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Sample records for bioluminescent bioreporter integrated

  1. Remote sensing of microbial volatile organic compounds with a bioluminescent bioreporter integrated circuit

    Science.gov (United States)

    Ripp, Steven A.; Daumer, Kathleen A.; Garland, Jay L.; Simpson, Michael L.; Sayler, Gary S.

    2004-03-01

    As a means towards advanced, early-warning detection of microbial growth in enclosed structures, we have constructed a bioluminescent bioreporter for the detection of the microbial volatile organic compound (MVOC) p-cymene. MVOCs are produced as metabolic by-products of bacteria and fungi and are detectable before any visible signs of microbial growth appear, thereby serving as very early indicators of potential biocontamination problems. The bioreporter, designated Pseudomonas putida UT93, contains a Vibrio fischeri luxCDABE gene fusion to a p-cymene/p-cumate inducible promoter. Exposure of strain UT93 to p-cymene from approximately 0.02 to 850 ppm produced self-generated bioluminescence in less than 1.5 hours. The bioreporter was also interfaced with an integrated circuit microluminometer to create a miniaturized hybrid sensor for remote monitoring of p-cymene signatures. This bioluminescent bioreporter integrated circuit (BBIC) device was capable of detecting fungal presence within approximately 3.5 hours of initial exposure to Penicillium roqueforti.

  2. Integrated CMOS photodetectors and signal processing for very low-level chemical sensing with the bioluminescent bioreporter integrated circuit

    Science.gov (United States)

    Bolton, Eric K.; Sayler, Gary S.; Nivens, David E.; Rochelle, James M.; Ripp, Steven; Simpson, Michael L.

    2002-01-01

    We report an integrated CMOS microluminometer optimized for the detection of low-level bioluminescence as part of the bioluminescent bioreporter integrated circuit (BBIC). This microluminometer improves on previous devices through careful management of the sub-femtoampere currents, both signal and leakage, that flow in the front-end processing circuitry. In particular, the photodiode is operated with a reverse bias of only a few mV, requiring special attention to the reset circuitry of the current-to-frequency converter (CFC) that forms the front-end circuit. We report a sub-femtoampere leakage current and a minimum detectable signal (MDS) of 0.15 fA (1510 s integration time) using a room temperature 1.47 mm2 CMOS photodiode. This microluminometer can detect luminescence from as few as 5000 fully induced Pseudomonas fluorescens 5RL bacterial cells. c2002 Elsevier Science B.V. All rights reserved.

  3. Bioluminescent bioreporter for assessment of arsenic contamination ...

    Indian Academy of Sciences (India)

    The bioreporter sensor system developed in this study can measure the estimated range of 0.74–60 g of As/L and is both specific and selective for sensing bioavailable As. The constructed bacterial biosensor was further used for the determination of arsenic ion concentration in different environmental samples of India.

  4. Preconcentration and Detection of Mercury with Bioluminescent Bioreporter E. coli ARL1.

    Czech Academy of Sciences Publication Activity Database

    Solovyev, Andrey; Koštejn, Martin; Kuncová, Gabriela; Dostálek, P.; Rohovec, Jan; Navrátil, Tomáš

    2015-01-01

    Roč. 99, č. 20 (2015), s. 8793-8802 ISSN 0175-7598 Institutional support: RVO:67985858 ; RVO:67985831 Keywords : mercury detection * bioreporters * biosorbents Subject RIV: CC - Organic Chemistry Impact factor: 3.376, year: 2015

  5. Effect of Naphthalene and Salicylate Analogues on the Bioluminescence of Bioreporter Pseudomonas Fluorescens HK44.

    Czech Academy of Sciences Publication Activity Database

    Trögl, Josef; Kuncová, Gabriela; Kubicová, L.; Pařík, P.; Hálová, Jaroslava; Demnerová, K.; Ripp, S.; Sayler, G. S.

    2007-01-01

    Roč. 52, 1 (2007) , s. 3-14 ISSN 0015-5632 R&D Projects: GA ČR(CZ) GA104/05/2637; GA ČR(CZ) GA203/06/1244 Institutional research plan: CEZ:AV0Z40720504; CEZ:AV0Z40320502 Keywords : pseudomonas fluorescens HK44 * bioluminescence * bioluminescence Subject RIV: CE - Biochemistry Impact factor: 0.989, year: 2007

  6. The Repetitive Detection of Toluene with Bioluminescence Bioreporter Pseudomonas putida TVA8 Encapsulated in Silica Hydrogel on an Optical Fiber.

    Czech Academy of Sciences Publication Activity Database

    Kuncová, Gabriela; Ishizaki, Takayuki; Solovyev, Andrey; Trögl, J.; Ripp, S.

    2016-01-01

    Roč. 9, č. 6 (2016), s. 467 ISSN 1996-1944 Institutional support: RVO:67985858 Keywords : bioluminescent biosensor * silica gel * encapsulation Subject RIV: CC - Organic Chemistry Impact factor: 2.654, year: 2016

  7. Remote detection of human toxicants in real time using a human-optimized, bioluminescent bacterial luciferase gene cassette bioreporter

    Science.gov (United States)

    Close, Dan; Webb, James; Ripp, Steven; Patterson, Stacey; Sayler, Gary

    2012-06-01

    Traditionally, human toxicant bioavailability screening has been forced to proceed in either a high throughput fashion using prokaryotic or lower eukaryotic targets with minimal applicability to humans, or in a more expensive, lower throughput manner that uses fluorescent or bioluminescent human cells to directly provide human bioavailability data. While these efforts are often sufficient for basic scientific research, they prevent the rapid and remote identification of potentially toxic chemicals required for modern biosecurity applications. To merge the advantages of high throughput, low cost screening regimens with the direct bioavailability assessment of human cell line use, we re-engineered the bioluminescent bacterial luciferase gene cassette to function autonomously (without exogenous stimulation) within human cells. Optimized cassette expression provides for fully endogenous bioluminescent production, allowing continuous, real time monitoring of the bioavailability and toxicology of various compounds in an automated fashion. To access the functionality of this system, two sets of bioluminescent human cells were developed. The first was programed to suspend bioluminescent production upon toxicological challenge to mimic the non-specific detection of a toxicant. The second induced bioluminescence upon detection of a specific compound to demonstrate autonomous remote target identification. These cells were capable of responding to μM concentrations of the toxicant n-decanal, and allowed for continuous monitoring of cellular health throughout the treatment process. Induced bioluminescence was generated through treatment with doxycycline and was detectable upon dosage at a 100 ng/ml concentration. These results demonstrate that leveraging autonomous bioluminescence allows for low-cost, high throughput direct assessment of toxicant bioavailability.

  8. Co-Cultured Continuously Bioluminescent Human Cells as Bioreporters for the Detection of Prodrug Therapeutic Impact Pre- and Post-Metabolism

    Directory of Open Access Journals (Sweden)

    Tingting Xu

    2017-12-01

    Full Text Available Modern drug discovery workflows require assay systems capable of replicating the complex interactions of multiple tissue types, but that can still function under high throughput conditions. In this work, we evaluate the use of substrate-free autobioluminescence in human cell lines to support the performance of these assays with reduced economical and logistical restrictions relative to substrate-requiring bioluminescent reporter systems. The use of autobioluminescence was found to support assay functionality similar to existing luciferase reporter targets. The autobioluminescent assay format was observed to correlate strongly with general metabolic activity markers such as ATP content and the presence of reactive oxygen species, but not with secondary markers such as glutathione depletion. At the transcriptional level, autobioluminescent dynamics were most closely associated with expression of the CYP1A1 phase I detoxification pathway. These results suggest constitutively autobioluminescent cells can function as general metabolic activity bioreporters, while pairing expression of the autobioluminescent phenotype to detoxification pathway specific promoters could create more specific sensor systems.

  9. Bioreporter Pseudomonas fluorescens HK44 Immobilized in a Silica Matrix

    Czech Academy of Sciences Publication Activity Database

    Trögl, Josef; Ripp, S.; Kuncová, Gabriela; Sayler, G. S.; Demnerová, K.

    2003-01-01

    Roč. 57, č. 12 (2003), s. 596-599 ISSN 0354-7531 R&D Projects: GA ČR GA104/01/0461 Institutional research plan: CEZ:AV0Z4072921 Keywords : bioluminescence bioreporter * optical sensor * naphthalene detection Subject RIV: CE - Biochemistry

  10. The Evolution of the Bacterial Luciferase Gene Cassette (lux as a Real-Time Bioreporter

    Directory of Open Access Journals (Sweden)

    Gary Sayler

    2012-01-01

    Full Text Available The bacterial luciferase gene cassette (lux is unique among bioluminescent bioreporter systems due to its ability to synthesize and/or scavenge all of the substrate compounds required for its production of light. As a result, the lux system has the unique ability to autonomously produce a luminescent signal, either continuously or in response to the presence of a specific trigger, across a wide array of organismal hosts. While originally employed extensively as a bacterial bioreporter system for the detection of specific chemical signals in environmental samples, the use of lux as a bioreporter technology has continuously expanded over the last 30 years to include expression in eukaryotic cells such as Saccharomyces cerevisiae and even human cell lines as well. Under these conditions, the lux system has been developed for use as a biomedical detection tool for toxicity screening and visualization of tumors in small animal models. As the technologies for lux signal detection continue to improve, it is poised to become one of the first fully implantable detection systems for intra-organismal optical detection through direct marriage to an implantable photon-detecting digital chip. This review presents the basic biochemical background that allows the lux system to continuously autobioluminesce and highlights the important milestones in the use of lux-based bioreporters as they have evolved from chemical detection platforms in prokaryotic bacteria to rodent-based tumorigenesis study targets. In addition, the future of lux imaging using integrated circuit microluminometry to image directly within a living host in real-time will be introduced and its role in the development of dose/response therapeutic systems will be highlighted.

  11. Bioluminescent bioreporter for assessment of arsenic contamination ...

    Indian Academy of Sciences (India)

    2013-03-01

    Mar 1, 2013 ... Chile, China, Ghana, Hungary, inner Mongolia, Mexico,. Nepal, New Zealand, Philippines, Taiwan, the United States and Vietnam (Berg et al. 2001; Wilson 2002). Consumption of water from these naturally contaminated aquifers led to chron- ic As poisoning in many of these locations, with perhaps the.

  12. Bioluminescent bioreporter for assessment of arsenic contamination ...

    Indian Academy of Sciences (India)

    In the present study the most efficient -factor controlling the ars operon was selected after screening of 39 Escherichia coli isolates by minimum inhibitory concentration test (MIC) studies from water samples of different geographical locations of India. Among all, strain isolated from Hooghly River (West Bengal) was found to ...

  13. Bioluminescent bioreporter for assessment of arsenic contamination ...

    Indian Academy of Sciences (India)

    2013-03-01

    Mar 1, 2013 ... In the present study the most efficient R-factor controlling the ars operon was selected after screening of 39. Escherichia coli isolates by minimum inhibitory concentration test (MIC) studies from water samples of different geographical locations of India. Among all, strain isolated from Hooghly River (West ...

  14. Cell-based bioreporter assay coupled to HPLC micro-fractionation in the evaluation of antimicrobial properties of the basidiomycete fungus Pycnoporus cinnabarinus.

    Science.gov (United States)

    Järvinen, Päivi; Nybond, Susanna; Marcourt, Laurence; Ferreira Queiroz, Emerson; Wolfender, Jean-Luc; Mettälä, Aila; Karp, Matti; Vuorela, Heikki; Vuorela, Pia; Hatakka, Annele; Tammela, Päivi

    2016-01-01

    Identification of bioactive components from complex natural product extracts can be a tedious process that aggravates the use of natural products in drug discovery campaigns. This study presents a new approach for screening antimicrobial potential of natural product extracts by employing a bioreporter assay amenable to HPLC-based activity profiling. A library of 116 crude extracts was prepared from fungal culture filtrates by liquid-liquid extraction with ethyl acetate, lyophilised, and screened against Escherichia coli using TLC bioautography. Active extracts were studied further with a broth microdilution assay, which was, however, too insensitive for identifying the active microfractions after HPLC separation. Therefore, an assay based on bioluminescent E. coli K-12 (pTetLux1) strain was coupled with HPLC micro-fractionation. Preliminary screening yielded six fungal extracts with potential antimicrobial activity. A crude extract from a culture filtrate of the wood-rotting fungus, Pycnoporus cinnabarinus (Jacq.) P. Karst. (Polyporaceae), was selected for evaluating the functionality of the bioreporter assay in HPLC-based activity profiling. In the bioreporter assay, the IC50 value for the crude extract was 0.10 mg/mL. By integrating the bioreporter assay with HPLC micro-fractionation, the antimicrobial activity was linked to LC-UV peak of a compound in the chromatogram of the extract. This compound was isolated and identified as a fungal pigment phlebiarubrone. HPLC-based activity profiling using the bioreporter-based approach is a valuable tool for identifying antimicrobial compound(s) from complex crude extracts, and offers improved sensitivity and speed compared with traditional antimicrobial assays, such as the turbidimetric measurement.

  15. Bioreporter pseudomonas fluorescens HK44 immobilized in a silica matrix

    Directory of Open Access Journals (Sweden)

    Trogl J.

    2003-01-01

    Full Text Available The bioluminescent bioreporter Pseudomonas fluorescens HK44, the whole cell bacterial biosensor that responds to naphthalene and its metabolites via the production of visible light, was immobilized into a silica matrix by the sol-gel technique. The bioluminescence intensities were measured in the maximum of the bioluminescence band at X = 500 nm. The immobilized cells (>105 cells per g silica matrix produced light after induction by salicylate (cone. > 10 g/l, naphthalene and aminobenzoic acid. The bioluminescence intensities induced by 2,3-dihydroxynaphthalene 3-hydroxybenzoic acid and 4-hydroxybenzoic acid were comparable to a negative control. The cells in the silica layers on glass slides produced light in response to the presence of an inductor at least 8 months after immobilization, and >50 induction cycles. The results showed that these test slides could be used as assays for the multiple determination of water pollution.

  16. Illuminating the detection chain of bacterial bioreporters

    NARCIS (Netherlands)

    Meer, J.R. van der; Tropel, D.; Jaspers, M.

    2004-01-01

    Engineering bacteria for measuring chemicals of environmental or toxicological concern (bioreporter bacteria) has grown slowly into a mature research area. Despite many potential advantages, current bioreporters do not perform well enough to comply with environmental detection standards. Basically,

  17. Integrating printed microfluidics with silicon photomultipliers for miniaturised and highly sensitive ATP bioluminescence detection.

    Science.gov (United States)

    Santangelo, M F; Libertino, S; Turner, A P F; Filippini, D; Mak, W C

    2018-01-15

    Bioluminescence has been widely used for important biosensing applications such as the measurement of adenosine triphosphate (ATP), the energy unit in biological systems and an indicator of vital processes. The current technology for detection is mainly based on large equipment such as readers and imaging systems, which require intensive and time-consuming procedures. A miniaturised bioluminescence sensing system, which would allow sensitive and continuous monitoring of ATP, with an integrated and low-cost disposable microfluidic chamber for handling of biological samples, is highly desirable. Here, we report the design, fabrication and testing of 3D printed microfluidics chips coupled with silicon photomultipliers (SiPMs) for high sensitive real-time ATP detection. The 3D microfluidic chip reduces reactant consumption and facilitates solution delivery close to the SiPM to increase the detection efficiency. Our system detects ATP with a limit of detection (LoD) of 8nM and an analytical dynamic range between 15nM and 1µM, showing a stability error of 3%, and a reproducibility error below of 20%. We demonstrate the dynamic monitoring of ATP in a continuous-flow system exhibiting a fast response time, ~4s, and a full recovery to the baseline level within 17s. Moreover, the SiPM-based bioluminescence sensing system shows a similar analytical dynamic range for ATP detection to that of a full-size PerkinElmer laboratory luminescence reader. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Wireless Luminescence Integrated Sensors (WLIS)

    Energy Technology Data Exchange (ETDEWEB)

    Simpson, M.L.; Sayler, G.S. (Univ. Tennessee)

    2003-11-10

    The goal of this project was the development of a family of wireless, single-chip, luminescence-sensing devices to solve a number of difficult distributed measurement problems in areas ranging from environmental monitoring and assessment to high-throughput screening of combinatorial chemistry libraries. These wireless luminescence integrated sensors (WLIS) consist of a microluminometer, wireless data transmitter, and RF power input circuit all realized in a standard integrated circuit (IC) process with genetically engineered, whole-cell, bioluminescent bioreporters encapsulated and deposited on the IC. The end product is a family of compact, low-power, rugged, low-cost sensors. As part of this program they developed an integrated photodiode/signal-processing scheme with an rms noise level of 175 electrons/second for a 13-minute integration time, and a quantum efficiency of 66% at the 490-nm bioluminescent wavelength. this performance provided a detection limit of < 1000 photons/second. Although sol-gel has previously been used to encapsulate yeast cells, the reaction conditions necessary for polymerization (primarily low pH) have beforehand proven too harsh for bacterial cell immobilizations. Utilizing sonication methods, they have were able to initiate polymerization under pH conditions conductive to cell survival. both a toluene bioreporter (Pseudomonas putida TVA8) and a naphthalene bioreporter (Pseudomonas fluorescens HK44) were successfully encapsulated in sol-gel and shown to produce a fairly significant bioluminescent response. In addition to the previously developed naphthalene- and toluene-sensitive bioreporters, they developed a yeast-based xenoestrogen reporter. This technology has been licensed by Micro Systems Technologies, a startup company in Dayton, Ohio for applications in environmental containments monitoring, and for detecting weapons of mass destruction (i.e. homeland security).

