical entities in biological matrices. While quantifying many biotherapeutics via ligand binding assay platforms can be fairly straightforward, biomarkers present some unique challenges that must be taken into account during assay development, validation and subsequent sample analysis. These challenges can be especially confounded by the relationship between two ligand binding assay tools: The regression curve and quality control samples. Due diligence must be performed to develop an assay that takes into account matrix vs. buffer effects and endogenous biomarker presence. Lack of diligence in these areas can lead to less than reliable results, thus potentially rendering the intended use of the assay moot.

  11. 6-sulfatoxymelatonin collected from infant diapers: feasibility and implications for urinary biochemical markers. (United States)

    Thomas, Karen A


    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.

  12. Biochemical and molecular characterization of hazelnut (Corylus avellana) seed lipoxygenases. (United States)

    Santino, Angelo; De Paolis, Angelo; Gallo, Antonia; Quarta, Angela; Casey, Rod; Mita, Giovanni


    Plant lipoxygenases (LOXs) are a class of dioxygenases which display diverse functions in several physiological processes such as growth, development and response to biotic and abiotic stresses. Even though LOXs have been characterized from several plant species, the physiological role of seed LOXs is still unclear. With the aim to better clarify the occurrence of LOXs and their influence on hazelnut seed quality, we carried out the biochemical and molecular characterization of the main LOX isoforms expressed during seed development. A genomic clone containing a complete LOX gene was isolated and fully characterized. The 9887 bp sequence reported contains an open reading frame of 5334 bp encoding a putative polypeptide of 99 kDa. Semiquantitative RT-PCR carried out from RNAs extracted from seeds at different maturation stages showed that LOXs are mainly expressed at early developmental stages. These results were confirmed by LOX activity assays. Biochemical characterization of the reaction products of the hazelnut LOX indicated that it is a 9-LOX. Two cDNAs were isolated by RT-PCR carried out on total RNA from immature hazelnut seeds. Sequence analysis indicated that the two cDNAs are highly homologous (91.9% degree of identity) and one of these corresponded exactly to the genomic clone. The deduced amino acid sequences of the hazelnut LOXs showed that they are closely related to a previously reported almond LOX (79.5% identity) and, to a lesser extent, to some LOXs involved in plant responses to pathogens (cotton and tobacco LOXs, 75.5 and 74.6% identity, respectively). The physiological role of hazelnut LOXs and their role in influencing seed quality are also discussed.

  13. Transporter assays and assay ontologies: useful tools for drug discovery. (United States)

    Zdrazil, Barbara; Chichester, Christine; Zander Balderud, Linda; Engkvist, Ola; Gaulton, Anna; Overington, John P


    Transport proteins represent an eminent class of drug targets and ADMET (absorption, distribution, metabolism, excretion, toxicity) associated genes. There exists a large number of distinct activity assays for transport proteins, depending on not only the measurement needed (e.g. transport activity, strength of ligand–protein interaction), but also due to heterogeneous assay setups used by different research groups. Efforts to systematically organize this (divergent) bioassay data have large potential impact in Public-Private partnership and conventional commercial drug discovery. In this short review, we highlight some of the frequently used high-throughput assays for transport proteins, and we discuss emerging assay ontologies and their application to this field. Focusing on human P-glycoprotein (Multidrug resistance protein 1; gene name: ABCB1, MDR1), we exemplify how annotation of bioassay data per target class could improve and add to existing ontologies, and we propose to include an additional layer of metadata supporting data fusion across different bioassays.

  14. Morpho-Physiological and Biochemical Criteria of Acantha-moeba spp. Isolated from the Egyptian Aquatic Environment

    Directory of Open Access Journals (Sweden)

    A Mohammed


    Full Text Available 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.Conclusio: 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.

  15. A novel 96-microwell-based high-throughput spectrophotometric assay for pharmaceutical quality control of crizotinib, a novel potent drug for the treatment of non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Tanveer Ahmed Wani


    Full Text Available This study describes the development and validation of a novel 96-microwell-based high throughput spectrophotometric assay for pharmaceutical quality control of crizotinib (CZT, a novel drug for the treatment of non-small cell lung cancer. We examined the reaction between CZT and 1,2-naphthoquinone-4-sulphonate, a chromogenic reagent. A red-colored product showing a maximum absorption peak (λmax at 490 nm was produced in an alkaline medium (pH 9. We examined stoichiometry of the reaction and postulated the reaction mechanism. To our knowledge, this is the first study to describe a color-developing reaction for the proposed assay. The reaction was performed in a 96-microwell plate, and the absorbance of the colored product was measured using an absorbance reader at 490 nm. Under optimized reaction conditions, Beer's law, which shows a correlation between absorbance and CZT concentration, was obeyed in the range of 4-50 µg/well with an appropriate correlation coefficient (0.999. The limits of detection and quantification were 1.73 and 5.23 µg/well, respectively. The assay showed high precision and accuracy. The proposed assay was applied successfully for the determination of CZT in capsules. Thus, the assay proposed in this study is practical and valuable for routine application in pharmaceutical quality control laboratories.

  16. Kombucha tea fermentation: Microbial and biochemical dynamics. (United States)

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


    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



    Bishnoi Kapil*, , ,; Kataria Mahesh; Singhal Vipin; Gupta Deepika


    Micronutrients added to foods are analyzed using various procedures depending on their nature and properties. The microbiological assays are better than chemical method because any suitable change in vitamin molecule which may not be detected by chemical method will be revealed by change in microbial activity. The microbiological assay of vitamins is based upon the comparison of the stimulation of growth of bacteria by measured concentration of vitamin with that produced by known concentratio...

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

    Directory of Open Access Journals (Sweden)

    Natalia Salvadores


    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.

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

    Directory of Open Access Journals (Sweden)

    R. Muthusamy


    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.

  20. 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

  1. Achilles tendinosis: Changes in biochemical composition and collagen turnover rate

    NARCIS (Netherlands)

    Mos, M. de; El, B. van; Groot, J. de; Jahr, H.; Schie, H.T.M. van; Arkel, E.R. van; Tol, H.; Heijboer, R.; Osch, G.J.V.M. van; Verhaar, J.A.N.


    Background: Understanding biochemical and structural changes of the extracellular matrix in Achilles tendinosis might be important for developing mechanism-based therapies. Hypothesis: In Achilles tendinosis, changes occur in biochemical composition and collagen turnover rate. Study Design: Descript

  2. Achilles tendinosis - Changes in biochemical composition and collagen turnover rate

    NARCIS (Netherlands)

    de Mos, Marieke; van El, Benno; DeGroot, Jeroen; Jahr, Holger; van Schie, Hans T. M.; van Arkel, Ewoud R.; Tol, Hans; Heijboer, Rien; van Osch, Gerjo J. V. M.; Verhaar, Jan A. N.


    Background: Understanding biochemical and structural changes of the extracellular matrix in Achilles tendinosis might be important for developing mechanism-based therapies. Hypothesis: In Achilles tendinosis, changes occur in biochemical composition and collagen turnover rate. Study Design: Descript

  3. Optical Slot-Waveguide Based Biochemical Sensors

    Directory of Open Access Journals (Sweden)

    Carlos Angulo Barrios


    Full Text Available Slot-waveguides allow light to be guided and strongly confined inside a nanometer-scale region of low refractive index. Thus stronger light-analyte interaction can be obtained as compared to that achievable by a conventional waveguide, in which the propagating beam is confined to the high-refractive-index core of the waveguide. In addition, slot-waveguides can be fabricated by employing CMOS compatible materials and technology, enabling miniaturization, integration with electronic, photonic and fluidic components in a chip, and mass production. These advantages have made the use of slot-waveguides for highly sensitive biochemical optical integrated sensors an emerging field. In this paper, recent achievements in slot-waveguide based biochemical sensing will be reviewed. These include slot-waveguide ring resonator based refractometric label-free biosensors, label-based optical sensing, and nano-opto-mechanical sensors.

  4. Biochemical changes in blood under Cr6+


    Kuzenko E.V.; Romaniuk A.M.; Butko H.Yu.; Logvinova H.V.


    Background. For the manufacture of dentures many different alloys containing chromium are used. Interaction with oral fluid, organic acids and food, results in formation of Cr3+, Cr6+ ions, but their influence on the whole organism is poorly investigated. Objective. To analyze the biochemical changes in blood plasma during the influence of Cr6+ ions. Methods. 15 animals of experimental group were receiving drinking water with potassium dichromate in a dose of 0,2 mol/l. Rats of control group ...

  5. Electronic modulation of biochemical signal generation (United States)

    Gordonov, Tanya; Kim, Eunkyoung; Cheng, Yi; Ben-Yoav, Hadar; Ghodssi, Reza; Rubloff, Gary; Yin, Jun-Jie; Payne, Gregory F.; Bentley, William E.


    Microelectronic devices that contain biological components are typically used to interrogate biology rather than control biological function. Patterned assemblies of proteins and cells have, however, been used for in vitro metabolic engineering, where coordinated biochemical pathways allow cell metabolism to be characterized and potentially controlled on a chip. Such devices form part of technologies that attempt to recreate animal and human physiological functions on a chip and could be used to revolutionize drug development. These ambitious goals will, however, require new biofabrication methodologies that help connect microelectronics and biological systems and yield new approaches to device assembly and communication. Here, we report the electrically mediated assembly, interrogation and control of a multi-domain fusion protein that produces a bacterial signalling molecule. The biological system can be electrically tuned using a natural redox molecule, and its biochemical response is shown to provide the signalling cues to drive bacterial population behaviour. We show that the biochemical output of the system correlates with the electrical input charge, which suggests that electrical inputs could be used to control complex on-chip biological processes.

  6. Haematological and biochemical analysis in canine enteritis

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    Abid Ali Bhat

    Full Text Available Aim: The present investigation screened eighteen clinical cases of canine enteritis for haematological and biochemical analyses. Materials and Methods: Eighteen dogs suffering from enteritis were selected and detailed clinical manifestations were noted. Hematological and biochemical parameters were estimated by using various kits. Blood was also collected from twelve healthy dogs for establishing control values and data obtained were subjected to statistical analysis. Results: The affected dogs showed anorexia, diarrhoea, depression, varying degree of dehydration and tachycardia. There were significant changes in packed cell volume, neutrophils, lymphocytes and mean corpuscular haemoglobin concentration. Biochemical investigation revealed significant decrease in plasma glucose, total plasma protein, albumin and albumin:globulin ratio (A:G ratio. The level of potassium and chloride was markedly decreased. Significant increase in alanine aminotransferase (ALT and blood urea nitrogen (BUN was observed. Conclusion: Packed Cell Volume (PCV and Total Erythrocyte Count (TEC remained almost similar between healthy dogs and dogs affected with diarrhoea. Mean Total Leukocyte Count (TLC value was significantly higher as compared to the control group. Hypoglycemia, hypoproteinemia, hypokalemia, hypochloremia and increase in blood urea nitrogen was observed in dogs suffering from enteritis. [Vet World 2013; 6(7.000: 380-383

  7. Explorations into Chemical Reactions and Biochemical Pathways. (United States)

    Gasteiger, Johann


    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.

  8. [Basic biochemical processes in glaucoma progression]. (United States)

    von Thun und Hohenstein-Blaul, N; Kunst, S; Pfeiffer, N; Grus, F H


    The term glaucoma summarizes a group of eye diseases that are accompanied by impairments of the optic nerve and related visual field deficits. An early diagnosis of glaucoma is currently not possible due to a lack of diagnostic tests; therefore, in most cases the disease is diagnosed many years after onset, which prevents an early therapy. The known risk factors for the development and progression of glaucomatous optic neuropathy comprise elevated intraocular pressure and a broad range of pressure fluctuations as well as lipometabolic disorders, genetic factor and diabetes. The consequences include the induction of anti-inflammatory proteins, elevated levels of oxidative stress and the destruction of retinal ganglion cells. Changes in the autoantibody repertoire have also been observed in the course of the disease. Basic ophthalmological research therefore focuses on the investigation of basic biochemical processes in the course of the disease. A better understanding of physiological and biochemical events is sought in order to develop new and more sensitive diagnostic options and to allow more targeted therapeutic measures. The understanding of biochemical processes allows a better insight into glaucoma progression to be gained, which will lead to improvements in diagnosis and therapy.

  9. Comparative evaluation of a chromogenic agar medium-PCR protocol with a conventional method for isolation of Vibrio parahaemolyticus strains from environmental and clinical samples. (United States)

    Canizalez-Roman, Adrian; Flores-Villaseñor, Héctor; Zazueta-Beltran, Jorge; Muro-Amador, Secundino; León-Sicairos, Nidia


    Screening for pathogenic Vibrio parahaemolyticus has become routine in certain areas associated with food-borne outbreaks. This study is an evaluation of the CHROMagar Vibrio (CV) medium-PCR protocol and the conventional method (TCBS (thiosulfate-citrate-bile salts-sucrose) agar plus biochemical and Wagatsuma agar tests) for detection of V. parahaemolyticus in shrimp, water, sediment, and stool samples collected for biosurveillance in an endemic area of northwestern Mexico. A total of 131 environmental and clinical samples were evaluated. The CV medium-PCR protocol showed a significantly improved ability (P TDH) in developing countries. In our results, Wagatsuma agar showed low sensitivity (65.4% at 24 h and 75.6% at 48 h) and specificity (52.4% at 48 h) for identifying V. parahaemolyticus positive for TDH. Overall, our data support the use of the CV medium-PCR protocol in place of the conventional method (TCBS-biochemical tests-Wagatsuma agar) for detection of pathogenic V. parahaemolyticus, both in terms of effectiveness and cost efficiency.

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

    Directory of Open Access Journals (Sweden)

    Carina Ladeira


    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.

  11. Cupric ion reducing antioxidant capacity assay for antioxidants in human serum and for hydroxyl radical scavengers. (United States)

    Apak, Reşat; Güçlü, Kubilay; Ozyürek, Mustafa; Bektaşoğlu, Burcu; Bener, Mustafa


    Tests measuring the combined antioxidant effect of the nonenzymatic defenses in biological fluids may be useful in providing an index of the organism's capability to counteract reactive species known as pro-oxidants, resist oxidative damage, and combat oxidative stress-related diseases. The selected chromogenic redox reagent for the assay of human serum should be easily accessible, stable, selective, and respond to all types of biologically important antioxidants such as ascorbic acid, alpha-tocopherol, beta-carotene, reduced glutathione (GSH), uric acid, and bilirubin, regardless of chemical type or hydrophilicity. Our recently developed cupric reducing antioxidant capacity (CUPRAC) spectrophotometric method for a number of polyphenols and flavonoids using the copper(II)-neocuproine reagent in ammonium acetate buffer is now applied to a complete series of plasma antioxidants for the assay of total antioxidant capacity of serum, and the resulting absorbance at 450 nm is recorded either directly (e.g., for ascorbic acid, alpha-tocopherol, and glutathione) or after incubation at 50 degrees C for 20 min (e.g., for uric acid, bilirubin, and albumin), quantitation being made by means of a calibration curve. The lipophilic antioxidants, alpha-tocopherol and beta-carotene, are assayed in dichloromethane. Lipophilic antioxidants of serum are extracted with n-hexane from an ethanolic solution of serum subjected to centrifugation. Hydrophilic antioxidants of serum are assayed in the centrifugate after perchloric acid precipitation of proteins. The CUPRAC molar absorptivities, linear ranges, and TEAC (trolox equivalent antioxidant capacity) coefficients of the serum antioxidants are established, and the results are evaluated in comparison with the findings of the ABTS/TEAC reference method. The intra- and inter-assay coefficients of variation (CVs) are 0.7 and 1.5%, respectively, for serum. The CUPRAC assay proved to be efficient for glutathione and thiol-type antioxidants

  12. Leishmania (Viannia panamensis expresses a nuclease with molecular and biochemical features similar to the Endonuclease G of higher eukaryotes*

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    Miguel A Toro-Londoño


    Full Text Available Objective: To characterize the molecular and biochemical features of the Endonuclease G of Leishmania (Viannia panamensis. Methods: The gene of the putative L. (V. panamensis Endonuclease G was amplified, cloned, and sequenced. The recombinant protein was produced in a heterologous expression system and biochemical assays were run to determine its ion, temperature, and pH preferences. Results: The L. (V. panamensis rENDOG has biochemical features similar to those found in other trypanosomatids and higher eukaryotes. In addition, phylogenetic analysis revealed a possible evolutionary relationship with metazoan ENDOG. Conclusions: L. (V. panamensis has a gene that codifies an ENDOG homologous to those of higher organisms. This enzyme can be produced in Escherichia coli and is able to degrade covalently closed circular double-stranded DNA. It has a magnesium preference, can be inhibited by potassium, and is able to function within a wide temperature and pH range.

  13. Leishmania (Viannia panamensis expresses a nuclease with molecular and biochemical features similar to the Endonuclease G of higher eukaryotes

    Directory of Open Access Journals (Sweden)

    Miguel A. Toro-Londoño


    Full Text Available Objective: To characterize the molecular and biochemical features of the Endonuclease G of Leishmania (Viannia panamensis.Methods: The gene of the putative L. (V. panamensis Endonuclease G was amplified, cloned, and sequenced. The recombinant protein was produced in a heterologous expression system and biochemical assays were run to determine its ion, temperature, and pH preferences.Results: The L. (V. panamensis rENDOG has biochemical features similar to those found in other trypanosomatids and higher eukaryotes. In addition, phylogenetic analysis revealed a possible evolutionary relationship with metazoan ENDOG.Conclusions: L. (V. panamensis has a gene that codifies an ENDOG homologous to those of higher organisms. This enzyme can be produced in Escherichia coli and is able to degrade covalently closed circular double-stranded DNA. It has a magnesium preference, can be inhibited by potassium, and is able to function within a wide temperature and pH range.

  14. The Use of Item Analysis for Improvement of Biochemical Teaching (United States)

    Nagata, Ryoichi


    Item analysis was used to find out which biochemical explanations need to be improved in biochemical teaching, not which items are to be discarded, improved, or reusable in biochemical examinations. The analysis revealed the basic facts of which less able students had more misunderstanding than able students. Identifying these basic facts helps…

  15. Cystatin C: A new biochemical marker in livestock sector

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    Pravas Ranjan Sahoo


    Full Text Available The livestock sector contributes largely to the economy of India. Different systemic diseases like renal diseases, neurological and cardiovascular diseases cause huge loss in production and productive potential of livestock in India, which is considered as a major concern for both small and large ruminants. Early detection of diseseses is essential to combat the economic loss. An efficient biochemical marker can be developed which would provide more specific, sensitive and reliable measurement of functions of different organs. Determination of endogenous marker Cystatin C may fulfill the above need which can provide a detection platform not only for Kidney function but also for assaying other organs' function. Cystatin C is a low molecular weight protein which is removed from the bloodstream by glomerular filtration in the kidneys. Thus, it may act as a potential biological tool in diagnosis of renal and other systemic diseases in livestock. This mini-review focuses on the Cystatin C and its clinical importance which can be extensively employed in the livestock sector. [J Adv Vet Anim Res 2016; 3(3.000: 200-205

  16. Balanced biochemical reactions: a new approach to unify chemical and biochemical thermodynamics.

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    Antonio Sabatini

    Full Text Available A novel procedure is presented which, by balancing elements and electric charge of biochemical reactions which occur at constant pH and pMg, allows assessing the thermodynamics properties of reaction Δ(rG'⁰, Δ(rH'⁰, Δ(rS'⁰ and the change in binding of hydrogen and magnesium ions of these reactions. This procedure of general applicability avoids the complex calculations required by the use of the Legendre transformed thermodynamic properties of formation Δ(fG'⁰, Δ(fH'⁰ and Δ(fS'⁰ hitherto considered an obligatory prerequisite to deal with the thermodynamics of biochemical reactions. As a consequence, the term "conditional" is proposed in substitution of "Legendre transformed" to indicate these thermodynamics properties. It is also shown that the thermodynamic potential G is fully adequate to give a criterion of spontaneous chemical change for all biochemical reactions and then that the use of the Legendre transformed G' is unnecessary. The procedure proposed can be applied to any biochemical reaction, making possible to re-unify the two worlds of chemical and biochemical thermodynamics, which so far have been treated separately.

  17. Biochemical and biological characterization of two serine proteinases from Colombian Crotalus durissus cumanensis snake venom. (United States)

    Patiño, Arley Camilo; Pereañez, Jaime Andrés; Gutiérrez, José María; Rucavado, Alexandra


    Two clotting serine proteinases, named Cdc SI and Cdc SII, were isolated and characterized for the first time from Colombian Crotalus durissus cumanensis snake venom. The enzymes were purified using two chromatographic steps: molecular exclusion on Sephacryl S-200 and RP-HPLC on C8 Column. The molecular masses of the proteins, determined by MALDI-TOF mass spectrometry, were 28,561.4 and 28,799.2 Da for Cdc SI and Cdc SII, respectively. The aim of the present study was to evaluate enzymatic, coagulant and toxic properties of the two enzymes. The serine proteinases hydrolyzed specific chromogenic substrate (BaPNA) and exhibited a Michaelis-Menten behavior. Cdc SI had V(max) of 0.038 ± 0.003 nmol/min and K(M) of 0.034 ± 0.017 mM, while Cdc SII displayed values of V(max) of 0.267 ± 0.011 nmol/min and K(M) of 0.145 ± 0.023 mM. N-terminal sequences were VIGGDEXNIN and VIGGDICNINEHNFLVALYE for Cdc SI and Cdc SII, respectively. Molecular masses, N-terminal sequences, inhibition assays, and enzymatic profile suggest that Cdc SI and Cdc SII belong to the family of snake venom thrombin-like enzymes. These serine proteinases differed in their clotting activity on human plasma, showing a minimum coagulant dose of 25 μg and 0.571 μg for Cdc SI and Cdc SII, respectively. Enzymes also showed coagulant activity on bovine fibrinogen and degraded chain α of this protein. Toxins lack hemorrhagic and myotoxic activities, but are capable to induce defibrin(ogen)ation, moderate edema, and an increase in vascular permeability. These serine proteinases may contribute indirectly to the local hemorrhage induced by metalloproteinases, by causing blood clotting disturbances, and might also contribute to cardiovascular alterations characteristic of patients envenomed by C. d. cumanensis in Colombia.

  18. Dual-color plasmonic enzyme-linked immunosorbent assay based on enzyme-mediated etching of Au nanoparticles (United States)

    Guo, Longhua; Xu, Shaohua; Ma, Xiaoming; Qiu, Bin; Lin, Zhenyu; Chen, Guonan


    Colorimetric enzyme-linked immunosorbent assay utilizing 3‧-3-5‧-5-tetramethylbenzidine(TMB) as the chromogenic substrate has been widely used in the hospital for the detection of all kinds of disease biomarkers. Herein, we demonstrate a strategy to change this single-color display into dual-color responses to improve the accuracy of visual inspection. Our investigation firstly reveals that oxidation state of 3‧-3-5‧-5-tetramethylbenzidine (TMB2+) can quantitatively etch gold nanoparticles. Therefore, the incorporation of gold nanoparticles into a commercial TMB-based ELISA kit could generate dual-color responses: the solution color varied gradually from wine red (absorption peak located at ~530 nm) to colorless, and then from colorless to yellow (absorption peak located at ~450 nm) with the increase amount of targets. These dual-color responses effectively improved the sensitivity as well as the accuracy of visual inspection. For example, the proposed dual-color plasmonic ELISA is demonstrated for the detection of prostate-specific antigen (PSA) in human serum with a visual limit of detection (LOD) as low as 0.0093 ng/mL.

  19. Barcoded microchips for biomolecular assays. (United States)

    Zhang, Yi; Sun, Jiashu; Zou, Yu; Chen, Wenwen; Zhang, Wei; Xi, Jianzhong Jeff; Jiang, Xingyu


    Multiplexed assay of analytes is of great importance for clinical diagnostics and other analytical applications. Barcode-based bioassays with the ability to encode and decode may realize this goal in a straightforward and consistent manner. We present here a microfluidic barcoded chip containing several sets of microchannels with different widths, imitating the commonly used barcode. A single barcoded microchip can carry out tens of individual protein/nucleic acid assays (encode) and immediately yield all assay results by a portable barcode reader or a smartphone (decode). The applicability of a barcoded microchip is demonstrated by human immunodeficiency virus (HIV) immunoassays for simultaneous detection of three targets (anti-gp41 antibody, anti-gp120 antibody, and anti-gp36 antibody) from six human serum samples. We can also determine seven pathogen-specific oligonucleotides by a single chip containing both positive and negative controls.

  20. Radioreceptor assay method for insulin

    Energy Technology Data Exchange (ETDEWEB)

    Mori, K.F.; Wood, R.J. (Bureau of Drug Research, Health and Welfare Canada, Ottawa, Ontario. Health Protection Branch)


    A sensitive practical radioreceptor assay method for pharmaceutical insulin products has been developed with partially purified rat liver plasma membranes and the optimal conditions under which the best overall assay performance is obtainable have been defined. Intra- and inter-assay variations of the method averaged 7.3 and 12.2% of the man, respectively, when expressed as the coefficient of variation. Potency estimates of an insulin product obtained with the proposed method correlated well with those determined by the mouse convulsion bioassay method. Liver membranes prepared according to the method could be stored for up to ten weeks at 4/sup 0/C and for 6 months or more at -18/sup 0/C without losing insulin-binding ability.

  1. Biochemical Markers for Assessing Aquatic Contamination

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    Zdeňka Svobodová


    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.

  2. BNDB – The Biochemical Network Database

    Directory of Open Access Journals (Sweden)

    Kaufmann Michael


    Full Text Available Abstract Background Technological advances in high-throughput techniques and efficient data acquisition methods have resulted in a massive amount of life science data. The data is stored in numerous databases that have been established over the last decades and are essential resources for scientists nowadays. However, the diversity of the databases and the underlying data models make it difficult to combine this information for solving complex problems in systems biology. Currently, researchers typically have to browse several, often highly focused, databases to obtain the required information. Hence, there is a pressing need for more efficient systems for integrating, analyzing, and interpreting these data. The standardization and virtual consolidation of the databases is a major challenge resulting in a unified access to a variety of data sources. Description We present the Biochemical Network Database (BNDB, a powerful relational database platform, allowing a complete semantic integration of an extensive collection of external databases. BNDB is built upon a comprehensive and extensible object model called BioCore, which is powerful enough to model most known biochemical processes and at the same time easily extensible to be adapted to new biological concepts. Besides a web interface for the search and curation of the data, a Java-based viewer (BiNA provides a powerful platform-independent visualization and navigation of the data. BiNA uses sophisticated graph layout algorithms for an interactive visualization and navigation of BNDB. Conclusion BNDB allows a simple, unified access to a variety of external data sources. Its tight integration with the biochemical network library BN++ offers the possibility for import, integration, analysis, and visualization of the data. BNDB is freely accessible at

  3. An assay for determining minimal concentrations of antibiotics that drive horizontal transfer of resistance. (United States)

    Jutkina, Jekaterina; Rutgersson, Carolin; Flach, Carl-Fredrik; Larsson, D G Joakim


    Ability to understand the factors driving horizontal transfer of antibiotic resistance from unknown, harmless bacteria to pathogens is crucial in order to tackle the growing resistance problem. However, current methods to measure effects of stressors on horizontal gene transfer have limitations and often fall short, as the estimated endpoints can be a mix of both the number of transfer events and clonal growth of transconjugants. Our aim was therefore to achieve a proper strategy for assessing the minimal concentration of a stressor (exemplified by tetracycline) that drives horizontal transfer of antibiotic resistance from a complex community to a model pathogen. Conditions were optimized to improve a culture-based approach using the bacterial community of treated sewage effluent as donor, and fluorescent, traceable Escherichia coli as recipient. Reduced level of background resistance, differentiation of isolates as well as decreased risk for measuring effects of selection were achieved through the use of chromogenic medium, optimization of conjugation time as well as applying a different antibiotic for isolation of transconjugants than the one tested for its ability to drive transfer. Using this assay, we showed that a very low concentration of tetracycline, 10μg/L i.e. 150 times below the minimal inhibitory concentration of the recipient, promoted horizontal transfer of multiple antibiotic-resistance determinants. Higher concentrations favoured selection of a tetracycline-resistance phenotype along with a decline in the number of detectable transfer events. The described method can be used to evaluate different environmental conditions and factors that trigger horizontal dissemination of mobile resistance elements, eventually resulting in the formation of drug-resistant pathogens.

  4. [Chronic fatigue syndrome: biochemical examination of blood]. (United States)

    Hakariya, Yukiko; Kuratsune, Hirohiko


    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.

  5. Azoospermia: clinical, hormonal, and biochemical investigation. (United States)

    Papadimas, J; Papadopoulou, F; Ioannidis, S; Spanos, E; Tarlatzis, B; Bontis, J; Mantalenakis, S


    The aim of this study was to evaluate the clinical, hormonal and biochemical characteristics of infertile men with azoospermia. A total of 187 azoospermic out of 2610 infertile men (7.2%) were studied. Mean testicular volume and basal plasma levels of FSH were the most useful parameters concerning the evaluation of azoospermia. Basal plasma levels of LH and T were useful only in azoospermic men with hypogonadism, whereas plasma PRL levels, semen volume, and seminal plasma fructose levels were not found to be of common use except in selected cases.

  6. Biochemical Disincentives to Fertilizing Cellulosic Ethanol Crops (United States)

    Gallagher, M. E.; Hockaday, W. C.; Snapp, S.; McSwiney, C.; Baldock, J.