  19. Increased bioassay sensitivity of bioactive molecule discovery using metal-enhanced bioluminescence

    International Nuclear Information System (INIS)

    Golberg, Karina; Elbaz, Amit; McNeil, Ronald; Kushmaro, Ariel; Geddes, Chris D.; Marks, Robert S.

    2014-01-01

    We report the use of bioluminescence signal enhancement via proximity to deposited silver nanoparticles for bioactive compound discovery. This approach employs a whole-cell bioreporter harboring a plasmid-borne fusion of a specific promoter incorporated with a bioluminescence reporter gene. The silver deposition process was first optimized to provide optimal nanoparticle size in the reaction time dependence with fluorescein. The use of silver deposition of 350 nm particles enabled the doubling of the bioluminescent signal amplitude by the bacterial bioreporter when compared to an untouched non-silver-deposited microtiter plate surface. This recording is carried out in the less optimal but necessary far-field distance. SEM micrographs provided a visualization of the proximity of the bioreporter to the silver nanoparticles. The electromagnetic field distributions around the nanoparticles were simulated using Finite Difference Time Domain, further suggesting a re-excitation of non-chemically excited bioluminescence in addition to metal-enhanced bioluminescence. The possibility of an antiseptic silver effect caused by such a close proximity was eliminated disregarded by the dynamic growth curves of the bioreporter strains as seen using viability staining. As a highly attractive biotechnology tool, this silver deposition technique, coupled with whole-cell sensing, enables increased bioluminescence sensitivity, making it especially useful for cases in which reporter luminescence signals are very weak

  20. Increased bioassay sensitivity of bioactive molecule discovery using metal-enhanced bioluminescence

    Energy Technology Data Exchange (ETDEWEB)

    Golberg, Karina, E-mail: karingo@bgu.ac.il; Elbaz, Amit [Ben-Gurion University of the Negev, Avram and Stella Goldstein-Goren Department of Biotechnology Engineering (Israel); McNeil, Ronald [The Institute of Fluorescence, University of Maryland Baltimore County (United States); Kushmaro, Ariel [Ben-Gurion University of the Negev, Avram and Stella Goldstein-Goren Department of Biotechnology Engineering (Israel); Geddes, Chris D. [The Institute of Fluorescence, University of Maryland Baltimore County (United States); Marks, Robert S., E-mail: rsmarks@bgu.ac.il [Ben-Gurion University of the Negev, Avram and Stella Goldstein-Goren Department of Biotechnology Engineering (Israel)

    2014-12-15

    We report the use of bioluminescence signal enhancement via proximity to deposited silver nanoparticles for bioactive compound discovery. This approach employs a whole-cell bioreporter harboring a plasmid-borne fusion of a specific promoter incorporated with a bioluminescence reporter gene. The silver deposition process was first optimized to provide optimal nanoparticle size in the reaction time dependence with fluorescein. The use of silver deposition of 350 nm particles enabled the doubling of the bioluminescent signal amplitude by the bacterial bioreporter when compared to an untouched non-silver-deposited microtiter plate surface. This recording is carried out in the less optimal but necessary far-field distance. SEM micrographs provided a visualization of the proximity of the bioreporter to the silver nanoparticles. The electromagnetic field distributions around the nanoparticles were simulated using Finite Difference Time Domain, further suggesting a re-excitation of non-chemically excited bioluminescence in addition to metal-enhanced bioluminescence. The possibility of an antiseptic silver effect caused by such a close proximity was eliminated disregarded by the dynamic growth curves of the bioreporter strains as seen using viability staining. As a highly attractive biotechnology tool, this silver deposition technique, coupled with whole-cell sensing, enables increased bioluminescence sensitivity, making it especially useful for cases in which reporter luminescence signals are very weak.

  1. U-SPECT-BioFluo : An integrated radionuclide, bioluminescence, and fluorescence imaging platform

    NARCIS (Netherlands)

    Van Oosterom, M.N.; Kreuger, R.; Buckle, T.; Mahn, W.A.; Bunschoten, A.; Josephson, L.; Van Leeuwen, F.W.B.; Beekman, F.J.

    2014-01-01

    Background: In vivo bioluminescence, fluorescence, and single-photon emission computed tomography (SPECT) imaging provide complementary information about biological processes. However, to date these signatures are evaluated separately on individual preclinical systems. In this paper, we introduce a

  2. On-line monitoring of aerobic bioremediation with bioluminescent reporter microbes. Final report, July 1991--December 1994

    Energy Technology Data Exchange (ETDEWEB)

    Sayler, G.S.

    1995-03-01

    A critical issue in the biological characterization of contaminated sites and in the evaluation of relative bioremediation treatment efficiencies is the development of appropriate monitoring methods for the assessment of pollutant bioavailability and microbial in situ activity potential. In nature, pollutants are found dispersed among the solid, liquid and gaseous phases of the complex environments rendering the analytical estimation of their bioavailability and degradation more difficult and irrelevant. Ex situ and extractive analytical techniques have only been misrepresentative of the natural conditions and often resulted in inaccurate estimates of pollutants mass transfer. In this project, the bioluminescent bioreporter bacterium P. Fluorescens HK44 was integrated to an optical device, capable of conducting emitted light, and used as an online biosensor of naphthalene and salicylate. The physiological requirements of the bacteria and the physical limitations of the biosensor were also determined.

  3. Integrated visualization of multi-angle bioluminescence imaging and micro CT

    NARCIS (Netherlands)

    Kok, P.; Dijkstra, J.; Botha, C.P.; Post, F.H.; Kaijzel, E.; Que, I.; Löwik, C.W.G.M.; Reiber, J.H.C.; Lelieveldt, B.P.F.

    2007-01-01

    This paper explores new methods to visualize and fuse multi-2D bioluminescence imaging (BLI) data with structural imaging modalities such as micro CT and MR. A geometric, back-projection-based 3D reconstruction for superficial lesions from multi-2D BLI data is presented, enabling a coarse estimate

  4. Whole-Cell Bioreporters for the Detection of Bioavailable Metals

    Science.gov (United States)

    Hynninen, Anu; Virta, Marko

    Whole-cell bioreporters are living microorganisms that produce a specific, quantifiable output in response to target chemicals. Typically, whole-cell bioreporters combine a sensor element for the substance of interest and a reporter element coding for an easily detectable protein. The sensor element is responsible for recognizing the presence of an analyte. In the case of metal bioreporters, the sensor element consists of a DNA promoter region for a metal-binding transcription factor fused to a promoterless reporter gene that encodes a signal-producing protein. In this review, we provide an overview of specific whole-cell bioreporters for heavy metals. Because the sensing of metals by bioreporter microorganisms is usually based on heavy metal resistance/homeostasis mechanisms, the basis of these mechanisms will also be discussed. The goal here is not to present a comprehensive summary of individual metal-specific bioreporters that have been constructed, but rather to express views on the theory and applications of metal-specific bioreporters and identify some directions for future research and development.

  5. Numerical modeling of the dynamic response of a bioluminescent bacterial biosensor.

    Science.gov (United States)

    Affi, Mahmoud; Solliec, Camille; Legentilhomme, Patrick; Comiti, Jacques; Legrand, Jack; Jouanneau, Sulivan; Thouand, Gérald

    2016-12-01

    Water quality and water management are worldwide issues. The analysis of pollutants and in particular, heavy metals, is generally conducted by sensitive but expensive physicochemical methods. Other alternative methods of analysis, such as microbial biosensors, have been developed for their potential simplicity and expected moderate cost. Using a biosensor for a long time generates many changes in the growth of the immobilized bacteria and consequently alters the robustness of the detection. This work simulated the operation of a biosensor for the long-term detection of cadmium and improved our understanding of the bioluminescence reaction dynamics of bioreporter bacteria inside an agarose matrix. The choice of the numerical tools is justified by the difficulty to measure experimentally in every condition the biosensor functioning during a long time (several days). The numerical simulation of a biomass profile is made by coupling the diffusion equation and the consumption/reaction of the nutrients by the bacteria. The numerical results show very good agreement with the experimental profiles. The growth model verified that the bacterial growth is conditioned by both the diffusion and the consumption of the nutrients. Thus, there is a high bacterial density in the first millimeter of the immobilization matrix. The growth model has been very useful for the development of the bioluminescence model inside the gel and shows that a concentration of oxygen greater than or equal to 22 % of saturation is required to maintain a significant level of bioluminescence. A continuous feeding of nutrients during the process of detection of cadmium leads to a biofilm which reduces the diffusion of nutrients and restricts the presence of oxygen from the first layer of the agarose (1 mm) and affects the intensity of the bioluminescent reaction. The main advantage of this work is to link experimental works with numerical models of growth and bioluminescence in order to provide a

  6. Detection of Buried Human Remains Using Bioreporter Fluorescence

    Energy Technology Data Exchange (ETDEWEB)

    Vass, A. Dr.; Singleton, G. B.

    2001-10-01

    The search for buried human remains is a difficult, laborious and time-consuming task for law enforcement agencies. This study was conducted as a proof of principle demonstration to test the concept of using bioreporter microorganisms as a means to cover large areas in such a search. These bioreporter microorganisms are affected by a particular component of decaying organic matter that is distinct from decaying vegetation. The diamino compounds cadaverine and putrescine were selected as target compounds for the proof-of-principle investigation, and a search for microorganisms and genes that are responsive to either of these compounds was conducted. One recombinant clone was singled out for characterization based on its response to putrescine. The study results show that small concentrations of putrescine increased expression from this bioreporter construct. Although the level of increase was small (making it difficult to distinguish the signal from background), the results demonstrate the principle that bioreporters can be used to detect compounds resulting from decaying human remains and suggest that a wider search for target compounds should be conducted.

  7. Controlled field release of a bioluminescent genetically engineered microorganism for bioremediation process monitoring and control

    Energy Technology Data Exchange (ETDEWEB)

    Ripp, S.; Nivens, D.E.; Ahn, Y.; Werner, C.; Jarrell, J. IV; Easter, J.P.; Cox, C.D.; Burlage, R.S.; Sayler, G.S.

    2000-03-01

    Pseudomonas fluorescens HK44 represents the first genetically engineered microorganism approved for field testing in the United States for bioremediation purposes. Strain HK44 harbors an introduced lux gene fused within a naphthalene degradative pathway, thereby allowing this recombinant microbe to bioluminescent as it degrades specific polyaromatic hydrocarbons such as naphthalene. The bioremediation process can therefore be monitored by the detection of light. P. fluorescens HK44 was inoculated into the vadose zone of intermediate-scale, semicontained soil lysimeters contaminated with naphthalene, anthracene, and phenanthrene, and the population dynamics were followed over an approximate 2-year period in order to assess the long-term efficacy of using strain HK44 for monitoring and controlling bioremediation processes. Results showed that P. fluorescens HK44 was capable of surviving initial inoculation into both hydrocarbon contaminated and uncontaminated soils and was recoverable from these soils 660 days post inoculation. It was also demonstrated that strain HK44 was capable of generating bioluminescence in response to soil hydrocarbon bioavailability. Bioluminescence approaching 166,000 counts/s was detected in fiber optic-based biosensor devices responding to volatile polyaromatic hydrocarbons, while a portable photomultiplier module detected bioluminescence at an average of 4300 counts/s directly from soil-borne HK44 cells within localized treatment areas. The utilization of lux-based bioreporter microorganisms therefore promises to be a viable option for in situ determination of environmental contaminant bioavailability and biodegradation process monitoring and control.

  8. A model system for pathogen detection using a two-component bacteriophage/bioluminescent signal amplification assay

    Science.gov (United States)

    Bright, Nathan G.; Carroll, Richard J.; Applegate, Bruce M.

    2004-03-01

    Microbial contamination has become a mounting concern the last decade due to an increased emphasis of minimally processed food products specifically produce, and the recognition of foodborne pathogens such as Campylobacter jejuni, Escherichia coli O157:H7, and Listeria monocytogenes. This research investigates a detection approach utilizing bacteriophage pathogen specificity coupled with a bacterial bioluminescent bioreporter utilizing the quorum sensing molecule from Vibrio fischeri, N-(3-oxohexanoyl)-homoserine lactone (3-oxo-C6-HSL). The 3-oxo-C6-HSL molecules diffuse out of the target cell after infection and induce bioluminescence from a population of 3-oxo-C6-HSL bioreporters (ROLux). E. coli phage M13, a well-characterized bacteriophage, offers a model system testing the use of bacteriophage for pathogen detection through cell-to-cell communication via a LuxR/3-oxo-C6-HSL system. Simulated temperate phage assays tested functionality of the ROLux reporter and production of 3-oxo-C6-HSL by various test strains. These assays showed detection limits of 102cfu after 24 hours in a varietry of detection formats. Assays incorporating the bacteriophage M13-luxI with the ROLux reporter and a known population of target cells were subsequently developed and have shown consistent detection limits of 105cfu target organisms. Measurable light response from high concentrations of target cells was almost immediate, suggesting an enrichment step to further improve detection limits and reduce assay time.

  9. Toxicity Testing of Pristine and Aged Silver Nanoparticles in Real Wastewaters Using Bioluminescent Pseudomonas putida

    Directory of Open Access Journals (Sweden)

    Florian Mallevre

    2016-03-01

    Full Text Available Impact of aging on nanoparticle toxicity in real matrices is scarcely investigated due to a lack of suitable methodologies. Herein, the toxicity of pristine and aged silver nanoparticles (Ag NPs to a bioluminescent Pseudomonas putida bioreporter was measured in spiked crude and final wastewater samples (CWs and FWs, respectively collected from four wastewater treatment plants (WWTPs. Results showed lower toxicity of pristine Ag NPs in CWs than in FWs. The effect of the matrix on the eventual Ag NP toxicity was related to multiple physico-chemical parameters (biological oxygen demand (BOD, chemical oxygen demand (COD, total suspended solids (TSS pH, ammonia, sulfide and chloride based on a multivariate analysis. However, no collection site effect was concluded. Aged Ag NPs (up to eight weeks were found less toxic than pristine Ag NPs in CWs; evident increased aggregation and decreased dissolution were associated with aging. However, Ag NPs exhibited consistent toxicity in FWs despite aging; comparable results were obtained in artificial wastewater (AW simulating effluent. The study demonstrates the potency of performing nanoparticle acute toxicity testing in real and complex matrices such as wastewaters using relevant bacterial bioreporters.

  10. Expression of a humanized viral 2A-mediated lux operon efficiently generates autonomous bioluminescence in human cells.

    Directory of Open Access Journals (Sweden)

    Tingting Xu

    Full Text Available Expression of autonomous bioluminescence from human cells was previously reported to be impossible, suggesting that all bioluminescent-based mammalian reporter systems must therefore require application of a potentially influential chemical substrate. While this was disproven when the bacterial luciferase (lux cassette was demonstrated to function in a human cell, its expression required multiple genetic constructs, was functional in only a single cell type, and generated a significantly reduced signal compared to substrate-requiring systems. Here we investigate the use of a humanized, viral 2A-linked lux genetic architecture for the efficient introduction of an autobioluminescent phenotype across a variety of human cell lines.The lux cassette was codon optimized and assembled into a synthetic human expression operon using viral 2A elements as linker regions. Human kidney, breast cancer, and colorectal cancer cell lines were both transiently and stably transfected with the humanized operon and the resulting autobioluminescent phenotype was evaluated using common imaging instrumentation. Autobioluminescent cells were screened for cytotoxic effects resulting from lux expression and their utility as bioreporters was evaluated through the demonstration of repeated monitoring of single populations over a prolonged period using both a modified E-SCREEN assay for estrogen detection and a classical cytotoxic compound detection assay for the antibiotic Zeocin. Furthermore, the use of self-directed bioluminescent initiation in response to target detection was assessed to determine its amenability towards deployment as fully autonomous sensors. In all cases, bioluminescent measurements were supported with traditional genetic and transcriptomic evaluations.Our results demonstrate that the viral 2A-linked, humanized lux genetic architecture successfully produced autobioluminescent phenotypes in all cell lines tested without the induction of cytotoxicity

  11. Silicon photomultiplier (SPM) detection of low-level bioluminescence for the development of deployable whole-cell biosensors: possibilities and limitations.

    Science.gov (United States)

    Li, Huaqing; Lopes, Nicholas; Moser, Scott; Sayler, Gary; Ripp, Steven

    2012-03-15

    Whole-cell bacterial bioreporters await miniaturized photon counting modules with high sensitivity and robust compatible hardware to fulfill their promise of versatile, on-site biosensor functionality. In this study, we explore the photon counting readout properties of the silicon photomultiplier (SPM) with a thermoelectric cooler and the possibilities of detecting low-level bioluminescent signals. Detection performance was evaluated through a simulated LED light source and the bioluminescence produced by the genetically engineered Pseudomonas fluorescens bacterial bioreporter 5RL. Compared with the conventional photomultiplier tube (PMT), the results revealed that the cooled SPM exhibits a wider linear response to inducible substrate concentrations (salicylate) ranging from 250 to 5000 ppb. Although cooling of the SPM lowered dark count rates and improved the minimum detectable signal, and the application of a digital filter enhanced the signal-to-noise ratio, the detection of very low light signals is still limited and remains a challenge in the design of compact photon counting systems. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Bioluminescent Bioreporter Pseudomonas putida TVA8 as a Detector of Water Pollution. Operational Conditions and Selectivity of Free Cells Sensor

    Czech Academy of Sciences Publication Activity Database

    Kuncová, Gabriela; Pazlarová, J.; Hlavatá, Alena; Ripp, S.; Sayler, G.S.

    2011-01-01

    Roč. 11, č. 3 (2011), s. 882-887 ISSN 1470-160X R&D Projects: GA MŠk ME 893 Institutional research plan: CEZ:AV0Z40720504 Keywords : whole-cell biosensor * bioluminiscence * pseudomonas putida TVA8 Subject RIV: EI - Biotechnology ; Bionics Impact factor: 2.695, year: 2011

  13. Prediction of Bioluminescent Proteins Using Auto Covariance Transformation of Evolutional Profiles

    Directory of Open Access Journals (Sweden)

    Yanxin Huang

    2012-03-01

    Full Text Available Bioluminescent proteins are important for various cellular processes, such as gene expression analysis, drug discovery, bioluminescent imaging, toxicity determination, and DNA sequencing studies. Hence, the correct identification of bioluminescent proteins is of great importance both for helping genome annotation and providing a supplementary role to experimental research to obtain insight into bioluminescent proteins’ functions. However, few computational methods are available for identifying bioluminescent proteins. Therefore, in this paper we develop a new method to predict bioluminescent proteins using a model based on position specific scoring matrix and auto covariance. Tested by 10-fold cross-validation and independent test, the accuracy of the proposed model reaches 85.17% for the training dataset and 90.71% for the testing dataset respectively. These results indicate that our predictor is a useful tool to predict bioluminescent proteins. This is the first study in which evolutionary information and local sequence environment information have been successfully integrated for predicting bioluminescent proteins. A web server (BLPre that implements the proposed predictor is freely available.

  14. Application of enzyme bioluminescence in ecology.

    Science.gov (United States)

    Esimbekova, Elena; Kratasyuk, Valentina; Shimomura, Osamu

    2014-01-01

    : This review examines the general principles of bioluminescent enzymatic toxicity bioassays and describes the applications of these methods and the implementation in commercial biosensors. Bioluminescent enzyme system technology (BEST) has been proposed in the bacterial coupled enzyme system, wherein NADH:FMN-oxidoreductase-luciferase substitutes for living organisms. BEST was introduced to facilitate and accelerate the development of cost-competitive enzymatic systems for use in biosensors for medical, environmental, and industrial applications. For widespread use of BEST, the multicomponent reagent "Enzymolum" has been developed, which contains the bacterial luciferase, NADH:FMN-oxidoreductase, and their substrates, co-immobilized in starch or gelatin gel. Enzymolum is the central part of Portable Laboratory for Toxicity Detection (PLTD), which consists of a biodetector module, a sampling module, a sample preparation module, and a reagent module. PLTD instantly signals chemical-biological hazards and allows us to detect a wide range of toxic substances. Enzymolum can be integrated as a biological module into the portable biodetector-biosensor originally constructed for personal use. Based on the example of Enzymolum and the algorithm for creating new enzyme biotests with tailored characteristics, a new approach was demonstrated in biotechnological design and construction. The examples of biotechnological design of various bioluminescent methods for ecological monitoring were provided. Possible applications of enzyme bioassays are seen in the examples for medical diagnostics, assessment of the effect of physical load on sportsmen, analysis of food additives, and in practical courses for higher educational institutions and schools. The advantages of enzymatic assays are their rapidity (the period of time required does not exceed 3-5 min), high sensitivity, simplicity and safety of procedure, and possibility of automation of ecological monitoring; the required

  15. Circular polarization observed in bioluminescence

    NARCIS (Netherlands)

    Wijnberg, Hans; Meijer, E.W.; Hummelen, J.C.; Dekkers, H.P.J.M.; Schippers, P.H.; Carlson, A.D.

    1980-01-01

    While investigating circular polarization in luminescence, and having found it in chemiluminescence, we have studied bioluminescence because it is such a widespread and dramatic natural phenomenon. We report here that left and right lanterns of live larvae of the fireflies, Photuris lucicrescens and

  16. Improvement of bioreporter bacteria-based test systems for the analysis of arsenic in drinking water and the rhizosphere

    International Nuclear Information System (INIS)

    Kuppardt, Anke

    2010-01-01

    Contamination of drinking water with arsenic can be measured in laboratories with atom absorption spectrometry (AAS), mass spectrometry with inductive coupled plasma (ICP-MS) or atom fluorescence spectrometry (AFS) at the relevant concentrations below 50 μg/L. Field test kits which easily and reliably measure arsenic concentrations are not yet available. Test systems on the basis of bioreporter bacteria offer an alternative. Based on the natural resistance mechanism of bacteria against arsenic compounds toxic for humans, bioreporter bacteria can be constructed that display arsenic concentrations with light emission (luminescence or fluorescence) or colour reactions. This is achieved by coupling the gene for the ArsR-protein and arsenic regulated promoters with suitable reporter genes. The resulting bioreporter bacteria report bioavailable arsenic in a dose dependent manner at the toxicologically relevant level of 2 to 80 μg/L and are therewith suitable both for the guideline levels of the WHO of 10 μg/L and for the national standards in South East Asia of 50 μg/L. This alternative method has the advantage of being independent from sophisticated apparatus as by eye detection is feasible and offers the possibility of measuring directly the bioavailable fraction. Bioreporter bacteria are also suitable for in situ research. Yet, in order to apply such bioreporter bacteria as a low-cost analytical tool in a regular manner, open questions exist regarding the preservation of the specific activity, the vitality of bioreporter bacteria and the improvement of bioreporter test systems for layman. The aim of this thesis hence was to optimize and improve bioreporter based test systems to allow easy conservation, storage and transport, and also an application without the need of a sophisticated infrastructure. For that purpose it was intended (i) to develop and validate a method that allows arsenic detection without external calibration (chapter 2) and (ii) to improve the

  17. Improvement of bioreporter bacteria-based test systems for the analysis of arsenic in drinking water and the rhizosphere

    Energy Technology Data Exchange (ETDEWEB)

    Kuppardt, Anke

    2010-02-05

    Contamination of drinking water with arsenic can be measured in laboratories with atom absorption spectrometry (AAS), mass spectrometry with inductive coupled plasma (ICP-MS) or atom fluorescence spectrometry (AFS) at the relevant concentrations below 50 {mu}g/L. Field test kits which easily and reliably measure arsenic concentrations are not yet available. Test systems on the basis of bioreporter bacteria offer an alternative. Based on the natural resistance mechanism of bacteria against arsenic compounds toxic for humans, bioreporter bacteria can be constructed that display arsenic concentrations with light emission (luminescence or fluorescence) or colour reactions. This is achieved by coupling the gene for the ArsR-protein and arsenic regulated promoters with suitable reporter genes. The resulting bioreporter bacteria report bioavailable arsenic in a dose dependent manner at the toxicologically relevant level of 2 to 80 {mu}g/L and are therewith suitable both for the guideline levels of the WHO of 10 {mu}g/L and for the national standards in South East Asia of 50 {mu}g/L. This alternative method has the advantage of being independent from sophisticated apparatus as by eye detection is feasible and offers the possibility of measuring directly the bioavailable fraction. Bioreporter bacteria are also suitable for in situ research. Yet, in order to apply such bioreporter bacteria as a low-cost analytical tool in a regular manner, open questions exist regarding the preservation of the specific activity, the vitality of bioreporter bacteria and the improvement of bioreporter test systems for layman. The aim of this thesis hence was to optimize and improve bioreporter based test systems to allow easy conservation, storage and transport, and also an application without the need of a sophisticated infrastructure. For that purpose it was intended (i) to develop and validate a method that allows arsenic detection without external calibration (chapter 2) and (ii) to

  18. Bioluminescence emissions of the firefly Luciola praeusta ...

    Indian Academy of Sciences (India)

    Prakash

    Bioluminescence is an enchanting process by which living organisms convert chemical energy into light. Fireflies are common organisms that exhibit this process. The enzyme luciferase catalyses the bioluminescence reaction, which uses luciferin, Mg-ATP and molecular oxygen to yield an electronically excited oxyluciferin ...

  19. Remote detection of buried explosives by fluorescent and bioluminescent microbial sensors (Conference Presentation)

    Science.gov (United States)

    Belkin, Shimshon; Yagur-Kroll, Sharon; Zohar, Cheinat; Rabinovitz, Zahi; Nussinovitch, Amos; Kabessa, Yossi; Agranat, Aharon J.

    2017-06-01

    Current landmine detection methodologies are not much different in principle from those employed 75 years ago, in that they require actual presence in the minefield, with obvious risks to personnel and equipment. Other limitations include an extremely large ratio of false positives, as well as a very limited ability to detect non-metallic landmines. In this lecture a microbial-based solution for the remote detection of buried landmines described. The small size requirements, rapid responses and sensing versatility of bacterial bioreporters allow their integration into diverse types of devices, for laboratory as well as field applications. The relative ease by which molecular sensing and reporting elements can be fused together to generate dose-dependent quantifiable physical (luminescent, fluorescent, colorimetric, electrochemical) responses to pre-determined conditions allows the construction of diverse classes of sensors. Over the last two decades we and others have employed this principle to design and construct microbial bioreporter strains for the sensitive detection of (a) specific chemicals of environmental concern (heavy metals, halogenated organics etc.) or (b) their deleterious biological effects on living systems (such as toxicity or genotoxicity). In many of these cases, additional molecular manipulations beyond the initial sensor-reporter fusion may be highly beneficial for enhancing the performance of the engineered sensor systems. This presentation highlights several of the approaches we have adopted over the years to achieve this aim, while focusing on the application of live cell microbeads for the remote detection of buried landmines and other explosive devices.

  20. Developing a Predictive Capability for Bioluminescence Signatures

    Science.gov (United States)

    2012-09-30

    stimulated bioluminescence in primarily laminar flows (Latz et al. 1994; Latz et al. 2004; Latz and Rohr 1999; Maldonado and Latz 2007). The primary...M. I., A. R. Juhl, A. M. Ahmed, S. E. Elghobashi, and J. Rohr . 2004. Hydrodynamic stimulation of dinoflagellate bioluminescence: a computational and...experimental study. J. Exp. Biol. 207: 1941-1951. Latz, M. I., and J. Rohr . 1999. Luminescent response of the red tide dinoflagellate Lingulodinium

  1. Toxicity assessment and modelling of Moringa oleifera seeds in water purification by whole cell bioreporter.

    Science.gov (United States)

    Al-Anizi, Ali Adnan; Hellyer, Maria Theresa; Zhang, Dayi

    2014-06-01

    Moringa oleifera has been used as a coagulation reagent for drinking water purification, especially in developing countries such as Malawi. This research revealed the cytoxicity and genotoxicity of M. oleifera by Acinetobacter bioreporter. The results indicated that significant cytoxicity effects were observed when the powdered M. oleifera seeds concentration is from 1 to 50 mg/L. Through direct contact, ethanolic-water extraction and hexane extraction, the toxic effects of hydrophobic and hydrophilic components in M. oleifera seeds were distinguished. It suggested that the hydrophobic lipids contributed to the dominant cytoxicity, consequently resulting in the dominant genotoxicity in the water-soluble fraction due to limited dissolution when the M. oleifera seeds granule concentration was from 10 to 1000 mg/L. Based on cytoxicity and genotoxicity model, the LC50 and LC90 of M. oleifera seeds were 8.5 mg/L and 300 mg/L respectively and their genotoxicity was equivalent to 8.3 mg mitomycin C per 1.0 g dry M. oleifera seed. The toxicity of M. oleifera has also remarkable synergistic effects, suggesting whole cell bioreporter as an appropriate and complementary tool to chemical analysis for environmental toxicity assessment. Copyright © 2014 Elsevier Ltd. All rights reserved.

  2. Whole-cell bioreporters and risk assessment of environmental pollution: A proof-of-concept study using lead.

    Science.gov (United States)

    Zhang, Xiaokai; Qin, Boqiang; Deng, Jianming; Wells, Mona

    2017-10-01

    As the world burden of environmental contamination increases, it is of the utmost importance to develop streamlined approaches to environmental risk assessment in order to prioritize mitigation measures. Whole-cell biosensors or bioreporters and speciation modeling have both become of increasing interest to determine the bioavailability of pollutants, as bioavailability is increasingly in use as an indicator of risk. Herein, we examine whether bioreporter results are able to reflect expectations based on chemical reactivity and speciation modeling, with the hope to extend the research into a wider framework of risk assessment. We study a specific test case concerning the bioavailability of lead (Pb) in aqueous environments containing Pb-complexing ligands. Ligands studied include ethylene diamine tetra-acetic acid (EDTA), meso-2,3 dimercaptosuccinic acid (DMSA), leucine, methionine, cysteine, glutathione, and humic acid (HA), and we also performed experiments using natural water samples from Lake Tai (Taihu), the third largest lake in China. We find that EDTA, DMSA, cysteine, glutathione, and HA amendment significantly reduced Pb bioavailability with increasing ligand concentration according to a log-sigmoid trend. Increasing dissolved organic carbon in Taihu water also had the same effect, whereas leucine and methionine had no notable effect on bioavailability at the concentrations tested. We find that bioreporter results are in accord with the reduction of aqueous Pb 2+ that we expect from the relative complexation affinities of the different ligands tested. For EDTA and HA, for which reasonably accurate ionization and complexation constants are known, speciation modeling is in agreement with bioreporter response to within the level of uncertainty recognised as reasonable by the United States Environmental Protection Agency for speciation-based risk assessment applications. These findings represent a first step toward using bioreporter technology to streamline

  3. Construction of a self- luminescent cyanobacterial bioreporter that detects a broad range of bioavailable heavy metals in aquatic environments

    Directory of Open Access Journals (Sweden)

    Keila eMartin-Betancor

    2015-03-01

    Full Text Available A self-luminescent bioreporter strain of the unicellular cyanobacterium Synechococcus sp. PCC 7942 was constructed by fusing the promoter region of the smt locus (encoding the transcriptional repressor SmtB and the metallothionein SmtA to luxCDABE from Photorhabdus luminescens; the sensor smtB gene controlling the expression of smtA was cloned in the same vector. The bioreporter performance was tested with a range of heavy metals and was shown to respond linearly to divalent Zn, Cd, Cu, Co, Hg and monovalent Ag. Chemical modelling was used to link bioreporter response with metal speciation and bioavailability. Limits of Detection (LODs, Maximum Permissive Concentrations (MPCs and dynamic ranges for each metal were calculated in terms of free ion concentrations. The ranges of detection varied from 11 to 72 pM for Hg2+ (the ion to which the bioreporter was most sensitive to 1.54-5.35 µM for Cd2+ with an order of decreasing sensitivity as follows: Hg2+ >> Cu2+ >> Ag+ > Co2+ ≥ Zn2+ > Cd2+. However, the maximum induction factor reached 75-fold in the case of Zn2+ and 56-fold in the case of Cd2+, implying that Zn2+ is the preferred metal in vivo for the SmtB sensor, followed by Cd2+, Ag+ and Cu2+ (around 45-50-fold induction, Hg2+ (30-fold and finally Co2+ (20-fold. The bioreporter performance was tested in real environmental samples with different water matrix complexity artificially contaminated with increasing concentrations of Zn, Cd, Ag and Cu, confirming its validity as a sensor of free heavy metal cations bioavailability in aquatic environments.

  4. Bioluminescence patterns among North American Armillaria species.

    Science.gov (United States)

    Mihail, Jeanne D

    2015-06-01

    Bioluminescence is widely recognized among white-spored species of Basidiomycota. Most reports of fungal bioluminescence are based upon visual light perception. When instruments such as photomultipliers have been used to measure fungal luminescence, more taxa have been discovered to produce light, albeit at a range of magnitudes. The present studies were undertaken to determine the prevalence of bioluminescence among North American Armillaria species. Consistent, constitutive bioluminescence was detected for the first time for mycelia of Armillaria calvescens, Armillaria cepistipes, Armillaria gemina, Armillaria nabsnona, and Armillaria sinapina and confirmed for mycelia of Armillaria gallica, Armillaria mellea, Armillaria ostoyae, and Armillaria tabescens. Emission spectra of mycelia representing all species had maximum intensity in the range 515-525 nm confirming that emitted light was the result of bioluminescence rather than chemiluminescence. Time series analysis of 1000 consecutive luminescence measurements revealed a highly significant departure from random variation. Mycelial luminescence of eight species exhibited significant, stable shifts in magnitude in response to a series of mechanical disturbance treatments, providing one mechanism for generating observed luminescence variation. Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  5. Near-infrared fluorescence imaging as an alternative to bioluminescent bacteria to monitor biomaterial-associated infections.