    Corn grain biofuel crops produce the highest yields when the cropping ecosystem is not nitrogen (N)-limited, achieved by application of fertilizer. There are environmental consequences for excessive fertilizer application to crops, including greenhouse gas emissions, hypoxic “dead zones,” and health problems from N runoff into groundwater. The increase in corn acreage in response to demand for alternative fuels (i.e. ethanol) could exacerbate these problems, and divert food supplies to fuel production. A potential substitute for grain ethanol that could reduce some of these impacts is cellulosic ethanol. Cellulosic ethanol feedstocks include grasses (switchgrass), hardwoods, and crop residues (e.g. corn stover, wheat straw). It has been assumed that these feedstocks will require similar N fertilization rates to grain biofuel crops to maximize yields, but carbohydrate yield versus N application has not previously been monitored. We report the biochemical stocks (carbohydrate, protein, and lignin in Mg ha-1) of a corn ecosystem grown under varying N levels. We measured biochemical yield in Mg ha-1 within the grain, leaf and stem, and reproductive parts of corn plants grown at seven N fertilization rates (0-202 kg N ha-1), to evaluate the quantity and quality of these feedstocks across a N fertilization gradient. The N fertilization rate study was performed at the Kellogg Biological Station-Long Term Ecological Research Site (KBS-LTER) in Michigan. Biochemical stocks were measured using 13C nuclear magnetic resonance spectroscopy (NMR), combined with a molecular mixing model (Baldock et al. 2004). Carbohydrate and lignin are the main biochemicals of interest in ethanol production since carbohydrate is the ethanol feedstock, and lignin hinders the carbohydrate to ethanol conversion process. We show that corn residue carbohydrate yields respond only weakly to N fertilization compared to grain. Grain carbohydrate yields plateau in response to fertilization at

  7. Biochemical Markers of Joint Tissue Turnover

    DEFF Research Database (Denmark)

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


    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...... available for the study of tissue turnover in each of the three compartments of the articular joint, that is the bone, the cartilage, and the synovium. Finally, we provide some perspective to future developments in biomarker discovery and discuss the potential impact such technologies could have on the drug...

  8. Biochemical composition of tangerine fruits under microfertilizers


    Oksana Belous; Yuliya Abilfazova


    The paper presents the long-term research and its results in the field of biochemical composition and mechanical analysis of dwarf tangerine fruits ('Miagava-Vase') growing in the subtropical zone of the Black Sea coast, Krasnodar region. The given results have been obtained after treatments with the following micro fertilizers: H3BO3 (at a concentration of 0.06%), MnSO4 H2O (0.4%), ZnSO4 H2O (0.3%) and CuSO4 H2O (0.06%), an option with foliar water spraying served as a control. It is shown t...

  9. Fluorescence monitoring of riboswitch transcription regulation using a dual molecular beacon assay


    Chinnappan, Raja; Dubé, Audrey; Lemay, Jean-François; Daniel A Lafontaine


    Riboswitches are mRNA elements that specifically bind cellular metabolites and control gene expression by modifying their structure. As riboswitches often control essential genes in pathogenic bacteria, riboswitches have been proposed as new targets for antibiotics. High-throughput screening provides a powerful approach to identify riboswitch ligand analogs that could act as powerful antibacterial drugs. Biochemical assays have already been used to find riboswitch-binding analogs, but those m...

  10. A rapid phospholipase D assay using zirconium precipitation of anionic substrate phospholipids

    DEFF Research Database (Denmark)

    Petersen, G.; Hansen, Harald S.; Chapman, K.D.


    Activation of phospholipase D (PLD) is involved in a number of signal transduction pathways in eukaryotic cells. The most common method for determination of PLD activity in vitro involves incubation with a radiolabeled substrate and lipid extraction followed by thin-layer chromatography in order ...... biochemical characterization of this anandamide-generating enzyme. - Petersen, G., K. D. Chapman, and H. S. Hansen. A rapid phospholipase D assay using zirconium precipitation of anionic substrate phospholipids: Application to N-acylethanolamine formation in vitro....

  11. Chromosome aberration assays in Allium

    Energy Technology Data Exchange (ETDEWEB)

    Grant, W.F.


    The common onion (Allium cepa) is an excellent plant for the assay of chromosome aberrations after chemical treatment. Other species of Allium (A. cepa var. proliferum, A. carinatum, A. fistulosum and A. sativum) have also been used but to a much lesser extent. Protocols have been given for using root tips from either bulbs or seeds of Allium cepa to study the cytological end-points, such as chromosome breaks and exchanges, which follow the testing of chemicals in somatic cells. It is considered that both mitotic and meiotic end-points should be used to a greater extent in assaying the cytogenetic effects of a chemical. From a literature survey, 148 chemicals are tabulated that have been assayed in 164 Allium tests for their clastogenic effect. Of the 164 assays which have been carried out, 75 are reported as giving a positive reaction, 49 positive and with a dose response, 1 positive and temperature-related, 9 borderline positive, and 30 negative; 76% of the chemicals gave a definite positive response. It is proposed that the Allium test be included among those tests routinely used for assessing chromosomal damage induced by chemicals.

  12. Disagreement between Human Papillomavirus Assays

    DEFF Research Database (Denmark)

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


    We aimed to determine the disagreement in primary cervical screening between four human papillomavirus assays: Hybrid Capture 2, cobas, CLART, and APTIMA. Material from 5,064 SurePath samples of women participating in routine cervical screening in Copenhagen, Denmark, was tested with the four ass...

  13. Rapid differentiation between clinically relevant mycobacteria in microscopy positive clinical specimens and mycobacterial isolates by line probe assay

    DEFF Research Database (Denmark)

    Johansen, Isik Somuncu; Lundgren, Bettina H; Thyssen, Jacob Pontoppidan;


    The Inno LiPA Mycobacteria assay, based on PCR amplification of the 16-23S rRNA spacer region of Mycobacterium species, has been designed for identification of mycobacteria grown in culture media and discrimination between Mycobacterium tuberculosis complex, M. avium, M. intracellulare, M. kansasii...... of conventional identification using 16S rDNA analysis and biochemical properties. The assay only misidentified one strain, which was found to be M. avium complex instead of M. intracellulare as found by the conventional tests. The assay allows rapid discrimination of the eight most clinically relevant...

  14. Pheochromocytoma-paraganglioma: Biochemical and genetic diagnosis. (United States)

    Cano Megías, Marta; Rodriguez Puyol, Diego; Fernández Rodríguez, Loreto; Sención Martinez, Gloria Lisette; Martínez Miguel, Patricia

    Pheochromocytomas and paragangliomas are tumours derived from neural crest cells, which can be diagnosed by biochemical measurement of metanephrine and methoxytyramine. Advances in genetic research have identified many genes involved in the pathogenesis of these tumours, suggesting that up to 35-45% may have an underlying germline mutation. These genes have a singular transcriptional signature and can be grouped into 2 clusters (or groups): cluster 1 (VHL and SHDx), involved in angiogenesis and hypoxia pathways; and cluster 2 (MEN2 and NF1), linked to the kinase signalling pathway. In turn, these genes are associated with a characteristic biochemical phenotype (noradrenergic and adrenergic), and clinical features (location, biological behaviour, age of presentation, etc.) in a large number of cases. Early diagnosis of these tumours, accompanied by a correct genetic diagnosis, should eventually become a priority to enable better treatment, early detection of complications, proper screening of family members and related tumours, as well as an improvement in the overall prognosis of these patients.

  15. Biotin: biochemical, physiological and clinical aspects. (United States)

    Said, Hamid M


    Significant progress has been made in our understanding of the biochemical, physiological and nutritional aspects of the water-soluble vitamin biotin (vitamin H). It is well know now that biotin plays important roles in a variety of critical metabolic reactions in the cell, and thus, is essential for normal human health, growth and development. This is underscored by the serious clinical abnormalities that occur in conditions of biotin deficiency, which include, among other things, growth retardation, neurological disorders, and dermatological abnormalities (reviewed in 1). Studies in animals have also shown that biotin deficiency during pregnancy leads to embryonic growth retardation, congenital malformation and death (Watanabe 1983; Cooper and Brown 1958; Mock et al. 2003; Zempleni and Mock 2000). The aim of this chapter is to provide coverage of current knowledge of the biochemical, physiological, and clinical aspects of biotin nutrition. Many sections of this chapter have been the subject of excellent recent reviews by others (Wolf 2001; McMahon 2002; Mock 2004; Rodriguez-Melendez and Zempleni 2003; Said 2004; Said et al. 2000; Said and Seetheram 2006), and thus, for more information the reader is advised to consider these additional sources.

  16. Biochemical research elucidating metabolic pathways in Pneumocystis*

    Directory of Open Access Journals (Sweden)

    Kaneshiro E.S.


    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.

  17. Biochemical Manifestation of HIV Lipodystrophy Syndrome

    Directory of Open Access Journals (Sweden)

    Kenneth Ihenetu, PhD


    Full Text Available Objectives: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.Methods: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.Results: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.Conclusions and Public Health Implications: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.

  18. [Biochemical principles of early saturnism recognition]. (United States)

    Tsimakuridze, M P; Mansuradze, E A; Zurashvili, D G; Tsimakuridze, M P


    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.

  19. Serum biochemical markers in carcinoma breast.

    Directory of Open Access Journals (Sweden)

    Seth R


    Full Text Available BACKGROUND: Despite the extensive research for many years throughout the world, the etiopathogenesis of cancer still remains obscure. For the early detection of carcinoma of various origins, a number of biochemical markers have been studied to evaluate the malignancy. AIM: To analyse serum gamma glutamyl transpeptidase (GGTP, lactate dehydrogenase (LDH and superoxide dismutase (SOD in carcinoma breast patients. SETTINGS & DESIGN: The serum biochemical markers were estimated in twenty five histopathologically confirmed patients with carcinoma breast and equal number of healthy age- matched individuals served as control. MATERIAL & METHODS: Serum gamma glutamyl transpeptidase (GGTP, lactate dehydrogenase (LDH and superoxide dismutase (SOD were estimated and their sensitivity determined. Statistics: Data was analysed with student′s ′t′-test and sensitivity score of these markers was determined. RESULTS & CONCLUSIONS: The mean serum GGTP, LDH and SOD activities in patients with carcinoma breast were tremendously increased as compared to controls, and a steady increase was observed in their activities from stage I through stage IV as well as following distant metastasis. Serum GGTP, LDH and SOD might prove to be most sensitive biomarkers in carcinoma breast in early detection of the disease.

  20. A High-Throughput Assay for Rho Guanine Nucleotide Exchange Factors Based on the Transcreener GDP Assay. (United States)

    Reichman, Melvin; Schabdach, Amanda; Kumar, Meera; Zielinski, Tom; Donover, Preston S; Laury-Kleintop, Lisa D; Lowery, Robert G


    Ras homologous (Rho) family GTPases act as molecular switches controlling cell growth, movement, and gene expression by cycling between inactive guanosine diphosphate (GDP)- and active guanosine triphosphate (GTP)-bound conformations. Guanine nucleotide exchange factors (GEFs) positively regulate Rho GTPases by accelerating GDP dissociation to allow formation of the active, GTP-bound complex. Rho proteins are directly involved in cancer pathways, especially cell migration and invasion, and inhibiting GEFs holds potential as a therapeutic strategy to diminish Rho-dependent oncogenesis. Methods for measuring GEF activity suitable for high-throughput screening (HTS) are limited. We developed a simple, generic biochemical assay method for measuring GEF activity based on the fact that GDP dissociation is generally the rate-limiting step in the Rho GTPase catalytic cycle, and thus addition of a GEF causes an increase in steady-state GTPase activity. We used the Transcreener GDP Assay, which relies on selective immunodetection of GDP, to measure the GEF-dependent stimulation of steady-state GTP hydrolysis by small GTPases using Dbs (Dbl's big sister) as a GEF for Cdc42, RhoA, and RhoB. The assay is well suited for HTS, with a homogenous format and far red fluorescence polarization (FP) readout, and it should be broadly applicable to diverse Rho GEF/GTPase pairs.

  1. Assays to measure nanomolar levels of the renin inhibitor CGP 38 560 in plasma

    Energy Technology Data Exchange (ETDEWEB)

    Cumin, F.; de Gasparo, M.; Wood, J.M.; Schnell, C.; Frueh, F.; Graf, P. (Ciba-Geigy Limited, Basel (Switzerland))


    A radioinhibitor binding assay and an enzyme inhibition assay have been developed to measure plasma levels of CGP 38 560, a potent human renin inhibitor. The detection limit of the assays was between 0.5 and 1 pmol/ml. There was a good correlation (r = 0.989) between the two assays for the measurement of human plasma spiked with CGP 38 560 in concentrations from 1.9 nM to 12 microM. Intra-assay variability was 6.1-17.3% and 4.4-27.2% for the radioinhibitor binding assay and the enzyme inhibition assay, respectively. Interassay variability was 6.0-28.2% and 3.8-28.4% for the radioinhibitor binding assay and the enzyme inhibition assay, respectively. Blood samples were collected during a pharmacological study performed in normotensive human volunteers on an unrestricted diet who were infused during a 30-minute period with CGP 38 560 A (50 micrograms/kg). Similar values for the concentrations of renin inhibitor in plasma were obtained with the radioinhibitor binding assay and the enzyme inhibitor assay, and there was a significant correlation between values obtained with the two different methodologies (r = 0.94). The plasma levels of renin inhibitor reached a maximum at the end of infusion and then decreased rapidly, indicating a short plasma half-life. The changes in biochemical parameters, plasma renin activity, and plasma concentration of active renin could be related to the concentrations of CGP 38 560 measured in the plasma.

  2. 21 CFR 225.158 - Laboratory assays. (United States)


    ... 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...

  3. Content Determination of Bacterial Endotoxin in Gatifloxacin by Kinetic Chromogenic Analysis%动态显色法定量测定加替沙星原料药中细菌内毒素含量

    Institute of Scientific and Technical Information of China (English)

    夏博; 郭锋


    Objective:To establish a method for the detection of bacterial endotoxins in gatifloxacin.Methods:Kinetic Chromogenic analysis was adopted to generate the standard curve and dilution rate of test sample.Bacterial endotoxin was added into test sample for in-terference test, and bacterial endotoxins in samples were determined.Results:The endotoxin test wasn′t interfered at concentrations be-low 0.312 mg・mL-1 .The recovery rate of bacterial endotoxin was 50%~200%.The concentration of endotoxin in the 3 batches of sam-ples met the requirements for bacterial endotoxins.Conlusion:The kinetic chromogenic analysis is suitable for bacterial endotoxin test of iodixanol injection.%目的:应用动态显色法定量测定加替沙星中的细菌内毒素含量。方法:建立动态显色法测定细菌内毒素标准曲线,通过外加内毒素测回收试验,对样品预处理方法进行了验证,证实供试液制备过程不会破坏样品本底的细菌内毒素;通过干扰试验确定样品浓度检测范围,并对样品的细菌内毒素进行定量测定。结果:样品在0.312 mg・mL-1以下浓度时,对试验无干扰作用,细菌内毒素回收率在50%~200%内,样品中的内毒素含量可定量测定。结论:动态显色法可用于定量检测加替沙星中的细菌内毒素。

  4. Haematological and Serum Biochemical Parameters of Local Turkey Poults Fed Diets Containing Two Varieties of Sorghum

    Directory of Open Access Journals (Sweden)

    E.B. Etuk


    Full Text Available These studies were conducted to determine the effects of two varieties of sorghum, Samsorg 17 and ICSV 400 on the haematological and serum biochemical parameters of local turkey breeds, reared in Nigeria. Two hundred and sixteen poults were divided into 9 treatment groups of 24 each, which were further replicated thrice and fed starter diets containing Samsorg 17 and ICSV 400. Similar (P > 0.05 RBC and PCV values were obtained with the two diets. Samsorg 17 fed poults produced lower, though not significantly (P > 0.05 serum albumin, glucose, urea, creatinine, sodium, chloride, ALP, SGPT and SGOT values than those on ICSV 400 diet. Higher RBC, MCHC, MCH, MCV and PCV values were observed with Samsorg 17 fed turkeys than those on ICSV 400 diets. Serum glucose and creatinine decreased and SGOT increased with dietary sorghum. Similar (p > 0.05 Hb, WBC, MCHC, MCV and PCV values were obtained in all groups. Values of serum biochemical indices assayed except urea, calcium, potassium and chloride showed no significant (p > 0.05 differences among the treatment groups. It was therefore concluded that Samsorg 17 and ICSV 400 sorghum varieties could sustain local turkey production without any on toward effects on their haematological and serum biochemical indices.

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

    Directory of Open Access Journals (Sweden)

    Kawahara Ryuji


    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

  6. The skin-blanching assay. (United States)

    Smit, P; Neumann, H A M; Thio, H B


    The skin-blanching assay is used for the determination and bioequivalence of dermatologic glucocorticoids (GCs). The exact mechanism of the production of blanching is not fully understood, but it is considered that local vasoconstriction of the skin microvasculature and the consequent blood-flow reduction cause this phenomenon. Several factors influence skin blanching, including drug concentration, duration of application, nature of vehicle, occlusion, posture and location. The intensity of vasoconstriction can be measured in several ways: visual or quantitative methods, such as reflectance spectroscopy, thermography, laser Doppler velocimetry and chromametry. In literature, contradicting results in the correlation of the skin-blanching assay with different tests to determine GC sensitivity have been reported, limiting its clinical usefulness.

  7. Comet Assay in Cancer Chemoprevention. (United States)

    Santoro, Raffaela; Ferraiuolo, Maria; Morgano, Gian Paolo; Muti, Paola; Strano, Sabrina


    The comet assay can be useful in monitoring DNA damage in single cells caused by exposure to genotoxic agents, such as those causing air, water, and soil pollution (e.g., pesticides, dioxins, electromagnetic fields) and chemo- and radiotherapy in cancer patients, or in the assessment of genoprotective effects of chemopreventive molecules. Therefore, it has particular importance in the fields of pharmacology and toxicology, and in both environmental and human biomonitoring. It allows the detection of single strand breaks as well as double-strand breaks and can be used in both normal and cancer cells. Here we describe the alkali method for comet assay, which allows to detect both single- and double-strand DNA breaks.

  8. Optical Assay of Erythrocyte Function in Banked Blood (United States)

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


    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.

  9. Protein binding assay for hyaluronate

    Energy Technology Data Exchange (ETDEWEB)

    Lacy, B.E.; Underhill, C.B.


    A relatively quick and simple assay for hyaluronate was developed using the specific binding protein, hyaluronectin. The hyaluronectin was obtained by homogenizing the brains of Sprague-Dawley rats, and then centrifuging the homogenate. The resulting supernatant was used as a source of crude hyaluronectin. In the binding assay, the hyaluronectin was mixed with (/sup 3/H)hyaluronate, followed by an equal volume of saturated (NH/sub 4/)/sub 2/SO/sub 4/, which precipitated the hyaluronectin and any (/sup 3/H)hyaluronate associated with it, but left free (/sup 3/H)hyaluronate in solution. The mixture was then centrifuged, and the amount of bound (/sup 3/H)hyaluronate in the precipitate was determined. Using this assay, the authors found that hyaluronectin specifically bound hyaluronate, since other glycosaminoglycans failed to compete for the binding protein. In addition, the interaction between hyaluronectin and hyaluronate was of relatively high affinity, and the size of the hyaluronate did not appear to substantially alter the amount of binding. To determine the amount of hyaluronate in an unknown sample, they used a competition assay in which the binding of a set amount of (/sup 3/H)hyaluronate was blocked by the addition of unlabeled hyaluronate. By comparing the degree of competition of the unknown samples with that of known amounts of hyaluronate, it was possible to determine the amount of hyaluronate in the unknowns. They have found that this method is sensitive to 1 or less of hyaluronate, and is unaffected by the presence of proteins.

  10. Bioluminescence assay for cell viability. (United States)

    Lomakina, G Yu; Modestova, Yu A; Ugarova, N N


    Theoretical aspects of the adenosine triphosphate bioluminescence assay based on the use of the firefly luciferin-luciferase system are considered, as well as its application for assessing cell viability in microbiology, sanitation, medicine, and ecology. Various approaches for the analysis of individual or mixed cultures of microorganisms are presented, and capabilities of the method for investigation of biological processes in live cells including necrosis, apoptosis, as well as for investigation of the dynamics of metabolism are described.

  11. Visual detection technique for efficient screening and isolation of Salmonella based on a novel enrichment assay using chromatography membrane. (United States)

    Tang, F; Xiong, Y; Zhang, H; Wu, K; Xiang, Y; Shao, J-B; Ai, H-W; Xiang, Y-P; Zheng, X-L; Lv, J-R; Sun, H; Bao, L-S; Zhang, Z; Hu, H-B; Zhang, J-Y; Chen, L; Lu, J; Liu, W-Y; Mei, H; Ma, Y; Xu, C-F; Fang, A-Y; Gu, M; Xu, C-Y; Chen, Y; Chen, Z; Sun, Z-Y


    To detect Salmonella more efficiently and isolate strains more easily, a novel and simple detection method that uses an enrichment assay and two chromogenic reactions on a chromatography membrane was developed. Grade 3 chromatography paper is used as functionalized solid phase support (SPS), which contains specially optimized medium. One reaction for screening is based on the sulfate-reducing capacity of Salmonella. Hydrogen sulfide (H2S) generated by Salmonella reacts with ammonium ferric citrate to produce black colored ferrous sulfide. Another reaction is based on Salmonella C8 esterase that is unique for Enterobacteriaceae except Serratia and interacts with 4-methylumbelliferyl caprylate (MUCAP) to produce fluorescent umbelliferone, which is visible under ultraviolet light. A very low detection limit (10(1) CFU ml(-1)) for Salmonella was achieved on the background of 10(5) CFU ml(-1) Escherichia coli. More importantly, testing with more than 1,000 anal samples indicated that our method has a high positive detection rate and is relatively low cost, compared with the traditional culture-based method. It took only 1 day for the preliminary screening and 2 days to efficiently isolate the Salmonella cells, indicating that the new assay is specific, rapid, and simple for Salmonella detection. In contrast to the traditional culture-based method, this method can be easily used to screen and isolate targeted strains with the naked eye. The results of quantitative and comparative experiments showed that the visual detection technique is an efficient alternative method for the screening of Salmonella spp. in many applications of large-sized samples related to public health surveillance.

  12. Hemoglobin variants: biochemical properties and clinical correlates. (United States)

    Thom, Christopher S; Dickson, Claire F; Gell, David A; Weiss, Mitchell J


    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.

  13. Simplifying biochemical models with intermediate species

    DEFF Research Database (Denmark)

    Feliu, Elisenda; Wiuf, Carsten


    canonical model that characterizes crucial dynamical properties, such as mono- and multistationarity and stability of steady states, of all models in the class. We show that if the core model does not have conservation laws, then the introduction of intermediates does not change the steady...... techniques, we study systematically the effects of intermediate, or transient, species in biochemical systems and provide a simple, yet rigorous mathematical classification of all models obtained from a core model by including intermediates. Main examples include enzymatic and post-translational modification...... systems, where intermediates often are considered insignificant and neglected in a model, or they are not included because we are unaware of their existence. All possible models obtained from the core model are classified into a finite number of classes. Each class is defined by a mathematically simple...

  14. On Biochemical Formation of Salt Deposits

    Institute of Scientific and Technical Information of China (English)


    A water/salt system in an evaporative environment is both a physicochemical region and a biological one. All the parameters of the system, such as the salinity, temperature and CO2 partial pressure, are affected by halophilic bacteria. The system controls salt deposition but is modified by an accompanying ecological system; therefore it should be called a water/salt/biological system. Salt minerals result from accumulation of the remains of bacteria/algae, namely, bacteria/algae formation; whereas biological, biophysical and biochemical processes provide full evidence for organic involvement. Consequently, salt deposits should not be called purely chemical but biological/chemical ones. This new argument supplements and develops the traditional idea and helps perfect the mineralization theory of salts and even general deposits, thus giving guidance to prospecting for salt deposits.

  15. The biochemical basis of hereditary fructose intolerance. (United States)

    Bouteldja, Nadia; Timson, David J


    Hereditary fructose intolerance is a rare, but potentially lethal, inherited disorder of fructose metabolism, caused by mutation of the aldolase B gene. Treatment currently relies solely on dietary restriction of problematic sugars. Biochemical study of defective aldolase B enzymes is key to revealing the molecular basis of the disease and providing a stronger basis for improved treatment and diagnosis. Such studies have revealed changes in enzyme activity, stability and oligomerisation. However, linking these changes to disease phenotypes has not always been straightforward. This review gives a general overview of the features of hereditary fructose intolerance, then concentrates on the biochemistry of the AP variant (Ala149Pro variant of aldolase B) and molecular pathological consequences of mutation of the aldolase B gene.

  16. Dynamic analysis of biochemical network using complex network method

    Directory of Open Access Journals (Sweden)

    Wang Shuqiang


    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. Transcription fluctuation effects on biochemical oscillations.

    Directory of Open Access Journals (Sweden)

    Ryota Nishino

    Full Text Available Some biochemical systems show oscillation. They often consist of feedback loops with repressive transcription regulation. Such biochemical systems have distinctive characteristics in comparison with ordinary chemical systems: i numbers of molecules involved are small, ii there are typically only a couple of genes in a cell with a finite regulation time. Due to the fluctuations caused by these features, the system behavior can be quite different from the one by deterministic rate equations, because the rate equations ignore molecular fluctuations and thus are exact only in the infinite molecular number limit. The molecular fluctuations on a free-running circadian system have been studied by Gonze et al. (2002 by introducing a scale parameter [Formula: see text] for the system size. They consider, however, only the first effect, assuming that the gene process is fast enough for the second effect to be ignored, but this has not been examined systematically yet. Here we study fluctuation effects due to the finite gene regulation time by introducing a new scale parameter [Formula: see text], which we take as the unbinding time of a nuclear protein from the gene. We focus on the case where the fluctuations due to small molecular numbers are negligible. In simulations on the same system studied by Gonze et al., we find the system is unexpectedly sensitive to the fluctuation in the transcription regulation; the period of oscillation fluctuates about 30 min even when the regulation time scale [Formula: see text] is around 30 s, that is even smaller than 1/1000 of its circadian period. We also demonstrate that the distribution width for the oscillation period and amplitude scales with [Formula: see text], and the correlation time scales with [Formula: see text] in the small [Formula: see text] regime. The relative fluctuations for the period are about half of that for the amplitude, namely, the periodicity is more stable than the amplitude.

  18. Biochemically enhanced methane production from coal (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

  19. Biochemical analysis of CTLA-4 immunoreactive material from human blood

    Directory of Open Access Journals (Sweden)

    Dennert Kate


    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.

  20. Inducible Sequential Oxidation Process in Water-Soluble Copper Nanoclusters for Direct Colorimetric Assay of Hydrogen Peroxide in a Wide Dynamic and Sampling Range. (United States)

    Du, Yibing; Fang, Jun; Wang, Hongli; Yang, Yang


    Direct and fast detection methods for H2O2 have great demand in materials science, biology, and medicine. Colorimetric assay of H2O2 has been regarded as one versatile approach that can avoid tedious operation and complicated setup. In this report, we provided a cost-effective and time-saving H2O2 colorimetric assay strategy based on a mercaptosuccinic acid (MSA)-stabilized Cu nanocluster (NC) probe without using any chromogenic reagent. Direct and fast colorimetric detection of H2O2 was realized based on the color change of MSA-capped Cu NCs in aqueous medium. It was found that the Cu NCs presented eligible resistance to natural oxidation either in concentrated solution or in the powder state. However, the dissolved oxygen in a highly diluted solution of the Cu NCs could trigger the aggregation of the Cu NCs and their further fusion into small Cu nanoparticles (NPs). When this diluted solution served as a probe solution for detecting H2O2, a sequential oxidation process occurred in the newly formed Cu NPs, including the cleavage of MSAs on the surface and conversion of Cu into Cu2O, leading to the probe with capacity for H2O2 assay in a wide dynamic and sampling range. The sensitive solution color change was attributed to the growth of the Cu NPs (fading of plasmonic absorption) upon the addition of low levels of H2O2 and the transition of the valence states of Cu (color reactions) upon the addition of high levels of H2O2. A concentration range of H2O2 from 1 μM to 1 M could be detected by a small dose of the probe. Moreover, the Cu NCs powder subsequent to storage for 10 months could maintain a similar sensitivity for H2O2 assay, which provides possibilities for a wide range of practical applications in water samples.

  1. Cross-resistance of bisultap resistant strain of Nilaparvata lugens and its biochemical mechanism. (United States)

    Ling, Shanfeng; Zhang, Runjie


    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.

  2. Development of on-line high performance liquid chromatography (HPLC)-biochemical detection methods as tools in the identification of bioactives. (United States)

    Malherbe, Christiaan J; de Beer, Dalene; Joubert, Elizabeth


    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.

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


    Carina Ladeira; Susana Viegas; Manuel C. Gomes


    The cytokinesis-block micronucleus cytome (CBMN) assay is a comprehensive system for measuring DNA damage; cytostasis and cytotoxicity-DNA damage events are scored specifically in once-divided binucleated cells. The endpoints possible to be measured are micronuclei (MN), a biomarker of chromosome breakage and/or whole chromosome loss, nucleoplasmic bridges (NPB), a biomarker of DNA misrepair and/or telomere end-fusions, and nuclear buds (NBUD), a biomarker of elimination of amplified DNA and/...

  4. Fluorescence-based assay as a new screening tool for toxic chemicals (United States)

    Moczko, Ewa; Mirkes, Evgeny M.; Cáceres, César; Gorban, Alexander N.; Piletsky, Sergey


    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.