    Science.gov (United States)

    Dinjaski, Nina; Suri, Shalu; Valle, Jaione; Lehman, Susan M; Lasa, Iñigo; Prieto, María Auxiliadora; García, Andrés J

    2014-07-01

    Biomaterial-associated infection is one of the most common complications related to the implantation of any biomedical device. Several in vivo imaging platforms have emerged as powerful diagnostic tools to longitudinally monitor biomaterial-associated infections in small animal models. In this study, we directly compared two imaging approaches: bacteria engineered to produce luciferase to generate bioluminescence and reactive oxygen species (ROS) imaging of the inflammatory response associated with the infected implant. We performed longitudinal imaging of bioluminescence associated with bacteria strains expressing plasmid-integrated luciferase driven by different promoters or a strain with the luciferase gene integrated into the chromosome. These luminescent strains provided an adequate signal for acute (0-4 days) monitoring of the infection, but the bioluminescence signal decreased over time and leveled off at 7 days post-implantation. This loss in the bioluminescence signal was attributed to changes in the metabolic activity of the bacteria. In contrast, near-infrared fluorescence imaging of ROS associated with inflammation to the implant provided sensitive and dose-dependent signals of biomaterial-associated bacteria. ROS imaging exhibited higher sensitivity than the bioluminescence imaging and was independent of the bacteria strain. Near-infrared fluorescence imaging of inflammatory responses represents a powerful alternative to bioluminescence imaging for monitoring biomaterial-associated bacterial infections. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  6. In vivo cell tracking with bioluminescence imaging

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Jung Eun; Kalimuthu, Senthilkumar; Ahn, Byeong Cheol [Dept. of Nuclear Medicine, Kyungpook National University School of Medicine and Hospital, Daegu (Korea, Republic of)

    2015-03-15

    Molecular imaging is a fast growing biomedical research that allows the visual representation, characterization and quantification of biological processes at the cellular and subcellular levels within intact living organisms. In vivo tracking of cells is an indispensable technology for development and optimization of cell therapy for replacement or renewal of damaged or diseased tissue using transplanted cells, often autologous cells. With outstanding advantages of bioluminescence imaging, the imaging approach is most commonly applied for in vivo monitoring of transplanted stem cells or immune cells in order to assess viability of administered cells with therapeutic efficacy in preclinical small animal models. In this review, a general overview of bioluminescence is provided and recent updates of in vivo cell tracking using the bioluminescence signal are discussed.

  7. Diketopiperazines produced by the halophilic archaeon, Haloterrigena hispanica, activate AHL bioreporters.

    Science.gov (United States)

    Tommonaro, Giuseppina; Abbamondi, Gennaro Roberto; Iodice, Carmine; Tait, Karen; De Rosa, Salvatore

    2012-04-01

    The generic term "quorum sensing" has been adopted to describe the bacterial cell-to-cell communication mechanism which coordinates gene expression when the population has reached a high cell density. Quorum sensing depends on the synthesis of small molecules that diffuse in and out of bacterial cells. There are few reports about this mechanism in Archaea. We report the isolation and chemical characterization of small molecules belonging to class of diketopiperazines (DKPs) in Haloterrigena hispanica, an extremely halophilic archaeon. One of the DKPs isolated, the compound cyclo-(L-prolyl-L-valine) activated N-acyl homoserine lactone (AHL) bioreporters, indicating that Archaea may have the ability to interact with AHL-producing bacteria within mixed communities.

  8. Chemistry and biology of insect bioluminescence

    International Nuclear Information System (INIS)

    Colepicolo Neto, P.; Bechara, E.J.H.

    1984-01-01

    Basic aspects on the Chemistry and Biology of bioluminescence are reviewed, with emphasis on insects. Data from the investigation of Lampyridae (fireflies) are collected from literature. With regard to Elateridae (click beetles) and Phengodidae (rail road worms), the least explored families of luminescent insects, new data are presented on the following aspects: (i) 'in vivo' emission spectra, (ii) chemical nature of the luciferin, (iii) conection between bioluminescence and 'oxygen toxicity' as a result of molecular oxygen storage and (iv) the role of light emission by larvae and pupae. (Author) [pt

  9. Pseudomonas fluorescens HK44: Lessons Lerned from a Model Whole-Cell Bioreporter with a Broad Application History

    Czech Academy of Sciences Publication Activity Database

    Trögl, J.; Chauhan, A.; Ripp, S.; Layton, A.C.; Kuncová, Gabriela; Sayler, G.S.

    2012-01-01

    Roč. 12, č. 2 (2012), s. 1544-1571 ISSN 1424-8220 R&D Projects: GA MŠk ME 892; GA MŠk ME 893 Grant - others:USDA(US) 2009-39210-20230 Institutional research plan: CEZ:AV0Z40720504 Keywords : bioluminiscence * bioreporter * biosensors Subject RIV: EI - Biotechnology ; Bionics Impact factor: 1.953, year: 2012

  10. Bioluminescent hydrocarbonclastic bacteria of the Niger Delta

    African Journals Online (AJOL)

    Administrator

    2007-02-19

    Feb 19, 2007 ... Utilization of three petroleum hydrocarbons (Mobil SAE 40 Engine Oil, Diesel and Bonny light Crude. Oil) by four ... growth of hydrocarbonoclastic bioluminescent bacteria which could serve as a potential tool for the remediation of petroleum ... lized TNT. In the Niger Delta, increasing petroleum exploration.

  11. Bioluminescence emissions of the firefly Luciola praeusta ...

    Indian Academy of Sciences (India)

    Prakash

    for the emission of light is much faster than was previously believed. [Barua A G, Hazarika S, Saikia N M and Baruah G D 2009 Bioluminescence emissions of the firefly Luciola praeusta Kiesenwetter 1874(Coleoptera : Lampyridae : Luciolinae); J. Biosci. 34 287–292]. Keywords. Firefly; emission spectrum; FWHM; ...

  12. Bioluminescence lights the way to food safety

    Science.gov (United States)

    Brovko, Lubov Y.; Griffiths, Mansel W.

    2003-07-01

    The food industry is increasingly adopting food safety and quality management systems that are more proactive and preventive than those used in the past which have tended to rely on end product testing and visual inspection. The regulatory agencies in many countries are promoting one such management tool, Hazard Analysis Critical Control Point (HACCP), as a way to achieve a safer food supply and as a basis for harmonization of trading standards. Verification that the process is safe must involve microbiological testing but the results need not be generated in real-time. Of all the rapid microbiological tests currently available, the only ones that come close to offering real-time results are bioluminescence-based methods. Recent developments in application of bioluminescence for food safety issues are presented in the paper. These include the use of genetically engineered microorganisms with bioluminescent and fluorescent phenotypes as a real time indicator of physiological state and survival of food-borne pathogens in food and food processing environments as well as novel bioluminescent-based methods for rapid detection of pathogens in food and environmental samples. Advantages and pitfalls of the methods are discussed.

  13. Bioluminescent hydrocarbonclastic bacteria of the Niger Delta ...

    African Journals Online (AJOL)

    Utilization of three petroleum hydrocarbons (Mobil SAE 40 Engine Oil, Diesel and Bonny light Crude Oil) by four bioluminescent bacteria (Vibrio harveyi, V. fisheri, Photobacterium leiognathi and P. Phosphoreum isolated from the Bonny estuary in the Niger Delta, Nigeria was investigated. Microbial utilization was monitored ...

  14. Automatic Formation of Hypotheses on the Relationships between Structure of Naphthalene Analogs and Bioluminescence Response of Bioreporter Pseudomonas fluorescens HK44

    Czech Academy of Sciences Publication Activity Database

    Trögl, J.; Hálová, J.; Kuncová, Gabriela; Pařík, P.

    2010-01-01

    Roč. 55, č. 5 (2010), s. 411-417 ISSN 0015-5632 R&D Projects: GA MŠk OC 121 Institutional research plan: CEZ:AV0Z40720504 Keywords : substituted naphthalenes * bioconversion * optical biosensor Subject RIV: CE - Biochemistry Impact factor: 0.977, year: 2010

  15. Isolation and development of bioluminescent reporter phages for bacterial dysentery.

    Science.gov (United States)

    Schofield, D A; Wray, D J; Molineux, I J

    2015-02-01

    Shigellosis is a significant cause of morbidity and mortality worldwide, most notably amongst children. Moreover, there is a global increase in the occurrence of multidrug-resistant isolates, including the epidemic and pandemic Shigella dysenteriae type 1 strain. We developed a bioluminescent reporter phage assay to facilitate detection and simultaneously determine antibiotic susceptibility. A Shigella flexneri phage (Shfl25875) was isolated from environmental wastewater and characterized by DNA sequencing. Shfl25875 is T4-like, harbors a 169,062-bp genome, and grows on most (28/29) S. flexneri strains and all 12 S. dysenteriae type 1 strains tested. The genes encoding bacterial luciferase were integrated into the Shfl25875 genome to create a "light-tagged" phage capable of transducing a bioluminescent phenotype to infected cells. Shfl25875::luxAB rapidly detects cultured isolates with high sensitivity. Specificity experiments indicate that the reporter does not respond to Shigella boydii, non-type 1 S. dysenteriae strains, and most non-Shigella Enterobacteriaceae. Shfl25875::luxAB generates ampicillin and ciprofloxacin susceptibility profiles that are similar to the standard Clinical and Laboratory Standards Institute (CLSI) growth microdilution method, but in a significantly shorter time. In addition, the reporter phage detects Shigella in mock-infected stool. This new reporter phage shows promise as a tool for the detection of cultured isolates or complex clinical samples.

  16. Integrated Heterodyne MOEMS for detection of low intensity signals

    Science.gov (United States)

    Elman, Noel M.; Krylov, Slava; Sternheim, Marek; Shacham-Diamand, Yosi

    2006-01-01

    A novel MEMS-based modulation scheme is presented as a method to enhance the signal-to-noise ratio (SNR) of silicon photodiodes adapted for the detection of light-emitting bio-reporter signals. Photodiodes are an attractive photodetector choice because they are VLSI compatible, easily miniaturized, highly scalable, and inexpensive. Silicon photodiodes exhibit a wide response range extending from the ultraviolet (UV) to the near infrared (IR) part of the spectrum, which in principle is appropriate for sensing low intensity optical signals. Silicon photodiodes, however, exhibit limited sensitivity to optical dc signals, as the magnitude of the low frequency noise is comparable to signal magnitude. Optical modulation prior to photodetection overcomes the inherent low frequency noise of photodetectors and system detection circuits. The enhancement scheme is based on a design of high frequency optical modulators that operate in the 1-2 kHz range in order to overcome the low frequency spectral noise. We have denominated this MEMS-based scheme Integrated Heterodyne Optical System (IHOS). The modulation efficiency of the proposed architecture can reach up to 50 percent. In order to implement the MOEMS optical modulators, a new two-mask fabrication process was developed that combines high-aspect ratio and low aspect ratio structures at the same device layer (aspect ratio is defined as a ratio between the structure height to its width). Long stroke electrostatic combdrive actuators integrated with folded flexures (high aspect-ratio) were fabricated together to drive large aperture shutters (low aspect ratio). We have denominated this process MASIS (Multiple Aspect Ratio Structural Integration). Under resonant excitation at approximately 1 kHz, MOEMS modulators demonstrated maximum displacement of about 40 microns at an actuation voltage of 15 V peak in air, and 3.5 V peak in vacuum (8 mTorr). Results of analytical solutions and finite element analysis (FEA) simulations are

  17. Use of Tunable Whole-Cell Bioreporters to Assess Bioavailable Cadmium and Remediation Performance in Soils.

    Directory of Open Access Journals (Sweden)

    Youngdae Yoon

    Full Text Available It is important to have tools to measure the bioavailability to assess the risks of pollutants because the bioavailability is defined as the portions of pollutants showing the biological effects on living organisms. This study described the construction of tunable Escherichia coli whole-cell bioreporter (WCB using the promoter region of zinc-inducible operon and its application on contaminated soils. It was verified that this WCB system showed specific and sensitive responses to cadmium rather than zinc in the experimental conditions. It was inferred that Cd(II associates stronger with ZntR, a regulatory protein of zinc-inducible operon, than other metal ions. Moreover, the expression of reporter genes, egfp and mcherry, were proportional to the concentration of cadmium, thereby being a quantitative sensor to monitor bioavailable cadmium. The capability to determine bioavailable cadmium was verified with Cd(II amended LUFA soils, and then the applicability on environmental systems was investigated with field soils collected from smelter area in Korea before and after soil-washing. The total amount of cadmium was decreased after soil washing, while the bioavailability was increased. Consequently, it would be valuable to have tools to assess bioavailability and the effectiveness of soil remediation should be evaluated in the aspect of bioavailability as well as removal efficiency.

  18. Use of Tunable Whole-Cell Bioreporters to Assess Bioavailable Cadmium and Remediation Performance in Soils

    Science.gov (United States)

    Yoon, Youngdae; Kim, Sunghoon; Chae, Yooeun; Kang, Yerin; Lee, Youngshim; Jeong, Seung-Woo; An, Youn-Joo

    2016-01-01

    It is important to have tools to measure the bioavailability to assess the risks of pollutants because the bioavailability is defined as the portions of pollutants showing the biological effects on living organisms. This study described the construction of tunable Escherichia coli whole-cell bioreporter (WCB) using the promoter region of zinc-inducible operon and its application on contaminated soils. It was verified that this WCB system showed specific and sensitive responses to cadmium rather than zinc in the experimental conditions. It was inferred that Cd(II) associates stronger with ZntR, a regulatory protein of zinc-inducible operon, than other metal ions. Moreover, the expression of reporter genes, egfp and mcherry, were proportional to the concentration of cadmium, thereby being a quantitative sensor to monitor bioavailable cadmium. The capability to determine bioavailable cadmium was verified with Cd(II) amended LUFA soils, and then the applicability on environmental systems was investigated with field soils collected from smelter area in Korea before and after soil-washing. The total amount of cadmium was decreased after soil washing, while the bioavailability was increased. Consequently, it would be valuable to have tools to assess bioavailability and the effectiveness of soil remediation should be evaluated in the aspect of bioavailability as well as removal efficiency. PMID:27171374

  19. Stimulated bioluminescence by fluid shear stress associated with pipe flow

    Energy Technology Data Exchange (ETDEWEB)

    Cao Jing; Wang Jiangan; Wu Ronghua, E-mail: caojing981@126.com [Col. of Electronic Eng., Naval University of Engineering, Wuhan 430033 (China)

    2011-01-01

    Dinoflagellate can be stimulated bioluminescence by hydrodynamic agitation. Two typical dinoflagellate (Lingulodinium polyedrum and Pyrocystis noctiluca) was choosed to research stimulated bioluminescence. The bioluminescence intensity and shear stress intensity were measured using fully developed pipe flow. There is shear stress threshold to agitate organism bioluminescence. From these experiment, the response thresholds of the stimulated bioluminscence always occurred in laminar flows at a shear stress level of 0.6-3 dyn/cm{sup 2}. At the same time, the spectral characteristc of dinoflagellate was recorded, the wavelength of them is about 470nm, and the full width at half maximum is approximate 30nm.

  20. Nanostructured biosensor using bioluminescence quenching technique for glucose detection.

    Science.gov (United States)

    Chen, Longyan; Chen, Longyi; Dotzert, Michelle; Melling, C W James; Zhang, Jin

    2017-08-22

    Most methods for monitoring glucose level require an external energy source which may limit their application, particularly in vivo test. Bioluminescence technique offers an alternative way to provide emission light without external energy source by using bioluminescent proteins found from firefly or marine vertebrates and invertebrates. For quick and non-invasive detection of glucose, we herein developed a nanostructured biosensor by applying the bioluminescence technique. Luciferase bioluminescence protein (Rluc) is conjugated with β-cyclodextrin (β-CD). The bioluminescence intensity of Rluc can be quenched by 8 ± 3 nm gold nanoparticles (Au NPs) when Au NPs covalently bind to β-CD. In the presence of glucose, Au NPs are replaced and leave far from Rluc through a competitive reaction, which results in the restored bioluminescence intensity of Rluc. A linear relationship is observed between the restored bioluminescence intensity and the logarithmic glucose concentration in the range of 1-100 µM. In addition, the selectivity of this designed sensor has been evaluated. The performance of the senor for determination of the concentration of glucose in the blood of diabetic rats is studied for comparison with that of the concentration of glucose in aqueous. This study demonstrates the design of a bioluminescence sensor for quickly detecting the concentration of glucose sensitively.

  1. Use of bioreporters and deletion mutants reveals ionic silver and ROS to be equally important in silver nanotoxicity.

    Science.gov (United States)

    Joshi, Nimisha; Ngwenya, Bryne T; Butler, Ian B; French, Chris E

    2015-04-28

    The mechanism of antibacterial action of silver nanoparticles (AgNp) was investigated by employing a combination of microbiology and geochemical approaches to contribute to the realistic assessment of nanotoxicity. Our studies showed that suspending AgNp in media with different levels of chloride relevant to environmental conditions produced low levels of ionic silver thereby suggesting that dissolution of silver ions from nanoparticulate surface could not be the sole mechanism of toxicity. An Escherichia coli based bioreporter strain responsive to silver ions together with mutant strains of E. coli lacking specific protective systems were tested against AgNp. Deletion mutants lacking silver ion efflux systems and resistance mechanisms against oxidative stress showed an increased sensitivity to AgNp. However, the bioreporter did not respond to silver nanoparticles. Our results suggest that oxidative stress is a major toxicity mechanism and that this is at least partially associated with ionic silver, but that bulk dissolution of silver into the medium is not sufficient to account for the observed effects. Chloride ions do not appear to offer significant protection, indicating that chloride in receiving waters will not necessarily protect environmental bacteria from the toxic effects of nanoparticles in effluents. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Assessment of tumor energy and oxygenation status by bioluminescence, nuclear magnetic resonance spectroscopy, and cryospectrophotometry.

    Science.gov (United States)

    Mueller-Klieser, W; Schaefer, C; Walenta, S; Rofstad, E K; Fenton, B M; Sutherland, R M

    1990-03-15

    The energy and oxygenation status of tumors from two murine sarcoma lines (KHT, RIF-1) and two human ovarian carcinoma xenograft lines (MLS, OWI) were assessed using three independent techniques. Tumor energy metabolism was investigated in vivo by 31P nuclear magnetic resonance spectroscopy. After nuclear magnetic resonance measurements, tumors were frozen in liquid nitrogen to determine the tissue ATP concentration by imaging bioluminescence and to register the intracapillary oxyhemoglobin (HbO2) saturation using the cryospectrophotometric method. There was a positive correlation between the nucleoside triphosphate beta/total resonance ratio or a negative correlation between the Pi/total resonance ratio and the model ATP concentration obtained by bioluminescence, respectively. This was true for small tumors with no extended necrosis irrespective of tumor type. Moreover, a positive correlation was obtained between the HbO2 saturations and the ATP concentration measured with bioluminescence. The results demonstrate the potential of combined studies using noninvasive, integrating methods and high-resolution imaging techniques for characterizing the metabolic milieu in tumors.

  3. Bioluminescence Assays for Monitoring Chondrogenic Differentiation and Cartilage Regeneration

    Directory of Open Access Journals (Sweden)

    Hyeon Jeong Je

    2017-06-01

    Full Text Available Since articular cartilage has a limited regeneration potential, for developing biological therapies for cartilage regeneration it is important to study the mechanisms underlying chondrogenesis of stem cells. Bioluminescence assays can visualize a wide range of biological phenomena such as gene expression, signaling, metabolism, development, cellular movements, and molecular interactions by using visible light and thus contribute substantially to elucidation of their biological functions. This article gives a concise review to introduce basic principles of bioluminescence assays and applications of the technology to visualize the processes of chondrogenesis and cartilage regeneration. Applications of bioluminescence assays have been highlighted in the methods of real-time monitoring of gene expression and intracellular levels of biomolecules and noninvasive cell tracking within animal models. This review suggests that bioluminescence assays can be applied towards a visual understanding of chondrogenesis and cartilage regeneration.

  4. Bioluminescence imaging of Chlamydia muridarum ascending infection in mice.

    Directory of Open Access Journals (Sweden)

    Jessica Campbell

    Full Text Available Chlamydial pathogenicity in the upper genital tract relies on chlamydial ascending from the lower genital tract. To monitor chlamydial ascension, we engineered a luciferase-expressing C. muridarum. In cells infected with the luciferase-expressing C. muridarum, luciferase gene expression and enzymatic activity (measured as bioluminescence intensity correlated well along the infection course, suggesting that bioluminescence can be used for monitoring chlamydial replication. Following an intravaginal inoculation with the luciferase-expressing C. muridarum, 8 of 10 mice displayed bioluminescence signal in the lower with 4 also in the upper genital tracts on day 3 after infection. By day 7, all 10 mice developed bioluminescence signal in the upper genital tracts. The bioluminescence signal was maintained in the upper genital tract in 6 and 2 mice by days 14 and 21, respectively. The bioluminescence signal was no longer detectable in any of the mice by day 28. The whole body imaging approach also revealed an unexpected airway infection following the intravaginal inoculation. Although the concomitant airway infection was transient and did not significantly alter the genital tract infection time courses, caution should be taken during data interpretation. The above observations have demonstrated that C. muridarum can not only achieve rapid ascending infection in the genital tract but also cause airway infection following a genital tract inoculation. These findings have laid a foundation for further optimizing the C. muridarum intravaginal infection murine model for understanding chlamydial pathogenic mechanisms.

  5. Estrogenic Activities of Food Supplements and Beers as Assessed by a Yeast Bioreporter Assay.

    Science.gov (United States)

    Omoruyi, Iyekhoetin Matthew; Pohjanvirta, Raimo

    2017-10-31

    Mounting evidence of the effects of endocrine-disrupting chemicals (EDCs) in humans has led to assaying a vast array of food items (processed or packaged) as possible sources of human exposure to estrogens. In this study, we investigated the current situation in this respect of different food supplements and beer brands. Eleven food supplements and 24 beer brands were obtained from Helsinki, Finland. Sample preparation was carried out by established methods while estrogenic activities were assessed by a yeast bioluminescent assay, using two recombinant yeast strains (Saccharomyces cerevisiae BMAEREluc/ERα and S. cerevisiae BMA64/luc). All the food supplements as well as 81% of the beer samples tested were found to be estrogenic, with estradiol equivalent concentrations of food supplements and beer brands ranging from 7.5 to 11.5 µg/ml and from below detection limits to 43.6 ng/ml, respectively. The estrogenic activities detected in beer samples were not dependent on the beer's alcoholic content, the country of production, or the size of the production brewery. The results of our study imply that both food supplements and beers can be a significant source of human exposure to estrogens. Therefore, further studies and regular surveillance are warranted.

  6. Bioluminescence in vivo imaging of autoimmune encephalomyelitis predicts disease

    Directory of Open Access Journals (Sweden)

    Steinman Lawrence

    2008-02-01

    Full Text Available Abstract Background Experimental autoimmune encephalomyelitis is a widely used animal model to understand not only multiple sclerosis but also basic principles of immunity. The disease is scored typically by observing signs of paralysis, which do not always correspond with pathological changes. Methods Experimental autoimmune encephalomyelitis was induced in transgenic mice expressing an injury responsive luciferase reporter in astrocytes (GFAP-luc. Bioluminescence in the brain and spinal cord was measured non-invasively in living mice. Mice were sacrificed at different time points to evaluate clinical and pathological changes. The correlation between bioluminescence and clinical and pathological EAE was statistically analyzed by Pearson correlation analysis. Results Bioluminescence from the brain and spinal cord correlates strongly with severity of clinical disease and a number of pathological changes in the brain in EAE. Bioluminescence at early time points also predicts severity of disease. Conclusion These results highlight the potential use of bioluminescence imaging to monitor neuroinflammation for rapid drug screening and immunological studies in EAE and suggest that similar approaches could be applied to other animal models of autoimmune and inflammatory disorders.

  7. Bacterial Bioluminescence: Spectral Study of the Emitters in the In Vivo Reaction

    NARCIS (Netherlands)

    Matheson, I.B.C.; Lee, J.; Muller, F.

    1981-01-01

    Transient fluorescent species are observed in the bioluminescent reactions of three reduced flavin mononucleotides with aliphatic aldehydes and oxygen, catalyzed by bacterial luciferase. In each case the fluorescence spectral distribution is similar to that of the bioluminescence but is readily

  8. Hv 1 Proton Channels in Dinoflagellates: Not Just for Bioluminescence?

    Science.gov (United States)

    Kigundu, Gabriel; Cooper, Jennifer L; Smith, Susan M E

    2018-04-26

    Bioluminescence in dinoflagellates is controlled by H V 1 proton channels. Database searches of dinoflagellate transcriptomes and genomes yielded hits with sequence features diagnostic of all confirmed H V 1, and show that H V 1 is widely distributed in the dinoflagellate phylogeny including the basal species Oxyrrhis marina. Multiple sequence alignments followed by phylogenetic analysis revealed three major subfamilies of H V 1 that do not correlate with presence of theca, autotrophy, geographic location, or bioluminescence. These data suggest that most dinoflagellates express a H V 1 which has a function separate from bioluminescence. Sequence evidence also suggests that dinoflagellates can contain more than one H V 1 gene. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  9. Influence of antibiotic pressure on bacterial bioluminescence, with emphasis on Staphylococcus aureus

    NARCIS (Netherlands)

    Daghighi, Seyedmojtaba; Sjollema, Jelmer; Harapanahalli, Akshay; Dijkstra, Rene J. B.; van der Mei, Henny C.; Busscher, Henk J.

    2015-01-01

    Bioluminescence imaging is used for longitudinal evaluation of bacteria in live animals. Clear relations exist between bacterial numbers and their bioluminescence. However, bioluminescence images of Staphylococcus aureus Xen29, S. aureus Xen36 and Escherichia coli Xen14 grown on tryptone soy agar in

  10. Repetitive Inductions of Bioluminescence of Pseudomonas putida TVA8 Immobilized by Adsorption on Optical Fibre.

    Czech Academy of Sciences Publication Activity Database

    Zajíc, J.; Bittner, M.; Brányik, T.; Solovyev, Andrey; Šabata, Stanislav; Kuncová, Gabriela; Pospíšilová, M.

    2016-01-01

    Roč. 70, č. 7 (2016), s. 877-887 ISSN 0366-6352 Institutional support: RVO:67985858 Keywords : whole-cell biosensor * bioluminiscent bioreporter * optical fibre sensor Subject RIV: CC - Organic Chemistry Impact factor: 1.258, year: 2016

  11. Random matrix-based dimensionality reduction for bioluminescence tomography reconstruction

    Science.gov (United States)

    Styles, Iain B.; Basevi, Hector R. A.; Guggenheim, James A.; Dehghani, Hamid

    2013-06-01

    We show how a random matrix can be used to reduce the dimensionality of the bioluminescence tomography reconstruction problem. A randomised low-rank approximation for the sensitivity matrix is computed, and we show how this can be used to reconstruct the bioluminescence source distribution on a randomised basis for the mesh nodes. The distribution on the original mesh can be found easily via a simple matrix multiplication. The majority of the computation required can be performed in advance of the reconstruction, and the reconstruction time itself is of the order milliseconds. This could allow for high frame rate real-time reconstructions to be performed.

  12. Feasibility Study for a Compact, Multi-Purpose Bioluminescence Detector

    Science.gov (United States)

    1998-09-30

    Symposium on Bioluminescence and Chemiluminescence. Eds. JW Hastings, LJ Kricka and PE Stanley. John Wiley & Sons Ltd, Sussex, UK. pp. 159-164...Lowenstine, M.R. Bowlby , and D.P. Cook. (1993) A new large volume bioluminescence bathyphotometer with defined turbulence excitation. Deep Sea Res. 40...and PE Stanley. John Wiley & Sons Ltd, Sussex, UK. pp. 159-164. Makemson, J.C., N.R. Fulayfil, W.L. Landry, L.M. Van Ert, C.F. Wimpee, E.A. Widder

  13. In vivo bioluminescence imaging using orthotopic xenografts towards patient's derived-xenograft Medulloblastoma models.

    Science.gov (United States)

    Asadzadeh, Fatemeh; Ferrucci, Veronica; DE Antonellis, Pasqualino; Zollo, Massimo

    2017-03-01

    Medulloblastoma is a cerebellar neoplasia of the central nervous system. Four molecular subgrups have been identified (MBWNT, MBSHH, MBgroup3 and MBgroup4) with distinct genetics and clinical outcome. Among these, MBgroup3-4 are highly metastatic with the worst prognosis. The current standard therapy includes surgery, radiation and chemotherapy. Thus, specific treatments adapted to cure those different molecular subgroups are needed. The use of orthotopic xenograft models, together with the non-invasive in vivo biolumiscence imaging (BLI) technology, is emerging during preclinical studies to test novel therapeutics for medulloblastoma treatment. Orthotopic MB xenografts were performed by injection of Daoy-luc cells, that had been previously infected with lentiviral particles to stably express luciferase gene, into the fourth right ventricle of the cerebellum of ten nude mice. For the implantation, specific stereotactic coordinates were used. Seven days after the implantation the mice were imaged by acquisitions of bioluminescence imaging (BLI) using IVIS 3D Illumina Imaging System (Xenogen). Tumor growth was evaluated by quantifying the bioluminescence signals using the integrated fluxes of photons within each area of interest using the Living Images Software Package 3.2 (Xenogen-Perkin Elmer). Finally, histological analysis using hematoxylin-eosin staining was performed to confirm the presence of tumorigenic cells into the cerebellum of the mice. We describe a method to use the in vivo bioluminescent imaging (BLI) showing the potential to be used to investigate the potential antitumorigenic effects of a drug for in vivo medulloblastoma treatment. We also discuss other studies in which this technology has been applied to obtain a more comprehensive knowledge of medulloblastoma using orthotopic xenograft mouse models. There is a need to develop patient's derived-xenograft (PDX) model systems to test novel drugs for medulloblastoma treatment within each molecular sub

  14. Bioluminescent system for dynamic imaging of cell and animal behavior

    Energy Technology Data Exchange (ETDEWEB)

    Hara-Miyauchi, Chikako [Department of Physiology, Keio University School of Medicine, Tokyo 160-8582 (Japan); Laboratory for Cell Function Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198 (Japan); Department of Biophysics and Biochemistry, Graduate School of Health Care Sciences, Tokyo Medical and Dental University, Tokyo 113-8510 (Japan); Tsuji, Osahiko [Department of Physiology, Keio University School of Medicine, Tokyo 160-8582 (Japan); Department of Orthopedic Surgery, Keio University School of Medicine, Tokyo 160-8582 (Japan); Hanyu, Aki [Division of Biochemistry, The Cancer Institute of the Japanese Foundation for Cancer Research, Tokyo 135-8550 (Japan); Okada, Seiji [Department of Advanced Medical Initiatives, Faculty of Medical Sciences, Kyushu University, Fukuoka 812-8582 (Japan); Yasuda, Akimasa [Department of Physiology, Keio University School of Medicine, Tokyo 160-8582 (Japan); Department of Orthopedic Surgery, Keio University School of Medicine, Tokyo 160-8582 (Japan); Fukano, Takashi [Laboratory for Cell Function Dynamics, Brain Science Institute, RIKEN, Saitama 351-0198 (Japan); Akazawa, Chihiro [Department of Biophysics and Biochemistry, Graduate School of Health Care Sciences, Tokyo Medical and Dental University, Tokyo 113-8510 (Japan); Nakamura, Masaya [Department of Orthopedic Surgery, Keio University School of Medicine, Tokyo 160-8582 (Japan); Imamura, Takeshi [Department of Molecular Medicine for Pathogenesis, Ehime University Graduate School of Medicine, Toon, Ehime 791-0295 (Japan); Core Research for Evolutional Science and Technology, The Japan Science and Technology Corporation, Tokyo 135-8550 (Japan); Matsuzaki, Yumi [Department of Physiology, Keio University School of Medicine, Tokyo 160-8582 (Japan); Okano, Hirotaka James, E-mail: hjokano@jikei.ac.jp [Department of Physiology, Keio University School of Medicine, Tokyo 160-8582 (Japan); Division of Regenerative Medicine Jikei University School of Medicine, Tokyo 150-8461 (Japan); and others

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer We combined a yellow variant of GFP and firefly luciferase to make ffLuc-cp156. Black-Right-Pointing-Pointer ffLuc-cp156 showed improved photon yield in cultured cells and transgenic mice. Black-Right-Pointing-Pointer ffLuc-cp156 enabled video-rate bioluminescence imaging of freely-moving animals. Black-Right-Pointing-Pointer ffLuc-cp156 mice enabled tracking real-time drug delivery in conscious animals. -- Abstract: The current utility of bioluminescence imaging is constrained by a low photon yield that limits temporal sensitivity. Here, we describe an imaging method that uses a chemiluminescent/fluorescent protein, ffLuc-cp156, which consists of a yellow variant of Aequorea GFP and firefly luciferase. We report an improvement in photon yield by over three orders of magnitude over current bioluminescent systems. We imaged cellular movement at high resolution including neuronal growth cones and microglial cell protrusions. Transgenic ffLuc-cp156 mice enabled video-rate bioluminescence imaging of freely moving animals, which may provide a reliable assay for drug distribution in behaving animals for pre-clinical studies.

  15. Unanimous Model for Describing the Fast Bioluminescence Kinetics of Ca

    NARCIS (Netherlands)

    Eremeeva, Elena V.; Bartsev, Sergey I.; Berkel, van Willem J.H.; Vysotski, Eugene S.

    2017-01-01

    Upon binding their metal ion cofactors, Ca2+-regulated photoproteins display a rapid increase of light signal, which reaches its peak within milliseconds. In the present study, we investigate bioluminescence kinetics of the entire photoprotein family. All five recombinant hydromedusan Ca2+-regulated

  16. Filtering and deconvolution for bioluminescence imaging of small animals

    International Nuclear Information System (INIS)

    Akkoul, S.