  5. Biochemical Mechanism of Chlorantraniliprole Resistance in the Diamondback Moth, Plutella xylostella Linnaeus

    Institute of Scientific and Technical Information of China (English)

    HU Zhen-di; FENG Xia; LIN Qing-sheng; CHEN Huan-yu; LI Zhen-yu; YIN Fei; LIANG Pei; GAO Xi-wu


    The insecticide chlorantraniliprole exhibits good efifcacy and plays an important role in controlling the diamondback moth, Plutella xylostella Linnaeus. However, resistance to chlorantraniliprole has been observed recently in some ifeld populations. At present study, diamondback moths with resistance to chlorantraniliprole (resistant ratio (RR) was 82.18) for biochemical assays were selected. The assays were performed to determine potential resistance mechanisms. The results showed that the selected resistant moths (GDLZ-R) and susceptible moth could be synergized by known metabolic inhibitors such as piperonyl butoxide (PBO), triphenyl phosphate (TPP) and diethyl-maleate (DEM) at different levels (1.68-5.50-fold and 2.20-2.89-fold, respectively), and DEM showed the maximum synergism in both strains. In enzymes assays, a high level of glutathione-S-transferase (GST) was observed in the resistant moth, in contrast, moths that are susceptible to the insecticide had only 1/3 the GST activity of the resistant moths. The analysis of short-term exposure of chlorantraniliprole on biochemical response in the resistant strain also showed that GST activity was signiifcantly elevated after exposure to a sub-lethal concentration of chlorantraniliprole (about 1/3 LC50, 12 mg L-1) 12 and 24 h, respectively. The results show that there is a strong correlation between the enzyme activity and resistance, and GST is likely the main detoxiifcation mechanism responsible for resistance to chlorantraniliprole in P. xylostella L., cytochrome P450 monooxygenase (P450) and carboxy-lesterase (CarE) are involved in to some extent.

  6. In vitro activity assays for MYST histone acetyltransferases and adaptation for high-throughput inhibitor screening (United States)

    McCullough, Cheryl E.; Marmorstein, Ronen


    Lysine acetylation is a post-translational modification that is carried out by acetyltransferases. The MYST proteins form the largest and most diverse family of acetyltransferases, which regulate gene expression, DNA repair, and cell cycle homeostasis, among other activities, by acetylating both histone and non-histone proteins. This chapter will describe methods for the preparation and biochemical characterization of MYST family acetyltransferases, including protocols for the preparation of recombinant protein, enzyme assays for measuring steady state parameters and binding assays to measure cofactor and inhibitor binding. We also provide details on adapting these assays for high throughput screening for small molecule MYST inhibitors. This chapter seeks to prepare researchers for some hurdles that they may encounter when studying the MYST proteins so that there may be better opportunity to plan appropriate controls and obtain high quality data. PMID:27372752

  7. Promoter competition assay for analyzing gene regulation in joint tissue engineering. (United States)

    Sun, Hui Bin; Malacinski, George M; Yokota, Hiroki


    We describe a new biochemical technique, "promoter competition assay," for examining the role of cis-acting DNA elements in tissue cultures. Recent advances in tissue engineering permit the culture of a variety of cells. Many tissues are engineered, however, without an appropriate understanding of molecular machinery that regulates gene expression and cellular growth. For elucidating the role of cis-acting regulatory elements in cellular differentiation and growth, we developed the promoter competition assay. This assay uses a transient transfer into cells of double-stranded DNA fragments consisting of cis-acting regulatory elements. The transferred DNA fragments act as a competitor and titrate the function of their genomic counterparts. Using synovial cells derived from a rheumatoid arthritis patient, we examined a role of NF-kappa B binding sites in the regulation of the expression of matrix metalloproteinase (MMP) genes. The results support a stimulatory role of NF-kappa B in transcriptional regulation of MMP-1 and MMP-13.

  8. Differentiation and identification of Shigella spp. and enteroinvasive Escherichia coli in environmental waters by a molecular method and biochemical test. (United States)

    Hsu, Bing-Mu; Wu, Shu-Fen; Huang, Shih-Wei; Tseng, Yu-Jung; Ji, Dar-Der; Chen, Jung-Sheng; Shih, Feng-Cheng


    Both Shigella spp. and enteroinvasive Escherichia coli (EIEC) are important human pathogens that are responsible for the majority of cases of endemic bacillary dysentery. However, they are difficult to identify and differentiate by biochemical tests or molecular methods alone. In this study, we developed a procedure to detect Shigella spp. and EIEC from environmental water samples using membrane filtration followed by nutrient broth enrichment, isolation using selective culture plates, and identification of the invasion plasmid antigen H (ipaH) gene by PCR amplification and DNA sequencing. Finally, we used a biochemical test and a serological assay to differentiate between Shigella and EIEC. Among the 93 water samples from nine reservoirs and one watershed, 76 (81.7%) water samples of culture plates had candidate colonies of Shigella and EIEC and 5 water samples were positive (5.4%) for a Shigella- and EIEC-specific polymerase chain reaction targeting the ipaH gene. Guided by the molecular method, the biochemical test, and the serological assay, 11 ipaH gene-positive isolates from 5 water samples were all identified as EIEC.

  9. A Biochemical Approach to the Problem of Dyslexia. (United States)

    Baker, Sidney McDonald


    The paper presents the case of a sixth-grade boy, labeled dyslexic, who responded positively to a biochemical approach. Remedy of iron, zinc, and Vitamin B-6 deficiencies as well as an imbalance of fatty acids resulted in improvements in hair and skin and also in reading. A biochemical approach to behavior problems is proposed. (Author/CL)

  10. Local biochemical and morphological differences in human Achilles tendinopathy

    DEFF Research Database (Denmark)

    J, Pingel; Fredberg, Ulrich; K, Qvortrup;


    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 ex...

  11. The physiological and biochemical bases of functional brain imaging



    Functional brain imaging is based on the display of computer-derived images of changes in physiological and/or biochemical functions altered by activation or depression of local functional activities in the brain. This article reviews the physiological and biochemical mechanisms involved.

  12. Biochemical evaluation of phenylketonuria (PKU: from diagnosis to treatment

    Directory of Open Access Journals (Sweden)

    Leticia Belmont-Martínez


    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.

  13. 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.


    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

  14. Editorial: ESBES - European Society of Biochemical Engineering Sciences. (United States)

    Ferreira, Guilherme; Jungbauer, Alois


    The latest ESBES special issue on "Biochemical Engineering Sciences" is edited by Prof. Guilherme Ferreira (Chairman, ESBES) and Prof. Alois Jungbauer (co-Editor-in-Chief, Biotechnology Journal). This special issue comprises the latest research in biochemical engineering science presented at the 9(th) ESBES Conference held in Istanbul, Turkey in 2012.

  15. Development of a system for characterizing biomass quality of lignocellulosic feedstocks for biochemical conversion (United States)

    Murphy, Patrick Thomas

    sugars, including non-structural carbohydrates (CN) (monosaccharides, starches, oligosaccharides), biochemically available carbohydrates (CB) (structural carbohydrates susceptible to enzymatic hydrolysis) with an associated 1st-order availability rate constant (k B) and unavailable carbohydrates (CU) (hemicellulose and cellulose in close association with lignin). The model partitions the noncarbohydrate dry matter into extractives, lignin, and ash. Quality parameters were determined using a biomass quality assay that combined established wet-chemistry analyses techniques, including total non-structural carbohydrates (TNC), alcohol insoluble residue (AIR), simultaneous saccharification and fermentation (SSCF), and Klason lignin. The next study evaluated multiple high-throughput (HTP) modifications to the original assay methods, including (i) using filter bags with batch sample processing, (ii) replacement of AIR with neutral detergent fiber (NDF) as a cell-wall isolation procedure, and (iii) elimination of the fermentation organism in the SSCF procedures used to determine biochemically available carbohydrates. The original and the HTP assay methods were compared using corn cobs, hybrid poplar, kenaf, and switchgrass. Biochemically available carbohydrates increased with the HTP methods in the corn cobs, hybrid poplar, and switchgrass, but remained the same in the kenaf. Total available carbohydrates increased and unavailable carbohydrates decreased with the HTP methods in the corn cobs and switchgrass and remained the same in the hybrid poplar and kenaf. There were no differences in total carbohydrates (CT) between the two methods. The final study evaluated the variability of biomass quality parameters in a set of corn stover samples, and developed calibration equations for determining parameter values using near infrared reflectance spectroscopy (NIRS). Fifty-two corn stover samples harvested in Iowa and Wisconsin in 2005 and 2006 were analyzed using the HTP assay for

  16. Assessing HTS performance using BioAssay Ontology: screening and analysis of a bacterial phospho-N-acetylmuramoyl-pentapeptide translocase campaign. (United States)

    Moberg, Andreas; Zander Balderud, Linda; Hansson, Eva; Boyd, Helen


    With the public availability of biochemical assays and screening data constantly increasing, new applications for data mining and method analysis are evolving in parallel. One example is BioAssay Ontology (BAO) for systematic classification of assays based on screening setup and metadata annotations. In this article we report a high-throughput screening (HTS) against phospho-N-acetylmuramoyl-pentapeptide translocase (MraY), an attractive antibacterial drug target involved in peptidoglycan synthesis. The screen resulted in novel chemistry identification using a fluorescence resonance energy transfer assay. To address a subset of the false positive hits, a frequent hitter analysis was performed using an approach in which MraY hits were compared with hits from similar assays, previously used for HTS. The MraY assay was annotated according to BAO and three internal reference assays, using a similar assay design and detection technology, were identified. Analyzing the assays retrospectively, it was clear that both MraY and the three reference assays all showed a high false positive rate in the primary HTS assays. In the case of MraY, false positives were efficiently identified by applying a method to correct for compound interference at the hit-confirmation stage. Frequent hitter analysis based on the three reference assays with similar assay method identified additional false actives in the primary MraY assay as frequent hitters. This article demonstrates how assays annotated using BAO terms can be used to identify closely related reference assays, and that analysis based on these assays clearly can provide useful data to influence assay design, technology, and screening strategy.

  17. Simple and sensitive method for the quantification of total bilirubin in human serum using 3-methyl-2-benzothiazolinone hydrazone hydrochloride as a chromogenic probe (United States)

    Nagaraja, Padmarajaiah; Avinash, Krishnegowda; Shivakumar, Anantharaman; Dinesh, Rangappa; Shrestha, Ashwinee Kumar


    We here describe a new spectrophotometric method for measuring total bilirubin in serum. The method is based on the cleavage of bilirubin giving formaldehyde which further reacts with diazotized 3-methyl-2-benzothiazolinone hydrazone hydrochloride giving blue colored solution with maximum absorbance at 630 nm. Sensitivity of the developed method was compared with Jendrassik-Grof assay procedure and its applicability has been tested with human serum samples. Good correlation was attained between both methods giving slope of 0.994, intercept 0.015, and R2 = 0.997. Beers law obeyed in the range of 0.068-17.2 μM with good linearity, absorbance y = 0.044 Cbil + 0.003. Relative standard deviation was 0.006872, within day precision ranged 0.3-1.2% and day-to-day precision ranged 1-6%. Recovery of the method varied from 97 to 102%. The proposed method has higher sensitivity with less interference. The obtained product was extracted and was spectrally characterized for structural confirmation with FT-IR, 1H NMR.

  18. BALL - biochemical algorithms library 1.3

    Directory of Open Access Journals (Sweden)

    Stöckel Daniel


    Full Text Available Abstract Background The Biochemical Algorithms Library (BALL is a comprehensive rapid application development framework for structural bioinformatics. It provides an extensive C++ class library of data structures and algorithms for molecular modeling and structural bioinformatics. Using BALL as a programming toolbox does not only allow to greatly reduce application development times but also helps in ensuring stability and correctness by avoiding the error-prone reimplementation of complex algorithms and replacing them with calls into the library that has been well-tested by a large number of developers. In the ten years since its original publication, BALL has seen a substantial increase in functionality and numerous other improvements. Results Here, we discuss BALL's current functionality and highlight the key additions and improvements: support for additional file formats, molecular edit-functionality, new molecular mechanics force fields, novel energy minimization techniques, docking algorithms, and support for cheminformatics. Conclusions BALL is available for all major operating systems, including Linux, Windows, and MacOS X. It is available free of charge under the Lesser GNU Public License (LPGL. Parts of the code are distributed under the GNU Public License (GPL. BALL is available as source code and binary packages from the project web site at Recently, it has been accepted into the debian project; integration into further distributions is currently pursued.

  19. Robustness analysis of stochastic biochemical systems. (United States)

    Ceska, Milan; Safránek, David; Dražan, Sven; Brim, Luboš


    We propose a new framework for rigorous robustness analysis of stochastic biochemical systems that is based on probabilistic model checking techniques. We adapt the general definition of robustness introduced by Kitano to the class of stochastic systems modelled as continuous time Markov Chains in order to extensively analyse and compare robustness of biological models with uncertain parameters. The framework utilises novel computational methods that enable to effectively evaluate the robustness of models with respect to quantitative temporal properties and parameters such as reaction rate constants and initial conditions. We have applied the framework to gene regulation as an example of a central biological mechanism where intrinsic and extrinsic stochasticity plays crucial role due to low numbers of DNA and RNA molecules. Using our methods we have obtained a comprehensive and precise analysis of stochastic dynamics under parameter uncertainty. Furthermore, we apply our framework to compare several variants of two-component signalling networks from the perspective of robustness with respect to intrinsic noise caused by low populations of signalling components. We have successfully extended previous studies performed on deterministic models (ODE) and showed that stochasticity may significantly affect obtained predictions. Our case studies demonstrate that the framework can provide deeper insight into the role of key parameters in maintaining the system functionality and thus it significantly contributes to formal methods in computational systems biology.

  20. 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: [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)


    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)

  1. PHA bioplastics, biochemicals, and energy from crops. (United States)

    Somleva, Maria N; Peoples, Oliver P; Snell, Kristi D


    Large scale production of polyhydroxyalkanoates (PHAs) in plants can provide a sustainable supply of bioplastics, biochemicals, and energy from sunlight and atmospheric CO(2). PHAs are a class of polymers with various chain lengths that are naturally produced by some microorganisms as storage materials. The properties of these polyesters make them functionally equivalent to many of the petroleum-based plastics that are currently in the market place. However, unlike most petroleum-derived plastics, PHAs can be produced from renewable feedstocks and easily degrade in most biologically active environments. This review highlights research efforts over the last 20 years to engineer the production of PHAs in plants with a focus on polyhydroxybutryrate (PHB) production in bioenergy crops with C(4) photosynthesis. PHB has the potential to be a high volume commercial product with uses not only in the plastics and materials markets, but also in renewable chemicals and feed. The major challenges of improving product yield and plant fitness in high biomass yielding C(4) crops are discussed in detail.

  2. A colorimetric assay for cytokinin oxidase. (United States)

    Libreros-Minotta, C A; Tipton, P A


    A simple and rapid colorimetric assay for cytokinin oxidase is described. The assay is based on the formation of a Schiff base between the enzymatic reaction product 3-methyl-2-butenal and p-aminophenol. The assay is effective in the submicromolar concentration range and can be used in crude plant extracts as well as in more highly purified preparations.

  3. Capillary Electrophoresis Analysis of Conventional Splicing Assays

    DEFF Research Database (Denmark)

    de Garibay, Gorka Ruiz; Acedo, Alberto; García-Casado, Zaida;


    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...

  4. 21 CFR 864.7525 - Heparin assay. (United States)


    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Heparin assay. 864.7525 Section 864.7525 Food and... HEMATOLOGY AND PATHOLOGY DEVICES Hematology Kits and Packages § 864.7525 Heparin assay. (a) Identification. A heparin assay is a device used to determine the level of the anticoagulant heparin in the...

  5. Multicentre comparison of a diagnostic assay

    DEFF Research Database (Denmark)

    Waters, Patrick; Reindl, Markus; Saiz, Albert;


    ) 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...

  6. 21 CFR 866.3210 - Endotoxin assay. (United States)


    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Endotoxin assay. 866.3210 Section 866.3210 Food... DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3210 Endotoxin assay. (a) Identification. An endotoxin assay is a device that uses serological techniques in whole blood. The device...

  7. Adapting Cell-Based Assays to the High Throughput Screening Platform: Problems Encountered and Lessons Learned. (United States)

    Maddox, Clinton B; Rasmussen, Lynn; White, E Lucile


    In recent years, cell-based phenotypic assays have emerged as an effective and robust addition to the array of assay technologies available for drug discovery in the high throughput screening arena. Previously, biochemical target-based assays have been the technology of choice. With the emergence of stem cells as a basis for a new screening technology, it is important to keep in mind the lessons that have been learned from the adaptation of existing stable cell lines onto the high throughput screening drug discovery platform, with special consideration being given to assay miniaturization, liquid handling complications and instrument-introduced artifacts. We present an overview of the problems encountered with the implementation of multiple cell-based assays at the High Throughput Screening Center at Southern Research Institute as well as empirically defined effective solutions to these problems. These include examples of artifacts induced by temperature differences throughout the screening campaign, cell plating conditions including the effect of room temperature incubation on assay consistency, DMSO carry-over, and incubator induced artifacts.

  8. Time-stretch microscopy on a DVD for high-throughput imaging cell-based assay. (United States)

    Tang, Anson H L; Yeung, P; Chan, Godfrey C F; Chan, Barbara P; Wong, Kenneth K Y; Tsia, Kevin K


    Cell-based assay based on time-stretch imaging is recognized to be well-suited for high-throughput phenotypic screening. However, this ultrafast imaging technique has primarily been limited to suspension-cell assay, leaving a wide range of solid-substrate assay formats uncharted. Moreover, time-stretch imaging is generally restricted to intrinsic biophysical phenotyping, but lacks the biomolecular signatures of the cells. To address these challenges, we develop a spinning time-stretch imaging assay platform based on the functionalized digital versatile disc (DVD). We demonstrate that adherent cell culture and biochemically-specific cell-capture can now be assayed with time-stretch microscopy, thanks to the high-speed DVD spinning motion that naturally enables on-the-fly cellular imaging at an ultrafast line-scan rate of >10MHz. As scanning the whole DVD at such a high speed enables ultra-large field-of-view imaging, it could be favorable for scaling both the assay throughput and content as demanded in many applications, e.g. drug discovery, and rare cancer cell screening.

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

    Directory of Open Access Journals (Sweden)

    Hampus Sunner


    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.

  10. Steroid assays in paediatric endocrinology. (United States)

    Honour, John W


    Most steroid disorders of the adrenal cortex come to clinical attention in childhood and in order to investigate these problems, there are many challenges to the laboratory which need to be appreciated to a certain extent by clinicians. The analysis of sex steroids in biological fluids from neonates, over adrenarche and puberty present challenges of specificities and concentrations often in small sample sizes. Different reference ranges are also needed for interpretations. For around 40 years, quantitative assays for the steroids and their regulatory peptide hormones have been possible using immunoassay techniques. Problems are recognised and this review aims to summarise the benefits and failings of immunoassays and introduce where tandem mass spectrometry is anticipated to meet the clinical needs for steroid analysis in paediatric endocrine investigations. It is important to keep a dialogue between clinicians and the laboratory, especially when any laboratory result does not make sense in the clinical investigation.

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

    Directory of Open Access Journals (Sweden)



    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.

  12. Evaluation of CHROMagar STEC and STEC O104 chromogenic agar media for detection of Shiga Toxin-producing Escherichia coli in stool specimens. (United States)

    Gouali, Malika; Ruckly, Corinne; Carle, Isabelle; Lejay-Collin, Monique; Weill, François-Xavier


    The performance of CHROMagar STEC and CHROMagar STEC O104 (CHROMagar Microbiology, Paris, France) media for the detection of Shiga toxin-producing Escherichia coli (STEC) was assessed with 329 stool specimens collected over 14 months from patients with suspected STEC infections (June 2011 to August 2012). The CHROMagar STEC medium, after an enrichment broth step, allowed the recovery of the STEC strain from 32 of the 39 (82.1%) Shiga toxin-positive stool specimens, whereas the standard procedure involving Drigalski agar allowed the recovery of only three additional STEC strains. The isolates that grew on CHROMagar STEC medium belonged to 15 serotypes, including the prevalent non-sorbitol-fermenting (NSF) O157:H7, O26:H11, and O104:H4 serotypes. The sensitivity, specificity, and positive and negative predictive values for the CHROMagar STEC medium were between 89.1% and 91.4%, 83.7% and 86.7%, 40% and 51.3%, and 98% and 98.8%, respectively, depending on whether or not stx-negative eae-positive E. coli was considered atypical enteropathogenic E. coli (EPEC) or STEC that had lost Shiga toxin genes during infection. In conclusion, the good performance of CHROMagar STEC agar medium, in particular, the high negative predictive value, and its capacity to identify NSF O157:H7 as well as common non-O157 STEC may be useful for clinical bacteriology, public health, and reference laboratories; it could be used in addition to a method targeting Shiga toxins (detection of stx genes by PCR, immunodetection of Shiga toxins in stool specimens, or Vero cell cytotoxicity assay) as an alternative to O157 culture medium. This combined approach should allow rapid visualization of both putative O157 and non-O157 STEC colonies for subsequent characterization, essential for real-time surveillance of STEC infections and investigations of outbreaks.

  13. Predictive Assay For Cancer Targets

    Energy Technology Data Exchange (ETDEWEB)

    Suess, A; Nguyen, C; Sorensen, K; Montgomery, J; Souza, B; Kulp, K; Dugan, L; Christian, A


    Early detection of cancer is a key element in successful treatment of the disease. Understanding the particular type of cancer involved, its origins and probable course, is also important. PhIP (2-amino-1-methyl-6 phenylimidazo [4,5-b]pyridine), a heterocyclic amine produced during the cooking of meat at elevated temperatures, has been shown to induce mammary cancer in female, Sprague-Dawley rats. Tumors induced by PhIP have been shown to contain discreet cytogenetic signature patterns of gains and losses using comparative genomic hybridization (CGH). To determine if a protein signature exists for these tumors, we are analyzing expression levels of the protein products of the above-mentioned tumors in combination with a new bulk protein subtractive assay. This assay produces a panel of antibodies against proteins that are either on or off in the tumor. Hybridization of the antibody panel onto a 2-D gel of tumor or control protein will allow for identification of a distinct protein signature in the tumor. Analysis of several gene databases has identified a number of rat homologs of human cancer genes located in these regions of gain and loss. These genes include the oncogenes c-MYK, ERBB2/NEU, THRA and tumor suppressor genes EGR1 and HDAC3. The listed genes have been shown to be estrogen-responsive, suggesting a possible link between delivery of bio-activated PhIP to the cell nucleus via estrogen receptors and gene-specific PhIP-induced DNA damage, leading to cell transformation. All three tumors showed similar silver staining patterns compared to each other, while they all were different than the control tissue. Subsequent screening of these genes against those from tumors know to be caused by other agents may produce a protein signature unique to PhIP, which can be used as a diagnostic to augment optical and radiation-based detection schemes.

  14. Assays for the biochemical and ultrastructural measurement of selective and nonselective types of autophagy in the yeast Saccharomyces cerevisiae

    NARCIS (Netherlands)

    Guimaraes, Rodrigo Soares; Delorme-Axford, Elizabeth; Klionsky, Daniel J.; Reggiori, Fulvio


    Autophagy is a conserved intracellular catabolic pathway that degrades unnecessary or dysfunctional cellular components. Components destined for degradation are sequestered into double-membrane vesicles called autophagosomes, which subsequently fuse with the vacuole/lysosome delivering their cargo i

  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


    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. Comparison of the cancer gene targeting and biochemical selectivities of all targeted kinase inhibitors approved for clinical use.

    Directory of Open Access Journals (Sweden)

    Joost C M Uitdehaag

    Full Text Available The anti-proliferative activities of all twenty-five targeted kinase inhibitor drugs that are in clinical use were measured in two large assay panels: (1 a panel of proliferation assays of forty-four human cancer cell lines from diverse tumour tissue origins; and (2 a panel of more than 300 kinase enzyme activity assays. This study provides a head-on comparison of all kinase inhibitor drugs in use (status Nov. 2013, and for six of these drugs, the first kinome profiling data in the public domain. Correlation of drug activities with cancer gene mutations revealed novel drug sensitivity markers, suggesting that cancers dependent on mutant CTNNB1 will respond to trametinib and other MEK inhibitors, and cancers dependent on SMAD4 to small molecule EGFR inhibitor drugs. Comparison of cellular targeting efficacies reveals the most targeted inhibitors for EGFR, ABL1 and BRAF(V600E-driven cell growth, and demonstrates that the best targeted agents combine high biochemical potency with good selectivity. For ABL1 inhibitors, we computationally deduce optimized kinase profiles for use in a next generation of drugs. Our study shows the power of combining biochemical and cellular profiling data in the evaluation of kinase inhibitor drug action.

  17. Activity assay of membrane transport proteins

    Institute of Scientific and Technical Information of China (English)

    Hao Xie


    Membrane transport proteins are integral membrane proteins and considered as potential drug targets. Activity assay of transport proteins is essential for developing drugs to target these proteins. Major issues related to activity assessment of transport proteins include availability of transporters,transport activity of transporters, and interactions between ligands and transporters. Researchers need to consider the physiological status of proteins (bound in lipid membranes or purified), availability and specificity of substrates, and the purpose of the activity assay (screening, identifying, or comparing substrates and inhibitors) before choosing appropriate assay strategies and techniques. Transport proteins bound in vesicular membranes can be assayed for transporting substrate across membranes by means of uptake assay or entrance counterflow assay. Alternatively, transport proteins can be assayed for interactions with ligands by using techniques such as isothermal titration calorimetry, nuclear magnetic resonance spectroscopy, or surface plasmon resonance. Other methods and techniques such as fluorometry, scintillation proximity assay, electrophysiological assay, or stopped-flow assay could also be used for activity assay of transport proteins. In this paper the major strategies and techniques for activity assessment of membrane transport proteins are reviewed.

  18. Biochemical evidence supporting the Cortina criteria. (United States)

    von Werder, K


    Whether acromegaly is inactive or active, respectively cured or not cured depends on the GH suppressibility and the basal IGF-I level. According to the Cortina criteria, GH suppression after oral ingestion of 75 g glucose to acromegaly. According to more recent publications, mortality, which is increased in active acromegaly, is normalized when GH and IGF-I levels have become normal as defined above. Morbidity, i.e. typical features of acromegaly like cardiac problems, carpal tunnel syndrome, carbohydrate intolerance, and excessive sweating may also improve though painful arthropathy, and coarse facial features usually remain unaltered even if the biochemistry has been completely normalized. Using more sensitive GH assays, a group of acromegalic patients was shown to have normal IGF-I levels after surgery, with post-glucose levels of GH 0.14 microg/l, which is the upper level of normal subjects and of a second group of successfully operated acromegalic patients. The latter group also had slightly lower IGF-I levels, though such levels were normal in both groups. Whether this may indicate that these patients who have higher GH levels after oral glucose measured with the more sensitive immunoradiometric assay (IRMA) will more likely develop recurrences remains to be demonstrated in a larger cohort. According to the criteria put forward in Cortina d'Ampezzo in February 1999, all patients who have post-glucose GH levels <1 microg/l and normal age-matched IGF-I levels have to be regarded as well controlled, i.e. sufficiently treated. Because of lack of evidence, there is at present no reason to change the consensus reached there.

  19. The Biochemical Prognostic Factors of Subclinical Hypothyroidism

    Directory of Open Access Journals (Sweden)

    Myung Won Lee


    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.

  20. Biochemical effects of Calotropis procera on hepatotoxicity

    Directory of Open Access Journals (Sweden)

    Ali Ismaiel Ali Abd Alrheam


    Full Text Available Introduction: Calotropis procera commonly known as Sodom apple is a 6-meter high shrub that belongs to the Aclepiadaceae plant family and is commonly found in West Africa and other tropical places. In Saudi Arabia the plant is commonly used in traditional medicine for the treatment of variety of diseases including fever, constipation, muscular spasm and joint pain. Aim: In the present study C. procera were investigated for the hepatoprotective activity. Material and Methods: Carbon tetrachloride is used to produce hepatotoxicity. Forty two male albino rats, weighting 150-200 gm divided into seven groups, each consisted of 6 rats. Carbon tetrachloride 2ml/kg was administered twice a week to all of the groups of animals except group I, which served as control and given the normal saline. Group II served as Carbon tetrachloirde control. Group III received Silymarin at 100 mg/kg/day dose, Group IV received aqueous leaves extracts C. procera 200mg/kg, Group V received chloroform leaves extracts C. procera 200mg/kg, Group VI received ethanol leaves extracts C. procera 200 mg/kg, Group VII received latex of C. procera 200mg/kg. The effect of aqueous, chloroform, ethanol leaves extract and latex C. procera on biochemical parameters of liver was measured. Results: The results showed that the aqueous, chloroform, ethanol leaves extract and latex C. procera produced significant decrease in Acid phosphatase, Alkaline phosphatase, Aspartate aminotransferase, Alanine aminotransferase, Total protein, Albumin and total bilirubin levels compared to the CCL4 treated group II. Conclusion: Calotropis procera appears to to have hepatoprotective activity and these may be due to enrich of the plant by phytoconstituents that activate and in hence a pharmacological response of different parts of the body and this study need further studies to shows the complete properties of the plant. [Biomed Res Ther 2015; 2(12.000: 446-453

  1. Chromogenic in situ hybridization to detect HER-2/neu gene amplification in histological and ThinPrep-processed breast cancer fine-needle aspirates: a sensitive and practical method in the trastuzumab era. (United States)

    Vocaturo, Amina; Novelli, Flavia; Benevolo, Maria; Piperno, Giulia; Marandino, Ferdinando; Cianciulli, Anna Maria; Merola, Roberta; Donnorso, Raffaele Perrone; Sperduti, Isabella; Buglioni, Simonetta; Mottolese, Marcella


    The increasing evidence of trastuzumab efficacy in breast cancer (BC) patients means that an accurate and reproducible evaluation of HER-2 statusis of paramount importance in histological and in cytological samples. Currently, the two main methods used to analyze HER-2 amplification or overexpression are fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC). Although the two methods are strongly correlated for histological tissue, the evaluation of tumor morphology through FISH may be difficult and fluorescence fades quickly. These limitations can be overcome by chromogenic in situ hybridization (CISH), which can visualize the amplification product along with morphological features. In view of this, in the present study, we analyzed the usefulness of CISH on formalin-fixed, paraffin-embedded (FFPE) BC specimens and investigated whether CISH can be a valid technique in the determination of HER-2 status for fine-needle aspirates (FNAs) processed by liquid-based cytology. The results we obtained in a retrospective series of 111 FFPE BC specimens demonstrated good concordance between CISH and IHC and between CISH and FISH. The former concordance was comparable with that observed between FISH and IHC. When CISH was applied to a prospective series of 53 FNAs, from surgically removed BC, our data showed evidence of a higher concordance of results between liquid-based cytology and the companion FFPE tissues using CISH rather than HercepTesttrade mark. Therefore, CISH analysis, which is avaluable and reproducible alternative to FISH for selecting breast cancer patients for trastuzumab therapy, can lower false-positive immunocytochemistry findings in ThinPrep-processed FNAs.