    2010-01-01

    This thesis is devoted to analysis of bioluminescence images applied to the small animal. This kind of imaging modality is used in cancerology studies. Nevertheless, some problems are related to the diffusion and the absorption of the tissues of the light of internal bioluminescent sources. In addition, system noise and the cosmic rays noise are present. This influences the quality of the images and makes it difficult to analyze. The purpose of this thesis is to overcome these disturbing effects. We first have proposed an image formation model for the bioluminescence images. The processing chain is constituted by a filtering stage followed by a deconvolution stage. We have proposed a new median filter to suppress the random value impulsive noise which corrupts the acquired images; this filter represents the first block of the proposed chain. For the deconvolution stage, we have performed a comparative study of various deconvolution algorithms. It allowed us to choose a blind deconvolution algorithm initialized with the estimated point spread function of the acquisition system. At first, we have validated our global approach by comparing our obtained results with the ground truth. Through various clinical tests, we have shown that the processing chain allows a significant improvement of the spatial resolution and a better distinction of very close tumor sources, what represents considerable contribution for the users of bioluminescence images. (author)

  17. Establishment of human cell lines showing circadian rhythms of bioluminescence.

    Science.gov (United States)

    Yoshikawa, Aki; Shimada, Hiroko; Numazawa, Kahori; Sasaki, Tsukasa; Ikeda, Masaaki; Kawashima, Minae; Kato, Nobumasa; Tokunaga, Katsushi; Ebisawa, Takashi

    2008-11-28

    We have established human retinal pigment epithelial cell lines stably expressing the luciferase gene, driven by the human Bmal1 promoter, to obtain human-derived cells that show circadian rhythms of bioluminescence after dexamethasone treatment. The average circadian period of bioluminescence for the obtained clones was 24.07+/-0.48 h. Lithium (10 mM) in the medium significantly lengthened the circadian period of bioluminescence, which is consistent with previous reports, while 2 mM or 5 mM lithium had no effect. This is the first report on the establishment of human-derived cell lines that proliferate infinitely and show circadian rhythms of bioluminescence, and also the first to investigate the effects of low-dose lithium on the circadian rhythms of human-derived cells in vitro. The established cells will be useful for various in vitro studies of human circadian rhythms and for the development of new therapies for human disorders related to circadian rhythm disturbances.

  18. Structure of fungal oxyluciferin, the product of the bioluminescence reaction.

    Science.gov (United States)

    Purtov, K V; Osipova, Z M; Petushkov, V N; Rodionova, N S; Tsarkova, A S; Kotlobay, A A; Chepurnykh, T V; Gorokhovatsky, A Yu; Yampolsky, I V; Gitelson, J I

    2017-11-01

    The structure of fungal oxyluciferin was determined, the enzymatic bioluminescence reaction under substrate saturation conditions with discrete monitoring of formed products was conducted, and the structures of the end products of the reaction were established. On the basis of these studies, the scheme of oxyluciferin degradation to the end products was developed. The structure of fungal oxyluciferin was confirmed by counter synthesis.

  19. Smartphone-based low light detection for bioluminescence application

    Science.gov (United States)

    We report a smartphone-based device and associated imaging-processing algorithm to maximize the sensitivity of standard smartphone cameras, that can detect the presence of single-digit pW of radiant flux intensity. The proposed hardware and software, called bioluminescent-based analyte quantitation ...

  20. Bioluminescent system for dynamic imaging of cell and animal behavior

    International Nuclear Information System (INIS)

    Hara-Miyauchi, Chikako; Tsuji, Osahiko; Hanyu, Aki; Okada, Seiji; Yasuda, Akimasa; Fukano, Takashi; Akazawa, Chihiro; Nakamura, Masaya; Imamura, Takeshi; Matsuzaki, Yumi; Okano, Hirotaka James

    2012-01-01

    Highlights: ► We combined a yellow variant of GFP and firefly luciferase to make ffLuc-cp156. ► ffLuc-cp156 showed improved photon yield in cultured cells and transgenic mice. ► ffLuc-cp156 enabled video-rate bioluminescence imaging of freely-moving animals. ► ffLuc-cp156 mice enabled tracking real-time drug delivery in conscious animals. -- Abstract: The current utility of bioluminescence imaging is constrained by a low photon yield that limits temporal sensitivity. Here, we describe an imaging method that uses a chemiluminescent/fluorescent protein, ffLuc-cp156, which consists of a yellow variant of Aequorea GFP and firefly luciferase. We report an improvement in photon yield by over three orders of magnitude over current bioluminescent systems. We imaged cellular movement at high resolution including neuronal growth cones and microglial cell protrusions. Transgenic ffLuc-cp156 mice enabled video-rate bioluminescence imaging of freely moving animals, which may provide a reliable assay for drug distribution in behaving animals for pre-clinical studies.

  1. Novel rat tail discitis model using bioluminescent Staphylococcus aureus.

    Science.gov (United States)

    Bostian, Phillip A; Karnes, Jonathan M; Cui, Shari; Robinson, Lisa J; Daffner, Scott D; Witt, Michelle R; Emery, Sanford E

    2017-09-01

    Management of spondylodiscitis is a challenging clinical problem requiring medical and surgical treatment strategies. The purpose of this study was to establish a rat model of spondylodiscitis that utilizes bioluminescent Staphylococcus aureus (S. aureus), thus permitting in vivo surveillance of infection intensity. Inocula of the bioluminescent S. aureus strain XEN36 were created in concentrations of 10 2 CFU/0.1 ml, 10 4  CFU/0.1 ml, and 10 6  CFU/0.1 ml. Three groups of rats were injected with the bacteria in the most proximal intervertebral tail segment. The third most proximal tail segment was injected with saline as a control. Bioluminescence was measured at baseline, 3 days, and weekly for a total of 6 weeks. Detected bioluminescence for each group peaked at day 3 and returned to baseline in 21 days. The average intensity was highest for the experimental group injected with the most concentrated bacterial solution (10 6  CFU/0.1 ml). Radiographic analysis revealed loss of intervertebral disc space and evidence of osseous bridging. Saline-injected spaces exhibited no decrease in intervertebral spacing as compared to distal sites. Histologic analysis revealed neutrophilic infiltrates, destruction of the annulus fibrosus and nucleus pulposus, destruction of vertebral endplates, and osseous bridging. Saline-injected discs exhibited preserved annulus fibrosus and nucleus pulposus on histology. This study demonstrates that injection of bioluminescent S. aureus into the intervertebral disc of a rat tail is a viable animal model for spondylodiscitis research. This model allows for real-time, in vivo quantification of infection intensity, which may decrease the number of animals required for infection studies of the intervertebral disc. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:2075-2081, 2017. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

  2. Semi-automated Image Processing for Preclinical Bioluminescent Imaging.

    Science.gov (United States)

    Slavine, Nikolai V; McColl, Roderick W

    Bioluminescent imaging is a valuable noninvasive technique for investigating tumor dynamics and specific biological molecular events in living animals to better understand the effects of human disease in animal models. The purpose of this study was to develop and test a strategy behind automated methods for bioluminescence image processing from the data acquisition to obtaining 3D images. In order to optimize this procedure a semi-automated image processing approach with multi-modality image handling environment was developed. To identify a bioluminescent source location and strength we used the light flux detected on the surface of the imaged object by CCD cameras. For phantom calibration tests and object surface reconstruction we used MLEM algorithm. For internal bioluminescent sources we used the diffusion approximation with balancing the internal and external intensities on the boundary of the media and then determined an initial order approximation for the photon fluence we subsequently applied a novel iterative deconvolution method to obtain the final reconstruction result. We find that the reconstruction techniques successfully used the depth-dependent light transport approach and semi-automated image processing to provide a realistic 3D model of the lung tumor. Our image processing software can optimize and decrease the time of the volumetric imaging and quantitative assessment. The data obtained from light phantom and lung mouse tumor images demonstrate the utility of the image reconstruction algorithms and semi-automated approach for bioluminescent image processing procedure. We suggest that the developed image processing approach can be applied to preclinical imaging studies: characteristics of tumor growth, identify metastases, and potentially determine the effectiveness of cancer treatment.

  3. In vivo bioluminescence imaging of Ca signalling in the brain of Drosophila.

    Directory of Open Access Journals (Sweden)

    Jean-René Martin

    Full Text Available Many different cells' signalling pathways are universally regulated by Ca(2+ concentration [Ca(2+] rises that have highly variable amplitudes and kinetic properties. Optical imaging can provide the means to characterise both the temporal and spatial aspects of Ca(2+ signals involved in neurophysiological functions. New methods for in vivo imaging of Ca(2+ signalling in the brain of Drosophila are required for probing the different dynamic aspects of this system. In studies here, whole brain Ca(2+ imaging was performed on transgenic flies with targeted expression of the bioluminescent Ca(2+ reporter GFP-aequorin (GA in different neural structures. A photon counting based technique was used to undertake continuous recordings of cytosolic [Ca(2+] over hours. Time integrals for reconstructing images and analysis of the data were selected offline according to the signal intensity. This approach allowed a unique Ca(2+ response associated with cholinergic transmission to be identified by whole brain imaging of specific neural structures. Notably, [Ca(2+] transients in the Mushroom Bodies (MBs following nicotine stimulation were accompanied by a delayed secondary [Ca(2+] rise (up to 15 min. later in the MB lobes. The delayed response was sensitive to thapsigargin, suggesting a role for intra-cellular Ca(2+ stores. Moreover, it was reduced in dunce mutant flies, which are impaired in learning and memory. Bioluminescence imaging is therefore useful for studying Ca(2+ signalling pathways and for functional mapping of neurophysiological processes in the fly brain.

  4. Use of a highly sensitive two-dimensional luminescence imaging system to monitor endogenous bioluminescence in plant leaves

    Directory of Open Access Journals (Sweden)

    Flor-Henry Michel

    2004-11-01

    Full Text Available Abstract Background All living organisms emit spontaneous low-level bioluminescence, which can be increased in response to stress. Methods for imaging this ultra-weak luminescence have previously been limited by the sensitivity of the detection systems used. Results We developed a novel configuration of a cooled charge-coupled device (CCD for 2-dimensional imaging of light emission from biological material. In this study, we imaged photon emission from plant leaves. The equipment allowed short integration times for image acquisition, providing high resolution spatial and temporal information on bioluminescence. We were able to carry out time course imaging of both delayed chlorophyll fluorescence from whole leaves, and of low level wound-induced luminescence that we showed to be localised to sites of tissue damage. We found that wound-induced luminescence was chlorophyll-dependent and was enhanced at higher temperatures. Conclusions The data gathered on plant bioluminescence illustrate that the equipment described here represents an improvement in 2-dimensional luminescence imaging technology. Using this system, we identify chlorophyll as the origin of wound-induced luminescence from leaves.

  5. BLProt: Prediction of bioluminescent proteins based on support vector machine and relieff feature selection

    KAUST Repository

    Kandaswamy, Krishna Kumar

    2011-08-17

    Background: Bioluminescence is a process in which light is emitted by a living organism. Most creatures that emit light are sea creatures, but some insects, plants, fungi etc, also emit light. The biotechnological application of bioluminescence has become routine and is considered essential for many medical and general technological advances. Identification of bioluminescent proteins is more challenging due to their poor similarity in sequence. So far, no specific method has been reported to identify bioluminescent proteins from primary sequence.Results: In this paper, we propose a novel predictive method that uses a Support Vector Machine (SVM) and physicochemical properties to predict bioluminescent proteins. BLProt was trained using a dataset consisting of 300 bioluminescent proteins and 300 non-bioluminescent proteins, and evaluated by an independent set of 141 bioluminescent proteins and 18202 non-bioluminescent proteins. To identify the most prominent features, we carried out feature selection with three different filter approaches, ReliefF, infogain, and mRMR. We selected five different feature subsets by decreasing the number of features, and the performance of each feature subset was evaluated.Conclusion: BLProt achieves 80% accuracy from training (5 fold cross-validations) and 80.06% accuracy from testing. The performance of BLProt was compared with BLAST and HMM. High prediction accuracy and successful prediction of hypothetical proteins suggests that BLProt can be a useful approach to identify bioluminescent proteins from sequence information, irrespective of their sequence similarity. 2011 Kandaswamy et al; licensee BioMed Central Ltd.

  6. Microluminometer chip and method to measure bioluminescence

    Science.gov (United States)

    Simpson, Michael L [Knoxville, TN; Paulus, Michael J [Knoxville, TN; Sayler, Gary S [Blaine, TN; Applegate, Bruce M [West Lafayette, IN; Ripp, Steven A [Knoxville, TN

    2008-05-13

    An integrated microluminometer includes an integrated circuit chip having at least one n-well/p-substrate junction photodetector for converting light received into a photocurrent, and a detector on the chip for processing the photocurrent. A distributed electrode configuration including a plurality of spaced apart electrodes disposed on an active region of the photodetector is preferably used to raise efficiency.

  7. Uptake kinetics and biodistribution of C-14-D-luciferin-a radiolabeled substrate for the firefly luciferase catalyzed bioluminescence reaction : impact on bioluminescence based reporter gene imaging

    NARCIS (Netherlands)

    Berger, Frank; Paulmurugan, Ramasamy; Bhaumik, Srabani; Gambhir, Sanjiv Sam

    2008-01-01

    Purpose Firefly luciferase catalyzes the oxidative decarboxylation of D-luciferin to oxyluciferin in the presence of cofactors, producing bioluminescence. This reaction is used in optical bioluminescence-based molecular imaging approaches to detect the expression of the firefly luciferase reporter

  8. Bioluminescent probe for detecting endogenous hypochlorite in living mice.

    Science.gov (United States)

    Tang, Chunchao; Gao, Yuqi; Liu, Tingting; Lin, Yuxing; Zhang, Xiaomeng; Zhang, Chaochao; Li, Xiang; Zhang, Tianchao; Du, Lupei; Li, Minyong

    2018-01-24

    As a kind of biologically important reactive oxygen species (ROS), hypochlorite (ClO - ) plays a crucial role in many physiological processes. As such, endogenous ClO - is a powerful antibacterial agent during pathogen invasion. Nonetheless, excessive endogenous ClO - could pose a health threat to mammalian animals including humans. However, the detection of endogenous ClO - by bioluminescence probes in vivo remains a considerable challenge. Herein, based on a caged strategy, we developed a turn-on bioluminescent probe 1 for the highly selective detection of ClO - in vitro and imaging endogenous ClO - in a mouse inflammation model. We anticipate that such a probe could help us understand the role of endogenous ClO - in a variety of physiological and pathological processes.

  9. Mutagenesis and Characterization Studies to Develop Novel Bioluminescent Systems

    Science.gov (United States)

    2010-05-12

    of detection by in vivo bioluminescence, Molecular Imaging 3 (2004) 324-332. [40] B.R. Branchini, R.A. Magyar , M.H. Murtiashaw, S.M. Anderson, M...diversity, and structure function relationships of insect luciferases. Cell. Mol. Life Sci. 59, 1833-1850. (28) Branchini, B. R., Magyar , R. A...active site. Biochemistry 37, 15311-15319. (29) Branchini, B. R., Magyar , R. A., Murtiashaw, M. H., Anderson, S. M., Helgerson, L. C., and Zimmer, M

  10. Bacterial bioluminescence and Gumbel statistics: From quorum sensing to correlation

    Science.gov (United States)

    Delle Side, Domenico; Velardi, Luciano; Nassisi, Vincenzo; Pennetta, Cecilia; Alifano, Pietro; Talà, Adelfia; Salvatore Tredici, Maurizio

    2013-12-01

    We show that, in particular experimental conditions, the time course of the radiant fluxes, measured from a bioluminescent emission of a Vibrio harveyi related strain, collapse after suitable rescaling onto the Gumbel distribution of extreme value theory. We argue that the activation times of the strain luminous emission follow the universal behavior described by this statistical law, in spite of the fact that no extremal process is known to occur.

  11. Bioluminescence imaging of estrogen receptor activity during breast cancer progression.

    Science.gov (United States)

    Vantaggiato, Cristina; Dell'Omo, Giulia; Ramachandran, Balaji; Manni, Isabella; Radaelli, Enrico; Scanziani, Eugenio; Piaggio, Giulia; Maggi, Adriana; Ciana, Paolo

    2016-01-01

    Estrogen receptors (ER) are known to play an important regulatory role in mammary gland development as well as in its neoplastic transformation. Although several studies highlighted the contribution of ER signaling in the breast transformation, little is known about the dynamics of ER state of activity during carcinogenesis due to the lack of appropriate models for measuring the extent of receptor signaling in time, in the same animal. To this aim, we have developed a reporter mouse model for the non-invasive in vivo imaging of ER activity: the ERE-Luc reporter mouse. ERE-Luc is a transgenic mouse generated with a firefly luciferase (Luc) reporter gene driven by a minimal promoter containing an estrogen responsive element (ERE). This model allows to measure receptor signaling in longitudinal studies by bioluminescence imaging (BLI). Here, we have induced sporadic mammary cancers by treating systemically ERE-Luc reporter mice with DMBA (9,10-dimethyl 1,2-benzanthracene) and measured receptor signaling by in vivo imaging in individual animals from early stage until a clinically palpable tumor appeared in the mouse breast. We showed that DMBA administration induces an increase of bioluminescence in the whole abdominal area 6 h after treatment, the signal rapidly disappears. Several weeks later, strong bioluminescence is observed in the area corresponding to the mammary glands. In vivo and ex vivo imaging analysis demonstrated that this bioluminescent signal is localized in the breast area undergoing neoplastic transformation. We conclude that this non-invasive assay is a novel relevant tool to identify the activation of the ER signaling prior the morphological detection of the neoplastic transformation.