  2. 40 CFR 158.2080 - Experimental use permit data requirements-biochemical pesticides. (United States)


    ... requirements-biochemical pesticides. 158.2080 Section 158.2080 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2080 Experimental use permit data requirements—biochemical pesticides. (a) Sections...

  3. Effects of organic solvents on the enzyme activity of Trypanosoma cruzi glyceraldehyde-3-phosphate dehydrogenase in calorimetric assays

    DEFF Research Database (Denmark)

    Wiggers, Henrik; Cheleski, J; Zottis, A


    is sixfold higher. The favorable effects of the organic solvents on the Michaelis-Menten enzyme-substrate complex formation ensure the consistency of the biological assays, structural integrity of the protein, and reproducibility over the measurement time. The reaction was also kinetically monitored......In drug discovery programs, dimethyl sulfoxide (DMSO) is a standard solvent widely used in biochemical assays. Despite the extensive use and study of enzymes in the presence of organic solvents, for some enzymes the effect of organic solvent is unknown. Macromolecular targets may be affected...... by the presence of different solvents in such a way that conformational changes perturb their active site structure accompanied by dramatic variations in activity when performing biochemical screenings. To address this issue, in this work we studied the effects of two organic solvents, DMSO and methanol (Me...

  4. A one-day, dispense-only IP-One HTRF assay for high-throughput screening of Galphaq protein-coupled receptors: towards cells as reagents. (United States)

    Bergsdorf, Christian; Kropp-Goerkis, Carmen; Kaehler, Irene; Ketscher, Lars; Boemer, Ulf; Parczyk, Karsten; Bader, Benjamin


    Abstract: Compared to biochemical high-throughput screening (HTS) assays, cell-based functional assays are generally thought to be more time consuming and complex because of additional efforts for running continuous cell cultures as well as the numerous assay steps when transferring media and compounds. A common strategy to compensate the anticipated reduction in overall throughput is to implement highly automated cell culture and screening systems. However, such systems require substantial investments in sophisticated hardware and highly specialized personnel. In trying to set up alternatives to increasing throughput in functional cell-based screening, we combined several approaches. By using (1) cryopreserved cell aliquots instead of continuous cell culture, (2) cells in suspension instead of adherent cells, and (3) "ready-to-screen" assay plates with nanoliter aliquots of test compounds, an assay procedure was developed that very much resembles a standard biochemical, enzymatic assay comprising only a few dispense steps. Chinese hamster ovary cells stably overexpressing a Galphaq-coupled receptor were used as a model system to measure receptor activation by detection of intracellular D-myo-inositol 1-phosphate with the help of homogeneous time-resolved fluorescence (HTRF, CISbio International, Bagnols-sur-Cèze, France). Initially established in 384-well adherent cell format, the assay was successfully transferred to 1,536-well format. The assay quality was sufficient to run HTS campaigns in both formats with good Z'-factors and excellent reproducibility of antagonists. Subsequently, the assay procedure was optimized for usage of suspension cells. The influences of cell culture media, plate type, cell number, and incubation time were assessed. Finally, the suspension cell assay was applied to pharmacological characterization of a small molecule antagonist by Schild plot analysis. Our data demonstrate not only the application of the IP-One HTRF assay (CISbio

  5. A Complete Optical Sensor System Based on a POF-SPR Platform and a Thermo-Stabilized Flow Cell for Biochemical Applications. (United States)

    Cennamo, Nunzio; Chiavaioli, Francesco; Trono, Cosimo; Tombelli, Sara; Giannetti, Ambra; Baldini, Francesco; Zeni, Luigi


    An optical sensor platform based on surface plasmon resonance (SPR) in a plastic optical fiber (POF) integrated into a thermo-stabilized flow cell for biochemical sensing applications is proposed. This device has been realized and experimentally tested by using a classic receptor-analyte assay. For this purpose, the gold surface of the POF was chemically modified through the formation of a self-assembling monolayer. The surface robustness of the POF-SPR platform has been tested for the first time thanks to the flow cell. The experimental results show that the proposed device can be successfully used for label-free biochemical sensing. The final goal of this work is to achieve a complete, small-size, simple to use and low cost optical sensor system. The whole system with the flow cell and the optical sensor are extensively described, together with the experimental results obtained with an immunoglobulin G (IgG)/anti-IgG assay.

  6. A Complete Optical Sensor System Based on a POF-SPR Platform and a Thermo-Stabilized Flow Cell for Biochemical Applications

    Directory of Open Access Journals (Sweden)

    Nunzio Cennamo


    Full Text Available An optical sensor platform based on surface plasmon resonance (SPR in a plastic optical fiber (POF integrated into a thermo-stabilized flow cell for biochemical sensing applications is proposed. This device has been realized and experimentally tested by using a classic receptor-analyte assay. For this purpose, the gold surface of the POF was chemically modified through the formation of a self-assembling monolayer. The surface robustness of the POF-SPR platform has been tested for the first time thanks to the flow cell. The experimental results show that the proposed device can be successfully used for label-free biochemical sensing. The final goal of this work is to achieve a complete, small-size, simple to use and low cost optical sensor system. The whole system with the flow cell and the optical sensor are extensively described, together with the experimental results obtained with an immunoglobulin G (IgG/anti-IgG assay.

  7. Structural, Biochemical, and Biophysical Characterization of Idelalisib Binding to Phosphoinositide 3-Kinase δ* (United States)

    Somoza, John R.; Koditek, David; Villaseñor, Armando G.; Novikov, Nikolai; Wong, Melanie H.; Liclican, Albert; Xing, Weimei; Lagpacan, Leanna; Wang, Ruth; Schultz, Brian E.; Papalia, Giuseppe A.; Samuel, Dharmaraj; Lad, Latesh; McGrath, Mary E.


    Idelalisib (also known as GS-1101, CAL-101, IC489666, and Zydelig) is a PI3Kδ inhibitor that has recently been approved for the treatment of several hematological malignancies. Given its use in human diseases, we needed a clear picture of how idelalisib binds to and inhibits PI3Kδ. Our data show that idelalisib is a potent and selective inhibitor of the kinase activity of PI3Kδ. A kinetic characterization clearly demonstrated ATP-competitive inhibition, and several additional biochemical and biophysical assays showed that the compound binds reversibly and noncovalently to the kinase. A crystal structure of idelalisib bound to the p110δ subunit of PI3Kδ furthers our understanding of the binding interactions that confer the potency and selectivity of idelalisib. PMID:25631052

  8. The biochemical mechanisms of the plant activation of promutagenic aromatic amines

    Energy Technology Data Exchange (ETDEWEB)

    Wagner, E.D.; Verdier, M.M.; Plewa, M.J. (Univ. of Illinois, Urbana (USA))


    Using specific monooxygenase and oxidase inhibitors in a plant cell/microbe coincubation assay, the biochemical mechanisms of the plant activation of two aromatic amines were compared. The biological endpoints included mutation induction, inhibition of mutagenicity, viability of the plant cells (activating system), and viability of the microbial cells. The activation of m-phenylenediamine by TX1 cells was mediated by enzyme systems that were inhibited by diethyldithiocarbamate, potassium cyanide, methimazole, (+)-catechin or acetaminophen. The inhibition by metyrapone was attended by toxicity in the plant cells. These data implicate a TX1 cell peroxidase and a FAD-dependent monoxygenase in the plant activation of m-phenylenediamine. The TX1 cell activation of 2-aminofluorene was inhibited by diethyldithiocarbamate, 7,8-benzoflavone, acetaminophen or (+)-catechin.


    Directory of Open Access Journals (Sweden)

    Luis E. Espinoza


    Full Text Available Streptomyces are common soil bacteria that produce secondary metabolites, including several antibiotics; however, the characteristics of marine Streptomyces are largely unknown. Sediment samples were taken from 3 sites in the Laguna Madre to isolate marine Streptomyces. Sediment was diluted, spread onto synthetic seawater media to estimate the total bacterial density of the samples and spread onto starch casein agar to isolate Streptomyces. Isolated Streptomyces were tested for salinity tolerance and optimal growth pH. Isolates were assayed using API 20E® test strips and BIOLOG™ plates to construct biochemical profiles and assess nutrient utilization abilities of the bacteria, respectively. Individual Streptomyces were tested for the ability to inhibit the growth of other isolated Streptomyces (i.e., interference competition and putatively identified by DNA sequencing. Results showed that there was no significant difference in microbial density in sediments from the 3 sampling sites. Eleven (11 Streptomyces pure cultures were obtained in total; most tolerated salinity up to 60 ppt and grew optimally at pH 7.5. Biochemical profile comparisons showed that the Streptomyces were only at least 74% similar; most (8/11 were >90% similar. Isolates could use between 87-95 carbon sources. Three (3 isolates displayed interference toward other isolates. Ten (10 isolates were identified as Streptomyces griseus by DNA sequencing. Laguna Madre Streptomyces organisms display some diverse characteristics with regards to their halotolerance, biochemical profiles, carbon source utilization and inhibition toward other organisms. Further investigations may yield greater understanding of these organisms in this and other marine environments and may be a reservoir of novel microorganisms and secondary metabolites.

  10. [Islet autoantibody assays in type I diabetes: superiority of passage from use of ICA to traditional tests]. (United States)

    Lazar, D; Weintrob, N; Abramov, N; Assa, S; Bloch, K; Ofan, R; Ben-Zaken, H; Vardi, P


    Islet cell antibodies (ICA) continue to serve as the basis of the principal serological test for definition of active autoimmunity of beta-cells. Its disadvantages are the need for human pancreatic tissue and difficulty in obtaining quantitative results. In the past decade biochemically-defined beta-cell antigens were described, leading to the development of sensitive and specific autoantibody assays, to predict insulin-dependent diabetes mellitus (IDDM). We examined the value of combined biochemically-based serological assays, such as autoantibodies to insulin (IAA), glutamic acid decarboxylase (GADA) and ICA512 (ICA512A) to replace the traditional ICA assay. Blood samples of 114 newly diagnosed IDDM patients, aged 12 +/- 5 yrs (range 2 months-29 years) were tested for ICA (indirect immunofluorescence), IAA, GADA and ICA512A (radiobinding assay). The latter 2 assays were performed using recombinant human [35S]-labeled antigen produced by in vitro transcription/translation. We found that fewer sera scored positive for ICA and/or IAA (80.7%, 92/114) than for 1 or more of IAA, GAD, or ICA512 (88.6%, 101/114). We conclude that combined testing for IAA, GAD and ICA512 can replace the traditional ICA/IAA test to predict IDDM and is helpful in the differential diagnosis of insulin-dependent and noninsulin-dependent diabetes.

  11. Novel patient cell-based HTS assay for identification of small molecules for a lysosomal storage disease.

    Directory of Open Access Journals (Sweden)

    Haifeng Geng

    Full Text Available Small molecules have been identified as potential therapeutic agents for lysosomal storage diseases (LSDs, inherited metabolic disorders caused by defects in proteins that result in lysosome dysfunctional. Some small molecules function assisting the folding of mutant misfolded lysosomal enzymes that are otherwise degraded in ER-associated degradation. The ultimate result is the enhancement of the residual enzymatic activity of the deficient enzyme. Most of the high throughput screening (HTS assays developed to identify these molecules are single-target biochemical assays. Here we describe a cell-based assay using patient cell lines to identify small molecules that enhance the residual arylsulfatase A (ASA activity found in patients with metachromatic leukodystrophy (MLD, a progressive neurodegenerative LSD. In order to generate sufficient cell lines for a large scale HTS, primary cultured fibroblasts from MLD patients were transformed using SV40 large T antigen. These SV40 transformed (SV40t cells showed to conserve biochemical characteristics of the primary cells. Using a specific colorimetric substrate para-nitrocatechol sulfate (pNCS, detectable ASA residual activity were observed in primary and SV40t fibroblasts from a MLD patient (ASA-I179S cultured in multi-well plates. A robust fluorescence ASA assay was developed in high-density 1,536-well plates using the traditional colorimetric pNCS substrate, whose product (pNC acts as "plate fluorescence quencher" in white solid-bottom plates. The quantitative cell-based HTS assay for ASA generated strong statistical parameters when tested against a diverse small molecule collection. This cell-based assay approach can be used for several other LSDs and genetic disorders, especially those that rely on colorimetric substrates which traditionally present low sensitivity for assay-miniaturization. In addition, the quantitative cell-based HTS assay here developed using patient cells creates an

  12. 2011 Biomass Program Platform Peer Review: Biochemical Conversion

    Energy Technology Data Exchange (ETDEWEB)

    Pezzullo, Leslie [Office of Energy Efficiency and Renewable Energy (EERE), Washington, DC (United States)


    This document summarizes the recommendations and evaluations provided by an independent external panel of experts at the 2011 U.S. Department of Energy Biomass Program’s Biochemical Conversion Platform Review meeting.

  13. Biochemical methane potential (BMP) of solid organic materials

    DEFF Research Database (Denmark)

    Raposo, Francisco; Fernández-Cegrí, V.; De la Rubia, M.A.


    This paper describes the results obtained for different participating research groups in an interlaboratory study related to the biochemical methane potential (BMP). In this research work, the full experimental conditions influencing the test such as inoculum, substrate characteristics and experi...

  14. Approaches to Chemical and Biochemical Information and Signal Processing (United States)

    Privman, Vladimir


    We outline models and approaches for error control required to prevent buildup of noise when ``gates'' and other ``network elements'' based on (bio)chemical reaction processes are utilized to realize stable, scalable networks for information and signal processing. We also survey challenges and possible future research. [4pt] [1] Control of Noise in Chemical and Biochemical Information Processing, V. Privman, Israel J. Chem. 51, 118-131 (2010).[0pt] [2] Biochemical Filter with Sigmoidal Response: Increasing the Complexity of Biomolecular Logic, V. Privman, J. Halamek, M. A. Arugula, D. Melnikov, V. Bocharova and E. Katz, J. Phys. Chem. B 114, 14103-14109 (2010).[0pt] [3] Towards Biosensing Strategies Based on Biochemical Logic Systems, E. Katz, V. Privman and J. Wang, in: Proc. Conf. ICQNM 2010 (IEEE Comp. Soc. Conf. Publ. Serv., Los Alamitos, California, 2010), pages 1-9.

  15. Assessment of biochemical concentrations of vegetation using remote sensing technology

    Institute of Scientific and Technical Information of China (English)


    The main biochemicals (such as lignin, protein, cellulose, sugar, starch, chlorophyll and water) of vegetation are directly or indirectly involved in major ecological processes, such as the functions of terrestrial ecosystems (i.e., nutrient-cycling processes, primary production, and decomposition). Remote sensing techniques provide a very convenient way of data acquisition capable of covering a large area several times during one season, so it can play a unique and essential role provided that we can relate remote sensing measurements to the biochemical characteristics of the Earth surface in a reliable and operational way. The application of remote sensing techniques for the estimation of canopy biochemicals was reviewed. Three methods of estimating biochemical concentrations of vegetation were included in this paper: index, stepwise multiple linear regression, and stepwise multiple linear regression based on a model of the forest crown. In addition, the vitality and potential applying value are stressed.

  16. Serum biochemical parameters of farmed carp (Cyprinus carpio

    Directory of Open Access Journals (Sweden)

    Tanţi Patriche


    Full Text Available Despite advances in ichthyo-pathology of recent years, interpretation of fish serum biochemical parameters is often difficult by lack of reference values. That is why to know the value of the serum biochemical parameters can be a useful tool for monitoring health status, detecting illnesses and responses to therapy. This paper provides data concerning biochemical composition of carp serum (Cyprinus carpio bred at Brateş Farm of Institute of Research and Development for Aquatic Ecology, Fishing and Aquaculture from Galaţi and Pleaşa Farm from Ploieşti, Romania. In research conducted onCyprinus carpio were determined following serum biochemical parameters: glucose (GLU, total proteins (TP, blood urea nitrogen (BUN, cholesterol (CHOL, triglyceride (TRIG, sodium (Na, potassium (K, calcium (Ca, magnesium (Mg, phosphorus (P, iron (Fe.

  17. Sublethal toxicity of carbofuran pesticide on the African catfish Clarias gariepinus (Burchell, 1822): hematological, biochemical and cytogenetic response. (United States)

    Harabawy, Ahmed S A; Ibrahim, Ahmed Th A


    The present work aimed to evaluate the toxic effects of two sublethal concentrations of carbofuran pesticide (0.16 and 0.49mg/L, for 35 days) on hematological and blood biochemical parameters of catfish, Clarias gariepinus, and to evaluate the genotoxic potential of carbofuran on the erythrocytes of C. gariepinus for the first time by micronucleus and erythrocyte alteration assays. The results revealed a significant (pgariepinus to carbofuran and allow us to consider that C. gariepinus as a good bioindicator to reflect the toxicity and the genotoxic potential of carbofuran that might be released into the aquatic ecosystems.

  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


    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......P-types which are epidemiologically unjustified, (ii) tests currently recommended for PhP-typing S. typhimurium may be somewhat unstable and not satisfactory for fingerprinting purposes, (iii) caution must be exercised when comparing data from different batches of PhP-48 plates, and (iv) best results...

  19. Evaluation of nutritional and biochemical parameters in spontaneously hypertensive rats following antihypertensive treatment

    Directory of Open Access Journals (Sweden)

    Joanna Suliburska


    Full Text Available Introduction. One side effect of antihypertensive drugs is their impact on nutritional status and metabolism. The purpose of this study was to assess the nutritional and biochemical parameters in spontaneously hypertensive rats following treatment with antihypertensive drugs. Material and methods. The experiment was performed on 50 male spontaneously hypertensive rats (SHR, which were assigned to fi ve groups: control (C, with perindopril (PR, with metoprolol (MT, with indapamide (ID, and with amlodipine (AM. All rats were provided ad libitum standard diet (with or without drugs and distilled water. After 45 days, the animals were weighed and killed. Liver, kidney, heart, spleen, pancreas, and blood samples were collected. Concentrations of glucose, cholesterol, triglycerides, and albumin were assayed in serum. Morphology parameters, such as white blood cell, red blood cell, hematocrit, and lymphocyte counts were measured in the blood. Blood pressure was measured using a tail-cuff plethysmograph. Results. The results obtained indicate that the hypotensive drugs under investigation had no effect on the selected nutritional parameters. Perindopril signifi cantly decreased the relative mass of the heart and amlodipine markedly decreased the relative mass of the pancreas. A markedly higher concentration of glucose in the group with indapamid, and a signifi cantly lower concentration of triglycerides in the group with metoprolol, were observed. Indapamide and amlodipine markedly increased the value of red blood cells and hematocrit in the blood of SHR. Conclusions. Long-term therapy with antihypertension drugs may infl uence tissue mass and biochemical and morphological status in the body.

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

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    Carles eBosch


    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.

  1. Biochemical and toxicological studies of aqueous extract of Syzigium aromaticum (L.) Merr. & Perry (Myrtaceae) in rodents. (United States)

    Agbaje, E O; Adeneye, A A; Daramola, A O


    The effects of long-term administration of boiled aqueous extract of Syzigium aromaticum (SYZ), commonly known as clove (which has been locally employed for treating gastrointestinal tract diseases and also used as food spices), on some biochemical indices, such as body weight, liver functions and blood parameters were studied in adult albino rats of both sexes. Selected doses of 300 and 700 mg kg(-1) were given orally through cannular to groups of animals for a period of 90 days, while the control group received distilled water throughout the duration of study via the same route. Blood samples collected after therapy and assayed for activities of some liver enzymes recorded a significant (p<0.05) and prominent effect on ALP and AST. Measurement of haematological parameters also revealed significant effects (p<0.05; p<0.001) on Hb, RBC (p<0.05), PCV (p<0.001), platelets (p<0.001) and granulocytes (p<0.001). An insignificant reduction was recorded for total WBC. The histopathological study conducted was in consonance with the results of the biochemical investigations that the aqueous extract of SYZ even at moderate doses, significantly affects body organs, their enzymes as well as the various functions. LD(50) for both intraperitoneal and oral routes of SYZ were 263 and 2500 mg kg(-1) respectively. The present work has revealed the toxicity of sub chronic administration of SYZ, which suggests that its prolonged usage must be avoided.


    Directory of Open Access Journals (Sweden)

    Arkadiusz Telesiński


    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.

  3. Biochemical characterization and antioxidant and antiproliferative activities of different Ganoderma collections. (United States)

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


    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.

  4. Biochemical basis of permethrin resistance in Anopheles arabiensis from Lower Moshi, north-eastern Tanzania

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    Oxborough Richard M


    Full Text Available Abstract Background Development of resistance to different classes of insecticides is a potential threat to malaria control. With the increasing coverage of long-lasting insecticide-treated nets in Tanzania, the continued monitoring of resistance in vector populations is crucial. It may facilitate the development of novel strategies to prevent or minimize the spread of resistance. In this study, metabolic-based mechanisms conferring permethrin (pyrethroid resistance were investigated in Anopheles arabiensis of Lower Moshi, Kilimanjaro region of north-eastern Tanzania. Methods WHO susceptibility test kits were used to detect resistance to permethrin in An. arabiensis. The levels and mechanisms of permethrin resistance were determined using CDC bottle bioassays and microplate (biochemical assays. In bottle bioassays, piperonyl butoxide (PBO and s,s,s-tributyl phosphorotrithioate (DEF were used as synergists to inhibit mixed function oxidases and non-specific esterases respectively. Biochemical assays were carried out in individual mosquitoes to detect any increase in the activity of enzymes typically involved in insecticide metabolism (mixed function oxidases, α- and β-esterases. Results Anopheles arabiensis from the study area was found to be partially resistant to permethrin, giving only 87% mortality in WHO test kits. Resistance ratios at KT50 and KT95 were 4.0 and 4.3 respectively. The permethrin resistance was partially synergized by DEF and by PBO when these were mixed with permethrin in bottle bioassays and was fully synergized when DEF and PBO were used together. The levels of oxidase and β-esterase activity were significantly higher in An. arabiensis from Lower Moshi than in the laboratory susceptible strain. There was no difference in α-esterase activity between the two strains. Conclusion Elevated levels of mixed function oxidases and β-esterases play a role in detoxification of permethrin in the resistant An. arabiensis population

  5. Anatomical and biochemical investigation of primary brain tumours

    Energy Technology Data Exchange (ETDEWEB)

    Del Sole, A. [Univ. di Milano (Italy); Falini, A. [Univ. Vita e Salute (Italy). IRCCS; Ravasi, L.; Ottobrini, L.; Lucignani, G. [Univ. di Milano (Italy). Ist. di Scienze Radiologiche; De Marchis, D. [Univ. di Milano-Bicocca (Italy); Bombardieri, E. [Istituto Nazionale dei Tumori, Milano (Italy)


    Cancerous transformation entails major biochemical changes including modifications of the energy metabolism of the cell, e.g. utilisation of glucose and other substrates, protein synthesis, and expression of receptors and antigens. Tumour growth also leads to heterogeneity in blood flow owing to focal necrosis, angiogenesis and metabolic demands, as well as disruption of transport mechanisms of substrates across cell membranes and other physiological boundaries such as the blood-brain barrier. All these biochemical, histological and anatomical changes can be assessed with emission tomography, X-ray computed tomography (CT), magnetic resonance imaging (MRI) and magnetic resonance spectroscopy (MRS). Whereas anatomical imaging is aimed at the diagnosis of brain tumours, biochemical imaging is better suited for tissue characterisation. The identification of a tumoural mass and the assessment of its size and vascularisation are best achieved with X-ray CT and MRI, while biochemical imaging can provide additional information that is crucial for tumour classification, differential diagnosis and follow-up. As the assessment of variables such as water content, appearance of cystic lesions and location of the tumour are largely irrelevant for tissue characterisation, a number of probes have been employed for the assessment of the biochemical features of tumours. Since biochemical changes may be related to the growth rate of cancer cells, they can be thought of as markers of tumour cell proliferation. Biochemical imaging with radionuclides of processes that occur at a cellular level provides information that complements findings obtained by anatomical imaging aimed at depicting structural, vascular and histological changes. This review focusses on the clinical application of anatomical brain imaging and biochemical assessment with positron emission tomography, single-photon emission tomography and MRS in the diagnosis of primary brain tumours, as well as in follow-up. (orig.)

  6. Diabetes, Biochemical Markers of Bone Turnover, Diabetes Control, and Bone


    Starup-Linde, Jakob


    Diabetes mellitus is known to have late complications including micro vascular and macro vascular disease. This review focuses on another possible area of complication regarding diabetes; bone. Diabetes may affect bone via bone structure, bone density, and biochemical markers of bone turnover. The aim of the present review is to examine in vivo from humans on biochemical markers of bone turnover in diabetics compared to non-diabetics. Furthermore, the effect of glycemic control on bone marker...

  7. Development and application of assays for serotonin

    Energy Technology Data Exchange (ETDEWEB)

    Gow, I.F.


    In this thesis, two assays for serotonin were developed, validated, and used to investigate the relationship between platelet aggregation, serotonin levels and sodium status and serotonin levels and platelet function in patients with cardiovascular disease. A radioimmunoassay (RIA) using an (/sup 125/I)-labelled tracer was developed and validated for the measurement of serotonin in human platelet-rich plasma (PRP) and rat serum. Antisera were raised against N-succinamylserotonin conjugated to bovine albumin and, to improve assay sensitivity, the analyte was made chemically similar to the immunogen by conversion to N-acetylserotonin prior to assay, using the specific amino reagent N-acetoxysuccinimide. An assay for serotonin using high-pressure liquid chromatography with electrochemical detection (HPLC-ECD) was developed, and used to validate the RIA. The RIA can be used to assay up to 100 samples/day compared with 10-20/day by the HPLC-ECD assay.

  8. [Fifty years of cooperation--FEBS and Polish Biochemical Society]. (United States)

    Barańska, Jolanta


    This year, the Federation of European Biochemical Societies (FEBS) celebrates its 50th anniversary. The Polish Biochemical Society, represented by the Society's President, Kazimierz Zakrzewski, was a founding member of the organization. The text presents a history of collaboration between FEBS and Polish Biochemical Society, the participation of Polish Biochemical Society members in different FEBS activities, as well as the role they played in running the Federation. Author describes FEBS Congresses which taken place in Warsaw, the first 3rd FEBS Meeting in 1966 and then 29th Congress in 2004. The profiles of Jakub Karol Parnas, the founding father of the Polish biochemistry and some crucial Presidents of the Society, are also presented. The text describes Parnas Conferences, organized jointly by Polish and Ukrainian Biochemical Societies from 1996, and growing from 2011 into three-nation event with participation of Ukrainian, Israeli and Polish scientists, largely due to significant help from FEBS. Summarizing the last few years, author judge the cooperation between the Federation and the Polish Biochemical Society as optimal.

  9. A novel multiplex cell viability assay for high-throughput RNAi screening.

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    Daniel F Gilbert

    Full Text Available Cell-based high-throughput RNAi screening has become a powerful research tool in addressing a variety of biological questions. In RNAi screening, one of the most commonly applied assay system is measuring the fitness of cells that is usually quantified using fluorescence, luminescence and absorption-based readouts. These methods, typically implemented and scaled to large-scale screening format, however often only yield limited information on the cell fitness phenotype due to evaluation of a single and indirect physiological indicator. To address this problem, we have established a cell fitness multiplexing assay which combines a biochemical approach and two fluorescence-based assaying methods. We applied this assay in a large-scale RNAi screening experiment with siRNA pools targeting the human kinome in different modified HEK293 cell lines. Subsequent analysis of ranked fitness phenotypes assessed by the different assaying methods revealed average phenotype intersections of 50.7±2.3%-58.7±14.4% when two indicators were combined and 40-48% when a third indicator was taken into account. From these observations we conclude that combination of multiple fitness measures may decrease false-positive rates and increases confidence for hit selection. Our robust experimental and analytical method improves the classical approach in terms of time, data comprehensiveness and cost.

  10. A rapid and quantitative coat protein complex II vesicle formation assay using luciferase reporters. (United States)

    Fromme, J Chris; Kim, Jinoh


    The majority of protein export from the endoplasmic reticulum (ER) is facilitated by coat protein complex II (COPII). The COPII proteins deform the ER membrane into vesicles at the ER exit sites. During the vesicle formation step, the COPII proteins load cargo molecules into the vesicles. Formation of COPII vesicles has been reconstituted in vitro in yeast and in mammalian systems. These in vitro COPII vesicle formation assays involve incubation of microsomal membranes and purified COPII proteins with nucleotides. COPII vesicles are separated from the microsomes by differential centrifugation. Interestingly, the efficiency of the COPII vesicle formation with purified recombinant mammalian COPII proteins is lower than that with cytosol, suggesting that an additional cytosolic factor(s) is involved in this process. Indeed, other studies have also implicated additional factors. To facilitate biochemical identification of such regulators, a rapid and quantitative COPII vesicle formation assay is necessary because the current assay is lengthy. To expedite this assay, we generated luciferase reporter constructs. The reporter proteins were packaged into COPII vesicles and yielded quantifiable luminescent signals, resulting in a rapid and quantitative COPII vesicle formation assay.

  11. A robotics-based automated assay for inorganic and organic phosphates. (United States)

    Cogan, E B; Birrell, G B; Griffith, O H


    Phosphate analyses are fundamental to a broad range of biochemical applications involving inorganic phosphate and organic phosphoesters such as phospholipids, phosphorylated proteins, and nucleic acids. A practical automated method utilizing robotics is described in this report. Five colorimetric methods of phosphate analyses based on formation of a phosphomolybdate complex and compatible with the automated assay were tested, and the fundamental chemistry is discussed. The relative sensitivities are malachite green > crystal violet > quinaldine red > ascorbate reduction > antimony-modified ascorbate reduction, although only a fourfold improvement was observed in going from the modified ascorbate procedure to malachite green. Malachite green was selected to optimize the assay because this dye provided the highest sensitivity. However, where color stability and low blanks are more important than sensitivity, the ascorbate reduction and quinaldine red methods were found to be better choices than malachite green. Automation using a robotic liquid-handling system substantially reduces the labor required to process large arrays of samples. The result is a sensitive, nonradioactive assay of inorganic phosphate with high throughput. A digestion step in an acid-resistant 96-well plate was developed to extend the assay to phosphate esters. The robotic-based assay was demonstrated with inorganic phosphate and a common phospholipid, phosphatidylcholine.

  12. A novel multiplex cell viability assay for high-throughput RNAi screening. (United States)

    Gilbert, Daniel F; Erdmann, Gerrit; Zhang, Xian; Fritzsche, Anja; Demir, Kubilay; Jaedicke, Andreas; Muehlenberg, Katja; Wanker, Erich E; Boutros, Michael


    Cell-based high-throughput RNAi screening has become a powerful research tool in addressing a variety of biological questions. In RNAi screening, one of the most commonly applied assay system is measuring the fitness of cells that is usually quantified using fluorescence, luminescence and absorption-based readouts. These methods, typically implemented and scaled to large-scale screening format, however often only yield limited information on the cell fitness phenotype due to evaluation of a single and indirect physiological indicator. To address this problem, we have established a cell fitness multiplexing assay which combines a biochemical approach and two fluorescence-based assaying methods. We applied this assay in a large-scale RNAi screening experiment with siRNA pools targeting the human kinome in different modified HEK293 cell lines. Subsequent analysis of ranked fitness phenotypes assessed by the different assaying methods revealed average phenotype intersections of 50.7±2.3%-58.7±14.4% when two indicators were combined and 40-48% when a third indicator was taken into account. From these observations we conclude that combination of multiple fitness measures may decrease false-positive rates and increases confidence for hit selection. Our robust experimental and analytical method improves the classical approach in terms of time, data comprehensiveness and cost.

  13. Cupric ion reducing antioxidant capacity assay for food antioxidants: vitamins, polyphenolics, and flavonoids in food extracts. (United States)

    Apak, Reşat; Güçlü, Kubilay; Ozyürek, Mustafa; Bektas Oğlu, Burcu; Bener, Mustafa


    Antioxidants are health beneficial compounds through their combat with reactive oxygen and nitrogen species and free radicals that may cause tissue damage leading to various diseases. This work reports the development of a simple and widely applicable antioxidant capacity index for dietary polyphenols, vitamins C and E, and plasma antioxidants utilizing the copper(II)-neocuproine (Cu(II)-Nc) reagent as the chromogenic oxidizing agent. This novel method based on an electron-transfer mechanism was named by our research group as 'cupric reducing antioxidant capacity', abbreviated as the CUPRAC method. The method is comprised of mixing the antioxidant solution with aqueous copper(II) chloride, alcoholic neocuproine, and ammonium acetate aqueous buffer at pH 7, and subsequently measuring the developed absorbance at 450 nm after 30 min. Since the color development is fast for compounds like ascorbic acid, gallic acid, and quercetin but slow for naringin and naringenin, the latter compounds are assayed after incubation at 50 degrees C on a water bath for 20 min. The flavonoid glycosides are hydrolyzed to their corresponding aglycones by refluxing in 1.2 M: HCl-containing 50% MeOH so as to exert maximal reducing power towards Cu(II)-Nc. The CUPRAC antioxidant capacities of synthetic mixtures are equal to the sum of individual capacities of antioxidant constituents, indicating lack of chemical deviations from Beer's law. Tests on antioxidant polyphenols demonstrate that the highest CUPRAC capacities are observed for epicatechin gallate, epigallocatechin gallate, quercetin, fisetin, epigallocatechin, catechin, and caffeic acid in this order, in accord with the number and position of the -OH groups as well the conjugation level of the molecule. The parallelism of the linear calibration curves of pure antioxidants in water and in a given complex matrix (plant extract) demonstrates that there are no chemical interactions of interferent nature among the solution constituents

  14. Direct Spectrophotometric Assay for Benzaldehyde Lyase Activity

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    Dessy Natalia


    Full Text Available Benzaldehyde lyase from Pseudomonas fluorescens Biovar I. (BAL, EC is a versatile catalyst for the organic synthesis of chiral α-hydroxy ketones. To allow fast assessment of enzyme activity, a direct spectrophotometric assay is desirable. Here, a new robust and easy-to-handle assay based on UV absorption is presented. The assay developed is based on the ligation of the α-hydroxy ketone (R-2,2′-furoin from 2-furaldehyde. A robust assay with direct monitoring of the product is facilitated with a convenient concentration working range minimising experimental associated with low concentrations.

  15. Bioassay and biochemical studies of the status of pirimiphos-methyl and cypermethrin resistance in Aedes (Stegomyia) aegypti and Aedes (Stegomyia) albopictus (Diptera: Culicidae) in Singapore. (United States)

    Lee, R M L; Choong, C T H; Goh, B P L; Ng, L C; Lam-Phua, S G


    Aedes (Stegomyia) aegypti (Linnaeus) and Ae. (Stegomyia) albopictus (Skuse) were sampled from five regions of Singapore (Central, North East, North West, South East and South West) and tested with diagnostic concentrations of the technical grade insecticides, pirimiphos-methyl and cypermethrin. Biochemical assays were performed on the same populations of Ae. aegypti and Ae. albopictus to determine activities of detoxifying enzymes, including non-specific esterase (EST), monooxygenase (MFO) and acetylcholinesterase (AChE). The diagnostic test showed that all Ae. aegypti populations were susceptible to pirimiphos-methyl (mortality = 99 to 100%), but resistant to cypermethrin (mortality = 11 to 76%). Resistance to pirimiphos-methyl was observed in all Ae. albopictus populations (mortality = 49 to 74%) while cypermethrin resistance was detected in most Ae. albopictus populations (mortality = 40 to 75%), except those from Central (mortality = 86%) and South East (mortality = 94%) showing incipient resistance. The biochemical assays showed that there was significant enhancement (P aegypti populations. The biochemical assay results suggested that AChE could play a role in pirimiphos-methyl resistance of Ae. albopictus in South West, South East and North East regions. The small but significant increase in EST activities in Ae. aegypti from all regions suggest that it may play a role in the observed cypermethrin resistance.

  16. Evaluation of the mitochondrial respiratory chain and oxidative phosphorylation system using polarography and spectrophotometric enzyme assays. (United States)

    Barrientos, Antoni; Fontanesi, Flavia; Díaz, Francisca


    The oxidative phosphorylation (OXPHOS) system consists of five multimeric complexes embedded in the mitochondrial inner membrane. They work in concert to drive the aerobic synthesis of ATP. Mitochondrial and nuclear DNA mutations affecting the accumulation and function of these enzymes are the most common cause of mitochondrial diseases and have also been associated with neurodegeneration and aging. For this reason, several approaches for the assessment of the OXPHOS system enzymes have been developed. Based on the methods described elsewhere, the assays describe methods that form a biochemical characterization of the OXPHOS system in cells and mitochondria isolated from cultured cells or tissues.

  17. Biochemical and histological characterization of tomato mutants

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    Carolina C. Monteiro


    Full Text Available Biochemical responses inherent to antioxidant systems as well morphological and anatomical properties of photomorphogenic, hormonal and developmental tomato mutants were investigated. Compared to the non-mutant Micro-Tom (MT, we observed that the malondialdehyde (MDA content was enhanced in the diageotropica (dgt and lutescent (l mutants, whilst the highest levels of hydrogen peroxide (H2O2 were observed in high pigment 1 (hp1 and aurea (au mutants. The analyses of antioxidant enzymes revealed that all mutants exhibited reduced catalase (CAT activity when compared to MT. Guaiacol peroxidase (GPOX was enhanced in both sitiens (sit and notabilis (not mutants, whereas in not mutant there was an increase in ascorbate peroxidase (APX. Based on PAGE analysis, the activities of glutathione reductase (GR isoforms III, IV, V and VI were increased in l leaves, while the activity of superoxide dismutase (SOD isoform III was reduced in leaves of sit, epi, Never ripe (Nr and green flesh (gf mutants. Microscopic analyses revealed that hp1 and au showed an increase in leaf intercellular spaces, whereas sit exhibited a decrease. The au and hp1 mutants also exhibited a decreased in the number of leaf trichomes. The characterization of these mutants is essential for their future use in plant development and ecophysiology studies, such as abiotic and biotic stresses on the oxidative metabolism.Neste trabalho, analisamos as respostas bioquímicas inerentes ao sistema antioxidante, assim como propriedades morfológicas e anatômicas de mutantes fotomorfogenéticos e hormonais de tomateiro. Comparados ao não mutante Micro-Tom (MT, observamos que o conteúdo de malondialdeído (MDA aumentou nos mutantes diageotropica (dgt e lutescent (l, enquanto os maiores níveis de H2O2 foram encontrados nos mutantes high pigment 1 (hp1 e aurea (au. Análises de enzimas antioxidantes mostraram que todos os mutantes reduziram a atividade de catalase (CAT quando comparado a MT. A


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    Ekeanyanwu Raphael Chukwuma


    Full Text Available The nutritive constituents of the seeds of Monodora tenuifolia were analyzed to augment the available information on Monodora tenuifolia research. Blood glucose and lipid profile were investigated on the flavonoid rich fraction of M. tenuifolia in rats. The composition (gkg-1 of alkaloids, cyanogenic glycosides, tannins and flavonoids were 13.3±0.1, 21.2×10-2±0.6, 1.3±0.1, 1.7±0.1 and 11.7±1.1 respectively. The proximate composition (gkg-1 of M. tenuifolia seed were crude fibre (262.3±1.2, crude protein (82.6±1.0, crude fat (349.9±1.9, ash (49.9±0.6, moisture (190.0±0.00 and carbohydrate (65.5±4.7. Analysis of the minerals content (gkg-1 yielded calcium (864.0±29.38, sodium (2752.0±140.35, iron (3.34±0.06, zinc (5.26±0.08, potassium (326.4±13.06, magnesium (342.9±13.71 and phosphorus (9.52±0.17, while vitamin analysis yielded vitamin A (10.05±0.17 iu/100 g, C (56.40±0.14 gkg-1 and E (11.71±0.87 iu /100 g, thiamine (0.11±0.01 gkg-1, niacin (0.46±0.32 gkg-1 and riboflavin (0.04±0.01 gkg-1. The results of amino acid analysis showed the total amino acid of M. tenuifolia seed was 71.78 of crude protein. The total essential amino acid with Histidine was calculated to be 29.24 of the crude protein. The antinutrient analysis of M. tenuifolia shows it contained total phenol (0.8±0.0 gkg-1, oxalates (4.09±1.17 gkg-1, phytates (0.012±0.42 gkg-1 and trypsin inhibitor (0.230±0.42 iu/g. The main fatty acids of the seed oil are linoleic acid (401.7 g kg-1, oleic acid (346.1 g kg-1 and palmitic acid (122.61 g kg-1. The LD50 of the flavonoid-rich fraction was found to be above 5000 mg kg-1 b.w. After the day 14 study, biochemical markers such as triacylglycerol, very low density lipoprotein increased significantly (p0.05 effect was observed on the blood glucose and lipid profile of wistar albino rats compared with the control. The result shows that M. tenuifolia seed is rich

  19. Alterations of biliary biochemical constituents and cytokines in infantile hepatitis syndrome

    Institute of Scientific and Technical Information of China (English)

    Yan Ding; Lei Zhao; Hong Mei; Zhi-Hua Huang; Shu-Ling Zhang


    AIM: To investigate the biliary biochemical constituents and cytokines in infantile hepatitis syndrome (IHS).METHODS: From 42 IHS subjects and 21 controls,serum and biliary biochemical constituents, including total bilirubin (TBIL), direct bilirubin (DBIL), alanine aminotransferase (ALT), gamma-glutamyl transpeptidase (γ-GT), total bile acid (TBA), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) both in bile and serum,were assayed. The subjects with IHS were divided into a cholestasis group (n = 21) and a hepatitis group (n = 21).RESULTS: In the cholestasis group, serum TBIL, DBIL,ALT, γ-GT, TBA, IL-6 and TNF-α levels were higher than those in the control (P < 0.01); and also the biliary TBIL, DBIL, γ-GT and TBA levels were lower than those in the control, whereas biliary IL-6 and TNF-α levels were higher than those in the control (P < 0.01). In the cholestasis group, serum IL-6 and TNF-α levels were lower than those in bile (P < 0.01). In the hepatitis group, serum DBTL, ALT, γ-GT, TBA, IL-6 and TNF-αlevels were higher than those in the control (P < 0.01or 140.57 ± 70.32 vs 79.06 ± 35.25, P < 0.05), while biliary TBIL, DBIL, γ-GT and TBA levels were lower than those in the control (P < 0.01), and biliary IL-6 and TNF-α levels were higher than those in the control (P < 0.01). In the hepatitis group, serum IL-6 and TNF-α levels were also lower than those in bile (P <0.01). Serum TBIL, DBIL, γ-GT, IL-6 and TNF-α levels in the cholestasis group were higher than those in the hepatitis group, while biliary IL-6 and TNF-α levels in the cholestasis group were higher than those in the hepatitis group. Biliary IL-6 and TNF-α were found to be more significantly increased than serum IL-6 and TNF-α in IHS (P < 0.01). The biliary IL-6 and TNF-α levels were positively correlated with serum DBIL, TBA and γ-GT levels in IHS subjects.CONCLUSION: Biliary biochemical constituents alter in coincidence with pathological changes in

  20. Assessing sediment contamination using six toxicity assays

    Directory of Open Access Journals (Sweden)

    Allen G. BURTON Jr.


    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. Radioreceptor assay: theory and applications to pharmacology

    Energy Technology Data Exchange (ETDEWEB)

    Perret, G. (U.E.R. de Medecine, Sante et Biologie Humaine, 93 - Bobigny (France)); Simon, P. (Faculte de Medecine Pitie-Salpetriere, 75 - Paris (France))

    The aim of the first part of this work is to present the theory of the radioreceptor assay and to compare it to the other techniques of radioanalysis (radioimmunoassay, competitive protein binding assays). The technology of the radioreceptor assay is then presented and its components (preparation of the receptors, radioligand, incubation medium) are described. The analytical characteristics of the radioreceptor assay (specificity, sensitivity, reproductibility, accuracy) and the pharmacological significance of the results are discussed. The second part is devoted to the description of the radioreceptor assays of some pharmacological classes (neuroleptics, tricyclic antidepressants, benzodiazepines, ..beta..-blockers, anticholinergic drugs) and to their use in therapeutic drug monitoring. In conclusion, by their nature, radioreceptor assays are highly sensitive, reliable, precise, accurate and simple to perform. Their chief disadvantage relates to specificity, since any substance having an appreciable affinity to the receptor site will displace the specifically bound radioligand. Paradoxically in some cases, this lack of specificity may be advantageous in that it allows for the detection of not only the apparent compound but of active metabolites and endogenous receptor agonists as well and in that radioreceptors assays can be devised for a whole pharmacological class and not only for one drug as it is the case for classical physico-chemical techniques. For all these reasons future of radioreceptor assay in pharmacology appears promising.


    NARCIS (Netherlands)



    Receptor assays occupy a particular position in the methods used in bioanalysis, as they do not exploit the physico-chemical properties of the analyte. These assays make use of the property of the analyte to bind to the specific binding site (receptor) and to competitively replace a labelled ligand

  3. Study on a new chromogenic substrate in detection of glucose in serum by glucose oxidase%血清葡萄糖氧化酶法新色原物的实验研究

    Institute of Scientific and Technical Information of China (English)

    王宝占; 李立和; 魏志斌; 刘冰


    Objective To establish the glucose oxidase ( GOD ) method using a new chromogenic substrate N-( 2-hydroxy-3-sulfopropyl )-3 ,5-dimethoxyaniline sodium salt ( HDAOS ) monohydrate in the detection of glucose in serum. Methods The tetrachlorophenol was replaced by HDAOS, the blue quinone-imine was formed, and it was measured at 600 nm. The GOD method, glucose oxidase-peroxidase-4-aminoantipyrene-phenol ( GOD-PAP ) and high performance liquid chromatography ( HPLC ) were used to determine serum hyper lipidemia samples ( 24 samples ), haemolysis samples ( 24 samples ), choloplania samples ( 24 samples ) and normal samples ( 24 samples ), and the results were compared. The methodology evaluation was performed. Results There was no statistical significance among the 3 methods, when the normal serum group was measured ( P >0. 05 ). Statistical significance only existed between the GOD method and GOD-PAP, when the serum groups of hyperlipidemia, hemolysis and choloplania were measured ( P 0.05 ). The between-run and within-run coefficients of variation ( CV) were 0.05);在测定脂血、溶血、黄疸血清时,本法葡萄糖测定结果与GOD-PAP法比较,差异有统计学意义(P<0.05);与HPLC法比较,差异无统计学意义(P>0.05).本法批内、批间变异系数(CV)均<3.0%;与HPLC法呈良好相关性(r2=0.995 8);在5 min内均可达到反应终点;线性范围为0.20~28.00 mmol/L;血红蛋白<2.0 g/L、乳糜<2.2%、总胆红素<200 μmol/L对本法的干扰误差<3%;参考范围为4.10~6.20 mmol/L(男)、4.04~6.15 mmol/L(女);最大吸收波长为595 nm.结论 以HDAOS代替4-氯酚生成蓝色醌亚胺,能通过降低脂血、溶血、黄疸光谱吸收的方法消除脂血、溶血和黄疸的干扰,提高GOD法测定葡萄糖的准确性,其使用方法与原有GOD终点法相同.

  4. The international standard ISO/TS 21872-1 to study the occurence of total and pathogenic Vibrio parahaemolyticus and Vibrio cholerae in seafood: ITS improvement by use of a chromogenic medium and PCR. (United States)

    Rosec, Jean-Philippe; Causse, Véronique; Cruz, Barbara; Rauzier, Jean; Carnat, Laurence


    During two surveys conducted in 2008 and 2009, the culture method described in the international standard ISO/TS 21872-1 was applied to the detection of Vibrio parahaemolyticus and Vibrio cholerae in 112 living bivalve mollusc samples, with a chromogenic medium used in addition to the TCBS agar, as second selective isolation medium and for enumeration of V. parahaemolyticus and V. cholerae by surface inoculation. A PCR method for detection of these 2 Vibrio species and the hemolysin genes tdh and trh, was applied in parallel. In 2009, the survey was extended to finfish fillets and crustaceans. PCR was also used for species confirmation of characteristic colonies. The identity of the PCR products, specifically targeting V. parahaemolyticus, was checked by sequencing. Occurrence of V. parahaemolyticus and V. cholerae isolates in living bivalve molluscs ranged from 30.4% to 32.6% and from 1.4% to 4.7% respectively. In frozen crustaceans (2009 survey) V. parahaemolyticus and V. cholerae isolates were respectively found in 45% and 10% of the samples. No V. parahaemolyticus or V. cholerae was detected in frozen fish fillets, neither by the ISO method nor by PCR. In 2009, enteropathogenic V. parahaemolyticus (trh+) was isolated from 4 out of 43 oyster samples while the trh gene was present in V. alginolyticus strains and in samples where V. parahaemolyticus was not detected (9 over 112 samples). The ISO method failed to isolate V. parahaemolyticus in 44% to 53% of the living bivalve molluscs where PCR detected the toxR gene specific of V. parahaemolyticus (Vp-toxR). Our results highlighted the need for a revision of the ISO/TS 21872-1 standard, at least, for analysis of living bivalve molluscs, and confirmed the increasing concern of enteropathogenic V. parahaemolyticus in French bivalve molluscs. Enrichment at 41.5°C was questioned and some reliable solutions for the improvement of the ISO/TS 21872-1 method, such as the PCR method for screening of positive samples and

  5. 铬黑T分光光度法测定饮料中痕量镍%Spectrophotometric Determination of Trace Amount of Nickel in Drinks with Eriochrome Black T as Chromogenic Reagent

    Institute of Scientific and Technical Information of China (English)

    斯琴高娃; 王立华; 王宁; 陈梦琪; 陈葛


    In a H2B4O7-KCl-NaOH buffer solution of pH 10.0,a stable coordination complex was formed by the reaction of Ni(Ⅱ) with sodium 1-(1-hydroxy-2-naphthylazo-6-nitro-2-naphthalene-4-sulfonate as chromogenic reagent,having its absorption maximum at the wavelength of 533 nm.Value of apparent molar absorptivity was found tobe 5.19× 104 L · mol 1 · cm-1.Linear relationship between values of absorbance and mass concentration of Ni(Ⅱ) was in the range within 1.2 mg · L 1.The proposed method was applied to the determination of Ni(Ⅱ)in drink samples,giving results in consistency with those obtained by AAS.Values of recovery and RSD's (n=9)found were in the ranges of 99.6%-100% and 1.4%-1.9%,respectively.%在pH 10.0的硼酸-氯化钾-氢氧化钠缓冲溶液中,显色剂铬黑T[1-(1-羟基-2-萘偶氮)-6-硝基-2-萘酚-4-磺酸钠]与镍(Ⅱ)形成稳定络合物.其最大吸收波长位于533 nm处,表观摩尔吸光率为5.19×104L·mol-1·cm-1.镍(Ⅱ)的质量浓度在1.2mg· L-1范围内与吸光度呈线性关系.方法用于饮料样品中微量镍(Ⅱ)的测定,测定值与原子吸收光谱法测定结果一致,回收率在99.6%~100%之间,相对标准偏差(n=9)在1.4%~1.9%之间.

  6. Acellular comet assay: a tool for assessing variables influencing the alkaline comet assay. (United States)

    Kennedy, Erin K; McNamee, James P; Prud'homme Lalonde, Louise; Jones, Trevor; Wilkinson, Diana


    In this study, an acellular modification to the alkaline comet assay to further evaluate key variables within the assay that may influence the outcome of genotoxicity studies is described. This acellular comet assay can detect differences of 0.2 Gy of (60)Co gamma-ray radiation between 0 and 1 Gy and differences of 1 Gy between 0 and 8 Gy; thus, this assay is applicable for a wide range of DNA damage levels. It is also shown that DNA damage from different radiation energies was not significantly different from (60)Co gamma-ray. This assay displayed a statistical increase in DNA damage due to uncontrolled exposure to natural light; however, the slope of the dose-response curve for light-exposed samples was similar to that for samples protected from light. A comparison of the alkaline comet assay with the acellular comet assay allowed for the intrinsic repair capacity of the alkaline comet assay to be quantified.

  7. Analysis of protein stability and ligand interactions by thermal shift assay. (United States)

    Huynh, Kathy; Partch, Carrie L


    Purification of recombinant proteins for biochemical assays and structural studies is time-consuming and presents inherent difficulties that depend on the optimization of protein stability. The use of dyes to monitor thermal denaturation of proteins with sensitive fluorescence detection enables rapid and inexpensive determination of protein stability using real-time PCR instruments. By screening a wide range of solution conditions and additives in a 96-well format, the thermal shift assay easily identifies conditions that significantly enhance the stability of recombinant proteins. The same approach can be used as an initial low-cost screen to discover new protein-ligand interactions by capitalizing on increases in protein stability that typically occur upon ligand binding. This unit presents a methodological workflow for small-scale, high-throughput thermal denaturation of recombinant proteins in the presence of SYPRO Orange dye.

  8. Photonic Crystal Surfaces as a General Purpose Platform for Label-Free and Fluorescent Assays. (United States)

    Cunningham, Brian T


    Photonic crystal surfaces can be designed to provide a wide range of functions that are used to perform biochemical and cell-based assays. Detection of the optical resonant reflections from photonic crystal surfaces enables high sensitivity label-free biosensing, while the enhanced electromagnetic fields that occur at resonant wavelengths can be used to enhance the detection sensitivity of any surface-based fluorescence assay. Fabrication of photonic crystals from inexpensive plastic materials over large surface areas enables them to be incorporated into standard formats that include microplates, microarrays, and microfluidic channels. This report reviews the design of photonic crystal biosensors, their associated detection instrumentation, and biological applications. Applications including small molecule high throughput screening, cell membrane integrin activation, gene expression analysis, and protein biomarker detection are highlighted. Recent results in which photonic crystal surfaces are used for enhancing the detection of Surface-Enhanced Raman Spectroscopy, and the development of high resolution photonic crystal-based laser biosensors are also described.

  9. A genetic assay for transcription errors reveals multilayer control of RNA polymerase II fidelity.

    Directory of Open Access Journals (Sweden)

    Jordan D Irvin


    Full Text Available We developed a highly sensitive assay to detect transcription errors in vivo. The assay is based on suppression of a missense mutation in the active site tyrosine in the Cre recombinase. Because Cre acts as tetramer, background from translation errors are negligible. Functional Cre resulting from rare transcription errors that restore the tyrosine codon can be detected by Cre-dependent rearrangement of reporter genes. Hence, transient transcription errors are captured as stable genetic changes. We used this Cre-based reporter to screen for mutations of Saccharomyces cerevisiae RPB1 (RPO21 that increase the level of misincorporation during transcription. The mutations are in three domains of Rpb1, the trigger loop, the bridge helix, and in sites involved in binding to TFIIS. Biochemical characterization demonstrates that these variants have elevated misincorporation, and/or ability to extend mispaired bases, or defects in TFIIS mediated editing.

  10. A genetic assay for transcription errors reveals multilayer control of RNA polymerase II fidelity. (United States)

    Irvin, Jordan D; Kireeva, Maria L; Gotte, Deanna R; Shafer, Brenda K; Huang, Ingold; Kashlev, Mikhail; Strathern, Jeffrey N


    We developed a highly sensitive assay to detect transcription errors in vivo. The assay is based on suppression of a missense mutation in the active site tyrosine in the Cre recombinase. Because Cre acts as tetramer, background from translation errors are negligible. Functional Cre resulting from rare transcription errors that restore the tyrosine codon can be detected by Cre-dependent rearrangement of reporter genes. Hence, transient transcription errors are captured as stable genetic changes. We used this Cre-based reporter to screen for mutations of Saccharomyces cerevisiae RPB1 (RPO21) that increase the level of misincorporation during transcription. The mutations are in three domains of Rpb1, the trigger loop, the bridge helix, and in sites involved in binding to TFIIS. Biochemical characterization demonstrates that these variants have elevated misincorporation, and/or ability to extend mispaired bases, or defects in TFIIS mediated editing.

  11. Relationship between ER-ICA and conventional steroid receptor assays in human breast cancer. (United States)

    Di Fronzo, G; Clement, C; Cappelletti, V; Miodini, P; Coradini, D; Ronchi, E; Andreola, S; Rilke, F


    We applied a new immunocytochemical assay for estrogen receptors (ER-ICA) to 82 human breast tumors. Results were correlated with cytosolic estrogen receptors (ERc) and nuclear ER (ERn) determined on the same sample respectively by the radioligand binding assay and by an ER enzyme immunoassay (ER-EIA). All ER-ICA-positive tumors contained more than 10 fmol/mg of protein of ERc and were therefore considered as ERc positive. In contrast, 15.4% of ERc-positive cases were ER-ICA negative. Comparison of ER-ICA results with ERn showed extensive agreement of negativity (92%), whereas 38% of ER-ICA-positive tumors were ER-EIA negative. However, the latter had ERc levels above the positivity threshold. Quantitative features of the immunocytochemical staining such as intensity and percentage of labelled cells, considered separately, did not reflect the amount of ERc or ERn. Cellularity was not significantly correlated with ER-ICA and biochemical results.

  12. Mesotrione herbicide promotes biochemical changes and DNA damage in two fish species

    Directory of Open Access Journals (Sweden)

    L.D.S. Piancini


    Full Text Available Mesotrione is one of the new herbicides that have emerged as an alternative after the ban of atrazine in the European Union. To our knowledge, any work using genetic or biochemical biomarkers was performed in any kind of fish evaluating the toxicity of this compound. The impact of acute (96 h exposure to environmentally relevant mesotrione concentrations (1.8, 7, 30, 115 e 460 μg L−1 were evaluated on the liver of Oreochorimis niloticus and Geophagus brasiliensis by assessing the activity of superoxide dismutase (SOD, glutathione peroxidase (GPx and glutathione-S- transferase (GST, the levels of reduced glutathione (GSH, carbonyl assays (PCO and lipid peroxide (LPO as well as the DNA damage to erithrocytes, liver and gills through the comet assay. We observed an increase in the concentration of GSH and the GPx activity in O. niloticus, and the GST and SOD activity in G. brasiliensis. We found significant increase in DNA damage in all tissues in both species. The results indicated that the acute exposure to mesotrione can induce oxidative stress and DNA damage in both species.

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

    Directory of Open Access Journals (Sweden)

    Aishwarya Singh Chauhan


    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.

  14. Acute Toxicity and Cytotoxicity Effect of Ethanolic Extract of Spondias tuberosa Arruda Bark: Hematological, Biochemical and Histopathological Evaluation

    Directory of Open Access Journals (Sweden)


    Full Text Available ABSTRACT Spondias tuberosa Arruda, popularly named as umbu, is native from savanna-like vegetation and widely used for medicinal purposes, however, the toxicological profile is not available yet. This study evaluated the phytochemical profile and acute toxicity and citoxicity of Ethanolic Extract of Spondias tuberosa Arruda Bark (EEStb in hematological, biochemical and histopathological parameters. Female Wistar rats were divided into: control (C and animal treated single doses of 300mg/Kg (EEStb300 or 2.000mg/kg body weight (ESStb2.000 of the EEStb. After 24 hours and 14 days from gavage, the behavior, hematological, biochemical and histopathological parameters were assayed. Cytotoxicity effect was evaluated on HEp-2 cell lines. Neither EEStb300 nor EEStb2.000 produced mortality nor changes in body weight during the 14-days of observation, but EEStb2.000 reduced quietly the food and water intake as well as locomotor activity at first day. There were no changes in macroscopic, histopathological, biochemical and hematological parameters. EEStb in concentrations of 6.25- 50μg ml−1 on HEp-2 cell did not produce cytotoxic effect. These results suggest that EEStb did not cause acute toxicity and cytotoxic, suggesting a good safety rate for Spondias tuberosa Arruda.