  12. Bioluminescence imaging of estrogen receptor activity during breast cancer progression

    Science.gov (United States)

    Vantaggiato, Cristina; Dell’Omo, Giulia; Ramachandran, Balaji; Manni, Isabella; Radaelli, Enrico; Scanziani, Eugenio; Piaggio, Giulia; Maggi, Adriana; Ciana, Paolo

    2016-01-01

    Estrogen receptors (ER) are known to play an important regulatory role in mammary gland development as well as in its neoplastic transformation. Although several studies highlighted the contribution of ER signaling in the breast transformation, little is known about the dynamics of ER state of activity during carcinogenesis due to the lack of appropriate models for measuring the extent of receptor signaling in time, in the same animal. To this aim, we have developed a reporter mouse model for the non-invasive in vivo imaging of ER activity: the ERE-Luc reporter mouse. ERE-Luc is a transgenic mouse generated with a firefly luciferase (Luc) reporter gene driven by a minimal promoter containing an estrogen responsive element (ERE). This model allows to measure receptor signaling in longitudinal studies by bioluminescence imaging (BLI). Here, we have induced sporadic mammary cancers by treating systemically ERE-Luc reporter mice with DMBA (9,10-dimethyl 1,2-benzanthracene) and measured receptor signaling by in vivo imaging in individual animals from early stage until a clinically palpable tumor appeared in the mouse breast. We showed that DMBA administration induces an increase of bioluminescence in the whole abdominal area 6 h after treatment, the signal rapidly disappears. Several weeks later, strong bioluminescence is observed in the area corresponding to the mammary glands. In vivo and ex vivo imaging analysis demonstrated that this bioluminescent signal is localized in the breast area undergoing neoplastic transformation. We conclude that this non-invasive assay is a novel relevant tool to identify the activation of the ER signaling prior the morphological detection of the neoplastic transformation. PMID:27069764

  13. Bioluminescence determination of active caspase-3 in single apoptotic cells

    Czech Academy of Sciences Publication Activity Database

    Lišková, Marcela; Klepárník, Karel; Matalová, Eva; Hegrová, Jitka; Přikryl, Jan; Švandová, Eva; Foret, František

    2013-01-01

    Roč. 34, č. 12 (2013), s. 1772-1777 ISSN 0173-0835 R&D Projects: GA ČR GAP206/11/2377 Grant - others:GA ČR(CZ) GAP502/12/1285 Program:GA Institutional support: RVO:68081715 ; RVO:67985904 Keywords : apoptosis * bioluminescence * caspase-3 Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.161, year: 2013

  14. Robust image modeling technique with a bioluminescence image segmentation application

    Science.gov (United States)

    Zhong, Jianghong; Wang, Ruiping; Tian, Jie

    2009-02-01

    A robust pattern classifier algorithm for the variable symmetric plane model, where the driving noise is a mixture of a Gaussian and an outlier process, is developed. The veracity and high-speed performance of the pattern recognition algorithm is proved. Bioluminescence tomography (BLT) has recently gained wide acceptance in the field of in vivo small animal molecular imaging. So that it is very important for BLT to how to acquire the highprecision region of interest in a bioluminescence image (BLI) in order to decrease loss of the customers because of inaccuracy in quantitative analysis. An algorithm in the mode is developed to improve operation speed, which estimates parameters and original image intensity simultaneously from the noise corrupted image derived from the BLT optical hardware system. The focus pixel value is obtained from the symmetric plane according to a more realistic assumption for the noise sequence in the restored image. The size of neighborhood is adaptive and small. What's more, the classifier function is base on the statistic features. If the qualifications for the classifier are satisfied, the focus pixel intensity is setup as the largest value in the neighborhood.Otherwise, it will be zeros.Finally,pseudo-color is added up to the result of the bioluminescence segmented image. The whole process has been implemented in our 2D BLT optical system platform and the model is proved.

  15. Triple Bioluminescence Imaging for In Vivo Monitoring of Cellular Processes

    Directory of Open Access Journals (Sweden)

    Casey A Maguire

    2013-01-01

    Full Text Available Bioluminescence imaging (BLI has shown to be crucial for monitoring in vivo biological processes. So far, only dual bioluminescence imaging using firefly (Fluc and Renilla or Gaussia (Gluc luciferase has been achieved due to the lack of availability of other efficiently expressed luciferases using different substrates. Here, we characterized a codon-optimized luciferase from Vargula hilgendorfii (Vluc as a reporter for mammalian gene expression. We showed that Vluc can be multiplexed with Gluc and Fluc for sequential imaging of three distinct cellular phenomena in the same biological system using vargulin, coelenterazine, and D-luciferin substrates, respectively. We applied this triple imaging system to monitor the effect of soluble tumor necrosis factor-related apoptosis-inducing ligand (sTRAIL delivered using an adeno-associated viral vector (AAV on brain tumors in mice. Vluc imaging showed efficient sTRAIL gene delivery to the brain, while Fluc imaging revealed a robust antiglioma therapy. Further, nuclear factor-κB (NF-κB activation in response to sTRAIL binding to glioma cells death receptors was monitored by Gluc imaging. This work is the first demonstration of trimodal in vivo bioluminescence imaging and will have a broad applicability in many different fields including immunology, oncology, virology, and neuroscience.

  16. In vitro and in vivo bioluminescent quantification of viable stem cells in engineered constructs.

    Science.gov (United States)

    Logeart-Avramoglou, Delphine; Oudina, Karim; Bourguignon, Marianne; Delpierre, Laetitia; Nicola, Marie-Anne; Bensidhoum, Morad; Arnaud, Eric; Petite, Herve

    2010-06-01

    Bioluminescent quantification of viable cells inside three-dimensional porous scaffolds was performed in vitro and in vivo. The assay quantified the bioluminescence of murine stem (C3H10T1/2) cells tagged with the luciferase gene reporter and distributed inside scaffolds of either soft, translucent, AN69 polymeric hydrogel or hard, opaque, coral ceramic materials. Quantitative evaluation of bioluminescence emitted from tagged cells adhering to these scaffolds was performed in situ using either cell lysates and a luminometer or intact cells and a bioluminescence imaging system. Despite attenuation of the signal when compared to cells alone, the bioluminescence correlated with the number of cells (up to 1.5 x 10(5)) present on each material scaffold tested, both in vitro and noninvasively in vivo (subcutaneous implants in the mouse model). The noninvasive bioluminescence measurement technique proved to be comparable to the cell-destructive bioluminescence measurement technique. Monitoring the kinetics of luciferase expression via bioluminescence enabled real-time assessment of cell survival and proliferation on the scaffolds tested over prolonged (up to 59 days) periods of time. This novel, sensitive, easy, fast-to-implement, quantitative bioluminescence assay has great, though untapped, potential for screening and determining noninvasively the presence of viable cells on biomaterial constructs in the tissue engineering and tissue regeneration fields.

  17. An adaptive regularization parameter choice strategy for multispectral bioluminescence tomography

    Energy Technology Data Exchange (ETDEWEB)

    Feng Jinchao; Qin Chenghu; Jia Kebin; Han Dong; Liu Kai; Zhu Shouping; Yang Xin; Tian Jie [Medical Image Processing Group, Institute of Automation, Chinese Academy of Sciences, P. O. Box 2728, Beijing 100190 (China); College of Electronic Information and Control Engineering, Beijing University of Technology, Beijing 100124 (China); Medical Image Processing Group, Institute of Automation, Chinese Academy of Sciences, P. O. Box 2728, Beijing 100190 (China); Medical Image Processing Group, Institute of Automation, Chinese Academy of Sciences, P. O. Box 2728, Beijing 100190 (China) and School of Life Sciences and Technology, Xidian University, Xi' an 710071 (China)

    2011-11-15

    Purpose: Bioluminescence tomography (BLT) provides an effective tool for monitoring physiological and pathological activities in vivo. However, the measured data in bioluminescence imaging are corrupted by noise. Therefore, regularization methods are commonly used to find a regularized solution. Nevertheless, for the quality of the reconstructed bioluminescent source obtained by regularization methods, the choice of the regularization parameters is crucial. To date, the selection of regularization parameters remains challenging. With regards to the above problems, the authors proposed a BLT reconstruction algorithm with an adaptive parameter choice rule. Methods: The proposed reconstruction algorithm uses a diffusion equation for modeling the bioluminescent photon transport. The diffusion equation is solved with a finite element method. Computed tomography (CT) images provide anatomical information regarding the geometry of the small animal and its internal organs. To reduce the ill-posedness of BLT, spectral information and the optimal permissible source region are employed. Then, the relationship between the unknown source distribution and multiview and multispectral boundary measurements is established based on the finite element method and the optimal permissible source region. Since the measured data are noisy, the BLT reconstruction is formulated as l{sub 2} data fidelity and a general regularization term. When choosing the regularization parameters for BLT, an efficient model function approach is proposed, which does not require knowledge of the noise level. This approach only requests the computation of the residual and regularized solution norm. With this knowledge, we construct the model function to approximate the objective function, and the regularization parameter is updated iteratively. Results: First, the micro-CT based mouse phantom was used for simulation verification. Simulation experiments were used to illustrate why multispectral data were used

  18. BLProt: prediction of bioluminescent proteins based on support vector machine and relieff feature selection

    Directory of Open Access Journals (Sweden)

    Hazrati Mehrnaz

    2011-08-01

    Full Text Available Abstract Background Bioluminescence is a process in which light is emitted by a living organism. Most creatures that emit light are sea creatures, but some insects, plants, fungi etc, also emit light. The biotechnological application of bioluminescence has become routine and is considered essential for many medical and general technological advances. Identification of bioluminescent proteins is more challenging due to their poor similarity in sequence. So far, no specific method has been reported to identify bioluminescent proteins from primary sequence. Results In this paper, we propose a novel predictive method that uses a Support Vector Machine (SVM and physicochemical properties to predict bioluminescent proteins. BLProt was trained using a dataset consisting of 300 bioluminescent proteins and 300 non-bioluminescent proteins, and evaluated by an independent set of 141 bioluminescent proteins and 18202 non-bioluminescent proteins. To identify the most prominent features, we carried out feature selection with three different filter approaches, ReliefF, infogain, and mRMR. We selected five different feature subsets by decreasing the number of features, and the performance of each feature subset was evaluated. Conclusion BLProt achieves 80% accuracy from training (5 fold cross-validations and 80.06% accuracy from testing. The performance of BLProt was compared with BLAST and HMM. High prediction accuracy and successful prediction of hypothetical proteins suggests that BLProt can be a useful approach to identify bioluminescent proteins from sequence information, irrespective of their sequence similarity. The BLProt software is available at http://www.inb.uni-luebeck.de/tools-demos/bioluminescent%20protein/BLProt

  19. Bioluminescence Tomography–Guided Radiation Therapy for Preclinical Research

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Bin [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland (United States); Wang, Ken Kang-Hsin, E-mail: kwang27@jhmi.edu [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland (United States); Yu, Jingjing [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland (United States); School of Physics and Information Technology, Shaanxi Normal University, Shaanxi (China); Eslami, Sohrab; Iordachita, Iulian [Laboratory for Computational Sensing and Robotics, Johns Hopkins University, Baltimore, Maryland (United States); Reyes, Juvenal; Malek, Reem [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland (United States); Tran, Phuoc T. [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland (United States); Department of Oncology and Urology, Brady Urological Institute, Johns Hopkins University, Baltimore, Maryland (United States); Patterson, Michael S. [Department of Medical Physics and Applied Radiation Sciences, McMaster University, Hamilton, Ontario (Canada); Wong, John W. [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland (United States)

    2016-04-01

    Purpose: In preclinical radiation research, it is challenging to localize soft tissue targets based on cone beam computed tomography (CBCT) guidance. As a more effective method to localize soft tissue targets, we developed an online bioluminescence tomography (BLT) system for small-animal radiation research platform (SARRP). We demonstrated BLT-guided radiation therapy and validated targeting accuracy based on a newly developed reconstruction algorithm. Methods and Materials: The BLT system was designed to dock with the SARRP for image acquisition and to be detached before radiation delivery. A 3-mirror system was devised to reflect the bioluminescence emitted from the subject to a stationary charge-coupled device (CCD) camera. Multispectral BLT and the incomplete variables truncated conjugate gradient method with a permissible region shrinking strategy were used as the optimization scheme to reconstruct bioluminescent source distributions. To validate BLT targeting accuracy, a small cylindrical light source with high CBCT contrast was placed in a phantom and also in the abdomen of a mouse carcass. The center of mass (CoM) of the source was recovered from BLT and used to guide radiation delivery. The accuracy of the BLT-guided targeting was validated with films and compared with the CBCT-guided delivery. In vivo experiments were conducted to demonstrate BLT localization capability for various source geometries. Results: Online BLT was able to recover the CoM of the embedded light source with an average accuracy of 1 mm compared to that with CBCT localization. Differences between BLT- and CBCT-guided irradiation shown on the films were consistent with the source localization revealed in the BLT and CBCT images. In vivo results demonstrated that our BLT system could potentially be applied for multiple targets and tumors. Conclusions: The online BLT/CBCT/SARRP system provides an effective solution for soft tissue targeting, particularly for small, nonpalpable, or

  20. Experimental Study on Bioluminescence Tomography with Multimodality Fusion

    Directory of Open Access Journals (Sweden)

    Yujie Lv

    2007-01-01

    Full Text Available To verify the influence of a priori information on the nonuniqueness problem of bioluminescence tomography (BLT, the multimodality imaging fusion based BLT experiment is performed by multiview noncontact detection mode, which incorporates the anatomical information obtained by the microCT scanner and the background optical properties based on diffuse reflectance measurements. In the reconstruction procedure, the utilization of adaptive finite element methods (FEMs and a priori permissible source region refines the reconstructed results and improves numerical robustness and efficiency. The comparison between the absence and employment of a priori information shows that multimodality imaging fusion is essential to quantitative BLT reconstruction.

  1. Application of ATP-based bioluminescence for bioaerosol quantification: effect of sampling method.

    Science.gov (United States)

    Han, Taewon; Wren, Melody; DuBois, Kelsey; Therkorn, Jennifer; Mainelis, Gediminas

    2015-12-01

    An adenosine triphosphate (ATP)-based bioluminescence has potential to offer a quick and affordable method for quantifying bioaerosol samples. Here we report on our investigation into how different bioaerosol aerosolization parameters and sampling methods affect bioluminescence output per bacterium, and implications of that effect for bioaerosol research. Bacillus atrophaeus and Pseudomonas fluorescens bacteria were aerosolized by using a Collison nebulizer (BGI Inc., Waltham, MA) with a glass or polycarbonate jar and then collected for 15 and 60 min with: (1) Button Aerosol Sampler (SKC Inc., Eighty Four, PA) with polycarbonate, PTFE, and cellulose nitrate filters, (2) BioSampler (SKC Inc.) with 5 and 20 mL of collection liquid, and (3) our newly developed Electrostatic Precipitator with Superhydrophobic Surface (EPSS). For all aerosolization and sampling parameters we compared the ATP bioluminescence output per bacterium relative to that before aerosolization and sampling. In addition, we also determined the ATP reagent storage and preparation conditions that that do not affect the bioluminescence signal intensity. Our results show that aerosolization by a Collison nebulizer with a polycarbonate jar yields higher bioluminescence output per bacterium compared to the glass jar. Interestingly enough, the bioluminescence output by P. fluorescens increased substantially after its aerosolization compared to the fresh liquid suspension. For both test microorganisms, the bioluminescence intensity per bacterium after sampling was significantly lower than that before sampling suggesting negative effect of sampling stress on bioluminescence output. The decrease in bioluminescence intensity was more pronounces for longer sampling times and significantly and substantially depended on the sampling method. Among the investigated method, the EPSS was the least injurious for both microorganisms and sampling times. While the ATP-based bioluminescence offers a quick bioaerosol

  2. Symplectin evolved from multiple duplications in bioluminescent squid

    Directory of Open Access Journals (Sweden)

    Warren R. Francis

    2017-07-01

    Full Text Available The squid Sthenoteuthis oualaniensis, formerly Symplectoteuthis oualaniensis, generates light using the luciferin coelenterazine and a unique enzyme, symplectin. Genetic information is limited for bioluminescent cephalopod species, so many proteins, including symplectin, occur in public databases only as sequence isolates with few identifiable homologs. As the distribution of the symplectin/pantetheinase protein family in Metazoa remains mostly unexplored, we have sequenced the transcriptomes of four additional luminous squid, and make use of publicly available but unanalyzed data of other cephalopods, to examine the occurrence and evolution of this protein family. While the majority of spiralians have one or two copies of this protein family, four well-supported groups of proteins are found in cephalopods, one of which corresponds to symplectin. A cysteine that is critical for symplectin functioning is conserved across essentially all members of the protein family, even those unlikely to be used for bioluminescence. Conversely, active site residues involved in pantetheinase catalysis are also conserved across essentially all of these proteins, suggesting that symplectin may have multiple functions including hydrolase activity, and that the evolution of the luminous phenotype required other changes in the protein outside of the main binding pocket.

  3. Bioluminescence Imaging to Detect Late Stage Infection of African Trypanosomiasis.

    Science.gov (United States)

    Burrell-Saward, Hollie; Ward, Theresa H

    2016-05-18

    Human African trypanosomiasis (HAT) is a multi-stage disease that manifests in two stages; an early blood stage and a late stage when the parasite invades the central nervous system (CNS). In vivo study of the late stage has been limited as traditional methodologies require the removal of the brain to determine the presence of the parasites. Bioluminescence imaging is a non-invasive, highly sensitive form of optical imaging that enables the visualization of a luciferase-transfected pathogen in real-time. By using a transfected trypanosome strain that has the ability to produce late stage disease in mice we are able to study the kinetics of a CNS infection in a single animal throughout the course of infection, as well as observe the movement and dissemination of a systemic infection. Here we describe a robust protocol to study CNS infections using a bioluminescence model of African trypanosomiasis, providing real time non-invasive observations which can be further analyzed with optional downstream approaches.