  15. Acute Toxicity and Cytotoxicity Effect of Ethanolic Extract of Spondias tuberosa Arruda Bark: Hematological, Biochemical and Histopathological Evaluation. (United States)

    Barbosa, Humberto M; Nascimento, Jailson N DO; Araújo, Thiago A S; Duarte, Filipe S; Albuquerque, Ulysses P; Vieira, Jeymesson R C; Santana, Edson R B DE; Yara, Ricardo; Lima, Cláudia S A; Gomes, Dayane A; Lira, Eduardo C


    Spondias tuberosa Arruda, popularly named as umbu, is native from savanna-like vegetation and widely used for medicinal purposes, however, the toxicological profile is not available yet. This study evaluated the phytochemical profile and acute toxicity and citoxicity of Ethanolic Extract of Spondias tuberosa Arruda Bark (EEStb) in hematological, biochemical and histopathological parameters. Female Wistar rats were divided into: control (C) and animal treated single doses of 300mg/Kg (EEStb300) or 2.000mg/kg body weight (ESStb2.000) of the EEStb. After 24 hours and 14 days from gavage, the behavior, hematological, biochemical and histopathological parameters were assayed. Cytotoxicity effect was evaluated on HEp-2 cell lines. Neither EEStb300 nor EEStb2.000 produced mortality nor changes in body weight during the 14-days of observation, but EEStb2.000 reduced quietly the food and water intake as well as locomotor activity at first day. There were no changes in macroscopic, histopathological, biochemical and hematological parameters. EEStb in concentrations of 6.25- 50μg ml-1 on HEp-2 cell did not produce cytotoxic effect. These results suggest that EEStb did not cause acute toxicity and cytotoxic, suggesting a good safety rate for Spondias tuberosa Arruda.

  16. Biochemical Changes in the Serum of Patients with Chronic Toxigenic Mold Exposures: A Risk Factor for Multiple Renal Dysfunctions

    Directory of Open Access Journals (Sweden)

    Ebere Anyanwu


    Full Text Available This paper analyzes and presents the biochemical abnormalities in the sera of patients presenting with chronic mycosis in order to investigate the relationship with the risks of multiple renal disorders. The study population (n = 10 consisted of six females and four males (mean age 36.3 years exposed by toxic molds in their homes and offices for an average of 2.8 years. The control group comprised ten people, five males and five females (mean age 35.9 years without any known exposures to toxic molds. Blood samples were obtained from both the patients and the controls and were processed using specific biochemical methods that included enzyme-linked immunoabsorbent assay (ELISA. There were biochemical abnormal concentrations in creatinine, uric acid, phosphorus, alkaline phosphotase, cholesterol, HDH, SGOT/AST, segmented neutrophils, lymphocytes, total T3, IgG and IgA immunoglobulins with significant differences between patients and controls. These abnormalities were consistent with multiple renal disorders. The major complaints of the mycosis patients were headaches, pulmonary symptoms, allergic reactions, memory loss, skin rashes, blurred vision symptoms, fatigue, and runny nose. These findings were depictive of a strong association of chronic mycosis with abnormal renal indicators. It was concluded that, although this research was a pilot investigation, based on the overall results, people exposed to chronic indoor environmental toxic molds were at risk of multiple renal complications.

  17. Improved benzodiazepine radioreceptor assay using the MultiScreen (R) Assay System

    NARCIS (Netherlands)

    Janssen, MJ; Ensing, K; de Zeeuw, RA


    In this article, an improved benzodiazepine radioreceptor assay is described, which allows substantial reduction in assay time, The filtration in this method was performed by using the MultiScreen(R) Assay System. The latter consists of a 96-well plate with glass fibre filters sealed at the bottom,

  18. Identification of biochemical network modules based on shortest retroactive distances.

    Directory of Open Access Journals (Sweden)

    Gautham Vivek Sridharan


    Full Text Available Modularity analysis offers a route to better understand the organization of cellular biochemical networks as well as to derive practically useful, simplified models of these complex systems. While there is general agreement regarding the qualitative properties of a biochemical module, there is no clear consensus on the quantitative criteria that may be used to systematically derive these modules. In this work, we investigate cyclical interactions as the defining characteristic of a biochemical module. We utilize a round trip distance metric, termed Shortest Retroactive Distance (ShReD, to characterize the retroactive connectivity between any two reactions in a biochemical network and to group together network components that mutually influence each other. We evaluate the metric on two types of networks that feature feedback interactions: (i epidermal growth factor receptor (EGFR signaling and (ii liver metabolism supporting drug transformation. For both networks, the ShReD partitions found hierarchically arranged modules that confirm biological intuition. In addition, the partitions also revealed modules that are less intuitive. In particular, ShReD-based partition of the metabolic network identified a 'redox' module that couples reactions of glucose, pyruvate, lipid and drug metabolism through shared production and consumption of NADPH. Our results suggest that retroactive interactions arising from feedback loops and metabolic cycles significantly contribute to the modularity of biochemical networks. For metabolic networks, cofactors play an important role as allosteric effectors that mediate the retroactive interactions.

  19. Biochemical characterisation during seed development of oil palm (Elaeis guineensis). (United States)

    Kok, Sau-Yee; Namasivayam, Parameswari; Ee, Gwendoline Cheng-Lian; Ong-Abdullah, Meilina


    Developmental biochemical information is a vital base for the elucidation of seed physiology and metabolism. However, no data regarding the biochemical profile of oil palm (Elaeis guineensis Jacq.) seed development has been reported thus far. In this study, the biochemical changes in the developing oil palm seed were investigated to study their developmental pattern. The biochemical composition found in the seed differed significantly among the developmental stages. During early seed development, the water, hexose (glucose and fructose), calcium and manganese contents were present in significantly high levels compared to the late developmental stage. Remarkable changes in the biochemical composition were observed at 10 weeks after anthesis (WAA): the dry weight and sucrose content increased significantly, whereas the water content and hexose content declined. The switch from a high to low hexose/sucrose ratio could be used to identify the onset of the maturation phase. At the late stage, dramatic water loss occurred, whereas the content of storage reserves increased progressively. Lauric acid was the most abundant fatty acid found in oil palm seed starting from 10 WAA.

  20. Elispot assay检测牛奶中庆大霉素%Establishment of Elispot Assay for Detection of Gentamicin in Milk

    Institute of Scientific and Technical Information of China (English)

    王丹; 许杨; 何庆华; 黄志兵; 康敏


    This manuscript developt a rapid detection method Elispot assay for Gentamicin sulfate,for this, Anti-GM polyclonal antibody through GM immunogen was obtained;Establishing milk GM-Elispot assay method by spotting coating antigen on PVDF membrane, and the coating antigen and GM in samples competitive binding the site of anti-GM polyclonal, then the HRP catalyzed chromogenic substrate, according to presence or absence of color and depth for interpretation of results.Results: The detection thre-shold of this method was 10 ng/mL,detection time was within 40 minutes and it can achieve semiquantitative detection.The strip of which was stored at 4 ℃ for 90 days could be used and no cross-reaction was found with several structural analogues.The results of samples analysis obtained from the Elispot assay were in a good agreement with that of ELISA method.%为了建立快速检测牛奶中庆大霉素的Elispot assay,采用制备GM免疫抗原获得抗GM 多克隆抗体.将检测抗原点阵在PVDF膜上,通过检测抗原和样品中GM竞争结合抗GM多克隆抗体,酶标结合物催化底物显色,根据颜色的有无及深浅判读结果,从而建立了检测牛奶中GM的Elispot assay.该方法的检测阈值为10 ng/mL,检测时间为40 min,可对样品实现半定量检测.该方法制备的试纸条于4℃密封保存90 d仍可用于检测;与多种结构类似物未见交叉反应;其结果与酶联免疫方法一致.

  1. Serum Bactericidal Assay: New Role in Salmonella Detection. (United States)

    Chen, Yu; Wu, Da; Sun, Min; Deng, Mingjun; Cui, Shuhua; Liang, Chengzhu; Geng, Juan; Sun, Tao; Long, Ling; Xiao, Xizhi


    While inspecting animal feed for Salmonella contamination, we routinely observed bacterial colonies on selective agars that were similar in appearance to those formed by Salmonella. These were identified as Citrobacter freundii, Proteus mirabilis, and Serratia fonticola using biochemical and serological techniques. Because the presence of these bacterial species confounds identification of Salmonella, we refer to them as "interference bacteria." Polyvalent antisera against these interference bacteria were prepared by immunizing rabbits with a mixture of all three organisms. To minimize or eliminate interference by these bacteria, the polyvalent antisera were introduced between the steps of selective enrichment and Salmonella-selective plating. The antisera raised against the interference bacteria, when combined with neonatal rabbit complement, exhibited specific bactericidal activity against C. freundii, P. mirabilis, and S. fonticola. The respective serum bactericidal assay titers were 2(9), 2(8), and 2(10). In selective broth, polyvalent antisera could also kill the target bacterial cells effectively. We tested 526 samples (186 white fishmeal, 97 red fishmeal, and 243 cattle bone powder) using the polyvalent antisera and found that the rates of contamination of each species of the three respective foods decreased by 58.8, 100, and 83%. Our data indicates that polyvalent sera against C. freundii, P. mirabilis, and S. fonticola can be used as inhibitors to increase the accuracy of Salmonella detection.

  2. Mobile non-destructive assay system

    Energy Technology Data Exchange (ETDEWEB)

    Colarusso, A.P.; Audas, J.H.; Bieri, J.M.; Herrera, G.C.; Hastings, R.D.; Horton, W.S.; Kuckertz, T.H.; Kunz, W.E.; Medvick, P.A.; Vogel, P.A.


    A mobile system for non-destructive assay (NDA), developed at the Los Alamos National Laboratory, provides accurate and sensitive measurements for transuranic (TRU) isotopes contained in 208-iota drums of miscellaneous nuclear wastes. The NDA unit consists of four major subsystems: an assay chamber, counting and digital electronics, data acquisition, and a neutron generator. It performs both active and passive neutron waste measurements. The former determines the amount of fissile isotopes at a sensitivity level of 1 mg plutonium. The latter determines spontaneous fission and ..cap alpha..,n) isotopes at a comparable level. A complete assay consists of sequential active and passive measurements. The assay measurement and other supporting data are incorporated in a commercial spreadsheet program (Lotus 1,2,3) for further analysis, which includes various matrix corrections and a determination of whether or not the drum exceeds the 100-nCi/g threshold for TRU wastes. Field tests have been performed on three separate occasions, accomplishing more than 1800 waste drum assays. These waste drum assays are discussed, especially those comparing passive and active neutron measurements with independent segmented gamma scan assays. Results obtained with a set of 15 drums containing plutonium prepared from standards and actual hot waste matrices are also reviewed.

  3. Nano-immunosafety: issues in assay validation

    Energy Technology Data Exchange (ETDEWEB)

    Boraschi, Diana; Italiani, Paola [Institute of Biomedical Technologies, National Research Council, Via G. Moruzzi 1, 56124 Pisa (Italy); Oostingh, Gertie J; Duschl, Albert [Department of Molecular Biology, University of Salzburg, Hellbrunnerstrasse 34, 5020 Salzburg (Austria); Casals, Eudald; Puntes, Victor F [Institut Catala de Nanotecnologia, Campus de la UAB - Facultat de Ciencies, Edifici CM7, 08193 Bellaterra (Spain); Nelissen, Inge, E-mail: [VITO NV, Boeretang 200, BE-2400 Mol (Belgium)


    Assessing the safety of engineered nanomaterials for human health must include a thorough evaluation of their effects on the immune system, which is responsible for defending the integrity of our body from damage and disease. An array of robust and representative assays should be set up and validated, which could be predictive of the effects of nanomaterials on immune responses. In a trans-European collaborative work, in vitro assays have been developed to this end. In vitro tests have been preferred for their suitability to standardisation and easier applicability. Adapting classical assays to testing the immunotoxicological effects of nanoparticulate materials has raised a series of issues that needed to be appropriately addressed in order to ensure reliability of results. Besides the exquisitely immunological problem of selecting representative endpoints predictive of the risk of developing disease, assay results turned out to be significantly biased by artefactual interference of the nanomaterials or contaminating agents with the assay protocol. Having addressed such problems, a series of robust and representative assays have been developed that describe the effects of engineered nanoparticles on professional and non-professional human defence cells. Two of such assays are described here, one based on primary human monocytes and the other employing human lung epithelial cells transfected with a reporter gene.

  4. Characterization of Bacterial Strains Isolated Through Microbial Profiling of Urine Samples

    Directory of Open Access Journals (Sweden)

    Poulomi Nandy


    Full Text Available The present study was conducted to determine the microbial profile in urine samples. Differential and selective chromogenic culture media were used for the rapid detection, identification and enumeration of urinary tract pathogens namely, E. coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus aureus, Enterococcus faecalis, Proteus mirabilis. Urine samples of normal healthy individuals as well as patients with Urinary Tract Infection (UTI were screened on hicrome agar plates. The cultivable bacteria present in urine were isolated based on chromogenic detection. Antibiotic sensitivity assay, morphological characterization and biochemical tests, namely protease, oxidase, catalase, lipase, DNase and lecithinase assay were performed with the 15 isolates obtained from urine samples. The molecular analyses of the isolates were done through partial sequencing of the 16SrDNA gene; six of them were found to be novel and submitted in GenBank under the accession numbers EF644491-96. Phylogenetic tree of the isolates were constructed by neighbour joining method.

  5. SABRE: A Tool for Stochastic Analysis of Biochemical Reaction Networks

    CERN Document Server

    Didier, Frederic; Mateescu, Maria; Wolf, Verena


    The importance of stochasticity within biological systems has been shown repeatedly during the last years and has raised the need for efficient stochastic tools. We present SABRE, a tool for stochastic analysis of biochemical reaction networks. SABRE implements fast adaptive uniformization (FAU), a direct numerical approximation algorithm for computing transient solutions of biochemical reaction networks. Biochemical reactions networks represent biological systems studied at a molecular level and these reactions can be modeled as transitions of a Markov chain. SABRE accepts as input the formalism of guarded commands, which it interprets either as continuous-time or as discrete-time Markov chains. Besides operating in a stochastic mode, SABRE may also perform a deterministic analysis by directly computing a mean-field approximation of the system under study. We illustrate the different functionalities of SABRE by means of biological case studies.

  6. The Development of a Biochemical Profile of Acacia Honey by Identifying Biochemical Determinants of its Quality

    Directory of Open Access Journals (Sweden)

    Liviu Alexandru MARGHITAS


    Full Text Available Codex Alimentarius Standard, EU Legislation and National Standards state honey authenticity. Authenticity in respect of production (to prevent adulteration and authenticity in respect of geographical and botanical origin are the two main aspects of general honey authenticity. Quality of honey depends on the plant source, the chemical composition of these plants as well, as on the climatic conditions and soil mineral composition. Romanian acacia (Robinia pseudoacacia honey that came from the most important Transylvanian massif (Valea lui Mihai, Bihor County, Romania was evaluated for authenticity by pollen-analysis, several physico-chemical analyses, including sugar profile and mineral content. As polyphenolic content could be also an important factor for botanical authentification, HPLC-DAD-MS analyses were performed to assess the fingerprint of this important secondary plant metabolite. Statistical data were processed in order to develop a biochemical profile of this type of honey and the main quality categories identification. The results of physico-chemical analysis demonstrated that the tested honey samples could be framed into monofloral type of acacia honeys. The analysis of acacia honeys originating from Valea lui Mihai, Romania, showed that polyphenolic profile (phenolic acids and flavonoids could be used as a complementary method for authenticity determination together with pollen analysis and other physico-chemical analysis.

  7. Enzyme-linked immunoassay for plasma-free metanephrines in the biochemical diagnosis of phaeochromocytoma in adults is not ideal.

    LENUS (Irish Health Repository)


    Abstract Background: The aim of the study was to define the analytical and diagnostic performance of the Labor Diagnostica Nord (LDN) 2-Met plasma ELISA assay for fractionated plasma metanephrines in the biochemical diagnosis of phaeochromocytoma. Methods: The stated manufacturer\\'s performance characteristics were assessed. Clinical utility was evaluated against liquid chromatography tandem mass spectrometry (LC-MS\\/MS) using bias, sensitivity and specificity outcomes. Samples (n=73) were collected from patients in whom phaeochromocytoma had been excluded (n=60) based on low probability of disease, repeat negative testing for urinary fractionated catecholamines and metanephrines, lack of radiological and histological evidence of a tumour and from a group (n=13) in whom the tumour had been histologically confirmed. Blood collected into k(2)EDTA tubes was processed within 30 min. Separated plasma was aliquoted (x2) and frozen at -40 degrees C prior to analyses. One aliquot was analysed for plasma metanephrines using the LDN 2-Met ELISA and the other by LC-MS\\/MS. Results: The mean bias of -32% for normetanephrine (ELISA) when compared to the reference method (LC-MS\\/MS) makes under-diagnosis of phaeochromocytoma likely. The sensitivity of the assay (100%) was equal to the reference method, but specificity (88.3%) lower than the reference method (95%), making it less than optimum for the biochemical diagnosis of phaeochromocytoma. Conclusions: Plasma-free metanephrines as measured by Labor Diagnostica Nord (LDN) 2-Met ELISA do not display test characteristics that would support their introduction or continuation as part of a screening protocol for the biochemical detection of phaeochromocytoma unless the calibration problem identified is corrected and other more accurate and analytically specific methods remain unavailable.

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

    Directory of Open Access Journals (Sweden)

    Hsien-Chin Wei


    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.

  9. A Multichannel Calorimetric Simultaneous Assay Platform Using a Microampere Constant-Current Looped Enthalpy Sensor Array (United States)

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


    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. PMID:28165412

  10. Determination of cell survival after irradiation via clonogenic assay versus multiple MTT Assay - A comparative study

    Directory of Open Access Journals (Sweden)

    Buch Karl


    Full Text Available Abstract For studying proliferation and determination of survival of cancer cells after irradiation, the multiple MTT assay, based on the reduction of a yellow water soluble tetrazolium salt to a purple water insoluble formazan dye by living cells was modified from a single-point towards a proliferation assay. This assay can be performed with a large number of samples in short time using multi-well-plates, assays can be performed semi-automatically with a microplate reader. Survival, the calculated parameter in this assay, is determined mathematically. Exponential growth in both control and irradiated groups was proven as the underlying basis of the applicability of the multiple MTT assay. The equivalence to a clonogenic survival assay with its disadvantages such as time consumption was proven in two setups including plating of cells before and after irradiation. Three cell lines (A 549, LN 229 and F 98 were included in the experiment to study its principal and general applicability.

  11. Physiological and biochemical basis of salmon young ifshes migratory behavior

    Institute of Scientific and Technical Information of China (English)

    Vladimir Ivanovich Martemyanov


    The review presents data on structural changes, physiological and biochemical reactions occurring at salmon young fishes during smoltification. It is shown, that young salmon fishes located in fresh water, in the process of smoltification undergo a complex of structural, physiological and biochemical changes directed on preparation of the organism for living in the sea. These changes cause stress reaction which excites young fishes to migrate down the river towards the sea. Measures to improve reproduction of young salmon fishes at fish farms are offered.

  12. The Biochemical Properties of Antibodies and Their Fragments. (United States)

    Hnasko, Robert M


    Immunoglobulins (Ig) or antibodies are powerful molecular recognition tools that can be used to identify minute quantities of a given target analyte. Their antigen-binding properties define both the sensitivity and selectivity of an immunoassay. Understanding the biochemical properties of this class of protein will provide users with the knowledge necessary to select the appropriate antibody composition to maximize immunoassay results. Here we define the general biochemical properties of antibodies and their similarities and differences, explain how these properties influence their functional relationship to an antigen target, and describe a method for the enzymatic fragmentation of antibodies into smaller functional parts.

  13. Biochemical methane potential, biodegradability, alkali treatment and influence of chemical composition on methane yield of yard wastes. (United States)

    Gunaseelan, Victor Nallathambi


    In this study, the biochemical CH4 potential, rate, biodegradability, NaOH treatment and the influence of chemical composition on CH4 yield of yard wastes generated from seven trees were examined. All the plant parts were sampled for their chemical composition and subjected to the biochemical CH4 potential assay. The component parts exhibited significant variation in biochemical CH4 potential, which was reflected in their ultimate CH4 yields that ranged from 109 to 382 ml g(-1) volatile solids added and their rate constants that ranged from 0.042 to 0.173 d(-1). The biodegradability of the yard wastes ranged from 0.26 to 0.86. Variation in the biochemical CH4 potential of the yard wastes could be attributed to variation in the chemical composition of the different fractions. In the Thespesia yellow withered leaf, Tamarindus fruit pericarp and Albizia pod husk, NaOH treatment enhanced the ultimate CH4 yields by 17%, 77% and 63%, respectively, and biodegradability by 15%, 77% and 61%, respectively, compared with the untreated samples. The effectiveness of NaOH treatment varied for different yard wastes, depending on the amounts of acid detergent fibre content. Gliricidia petals, Prosopis leaf, inflorescence and immature pod, Tamarindus seeds, Albizia seeds, Cassia seeds and Delonix seeds exhibited CH4 yields higher than 300 ml g(-1) volatile solids added. Multiple linear regression models for predicting the ultimate CH4 yield and biodegradability of yard wastes were designed from the results of this work.

  14. 21 CFR 864.7250 - Erythropoietin assay. (United States)


    ... erythropoietin (an enzyme that regulates the production of red blood cells) in serum or urine. This assay provides diagnostic information for the evaluation of erythrocytosis (increased total red cell mass)...

  15. Variables Affecting Two Electron Transport System Assays


    Burton, G. Allen; Lanza, Guy R.


    Several methodological variables were critical in two commonly used electron transport activity assays. The dehydrogenase assay based on triphenyl formazan production exhibited a nonlinear relationship between formazan production (dehydrogenase activity) and sediment dilution, and linear formazan production occurred for 1 h in sediment slurries. Activity decreased with increased time of sediment storage at 4°C. Extraction efficiencies of formazan from sediment varied with alcohol type; methan...

  16. Electrochemical Assay of Gold-Plating Solutions (United States)

    Chiodo, R.


    Gold content of plating solution is assayed by simple method that required only ordinary electrochemical laboratory equipment and materials. Technique involves electrodeposition of gold from solution onto electrode, the weight gain of which is measured. Suitable fast assay methods are economically and practically necessary in electronics and decorative-plating industries. If gold content in plating bath is too low, poor plating may result, with consequent economic loss to user.

  17. The comet assay: a heavenly method! (United States)

    Collins, Andrew R


    The contributions to this special issue of Mutagenesis have been selected to cover the main research areas served by the comet assay, namely genotoxicology, environmental toxicology, human biomonitoring and fundamental investigations into mechanisms of DNA damage and repair. Innovative methods are described, technical issues are explored, and guidelines are given for venturing into relatively new or unexploited areas of research. The popularity of the comet assay in a historical context is illustrated by a bibliometric survey.

  18. Biosensors and bioelectronics on smartphone for portable biochemical detection. (United States)

    Zhang, Diming; Liu, Qingjun


    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.

  19. Polynomial analysis of canopy spectra and biochemical component content inversion

    Institute of Scientific and Technical Information of China (English)

    YAN Chunyan; LIU Qiang; NIU Zheng; WANG Jihua; HUANG Wenjiang; LIU Liangyun


    A polynomial expression model was developed in this paper to describe directional canopy spectra, and the decomposition of the polynomial expression was used as a tool for retrieving biochemical component content from canopy multi-angle spectra. First, the basic formula of the polynomial expression was introduced and the physical meaning of its terms and coefficients was discussed. Based on this analysis, a complete polynomial expression model and its decomposition method were given. By decomposing the canopy spectra simulated with SAILH model, it shows that the polynomial expression can not only fit well the canopy spectra, but also show the contribution of every order scattering to the whole reflectance. Taking the first scattering coefficients a10 and a01 for example, the test results show that the polynomial coefficients reflect very well the hot spot phenomenon and the effects of viewing angles, LAI and leaf inclination angle on canopy spectra. By coupling the polynomial expression with leaf model PROSPECT, a canopy biochemical component content inversion model was given. In the simulated test, the canopy multi-angle spectra were simulated by two different models, SAILH and 4-SCALE respectively, then the biochemical component content was retrieved by inverting the coupled polynomial expression + PROSPECT model. Results of the simulated test are promising, and when applying the algorithm to measured corn canopy multi-angle spectra, we also get relatively accurate chlorophyll content. It shows that the polynomial analysis provides a new method to get biochemical component content independent of any specific canopy model.

  20. Saliva as research material: Biochemical, physicochemical and practical aspects

    NARCIS (Netherlands)

    Schipper, R.G.; Silletti, E.; Vingerhoeds, M.H.


    Whole saliva is a complex mixture of proteins and other molecules which originate from several sources. The biochemical and physicochemical properties of saliva contribute to the numerous functions of saliva in, e.g., speech, maintaining oral and general health, and food processing. Interest in sali

  1. MATLAB-Based Teaching Modules in Biochemical Engineering (United States)

    Lee, Kilho; Comolli, Noelle K.; Kelly, William J.; Huang, Zuyi


    Mathematical models play an important role in biochemical engineering. For example, the models developed in the field of systems biology have been used to identify drug targets to treat pathogens such as Pseudomonas aeruginosa in biofilms. In addition, competitive binding models for chromatography processes have been developed to predict expanded…

  2. Biochemical aspects of pressure tolerance in marine mammals. (United States)

    Castellini, Michael A; Rivera, Patricia M; Castellini, Judith M


    Some marine mammals can dive to depths approaching 2000 m. At these hydrostatic pressures (200 atm), some fish species show alterations in enzyme structure and function that make them pressure-tolerant. Do marine mammals also possess biochemical adaptations to withstand such pressures? In theory, biochemical alterations might occur at the control of enzymatic pathways, by impacting cell membrane fluidity changes or at a higher level, such as cellular metabolism. Studies of marine mammal tissues show evidence of all of these changes, but the results are not consistent across species or diving depth. This review discusses whether the elevated body temperature of marine mammals imparts pressure tolerance at the biochemical level, whether there are cell membrane structural differences in marine mammals and whether whole, living cells from marine mammals alter their metabolism when pressure stressed. We conclude that temperature alone is probably not protective against pressure and that cell membrane composition data are not conclusive. Whole cell studies suggest that marine mammals either respond positively to pressure or are not impacted by pressure. However, the range of tissue types and enzyme systems that have been studied is extremely limited and needs to be expanded before more general conclusions about how these mammals tolerate elevated pressures on a biochemical level can be drawn.

  3. The Stereochemistry of Biochemical Molecules: A Subject to Revisit (United States)

    Centelles, Josep J.; Imperial, Santiago


    Although Fischer's convention for stereoisomers is useful for simple molecules, the stereochemistry of complex biochemical molecules is often poorly indicated in textbooks. This article reports on errors in stereochemistry of complex hydrosoluble vitamin B12 molecule. Twenty-five popular biochemistry textbooks were examined for their treatment of…

  4. [Experiments using rats on Kosmos biosatellites: morphologic and biochemical studies]. (United States)

    Il'in, E A; Kaplanskiĭ, A S; Savina, E A


    Results of morphological and biochemical investigations of rats flown on Cosmos biosatellites are discussed. It is emphasized that most changes occurring during exposure to microgravity are directly or indirectly related to lower musculoskeletal loads which in turn produce deconditioning of different physiological systems and organism as a whole. It is concluded that this deconditioning is associated with both metabolic and structural changes.

  5. Biochemical Parameters of Orienteers Competing in a Long Distance Race. (United States)

    Mikan, Vladimir; And Others


    Measured important biochemical parameters in a group of orienteers two hours before beginning and immediately after an orienteering marathon. Found levels of dehydration. Suggests a drinking regimen which is designed for orienteering races. Concludes that no runner having kidney or liver abnormalities or changes in the urine should be allowed to…

  6. Model Based Monitoring and Control of Chemical and Biochemical Processes

    DEFF Research Database (Denmark)

    Huusom, Jakob Kjøbsted

    This presentation will give an overview of the work performed at the department of Chemical and Biochemical Engineering related to process control. A research vision is formulated and related to a number of active projects at the department. In more detail a project describing model estimation...

  7. Evaluation of biological and biochemical quality of whey protein. (United States)

    Haraguchi, Fabiano Kenji; Pedrosa, Maria Lucia; Paula, Heberth de; Santos, Rinaldo Cardoso dos; Silva, Marcelo Eustáquio


    Nutritional and biochemical properties of noncommercial whey protein have been described since 1950. However, comparisons between commercial whey protein for human consumption and casein are rarely found. The aim of this study was to compare biological quality of a commercial whey protein with casein and its effect on biochemical parameters of rats. Thirty-two weanling Fisher rats were divided into three groups and given the following diets: casein group, standard diet (AOAC); whey protein group, modified AOAC diet with whey protein instead of casein; and casein:whey group, modified AOAC diet with 70%:30% casein:whey. A protein-free group was used for determination of endogenous nitrogen losses. Net protein ratio, protein efficiency ratio, and true digestibility were determined, and blood was collected for biochemical analysis. When compared with casein, whey protein showed significant differences for all biological parameters evaluated, as well as for albumin, total protein, total cholesterol, and glucose concentrations. Replacing 30% of casein with whey protein did not affect these parameters. A positive relation among whey protein, high-density lipoprotein-cholesterol, and paraoxonase activity was found. Hepatic or renal dysfunctions were not observed. In conclusion, in comparison with casein, commercial whey protein had higher values of biological parameters, and biochemical evaluation revealed it improved glycemic homeostasis, lipid status, and paraoxonase activity in rats.

  8. Development of a new first-aid biochemical detector (United States)

    Hu, Jingfei; Liao, Haiyang; Su, Shilin; Ding, Hao; Liu, Suquan


    The traditional biochemical detector exhibits poor adaptability, inconvenient carrying and slow detection, which can't meet the needs of first-aid under field condition like natural or man-made disasters etc. Therefore a scheme of first-aid biochemical detector based on MOMES Micro Spectrometer, UV LED and Photodiode was proposed. An optical detection structure combined continuous spectrum sweep with fixed wavelength measurement was designed, which adopted mobile detection optical path consisting of Micro Spectrometer and Halogen Lamp to detect Chloride (Cl-), Creatinine (Cre), Glucose (Glu), Hemoglobin (Hb). The UV LED and Photodiode were designed to detect Potassium (K-), Carbon dioxide (CO2), Sodium (Na+). According to the field diagnosis and treatment requirements, we designed the embedded control hardware circuit and software system, the prototype of first-aid biochemical detector was developed and the clinical trials were conducted. Experimental results show that the sample's absorbance repeatability is less than 2%, the max coefficient of variation (CV) in the batch repeatability test of all 7 biochemical parameters in blood samples is 4.68%, less than the clinical requirements 10%, the correlation coefficient (R2) in the clinical contrast test with AU5800 is almost greater than 0.97. To sum up, the prototype meets the requirements of clinical application.

  9. Integrating Carbon Nanotubes into Microfluidic Chip for Separating Biochemical Compounds

    DEFF Research Database (Denmark)

    Chen, Miaoxiang Max; Mogensen, Klaus Bo; Bøggild, Peter


    We present a new type of device to separate biochemical compounds wherein carbon nanotubes (CNTs) are integrated as chromatographic stationary phase. The CNTs were directly grown on the bottom of microfluidic channels on Si/SiO2 substrates by chemical vapor deposition (CVD). Acetylene was used...

  10. Physicochemical and biochemical characterization of biosurfactants released by Lactobacillus strains

    NARCIS (Netherlands)

    Velraeds, MMC; vanderMei, HC; Reid, G; Busscher, HJ


    Biosurfactants from Lactobacillus casei subsp. rhamnosus 36 and ATCC 7469, Lactobacillus fermentum B54 and Lactobacillus acidophilus RC14 were isolated from bacteria in their mid-exponential (4-5 h) and stationary growth phases (18 h) and physicochemical and biochemical properties of the freeze-drie

  11. Metstoich--Teaching Quantitative Metabolism and Energetics in Biochemical Engineering (United States)

    Wong, Kelvin W. W.; Barford, John P.


    Metstoich, a metabolic calculator developed for teaching, can provide a novel way to teach quantitative metabolism to biochemical engineering students. It can also introduce biochemistry/life science students to the quantitative aspects of life science subjects they have studied. Metstoich links traditional biochemistry-based metabolic approaches…

  12. Photography by Cameras Integrated in Smartphones as a Tool for Analytical Chemistry Represented by an Butyrylcholinesterase Activity Assay. (United States)

    Pohanka, Miroslav


    Smartphones are popular devices frequently equipped with sensitive sensors and great computational ability. Despite the widespread availability of smartphones, practical uses in analytical chemistry are limited, though some papers have proposed promising applications. In the present paper, a smartphone is used as a tool for the determination of cholinesterasemia i.e., the determination of a biochemical marker butyrylcholinesterase (BChE). The work should demonstrate suitability of a smartphone-integrated camera for analytical purposes. Paper strips soaked with indoxylacetate were used for the determination of BChE activity, while the standard Ellman's assay was used as a reference measurement. In the smartphone-based assay, BChE converted indoxylacetate to indigo blue and coloration was photographed using the phone's integrated camera. A RGB color model was analyzed and color values for the individual color channels were determined. The assay was verified using plasma samples and samples containing pure BChE, and validated using Ellmans's assay. The smartphone assay was proved to be reliable and applicable for routine diagnoses where BChE serves as a marker (liver function tests; some poisonings, etc.). It can be concluded that the assay is expected to be of practical applicability because of the results' relevance.

  13. Thermodynamically consistent Bayesian analysis of closed biochemical reaction systems

    Directory of Open Access Journals (Sweden)

    Goutsias John


    Full Text Available Abstract Background Estimating the rate constants of a biochemical reaction system with known stoichiometry from noisy time series measurements of molecular concentrations is an important step for building predictive models of cellular function. Inference techniques currently available in the literature may produce rate constant values that defy necessary constraints imposed by the fundamental laws of thermodynamics. As a result, these techniques may lead to biochemical reaction systems whose concentration dynamics could not possibly occur in nature. Therefore, development of a thermodynamically consistent approach for estimating the rate constants of a biochemical reaction system is highly desirable. Results We introduce a Bayesian analysis approach for computing thermodynamically consistent estimates of the rate constants of a closed biochemical reaction system with known stoichiometry given experimental data. Our method employs an appropriately designed prior probability density function that effectively integrates fundamental biophysical and thermodynamic knowledge into the inference problem. Moreover, it takes into account experimental strategies for collecting informative observations of molecular concentrations through perturbations. The proposed method employs a maximization-expectation-maximization algorithm that provides thermodynamically feasible estimates of the rate constant values and computes appropriate measures of estimation accuracy. We demonstrate various aspects of the proposed method on synthetic data obtained by simulating a subset of a well-known model of the EGF/ERK signaling pathway, and examine its robustness under conditions that violate key assumptions. Software, coded in MATLAB®, which implements all Bayesian analysis techniques discussed in this paper, is available free of charge at Conclusions Our approach provides an attractive statistical methodology for

  14. 尿道及其他显色培养基快速分离鉴定常见泌尿道感染致病菌的临床应用%Urethra and other chromogenic medium fast isolation and identification of common urinary tract infection pathogens in clinical application

    Institute of Scientific and Technical Information of China (English)

    翁杏华; 徐涛; 陈瑞湖; 关尚; 陈玉莲


    Objective Urethra and other chromogenic medium fast identification of common urinary tract pathogens isolated in order to avoid errors caused by mixed bacteria identification and susceptibility results. Methods Accordance with the "national clinical laboratory Procedures"3rd edition urine culture methods used;specimens inoculated directly on blood agar and urethra chromogenic media for culture and identification plates,etc. with the French bioMérieux atbexpression automatic identification system for identification of bacteria and / or verification. Results Major urinary tract infection escherichia coli bacteria 49. 1%,klebsiella pneumoniae 8. 9%,proteus 10. 8%. Conclu-sion Proven compliance rate of 90% to 100%. Urethral chromogenic medium can quickly and easily isolated and identified common urina-ry tract pathogens,saving the cost of reagents,suitable for all basic hospital application.%目的:总结尿道及其他显色培养基快速分离鉴定常见泌尿道致病菌以避免混合菌引起错误的鉴定及药敏结果。方法:按照《全国临床检验操作规程》第3版尿培养的方法;标本直接接种在血平板及尿道显色培养基等平板上进行培养鉴定,用法国生物梅里埃公司ATBExpression自动细菌鉴定仪进行鉴定和/或验证。结果:主要泌尿道感染菌49.1%为大肠埃希菌,8.9%为肺炎克雷伯菌,10.8%为变形杆菌。结论:经验证明总符合率为96%,尿道显色培养基能快速、方便分离鉴定常见泌尿道致病菌,节约试剂成本,适合各基层医院推广应用。

  15. Consumption of thermally oxidized palm oil diets alters biochemical indices in rats

    Directory of Open Access Journals (Sweden)

    Ayodeji Osmund Falade


    Full Text Available Palm oil is thermally oxidized to increase its palatability and this has been a usual practice in most homes. This study sought to assess the biochemical responses of rats to thermally oxidized palm oil diets. Therefore, Wistar strain albino rats (Rattus norveigicus were fed with fresh palm oil (control and thermally oxidized palm oil (test groups diets and water ad libitum for 30 days. Then, the malondialdehyde (MDA contents and total protein of the plasma and liver were determined. Subsequently, the plasma liver function markers [alanine transaminase (ALT, aspartate transaminase (AST, alkaline phosphatase (ALP, albumin (ALB and total bilirubin (TBIL ] and the lipid profile [triglyceride (TRIG, total cholesterol (T-CHOL, high density lipoprotein (HDL-CHOL and low density lipoprotein (LDL-CHOL ] were assayed. The results of the study revealed that there was a significant decrease (P < 0.05 in the plasma and liver total protein, ALB, TRIG and HDL-CHOL of the test groups when compared with the control. Conversely, there was a significant increase (P < 0.05 in the activities of ALT, AST and ALP, TBIL, T-CHOL, LDL-CHOL and plasma/liver MDA of the test groups when compared with the control. These effects were most pronounced in rats fed with 20 min-thermally oxidized palm oil diet. Hence, consumption of thermally oxidized palm oil diets had deleterious effects on biochemical indices in rats. Therefore, cooking with and/or consumption of palm oil subjected to heat treatment for several long periods of time should be discouraged in our homes as this might have deleterious effects on human health.

  16. Biochemical, cellular, and biophysical characterization of a potent inhibitor of mutant isocitrate dehydrogenase IDH1. (United States)

    Davis, Mindy I; Gross, Stefan; Shen, Min; Straley, Kimberly S; Pragani, Rajan; Lea, Wendy A; Popovici-Muller, Janeta; DeLaBarre, Byron; Artin, Erin; Thorne, Natasha; Auld, Douglas S; Li, Zhuyin; Dang, Lenny; Boxer, Matthew B; Simeonov, Anton


    Two mutant forms (R132H and R132C) of isocitrate dehydrogenase 1 (IDH1) have been associated with a number of cancers including glioblastoma and acute myeloid leukemia. These mutations confer a neomorphic activity of 2-hydroxyglutarate (2-HG) production, and 2-HG has previously been implicated as an oncometabolite. Inhibitors of mutant IDH1 can potentially be used to treat these diseases. In this study, we investigated the mechanism of action of a newly discovered inhibitor, ML309, using biochemical, cellular, and biophysical approaches. Substrate binding and product inhibition studies helped to further elucidate the IDH1 R132H catalytic cycle. This rapidly equilibrating inhibitor is active in both biochemical and cellular assays. The (+) isomer is active (IC50 = 68 nm), whereas the (-) isomer is over 400-fold less active (IC50 = 29 μm) for IDH1 R132H inhibition. IDH1 R132C was similarly inhibited by (+)-ML309. WT IDH1 was largely unaffected by (+)-ML309 (IC50 >36 μm). Kinetic analyses combined with microscale thermophoresis and surface plasmon resonance indicate that this reversible inhibitor binds to IDH1 R132H competitively with respect to α-ketoglutarate and uncompetitively with respect to NADPH. A reaction scheme for IDH1 R132H inhibition by ML309 is proposed in which ML309 binds to IDH1 R132H after formation of the IDH1 R132H NADPH complex. ML309 was also able to inhibit 2-HG production in a glioblastoma cell line (IC50 = 250 nm) and had minimal cytotoxicity. In the presence of racemic ML309, 2-HG levels drop rapidly. This drop was sustained until 48 h, at which point the compound was washed out and 2-HG levels recovered.

  17. Biochemical and biophysical investigations of the interaction between human glucokinase and pro-apoptotic BAD. (United States)

    Rexford, Alix; Zorio, Diego A R; Miller, Brian G


    The glycolytic enzyme glucokinase (GCK) and the pro-apoptotic protein BAD reportedly reside within a five-membered complex that localizes to the mitochondria of mammalian hepatocytes and pancreatic β-cells. Photochemical crosslinking studies using a synthetic analog of BAD's BH3 domain and in vitro transcription/translation experiments support a direct interaction between BAD and GCK. To investigate the biochemical and biophysical consequences of the BAD:GCK interaction, we developed a method for the production of recombinant human BAD. Consistent with published reports, recombinant BAD displays high affinity for Bcl-xL (KD = 7 nM), and phosphorylation of BAD at S118, within the BH3 domain, abolishes this interaction. Unexpectedly, we do not detect association of recombinant, full-length BAD with recombinant human pancreatic GCK over a range of protein concentrations using various biochemical methods including size-exclusion chromatography, chemical cross-linking, analytical ultracentrifugation, and isothermal titration calorimetry. Furthermore, fluorescence polarization assays and isothermal titration calorimetry detect no direct interaction between GCK and BAD BH3 peptides. Kinetic characterization of GCK in the presence of high concentrations of recombinant BAD show modest (BAD BH3 peptides. These results raise questions as to the mechanism of action of stapled peptide analogs modeled after the BAD BH3 domain, which reportedly enhance the Vmax value of GCK and stimulate insulin release in BAD-deficient islets. Based on our results, we postulate that the BAD:GCK interaction, and any resultant regulatory effect(s) upon GCK activity, requires the participation of additional members of the mitochondrial complex.

  18. Biochemical studies on Francisella tularensis RelA in (p)ppGpp biosynthesis. (United States)

    Wilkinson, Rachael C; Batten, Laura E; Wells, Neil J; Oyston, Petra C F; Roach, Peter L


    The bacterial stringent response is induced by nutrient deprivation and is mediated by enzymes of the RSH (RelA/SpoT homologue; RelA, (p)ppGpp synthetase I; SpoT, (p)ppGpp synthetase II) superfamily that control concentrations of the 'alarmones' (p)ppGpp (guanosine penta- or tetra-phosphate). This regulatory pathway is present in the vast majority of pathogens and has been proposed as a potential anti-bacterial target. Current understanding of RelA-mediated responses is based on biochemical studies using Escherichia coli as a model. In comparison, the Francisella tularensis RelA sequence contains a truncated regulatory C-terminal region and an unusual synthetase motif (EXSD). Biochemical analysis of F. tularensis RelA showed the similarities and differences of this enzyme compared with the model RelA from Escherichia coli. Purification of the enzyme yielded a stable dimer capable of reaching concentrations of 10 mg/ml. In contrast with other enzymes from the RelA/SpoT homologue superfamily, activity assays with F. tularensis RelA demonstrate a high degree of specificity for GTP as a pyrophosphate acceptor, with no measurable turnover for GDP. Steady state kinetic analysis of F. tularensis RelA gave saturation activity curves that best fitted a sigmoidal function. This kinetic profile can result from allosteric regulation and further measurements with potential allosteric regulators demonstrated activation by ppGpp (5',3'-dibisphosphate guanosine) with an EC50 of 60±1.9 μM. Activation of F. tularensis RelA by stalled ribosomal complexes formed with ribosomes purified from E. coli MRE600 was observed, but interestingly, significantly weaker activation with ribosomes isolated from Francisella philomiragia.


    Directory of Open Access Journals (Sweden)

    Yu. P. Sedlukho


    Full Text Available The paper considers problems and features of biochemical removal of hydrogen sulfide from ground water. The analysis of existing methods for purification of ground water from hydrogen sulfide has been given in the paper. The paper has established shortcomings of physical and chemical purification of ground water. While using aeration methods for removal of hydrogen sulfide formation of colloidal sulfur that gives muddiness and opalescence to water occurs due to partial chemical air oxidation. In addition to this violation of sulfide-carbonate equilibrium taking place in the process of aeration due to desorption of H2S and CO2, often leads to clogging of degasifier nozzles with formed CaCO3 that causes serious operational problems. Chemical methods require relatively large flow of complex reagent facilities, storage facilities and transportation costs.In terms of hydrogen sulfide ground water purification the greatest interest is given to the biochemical method. Factors deterring widespread application of the biochemical method is its insufficient previous investigation and necessity to execute special research in order to determine optimal process parameters while purifying groundwater of a particular water supply source. Biochemical methods for oxidation of sulfur compounds are based on natural biological processes that ensure natural sulfur cycle. S. Vinogradsky has established a two-stage mechanism for oxidation of hydrogen sulfide with sulfur bacteria (Beggiatoa. The first stage presupposes oxidation of hydrogen sulphide to elemental sulfur which is accumulating in the cytoplasm in the form of globules. During the second stage sulfur bacteria begin to oxidize intracellular sulfur to sulfuric acid due to shortage of hydrogen sulfide.The paper provides the results of technological tests of large-scale pilot plants for biochemical purification of groundwater from hydrogen sulfide in semi-industrial conditions. Dependences of water quality

  20. Accurate atom-mapping computation for biochemical reactions. (United States)

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


    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.

  1. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.


    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  2. Controlling variation in the comet assay

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    Andrew Richard Collins


    Full Text Available Variability of the comet assay is a serious issue, whether it occurs from experiment to experiment in the same laboratory, or between different laboratories analysing identical samples. Do we have to live with high variability, just because the comet assay is a biological assay rather than analytical chemistry? Numerous attempts have been made to limit variability by standardising the assay protocol, and the critical steps in the assay have been identified; agarose concentration, duration of alkaline incubation, and electrophoresis conditions (time, temperature and voltage gradient are particularly important. Even when these are controlled, variation seems to be inevitable. It is helpful to include in experiments reference standards, i.e. cells with a known amount of specific damage to the DNA. They can be aliquots frozen from a single large batch of cells, either untreated (negative controls or treated with, for example, H2O2 or X-rays to induce strand breaks (positive control for the basic assay, or photosensitiser plus light to oxidise guanine (positive control for Fpg- or OGG1-sensitive sites. Reference standards are especially valuable when performing a series of experiments over a long period - for example, analysing samples of white blood cells from a large human biomonitoring trial - to check that the assay is performing consistently, and to identify anomalous results necessitating a repeat experiment. The reference values of tail intensity can also be used to iron out small variations occurring from day to day. We present examples of the use of reference standards in human trials, both within one laboratory and between different laboratories, and describe procedures that can be used to control variation.

  3. The Comet Assay: Tails of the (Unexpected. Use of the comet assay in pharmaceutical development.

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    Bas-jan Van Der Leede


    Full Text Available In genotoxicity testing of pharmaceuticals the rodent alkaline comet assay is being increasingly used as a second in vivo assay in addition to the in vivo micronucleus assay to mitigate in vitro positive results as recommended by regulatory guidance. In this presentation we want to give insight into the circumstances in vivo comet assay is deployed in a Genetic Toxicology Department of a pharmaceutical company. As the in vivo comet assay is a salvage assay, it means that some events have occurred in an in vitro assay and that the compound (or metabolite responsible for this signal is potentially deselected for further development. More than often the decision to perform an in vivo comet assay is at a very early stage in development and the first time that the compound will be tested in vivo at high/toxic dose levels. As almost no toxicokinetic data and tissue distribution data are available a careful design with maximizes the chances for successful mitigation is necessary. Decisions on acute or repeated dosing need to be made and arrangements for combining the in vivo comet assay with the in vivo micronucleus assay are to be considered. Often synthesis methods need to be scaled up fast to provide the required amount of compound and information on suitable formulations needs to be in place. As exposure data is crucial for interpretation of results, analytical methods need to be brought in place rapidly. An experienced multi skilled and communicative team needs to be available to deploy successfully this kind of assays at an early stage of development. We will present a few scenarios on study conduct and demonstrate how this assay can make a difference for the further development of a new drug.


    Directory of Open Access Journals (Sweden)

    P. D. Demeshko


    Full Text Available Purpose. To evaluate influence of clinical, biochemical and histological factors to biochemical relapse free survival (BRFS following salvage radiotherapy (RT in men with biochemical recurrence after radical prostatectomy.Material and methods. 77 patients with newly diagnosed biochemical recurrence (BR after RPE were included into retrospective study. All of them underwent local salvage RT. Сlinical variables (age, serum prostate-specific antigen [PSA] level and PSA kinetics, time RPE-BR, Gleason grade, stage after RPE and clinical findings were evaluated using Cox proportional hazards regression analysis.Results. The median, 1- and 3-year BRFS were 19,9 months, 63,8 ± 6,5 % and 24,7 ± 8,5 % respectively. Significant variables in the multivariable model were age, PSA level before RT, prostatectomy T3b stage, PSA doubling time and positive digital rectal examination findings (p < 0,05. Several clinical parameters help predict the outcomes of men with PSA elevation after radical prostatectomy. These data may be useful in counseling men regarding the timing of administration of adjuvant therapies.

  5. Using the CPTAC Assay Portal to identify and implement highly characterized targeted proteomics assays (United States)

    Whiteaker, Jeffrey R; Halusa, Goran N; Hoofnagle, Andrew N; Sharma, Vagisha; MacLean, Brendan; Yan, Ping; Wrobel, John A; Kennedy, Jacob; Mani, DR; Zimmerman, Lisa J; Meyer, Matthew R.; Mesri, Mehdi; Abbatiello, Susan E; Boja, Emily; Carr, Steven A.; Chan, Daniel W.; Chen, Xian; Chen, Jing; Davies, Sherri R; Ellis, Matthew J. C.; Fenyö, David; Hiltke, Tara; Ketchum, Karen A.; Kinsinger, Chris; Kuhn, Eric; Liebler, Daniel C.; Lin, De; Liu, Tao; Loss, Michael; MacCoss, Michael J; Qian, Wei-Jun; Rivers, Robert; Rodland, Karin D.; Ruggles, Kelly V; Scott, Mitchell G; Smith, Richard D.; Thomas, Stefani; Townsend, R. Reid; Whiteley, Gordon; Wu, Chaochao; Zhang, Hui; Zhang, Zhen; Rodriguez, Henry; Paulovich, Amanda G


    Summary The Clinical Proteomic Tumor Analysis Consortium (CPTAC) of the National Cancer Institute (NCI) has launched an Assay Portal ( to serve as an open-source repository of well-characterized targeted proteomic assays. The portal is designed to curate and disseminate highly characterized, targeted mass spectrometry (MS)-based assays by providing detailed assay performance characterization data, standard operating procedures, and access to reagents. Assay content is accessed via the portal through queries to find assays targeting proteins associated with specific cellular pathways, protein complexes, or specific chromosomal regions. The position of the peptide analytes for which there are available assays are mapped relative to other features of interest in the protein, such as sequence domains, isoforms, single nucleotide polymorphisms, and post-translational modifications. The overarching goals are to enable robust quantification of all human proteins and to standardize the quantification of targeted MS-based assays to ultimately enable harmonization of results over time and across laboratories. PMID:26867747

  6. Using the CPTAC Assay Portal to identify and implement highly characterized targeted proteomics assays

    Energy Technology Data Exchange (ETDEWEB)

    Whiteaker, Jeffrey R.; Halusa, Goran; Hoofnagle, Andrew N.; Sharma, Vagisha; MacLean, Brendan; Yan, Ping; Wrobel, John; Kennedy, Jacob; Mani, DR; Zimmerman, Lisa J.; Meyer, Matthew R.; Mesri, Mehdi; Boja, Emily; Carr, Steven A.; Chan, Daniel W.; Chen, Xian; Chen, Jing; Davies, Sherri; Ellis, Matthew; Fenyo, David; Hiltket, Tara; Ketchum, Karen; Kinsinger, Christopher; Kuhn, Eric; Liebler, Daniel; Liu, Tao; Loss, Michael; MacCoss, Michael; Qian, Weijun; Rivers, Robert; Rodland, Karin D.; Ruggles, Kelly; Scott, Mitchell; Smith, Richard D.; Thomas, Stefani N.; Townsend, Reid; Whiteley, Gordon; Wu, Chaochao; Zhang, Hui; Zhang, Zhen; Rodriguez, Henry; Paulovich, Amanda G.


    The Clinical Proteomic Tumor Analysis Consortium (CPTAC) of the National Cancer Institute (NCI) has launched an Assay Portal ( to serve as an open-source repository of well-characterized targeted proteomic assays. The portal is designed to curate and disseminate highly characterized, targeted mass spectrometry (MS)-based assays by providing detailed assay performance characterization data, standard operating procedures, and access to reagents. Assay content is accessed via the portal through queries to find assays targeting proteins associated with specific cellular pathways, protein complexes, or specific chromosomal regions. The position of the peptide analytes for which there are available assays are mapped relative to other features of interest in the protein, such as sequence domains, isoforms, single nucleotide polymorphisms, and post-translational modifications. The overarching goals are to enable robust quantification of all human proteins and to standardize the quantification of targeted MS-based assays to ultimately enable harmonization of results over time and across laboratories.


    Directory of Open Access Journals (Sweden)

    V. A. Chernyaev


    Full Text Available Purpose. To reveal prognostic factors of PSA-failure following radical prostatectomy in patients with localized and locally-advanced prostate cancer.Materials and methods. Medical data of 386 consecutive patients with localized and locally-advanced prostate cancer who underwent radical prostatectomy from 1997 to 2011 were analyzed. Median age was 61.0 years. Median PSA before surgery – 10.3 ng/ml. Plasma levels of VEGF, VEGFR2, VEGFR3, TGF-β1, CD105, IL-6 were measured using Enzyme Linked-Immuno-Sorbent Assay (ELISA before radical prostatectomy in 77 patients. Postoperatively the tumours were categorized as pT2 in 288 (59.1 %, pT3 – in 144 (37.3 %, pT4 – in 14 (3.6; pN+ – in 34 (8.8 % cases. Gleason score < 7 was present in 254 (65.8 %,  7 – in 132 (34.2 % specimens. Perineural invasion was identified in 188 (48.7 %, angiolymphatic invasion – in 126 (32.6 cases.Results. Biochemical recurrence occurred in 64 (16.6 % out of 386 patients at a median follow-up of 30.5 (12−164 months. Independent predictors of biochemical recurrence were PSA (HR 0.161 (95% CI:0.058−0.449; р = 0.001, Gleason sum in surgical specimens (HR 0.496 (95 % CI:0.268−0.917; p = 0.025, pN (HR 0.415 (95 % CI:0.181−0.955; p = 0.039. The patients were divided into 3 prognostic groups: good (0 factor, intermediate (1 factor, poor (2 factors and very poor (3 factors (AUC – 0.720 (95% CI: 0.656−0.784. High preoperative levels VEGF ( 67 pg/ml (р = 0.005, VEGFR2 ( 3149 pg/ml (р = 0.036, VEGFR3 ( 2268 pg/ml (р = 0.001, TGF-β1 ( 14473 pg/ml (р = 0.052 were identified as unfavorable prognostic factors for survival without PSA-failure. Conclusion. Independent prognostic factors of biochemical recurrence after prostatectomy were PSA, Gleason sum and pN. Joint effect of the factors allows to predict PSA-relapse with accuracy 0.720. Preoperative serum levels VEGF, VEGFR2, VEGFR3, TGF-β1 potentially are perspective markers for PSA-failure after


    Directory of Open Access Journals (Sweden)

    V. A. Chernyaev


    Full Text Available Purpose. To reveal prognostic factors of PSA-failure following radical prostatectomy in patients with localized and locally-advanced prostate cancer.Materials and methods. Medical data of 386 consecutive patients with localized and locally-advanced prostate cancer who underwent radical prostatectomy from 1997 to 2011 were analyzed. Median age was 61.0 years. Median PSA before surgery – 10.3 ng/ml. Plasma levels of VEGF, VEGFR2, VEGFR3, TGF-β1, CD105, IL-6 were measured using Enzyme Linked-Immuno-Sorbent Assay (ELISA before radical prostatectomy in 77 patients. Postoperatively the tumours were categorized as pT2 in 288 (59.1 %, pT3 – in 144 (37.3 %, pT4 – in 14 (3.6; pN+ – in 34 (8.8 % cases. Gleason score < 7 was present in 254 (65.8 %,  7 – in 132 (34.2 % specimens. Perineural invasion was identified in 188 (48.7 %, angiolymphatic invasion – in 126 (32.6 cases.Results. Biochemical recurrence occurred in 64 (16.6 % out of 386 patients at a median follow-up of 30.5 (12−164 months. Independent predictors of biochemical recurrence were PSA (HR 0.161 (95% CI:0.058−0.449; р = 0.001, Gleason sum in surgical specimens (HR 0.496 (95 % CI:0.268−0.917; p = 0.025, pN (HR 0.415 (95 % CI:0.181−0.955; p = 0.039. The patients were divided into 3 prognostic groups: good (0 factor, intermediate (1 factor, poor (2 factors and very poor (3 factors (AUC – 0.720 (95% CI: 0.656−0.784. High preoperative levels VEGF ( 67 pg/ml (р = 0.005, VEGFR2 ( 3149 pg/ml (р = 0.036, VEGFR3 ( 2268 pg/ml (р = 0.001, TGF-β1 ( 14473 pg/ml (р = 0.052 were identified as unfavorable prognostic factors for survival without PSA-failure. Conclusion. Independent prognostic factors of biochemical recurrence after prostatectomy were PSA, Gleason sum and pN. Joint effect of the factors allows to predict PSA-relapse with accuracy 0.720. Preoperative serum levels VEGF, VEGFR2, VEGFR3, TGF-β1 potentially are perspective markers for PSA-failure after

  9. Research highlights: digital assays on chip. (United States)

    Kim, Donghyuk; Wei, Qingshan; Kong, Janay Elise; Ozcan, Aydogan; Di Carlo, Dino


    The ability to break up a volume of fluid into smaller pieces that are confined or separated to prevent molecular communication/transport is a key capability intrinsic to microfluidic systems. This capability has been used to develop or implement digital versions of traditional molecular analysis assays, including digital PCR and digital immunoassays/ELISA. In these digital versions, the concentration of the target analyte is in a range such that, when sampled into smaller fluid volumes, either a single molecule or no molecule may be present. Subsequent amplification is sensitive enough to obtain a digital readout of the presence of these target molecules. Advantages of such approaches that are claimed include quantification without calibration and robustness to variations in reaction conditions or times because the digital readout is less sensitive to absolute signal intensity levels. Weaknesses of digital approaches include a lower dynamic range of concentrations over which the assay is sensitive, which depends on the total volume that can be analyzed. We highlight recent efforts to expand the dynamic range of digital assays based on exploiting reaction/diffusion phenomena. A side-by-side study that evaluates the strengths of digital assays reveals that the majority of these claims are supported, with specific caveats. Finally, we highlight approaches to apply digital assays to analyze new types of reactions, including the active transport of protons across membranes by ATPases at the single protein level - perhaps opening up new biophysical understanding and screening opportunities, similar to widely deployed single-molecule ion channel analysis.

  10. Comet assay on mice testicular cells

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    Anoop Kumar Sharma


    Full Text Available Heritable mutations may result in a variety of adverse outcomes including genetic disease in the offspring. In recent years the focus on germ cell mutagenicity has increased and the “Globally Harmonized System of Classification and Labelling of Chemicals (GHS” has published classification criteria for germ cell mutagens (Speit et al., 2009. The in vivo Comet assay is considered a useful tool for investigating germ cell genotoxicity. In the present study DNA strand breaks in testicular cells of mice were investigated. Different classes of chemicals were tested in order to evaluate the sensitivity of the comet assay in testicular cells. The chemicals included environmentally relevant substances such as Bisphenol A, PFOS and Tetrabrombisphenol A. Statistical power calculations will be presented to aid in the design of future Comet assay studies on testicular cells. Power curves were provided with different fold changes in % tail DNA, different number of cells scored and different number of gels (Hansen et al., 2014. An example is shown in Figure 1. A high throughput version of the Comet assay was used. Samples were scored with a fully automatic comet assay scoring system that provided faster scoring of randomly selected cells.

  11. Biochemical and molecular characterization of the gentisate transporter GenK in Corynebacterium glutamicum.

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    Ying Xu

    Full Text Available BACKGROUND: Gentisate (2,5-dihydroxybenzoate is a key ring-cleavage substrate involved in various aromatic compounds degradation. Corynebacterium glutamicum ATCC13032 is capable of growing on gentisate and genK was proposed to encode a transporter involved in this utilization by its disruption in the restriction-deficient mutant RES167. Its biochemical characterization by uptake assay using [(14C]-labeled gentisate has not been previously reported. METHODOLOGY/PRINCIPAL FINDINGS: In this study, biochemical characterization of GenK by uptake assays with [(14C]-labeled substrates demonstrated that it specifically transported gentisate into the cells with V(max and K(m of 3.06 ± 0.16 nmol/min/mg of dry weight and 10.71 ± 0.11 µM respectively, and no activity was detected for either benzoate or 3-hydoxybenzoate. When GenK was absent in strain RES167 ΔgenK, it retained 85% of its original transport activity at pH 6.5 compared to that of strain RES167. However, it lost 79% and 88% activity at pH 7.5 and 8.0, respectively. A number of competing substrates, including 3-hydroxybenzoate, benzoate, protocatechuate and catechol, significantly inhibited gentisate uptake by more than 40%. Through site-directed mutagenesis, eight amino acid residues of GenK, Asp-54, Asp-57 and Arg-386 in the hydrophobic transmembrane regions and Arg-103, Trp-309, Asp-312, Arg-313 and Ile-317 in the hydrophilic cytoplasmic loops were shown to be important for gentisate transport. When conserved residues Asp-54 and Asp-57 respectively were changed to glutamate, both mutants retained approximately 50% activity and were able to partially complement the ability of strain RES167 ΔgenK to grow on gentisate. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that GenK is an active gentisate transporter in Corynebacterium glutamicum ATCC13032. The GenK-mediated gentisate transport was also shown to be a limiting step for the gentisate utilization by this strain. This enhances our

  12. Biochemical characterization of a recombinant Japanese encephalitis virus RNA-dependent RNA polymerase

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    Kim Chan-Mi


    Full Text Available Abstract Background Japanese encephalitis virus (JEV NS5 is a viral nonstructural protein that carries both methyltransferase and RNA-dependent RNA polymerase (RdRp domains. It is a key component of the viral RNA replicase complex that presumably includes other viral nonstructural and cellular proteins. The biochemical properties of JEV NS5 have not been characterized due to the lack of a robust in vitro RdRp assay system, and the molecular mechanisms for the initiation of RNA synthesis by JEV NS5 remain to be elucidated. Results To characterize the biochemical properties of JEV RdRp, we expressed in Escherichia coli and purified an enzymatically active full-length recombinant JEV NS5 protein with a hexahistidine tag at the N-terminus. The purified NS5 protein, but not the mutant NS5 protein with an Ala substitution at the first Asp of the RdRp-conserved GDD motif, exhibited template- and primer-dependent RNA synthesis activity using a poly(A RNA template. The NS5 protein was able to use both plus- and minus-strand 3'-untranslated regions of the JEV genome as templates in the absence of a primer, with the latter RNA being a better template. Analysis of the RNA synthesis initiation site using the 3'-end 83 nucleotides of the JEV genome as a minimal RNA template revealed that the NS5 protein specifically initiates RNA synthesis from an internal site, U81, at the two nucleotides upstream of the 3'-end of the template. Conclusion As a first step toward the understanding of the molecular mechanisms for JEV RNA replication and ultimately for the in vitro reconstitution of viral RNA replicase complex, we for the first time established an in vitro JEV RdRp assay system with a functional full-length recombinant JEV NS5 protein and characterized the mechanisms of RNA synthesis from nonviral and viral RNA templates. The full-length recombinant JEV NS5 will be useful for the elucidation of the structure-function relationship of this enzyme and for the

  13. Fungicide resistance assays for fungal plant pathogens. (United States)

    Secor, Gary A; Rivera, Viviana V


    Fungicide resistance assays are useful to determine if a fungal pathogen has developed resistance to a fungicide used to manage the disease it causes. Laboratory assays are used to determine loss of sensitivity, or resistance, to a fungicide and can explain fungicide failures and for developing successful fungicide recommendations in the field. Laboratory assays for fungicide resistance are conducted by measuring reductions in growth or spore germination of fungi in the presence of fungicide, or by molecular procedures. This chapter describes two techniques for measuring fungicide resistance, using the sugarbeet leaf spot fungus Cercospora beticola as a model for the protocol. Two procedures are described for fungicides from two different classes; growth reduction for triazole (sterol demethylation inhibitor; DMI) fungicides, and inhibition of spore germination for quinone outside inhibitor (QoI) fungicides.

  14. Comet assay on tetraploid yeast cells

    DEFF Research Database (Denmark)

    Rank, Jette; Syberg, Kristian; Jensen, Klara


    Tetraploid yeast cells (Saccharomyces cerevisiae) were used in the comet assay with the intention of developing a new, fast and easy assay for detecting environmental genotoxic agents without using higher organisms. Two DNA-damaging chemicals, H2O2 and acrylamide, together with wastewater from...... three municipal treatment plants were tested for their effect on the yeast-cell DNA. The main problem with using yeast in the comet assay is the necessity to degrade the cell wall. This was achieved by using Zymolase 100 T twice during the procedure, since Zymolase 20 T did not open the cell wall....... Analytical problems that arose due to the small amount of DNA in the yeast nuclei in haploid and diploid cells, which contain 13 Mbp and 26 Mbp DNA per cell, respectively, were solved by using tetraploid yeast cells (52 Mbp) instead. DNA damage was shown after exposure to H2O2 and acrylamide. The lowest dose...

  15. EDTA interference in electrochemiluminescence ACTH assay. (United States)

    Toprak, Burak; Yalcin, Hulya; Arı, Elif; Colak, Ayfer


    Background As plasma is the recommended sample type for Roche adrenocorticotropic hormone (ACTH) assay, we evaluated the effect of EDTA concentration on Cobas ACTH assay. Methods Samples containing twofold and fourfold higher concentrations of EDTA were prepared by adding plasma to empty K2EDTA tubes and by making under-filled EDTA tubes. All measurements were performed with four replicates. Results Increased EDTA concentration resulted in a significant decrease in ACTH concentration. Fifty-per cent-filled EDTA tube showed 19% decrease in ACTH concentration and 25% filled EDTA tube showed 50% decrease in ACTH concentration. Conclusion We recommend that inadequately filled EDTA specimens should be rejected when using Cobas ACTH assay.

  16. Determination of Anti-FIIαActivity in Compound Heparin Sodium Cream by a Trace Chromogenic Sub-strate Method%微量生色底物法测定复方肝素钠乳膏抗Ⅱα因子活性

    Institute of Scientific and Technical Information of China (English)

    刘艳芳; 马宏达; 颜鸣; 候明晓


    目的:建立复方肝素钠乳膏抗Ⅱα因子活性的测定方法。方法:采用微量生色底物法,按生物检定统计法中量反应平行线原理4×4法完全随机实验设计,测定复方肝素钠乳膏抗Ⅱα因子活性。结果:肝素钠抗Ⅱα因子活性在0.00504~0.021 IU· ml-1范围内线性关系良好(r=0.992),平均回收率为101.6%(RSD=2.76%, n=9)。结论:本法准确、可靠、重复性好,可有效控制复方肝素钠乳膏质量。%Objective:To establish a trace chromogenic substrate method for the determination of anti-FIIαactivity in compound heparin sodium cream .Methods: The anti-FIIαactivity in compound heparin sodium cream was determined by a trace chromogenic substrate method according to the completely random design of experiment based on the amount reaction principle of 4*4 parallel lines in the biological test statistics method .Results:The calibration curve was linear within the range of 0.005 04 IU· ml-1-0.021 IU· ml-1(r=0.992).The average recovery was 101.6% with RSD of 2.76% (n=9).Conclusion: The method is accurate, reliable and reproducible , and can be used for evaluating the quality of compound heparin sodium cream .

  17. Detection of Eperythrozoon wenyoni by PCR assay

    Institute of Scientific and Technical Information of China (English)

    Jian WANG; Yutao ZHU; Jianhua QIN; Fumei ZHANG; Yuelan ZHAO


    The objective of this research was to develop a detection method for Eperythrozoon wenyoni infection using polymerase chain reaction (PCR) assay technique. A pair of primers was designed and synthesized according to the conservative sequence 16S rRNA. The PCR assay was performed with the primers. A 985-bp fragment was amplified by using PCR. The amplified fragments with the expected size were identified by EcoR I restriction digestion. The crossing-reaction, specific-reaction and duplicate-reaction indicated that the PCR method is a specific, sensitive, fast and effective method for diagnosing E. Wenyoni infection at group level.

  18. A novel immunofluorescent assay to investigate oxidative phosphorylation deficiency in mitochondrial myopathy: understanding mechanisms and improving diagnosis. (United States)

    Rocha, Mariana C; Grady, John P; Grünewald, Anne; Vincent, Amy; Dobson, Philip F; Taylor, Robert W; Turnbull, Doug M; Rygiel, Karolina A


    Oxidative phosphorylation defects in human tissues are often challenging to quantify due to a mosaic pattern of deficiency. Biochemical assays are difficult to interpret due to the varying enzyme deficiency levels found in individual cells. Histochemical analysis allows semi-quantitative assessment of complex II and complex IV activities, but there is no validated histochemical assay to assess complex I activity which is frequently affected in mitochondrial pathology. To help improve the diagnosis of mitochondrial disease and to study the mechanisms underlying mitochondrial abnormalities in disease, we have developed a quadruple immunofluorescent technique enabling the quantification of key respiratory chain subunits of complexes I and IV, together with an indicator of mitochondrial mass and a cell membrane marker. This assay gives precise and objective quantification of protein abundance in large numbers of individual muscle fibres. By assessing muscle biopsies from subjects with a range of different mitochondrial genetic defects we have demonstrated that specific genotypes exhibit distinct biochemical signatures in muscle, providing evidence for the diagnostic use of the technique, as well as insight into the underlying molecular pathology. Stringent testing for reproducibility and sensitivity confirms the potential value of the technique for mechanistic studies of disease and in the evaluation of therapeutic approaches.

  19. Isolation and Biochemical Characterizations of Mid Gut Microbiota of Culex (Culex quinquefasciatus Mosquitoes in Some Urban Sub Urban & Rural Areas of West Bengal.

    Directory of Open Access Journals (Sweden)

    Rahul Kumar


    Full Text Available Mosquitoes, in general are medically important vectors of many diseases like Malaria, Dengue and Filariasis, which are a great challenge for public health in many countries. All animals and plants establish symbiotic relationship with microbes. Mosquitoes can be considered as an holobiont units in which the host (mosquito and its microbiota are involved in complex reciprocal multipartite interaction such as host reproduction and survival, protection against natural enemies. This naturally acquired microbial flora can modulate the mosquitos’ vectorial capacity by inhibiting the development of pathogen. But enough care has not been under taken regarding the biochemical characterization of Culex mosquitoes (Culex quinquefasciatus in West Bengal. Therefore a preliminary investigation have been undertaken for the determination of biochemical characterization such as gram staining, pattern of growth, detection of economically important enzyme as well as antibiotic susceptibility assay of midgut bacterial isolates of Culex (Culex quinquefasciatus in some urban, sub-urban and rural areas of West Bengal.

  20. Direct competition assay for transcription fidelity. (United States)

    Lubkowska, Lucyna; Kireeva, Maria L


    Accurate transcription is essential for faithful information flow from DNA to RNA and to the protein. Mechanisms of cognate substrate selection by RNA polymerases are currently elucidated by structural, genetic, and biochemical approaches. Here, we describe a fast and reliable approach to quantitative analyses of transcription fidelity, applicable to analyses of RNA polymerase selectivity against misincorporation, incorporation of dNMPs, and chemically modified rNMP analogues. The method is based on different electrophoretic mobility of RNA oligomers of the same length but differing in sequence.

  1. Instructions for Uploading Data to the Assay Portal - Instructions for Uploading Data to the Assay Portal (United States)

    This document provides instructions for configuring and uploading data files to the CPTAC Assay Portal. It is divided into sections, with an overview checklist provided at the end. If help is needed at any stage of the process, please use the support page:

  2. Alteration of biochemical parameters after supplementation of multivitamins and minerals of sprayers on grape gardens of Western Maharashtra (India

    Directory of Open Access Journals (Sweden)

    Jyotsna A. Patil


    Full Text Available Background: In the horticulture sector, the land used for cash crop like grapes is increasing particularly in Maharashtra state. There is an increasing use of the pesticides in an attempt to increase the yield and reduce the post harvest losses. The environmental pollution and poisoning due to wide use of pesticide during grape cultivation may be much more important factors to disturb the socio economical status of uneducated farm worker in rural areas. Aim and Objectives: The study aimed to evaluate the pesticide exposure of the population associated with grape cultivation with respect to biochemical effects, oxidative stress, antioxidants status and protecting them from such effects by supplementing vitamins A to Z. Material and Methods: For this study 30 subjects with occupational pesticides exposure i.e. sprayers of grape gardens having age in the range of 20 to 45 years from Tasgaon taluka, Sangli district, (Western Maharashtra India were taken and all these study group subject,500 mg vitamin A to Z tablet / day for fifteen days were supplemented. The blood was collected by venipuncture from all these subjects for biochemical parameters assay before and after fifteen days of vitamins A to Z supplementation and all biochemical parameters were estimated by using standard methods. Results: Fifteen days vitamin A to Z and multi minerals supplementation to sprayers of grape gardens, we observed significant increase in serum Acetyl Cholinesterase (P<0.001, 2.32%, and decreased Aspartate Transaminase (P<0.05, 8.36%, Alanine Transaminase (P<0.05, 14.16%, serum Lipid Peroxide (P<0.001, 24.39%, Glutathione S-Transferase (P<0.01, 29.84% and increased RBC-Superoxide Dismutase (P<0.05, 11.55%, RBC-Catalase (P<0.001, 25.7%, serum zinc (P<0.01, 3.74%, copper (P<0.001, 4.13%, blood lead (P<0.01, 3.33% as compared to before vitamin A to Z and multi minerals supplementation and other biochemical parameters such as serum C-Reactive Proteins, Total Proteins

  3. Demonstration of DSI-semen--A novel DNA methylation-based forensic semen identification assay. (United States)

    Wasserstrom, Adam; Frumkin, Dan; Davidson, Ariane; Shpitzen, Moshe; Herman, Yael; Gafny, Ron


    Determining whether the source tissue of biological material is semen is important in confirming sexual assaults, which account for a considerable percentage of crime cases. The gold standard for confirming the presence of semen is microscopic identification of sperm cells, however, this method is labor intensive and operator-dependent. Protein-based immunologic assays, such as PSA, are highly sensitive and relatively fast, but suffer from low specificity in some situations. In addition, proteins are less stable than DNA under most environmental insults. Recently, forensic tissue identification advanced with the development of several approaches based on mRNA and miRNA for identification of various body fluids. Herein is described DNA source identifier (DSI)-semen, a DNA-based assay that determines whether the source tissue of a sample is semen based on detection of semen-specific methylation patterns in five genomic loci. The assay is comprised of a simple single tube biochemical procedure, similar to DNA profiling, followed by automatic software analysis, yielding the identification (semen/non-semen) accompanied by a statistical confidence level. Three additional internal control loci are used to ascertain the reliability of the results. The assay, which aims to replace microscopic examination, can easily be integrated by forensic laboratories and is automatable. The kit was tested on 135 samples of semen, saliva, venous blood, menstrual blood, urine, and vaginal swabs and the identification of semen vs. non-semen was correct in all cases. In order to test the assay's applicability in "real-life" situations, 33 actual casework samples from the forensic biological lab of the Israeli police were analyzed, and the results were compared with microscopic examination performed by Israeli police personnel. There was complete concordance between both analyses except for one sample, in which the assay identified semen whereas no sperm was seen in the microscope. This

  4. 40 CFR 158.2084 - Experimental use permit biochemical pesticides nontarget organisms and environmental fate data... (United States)


    ... pesticides nontarget organisms and environmental fate data requirements table. 158.2084 Section 158.2084 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) PESTICIDE PROGRAMS DATA REQUIREMENTS FOR PESTICIDES Biochemical Pesticides § 158.2084 Experimental use permit biochemical...

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

    Directory of Open Access Journals (Sweden)

    Giuseppina Lima Pace Pereira


    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.

  6. Biochemical functionalization of peptide nanotubes with phage displayed peptides (United States)

    Swaminathan, Swathi; Cui, Yue


    The development of a general approach for the biochemical functionalization of peptide nanotubes (PNTs) could open up existing opportunities in both fundamental studies as well as a variety of applications. PNTs are spontaneously assembled organic nanostructures made from peptides. Phage display has emerged as a powerful approach for identifying selective peptide binding motifs. Here, we demonstrate for the first time the biochemical functionalization of PNTs via peptides identified from a phage display peptide library. The phage-displayed peptides are shown to recognize PNTs. These advances further allow for the development of bifunctional peptides for the capture of bacteria and the self-assembly of silver particles onto PNTs. We anticipate that these results could provide significant opportunities for using PNTs in both fundamental studies and practical applications, including sensors and biosensors nanoelectronics, energy storage devices, drug delivery, and tissue engineering.

  7. The application of information theory to biochemical signaling systems. (United States)

    Rhee, Alex; Cheong, Raymond; Levchenko, Andre


    Cell signaling can be thought of fundamentally as an information transmission problem in which chemical messengers relay information about the external environment to the decision centers within a cell. Due to the biochemical nature of cellular signal transduction networks, molecular noise will inevitably limit the fidelity of any messages received and processed by a cell's signal transduction networks, leaving it with an imperfect impression of its environment. Fortunately, Shannon's information theory provides a mathematical framework independent of network complexity that can quantify the amount of information that can be transmitted despite biochemical noise. In particular, the channel capacity can be used to measure the maximum number of stimuli a cell can distinguish based upon the noisy responses of its signaling systems. Here, we provide a primer for quantitative biologists that covers fundamental concepts of information theory, highlights several key considerations when experimentally measuring channel capacity, and describes successful examples of the application of information theoretic analysis to biological signaling.

  8. The free energy cost of accurate biochemical oscillations

    CERN Document Server

    Cao, Yuansheng; Ouyang, Qi; Tu, Yuhai


    Oscillation is an important cellular process that regulates timing of different vital life cycles. However, in the noisy cellular environment, oscillations can be highly inaccurate due to phase fluctuations. It remains poorly understood how biochemical circuits suppress phase fluctuations and what is the incurred thermodynamic cost. Here, we study four different types of biochemical oscillations representing three basic oscillation motifs shared by all known oscillatory systems. We find that the phase diffusion constant follows the same inverse dependence on the free energy dissipation per period for all systems studied. This relationship between the phase diffusion and energy dissipation is shown analytically in a model of noisy oscillation. Microscopically, we find that the oscillation is driven by multiple irreversible cycles that hydrolyze the fuel molecules such as ATP; the number of phase coherent periods is proportional to the free energy consumed per period. Experimental evidence in support of this un...


    Directory of Open Access Journals (Sweden)

    Ecaterina Emnova


    Full Text Available The paper deals with the evaluation of the intensity of certain soil biochemical processes (e.g. soil organic C mineralization at Organic and mixed Mineral+Organic fertilization of typical chernozem in crop rotation dynamics (for 6 years by use of eco-physiological indicators of biological soil quality: microbial biomass carbon, basal soil respiration, as well as, microbial and metabolic quotients. Soil sampling was performed from a long-term field crop experiment, which has been established in 1971 at the Balti steppe (Northern Moldova. The crop types had a more considerable impact on the soil microbial biomass accumulation and community biochemical activity compared to long-term Organic or mixed Mineral + Organic fertilizers amendments. The Org fertilization system doesn’t make it possible to avoid the loss of organic C in arable typical chernozem. The organic fertilizer (cattle manure is able to mitigate the negative consequences of long-term mineral fertilization.

  10. Electrolyte-Gated Graphene Ambipolar Frequency Multipliers for Biochemical Sensing. (United States)

    Fu, Wangyang; Feng, Lingyan; Mayer, Dirk; Panaitov, Gregory; Kireev, Dmitry; Offenhäusser, Andreas; Krause, Hans-Joachim


    In this Letter, the ambipolar properties of an electrolyte-gated graphene field-effect transistor (GFET) have been explored to fabricate frequency-doubling biochemical sensor devices. By biasing the ambipolar GFETs in a common-source configuration, an input sinusoidal voltage at frequency f applied to the electrolyte gate can be rectified to a sinusoidal wave at frequency 2f at the drain electrode. The extraordinary high carrier mobility of graphene and the strong electrolyte gate coupling provide the graphene ambipolar frequency doubler an unprecedented unity gain, as well as a detection limit of ∼4 pM for 11-mer single strand DNA molecules in 1 mM PBS buffer solution. Combined with an improved drift characteristics and an enhanced low-frequency 1/f noise performance by sampling at doubled frequency, this good detection limit suggests the graphene ambipolar frequency doubler a highly promising biochemical sensing platform.

  11. Biomphalaria prona (Gastropoda: Planorbidae: a morphological and biochemical study

    Directory of Open Access Journals (Sweden)

    W. Lobato Paraense


    Full Text Available Two samples of Biomphalaria prona (Martens, 1873 from Lake Valencia (type locality and seven from other Venezuelan localities were studied morphologically (shell and reproductive system and biochemically (allozyme electrophoresis. In spite of marked differences in shell characters, all of them proved indistinguishable under the anatomic and biochemical criteria. So far B. prona has been considered an endemic species, restricted to Lake Valencia. It is now demonstrated that the extralacustrine populations refered to Biomphalaria havanensis (Pfeiffer, 1839 by several authors correspond in shell characters to an extreme variant of B. prona from the Lake and really belong to the last*mentioned species. They may be regarded as the result of a process of directional selection favoring a shell phenotype other than those making up the modal class in the Lake.

  12. The Metabolic Syndrome and Biochemical Recurrence following Radical Prostatectomy

    Directory of Open Access Journals (Sweden)

    Jennifer M. Post


    Full Text Available Metabolic syndrome refers to a set of conditions that increases the risk of cardiovascular disease and has been associated with an increased risk of prostate cancer, particularly among African American men. This study aimed to estimate the association of metabolic syndrome with biochemical recurrence (BCR in a racially diverse population. Among 383 radical prostatectomy patients, 67 patients had documented biochemical recurrence. Hypertension was significantly, positively associated with the rate of BCR (hazard ratio (HR = 2.1; 95%  CI = 1.1, 3.8. There were distinct racial differences in the prevalence of individual metabolic syndrome components; however, the observed associations with BCR did not differ appreciably by race. We conclude that hypertension may contribute to a poorer prognosis in surgically treated prostate cancer patients. Our findings suggest that targeting components of the metabolic syndrome which are potentially modifiable through lifestyle interventions may be a viable strategy to reduce risk of BCR in prostate cancer.

  13. Effects of Substrate to Inoculum Ratio on the Biochemical Methane Potential of Piggery Slaughterhouse Wastes (United States)

    Yoon, Young-Man; Kim, Seung-Hwan; Shin, Kook-Sik; Kim, Chang-Hyun


    The aim of this study was to assess the effect of substrate to inoculum ratio (S/I ratio) on the biochemical methane potential (BMP) and anaerobic biodegradability (Ddeg) of different piggery slaughterhouse wastes, such as piggery blood, intestine residue, and digestive tract content. These wastes were sampled from a piggery slaughterhouse located in Kimje, South Korea. Cumulative methane production curves for the wastes were obtained from the anaerobic batch fermentation having different S/I ratios of 0.1, 0.5, 1.0, and 1.5. BMP and anaerobic biodegradabilities (Ddeg) of the wastes were calculated from cumulative methane production data for the tested conditions. At the lowest S/I ration of 0.1, BMPs of piggery blood, intestine residue, and digestive tract content were determined to be 0.799, 0.848, and 1.076 Nm3 kg−1-VSadded, respectively, which were above the theoretical methane potentials of 0.539, 0.644, and 0.517 Nm3 kg−1-VSadded for blood, intestine residue, and digestive tract content, respectively. However, BMPs obtained from the higher S/I ratios of 0.5, 1.0, and 1.5 were within the theoretical range for all three types of waste and were not significantly different for the different S/I ratios tested. Anaerobic biodegradabilities calculated from BMP data showed a similar tendency. These results imply that, for BMP assay in an anaerobic reactor, the S/I ratio of anaerobic reactor should be above 0.1 and the inoculum should be sufficiently stabilized to avoid further degradation during the assay. PMID:25049994


    the light of biochemical evolution , hemoproteids - resp. the porphyrin compounds they are based upon - must have existed on earth as early as... the CO2-assimilation, which means that the porphyrin catalyse of oxydative metabolism consequently derived from the second period of the early atmosphere. This aspect is discussed. (Author)...biological organisms with oxydative metabolism occurred. Since the primitive early atmosphere was followed by a CO2-containing

  15. Biochemical recurrence of prostate cancer: the controversial recognition and management

    Institute of Scientific and Technical Information of China (English)

    XIA Shu-jie; JING Yi-feng


    @@ Over the past decaade, more and more patients diagnosed as prostate cancer have received radical management attributing to the advent of prostate-specific antigen (PSA) based medical screening.Radical prostatectomy (RP) and radiation therapy (RT) are the most commonly used forms of definitive therapy for clinically localized prostate cancer.However, despite these technique advances, biochemical recurrence (BCR),as determined by subsequent rises in the serum PSA level,is still a challenge that urologists face.

  16. Biochemical Changes During Seed Germination of Sterculia urens Roxb.


    Botcha SATYANARAYANA; Prattipati Subhashini DEVI; Atluru ARUNDATHI


    The present study describes biochemical changes taking place during seed germination of Sterculia urens. The levels of proteins, total amino acids, reducing sugars, total soluble sugars and lipids were studied during various stages of seed germination (0-15 days). Total protein content was decreased in cotyledons during seed germination while free amino acid content increased to its maximum extent by 9th day of germination and reverse trend thereafter. The levels of reducing sugars and total ...

  17. Biological and biochemical properties in evaluation of forest soil quality


    Błońska Ewa; Lasota Jarosław


    The aim of this study was to assess the possibility of using biological and biochemical parameters in the evaluation of forest soil quality and changes caused by land use. The study attempted to determine a relationship between the enzymatic activity of soil, the number of earthworms and soil physico-chemical properties. The study was carried out in central Poland in adjoining Forest Districts (Przedbórz and Smardzewice). In soil samples taken from 12 research plots, basic physico-chemical pr...

  18. Biological and biochemical properties in evaluation of forest soil quality


    Błońska, Ewa; Lasota, Jarosław


    The aim of this study was to assess the possibility of using biological and biochemical parameters in the evaluation of forest soil quality and changes caused by land use. The study attempted to determine a relationship between the enzymatic activity of soil, the number of earthworms and soil physico-chemical properties. The study was carried out in central Poland in adjoining Forest Districts (Przedbórz and Smardzewice). In soil samples taken from 12 research plots, basic physico-chem...

  19. Molecular and biochemical studies on bovine ephemeral fever

    Directory of Open Access Journals (Sweden)

    Nahed S. Thabet; Emad W. Ghazy; Mohamed A. Nayel; Mohamed Abo-Elkhair


    Full Text Available Bovine ephemeral fever (BEF in cattle has been reported to be associated with a range of biochemical changes which are similar to those seen in milk fever. This study aimed to clarify the biochemical alterations that associate infection of cattle with BEF with special references to the mechanisms involved in the development of hypocalcemia. The study was conducted on 30 cases of cattle infected with BEF based on the characteristic clinical signs which were confirmed by isolation of virus and RT-PCR. Another 6 healthy cows were used in the study as control. The evaluated parameters included biochemical variables such as serum values of total protein (TP, albumin (Alb, glucose (Glu, total calcium (tCa, ionized calcium (iCa, inorganic phosphorus (P, magnesium (Mg, sodium (Na, potassium (K, chloride (Cl, creatinine (Cr, blood urea nitrogen (BUN and serum activity of alkaline phosphatase (ALP. Hormonal profile included parathyroid hormone (PTH, insulin (Ins, and cortisol (Cor. The results showed that BEF-infected animals demonstrated a significant decrease (P<0.05 in serum concentrations of TP, Glo, iCa, P, Na, K, BUN and ALP while the mean values of serum levels of Glu and Cl were significantly increased (P<0.05. The mean values of serum levels of PTH were significantly decreased (P<0.05 while serum concentrations of Ins and Cor showed a significant increase. It was concluded that the clinical signs of bovine ephemeral fever are related to the hypocalcemia resulting from suppression of parathyroid hormone which seems to be mediated by respiratory alkalosis caused by the disease. This explanation needs future studies to provide a direct link between measurement of blood indicators of acid-base status, blood biochemical parameters and urine analysis. However, this work can provide a good knowledge about the pathogenesis of the disease that can lead to better management and proper treatment.

  20. Accelerated maximum likelihood parameter estimation for stochastic biochemical systems

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    Daigle Bernie J


    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