  4. Quantification of bioluminescence from the surface to the deep sea demonstrates its predominance as an ecological trait

    Science.gov (United States)

    Martini, Séverine; Haddock, Steven H. D.

    2017-04-01

    The capability of animals to emit light, called bioluminescence, is considered to be a major factor in ecological interactions. Because it occurs across diverse taxa, measurements of bioluminescence can be powerful to detect and quantify organisms in the ocean. In this study, 17 years of video observations were recorded by remotely operated vehicles during surveys off the California Coast, from the surface down to 3,900 m depth. More than 350,000 observations are classified for their bioluminescence capability based on literature descriptions. The organisms represented 553 phylogenetic concepts (species, genera or families, at the most precise taxonomic level defined from the images), distributed within 13 broader taxonomic categories. The importance of bioluminescent marine taxa is highlighted in the water column, as we showed that 76% of the observed individuals have bioluminescence capability. More than 97% of Cnidarians were bioluminescent, and 9 of the 13 taxonomic categories were found to be bioluminescent dominant. The percentage of bioluminescent animals is remarkably uniform over depth. Moreover, the proportion of bioluminescent and non-bioluminescent animals within taxonomic groups changes with depth for Ctenophora, Scyphozoa, Chaetognatha, and Crustacea. Given these results, bioluminescence has to be considered an important ecological trait from the surface to the deep-sea.

  5. Foraging in the darkness of the Southern Ocean: influence of bioluminescence on a deep diving predator.

    Directory of Open Access Journals (Sweden)

    Jade Vacquié-Garcia

    Full Text Available How non-echolocating deep diving marine predators locate their prey while foraging remains mostly unknown. Female southern elephant seals (SES (Mirounga leonina have vision adapted to low intensity light with a peak sensitivity at 485 nm. This matches the wavelength of bioluminescence produced by a large range of marine organisms including myctophid fish, SES's main prey. In this study, we investigated whether bioluminescence provides an accurate estimate of prey occurrence for SES. To do so, four SES were satellite-tracked during their post-breeding foraging trip and were equipped with Time-Depth-Recorders that also recorded light levels every two seconds. A total of 3386 dives were processed through a light-treatment model that detected light events higher than ambient level, i.e. bioluminescence events. The number of bioluminescence events was related to an index of foraging intensity for SES dives deep enough to avoid the influence of natural ambient light. The occurrence of bioluminescence was found to be negatively related to depth both at night and day. Foraging intensity was also positively related to bioluminescence both during day and night. This result suggests that bioluminescence likely provides SES with valuable indications of prey occurrence and might be a key element in predator-prey interactions in deep-dark marine environments.

  6. [Detection of low-level microorganism by concomitant use of ATP amplification and bioluminescence assay].

    Science.gov (United States)

    Chen, Ying; Zou, Bingjie; Zhu, Shuhui; Ma, Yinjiao; Zhou, Guohua

    2009-06-01

    To detect low levels of microorganism by bioluminescence assay, the reaction of ATP amplification catalyzed by ADK (adenylate kinase) combined with PPK (polyphosphate kinase) can be employed. However, the endogenous ADP bound to PPK is a background source and interfere the effective detection of low levels of exogenous ATP. We expressed a fusion protein of PPK and ADK and established a new method to decrease the background signal. The genes of PPK and ADK were amplified by PCR and cloned into vector pET28a (+) to provide a recombinant expression plasmid pET28a (+)-PPKADK to prepare the fusion protein. Apyrase was immobilized on the surface of magnetic beads coated with polyurethane to provide Beads-apyrase to eliminate background caused by ADP bound to PPK-ADK. The exogenous ATP and microorganism were also detected by using ATP amplification reaction coupled with bioluminescence assay. The purified fusion protein showed both ADK and PPK activities. Beads-apyrase could eliminate ADP contamination conveniently and effectively, thus less than 1 fmol of ATP was detected by ATP amplification reaction coupled with bioluminescence assay. Using ATP amplification reaction, the sensitivity of bioluminescence assay was 100-fold than that of normal bioluminescence assay without ATP amplification. Beads-apyrase is an effective tool to eliminate the background of the reaction of ATP amplification. The sensitivity of bioluminescence assay was increased significantly with concomitant use of ATP amplification and bioluminescence assay.

  7. In vitro validation of bioluminescent monitoring of disease progression and therapeutic response in leukaemia model animals

    International Nuclear Information System (INIS)

    Inoue, Yusuke; Okubo, Toshiyuki; Tojo, Arinobu; Sekine, Rieko; Soda, Yasushi; Kobayashi, Seiichiro; Nomura, Akiko; Izawa, Kiyoko; Kitamura, Toshio; Ohtomo, Kuni

    2006-01-01

    The application of in vivo bioluminescence imaging to non-invasive, quantitative monitoring of tumour models relies on a positive correlation between the intensity of bioluminescence and the tumour burden. We conducted cell culture studies to investigate the relationship between bioluminescent signal intensity and viable cell numbers in murine leukaemia model cells. Interleukin-3 (IL-3)-dependent murine pro-B cell line Ba/F3 was transduced with firefly luciferase to generate cells expressing luciferase stably under the control of a retroviral long terminal repeat. The luciferase-expressing cells were transduced with p190 BCR-ABL to give factor-independent proliferation. The cells were cultured under various conditions, and bioluminescent signal intensity was compared with viable cell numbers and the cell cycle stage. The Ba/F3 cells showed autonomous growth as well as stable luciferase expression following transduction with both luciferase and p190 BCR-ABL, and in vivo bioluminescence imaging permitted external detection of these cells implanted into mice. The bioluminescence intensities tended to reflect cell proliferation and responses to imatinib in cell culture studies. However, the luminescence per viable cell was influenced by the IL-3 concentration in factor-dependent cells and by the stage of proliferation and imatinib concentration in factor-independent cells, thereby impairing the proportionality between viable cell number and bioluminescent signal intensity. Luminescence per cell tended to vary in association with the fraction of proliferating cells. Although in vivo bioluminescence imaging would allow non-invasive monitoring of leukaemia model animals, environmental factors and therapeutic interventions may cause some discrepancies between tumour burden and bioluminescence intensity. (orig.)

  8. In vitro validation of bioluminescent monitoring of disease progression and therapeutic response in leukaemia model animals

    Energy Technology Data Exchange (ETDEWEB)

    Inoue, Yusuke; Okubo, Toshiyuki [University of Tokyo, Department of Radiology, Institute of Medical Science, Tokyo (Japan); Tojo, Arinobu; Sekine, Rieko; Soda, Yasushi; Kobayashi, Seiichiro; Nomura, Akiko; Izawa, Kiyoko [University of Tokyo, Division of Molecular Therapy, Advanced Clinical Research Centre, Tokyo (Japan); Kitamura, Toshio [University of Tokyo, Division of Cellular Therapy, Advanced Clinical Research Centre, Tokyo (Japan); Ohtomo, Kuni [University of Tokyo, Department of Radiology, Graduate School of Medicine, Tokyo (Japan)

    2006-05-15

    The application of in vivo bioluminescence imaging to non-invasive, quantitative monitoring of tumour models relies on a positive correlation between the intensity of bioluminescence and the tumour burden. We conducted cell culture studies to investigate the relationship between bioluminescent signal intensity and viable cell numbers in murine leukaemia model cells. Interleukin-3 (IL-3)-dependent murine pro-B cell line Ba/F3 was transduced with firefly luciferase to generate cells expressing luciferase stably under the control of a retroviral long terminal repeat. The luciferase-expressing cells were transduced with p190 BCR-ABL to give factor-independent proliferation. The cells were cultured under various conditions, and bioluminescent signal intensity was compared with viable cell numbers and the cell cycle stage. The Ba/F3 cells showed autonomous growth as well as stable luciferase expression following transduction with both luciferase and p190 BCR-ABL, and in vivo bioluminescence imaging permitted external detection of these cells implanted into mice. The bioluminescence intensities tended to reflect cell proliferation and responses to imatinib in cell culture studies. However, the luminescence per viable cell was influenced by the IL-3 concentration in factor-dependent cells and by the stage of proliferation and imatinib concentration in factor-independent cells, thereby impairing the proportionality between viable cell number and bioluminescent signal intensity. Luminescence per cell tended to vary in association with the fraction of proliferating cells. Although in vivo bioluminescence imaging would allow non-invasive monitoring of leukaemia model animals, environmental factors and therapeutic interventions may cause some discrepancies between tumour burden and bioluminescence intensity. (orig.)

  9. Bioanalytical Applications of Real-Time ATP Imaging Via Bioluminescence

    Energy Technology Data Exchange (ETDEWEB)

    Gruenhagen, Jason Alan [Iowa State Univ., Ames, IA (United States)

    2003-01-01

    The research discussed within involves the development of novel applications of real-time imaging of adenosine 5'-triphosphate (ATP). ATP was detected via bioluminescence and the firefly luciferase-catalyzed reaction of ATP and luciferin. The use of a microscope and an imaging detector allowed for spatially resolved quantitation of ATP release. Employing this method, applications in both biological and chemical systems were developed. First, the mechanism by which the compound 48/80 induces release of ATP from human umbilical vein endothelial cells (HUVECs) was investigated. Numerous enzyme activators and inhibitors were utilized to probe the second messenger systems involved in release. Compound 48/80 activated a G{sub q}-type protein to initiate ATP release from HUVECs. Ca2+ imaging along with ATP imaging revealed that activation of phospholipase C and induction of intracellular Ca2+ signaling were necessary for release of ATP. Furthermore, activation of protein kinase C inhibited the activity of phospholipase C and thus decreased the magnitude of ATP release. This novel release mechanism was compared to the existing theories of extracellular release of ATP. Bioluminescence imaging was also employed to examine the role of ATP in the field of neuroscience. The central nervous system (CNS) was dissected from the freshwater snail Lymnaea stagnalis. Electrophysiological experiments demonstrated that the neurons of the Lymnaea were not damaged by any of the components of the imaging solution. ATP was continuously released by the ganglia of the CNS for over eight hours and varied from ganglion to ganglion and within individual ganglia. Addition of the neurotransmitters K+ and serotonin increased release of ATP in certain regions of the Lymnaea CNS. Finally, the ATP imaging technique was investigated for the study of drug release systems. MCM-41-type mesoporous nanospheres were loaded with ATP and end-capped with mercaptoethanol

  10. DEVELOPMENT OF A DUAL MODALITY TOMOGRAPHIC IMAGING SYSTEM FOR BIOLUMINESCENCE AND PET

    Energy Technology Data Exchange (ETDEWEB)

    CHATZIIOANNOU, ARION

    2011-12-21

    The goal of this proposal was to develop a new hybrid imaging modality capable to simultaneously image optical bioluminescence signals, as well as radionuclide emissions from the annihilation of positrons originating from molecular imaging probes in preclinical mouse models. This new technology enables the simultaneous in-vivo measurements of both emissions that could be produced from a single or a combination of two different biomarkers. It also facilitates establishing the physical limitations of bioluminescence imaging, its tomographic and spectral image reconstruction potential and the quantification of bioluminescence signals.

  11. Ultraweak bioluminescence dynamics and singlet oxygen correlations during injury repair in sweet potato

    Science.gov (United States)

    Hossu, Marius; Ma, Lun; Chen, Wei

    2011-03-01

    Ultraweak bioluminescence at the level of hundreds of photons per second per square centimeter after cutting injury of sweet potato was investigated. A small emission peak immediate after cutting and a later and higher peak were observed. Selective singlet oxygen inhibitors and sensors have been use to study the contribution of singlet oxygen during the curing process, demonstrating increased presence of singlet oxygen during and after the late bioemission peak. It was confirmed that singlet oxygen has direct contribution to ultraweak bioluminescence but also induces the formation of other exited luminescent species that are responsible for the recorded bioluminescence.

  12. Development of a Bioluminescent Nitroreductase Probe for Preclinical Imaging.

    Directory of Open Access Journals (Sweden)

    Anzhelika G Vorobyeva

    Full Text Available Bacterial nitroreductases (NTRs have been widely utilized in the development of novel antibiotics, degradation of pollutants, and gene-directed enzyme prodrug therapy (GDEPT of cancer that reached clinical trials. In case of GDEPT, since NTR is not naturally present in mammalian cells, the prodrug is activated selectively in NTR-transformed cancer cells, allowing high efficiency treatment of tumors. Currently, no bioluminescent probes exist for sensitive, non-invasive imaging of NTR expression. We therefore developed a "NTR caged luciferin" (NCL probe that is selectively reduced by NTR, producing light proportional to the NTR activity. Here we report successful application of this probe for imaging of NTR in vitro, in bacteria and cancer cells, as well as in vivo in mouse models of bacterial infection and NTR-expressing tumor xenografts. This novel tool should significantly accelerate the development of cancer therapy approaches based on GDEPT and other fields where NTR expression is important.

  13. Dive Activities for Bioluminescence 2009 - Office of Ocean Exploration and Research

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Information about dive activities were recorded by personnel during the "Bioluminescence 2009" expedition, July 20 through 31, 2009. Additional information was...

  14. Bioluminescent luciferase-modified magnetic nanoparticles as potential imaging agents for mammalian spermatozoa detection and tracking

    Science.gov (United States)

    Background: Nanoparticles have emerged as key materials for developing applications in nanomedicine, nanobiotechnology, bioimaging and theranostics. Existing bioimaging technologies include bioluminescent resonance energy transfer-conjugated quantum dots (BRET-QDs). Despite the current use of BRET-Q...

  15. Rapid Analysis of Eukaryotic Bioluminescence to Assess Potential Groundwater Contamination Events

    Directory of Open Access Journals (Sweden)

    Zacariah L. Hildenbrand

    2015-01-01

    Full Text Available Here we present data using a bioluminescent dinoflagellate, Pyrocystis lunula, in a toxicological bioassay to rapidly assess potential instances of groundwater contamination associated with natural gas extraction. P. lunula bioluminescence can be quantified using spectrophotometry as a measurement of organismal viability, with normal bioluminescent output declining with increasing concentration(s of aqueous toxicants. Glutaraldehyde and hydrochloric acid (HCl, components used in hydraulic fracturing and shale acidization, triggered significant toxicological responses in as little as 4 h. Conversely, P. lunula was not affected by the presence of arsenic, selenium, barium, and strontium, naturally occurring heavy metal ions potentially associated with unconventional drilling activities. If exogenous compounds, such as glutaraldehyde and HCl, are thought to have been introduced into groundwater, quantification of P. lunula bioluminescence after exposure to water samples can serve as a cost-effective detection and risk assessment tool to rapidly assess the impact of putative contamination events attributed to unconventional drilling activity.

  16. Facile synthesis of gold-silver alloy nanoparticles for application in metal enhanced bioluminescence.

    Science.gov (United States)

    Abhijith, K S; Sharma, Richa; Ranjan, Rajeev; Thakur, M S

    2014-07-01

    In the present study we explored metal enhanced bioluminescence in luciferase enzymes for the first time. For this purpose a simple and reproducible one pot synthesis of gold-silver alloy nanoparticles was developed. By changing the molar ratio of tri-sodium citrate and silver nitrate we could synthesize spherical Au-Ag colloids of sizes ranging from 10 to 50 nm with a wide range of localized surface plasmon resonance (LSPR) peaks (450-550 nm). The optical tunability of the Au-Ag colloids enabled their effective use in enhancement of bioluminescence in a luminescent bacterium Photobacterium leiognathi and in luciferase enzyme systems from fireflies and bacteria. Enhancement of bioluminescence was 250% for bacterial cells, 95% for bacterial luciferase and 52% for firefly luciferase enzyme. The enhancement may be a result of energy transfer or plasmon induced enhancement. Such an increase can lead to higher sensitivity in detection of bioluminescent signals with potential applications in bio-analysis.

  17. Effect of irradiation on detection of bacteria in dehydrated vegetables with ATP bioluminescence assay

    International Nuclear Information System (INIS)

    Xiao Huan; Luo Shishi; Wang Zegang; Feng Min; Zhu Jiating; Chen Xiulan; Zhai Jianqing

    2011-01-01

    ATP bioluminescence intensity of 4 kinds of irradiated dehydrated vegetables was inconsistent with the bacteria number, the reasons were investigated in this paper. Results showed that irradiation had little effect on background luminescence, and there was no effect on luciferase-luminous system. When irradiation killed the bacteria, the ATPase activity also decreased. As a result, the ATP content in bacteria didn't decreased with the killed of bacteria, which contributed to the increase of free ATP in ATP extract and finally led to the disagreement between the bioluminescence intensity and the actual number of bacteria. When the free ATP in the dehydrated vegetable was removed, the bioluminescence intensity of ATP extract was consistent with the actual number of bacteria in irradiated dehydrated vegetable and ATP bioluminescence technology could be used in bacteria detection of irradiated samples. (authors)

  18. An improved single-step lysis protocol to measure luciferase bioluminescence in Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Hasenkamp Sandra

    2012-02-01

    Full Text Available Abstract This report describes the optimization and evaluation of a simple single-step lysis protocol to measure luciferase bioluminescence from genetically modified Plasmodium falciparum. This protocol utilizes a modified commercial buffer to improve speed of assay and consistency in the bioluminescence signal measured by reducing the manipulation steps required to release the cytoplasmic fraction. The utility of this improved assay protocol is demonstrated in typical assays that explore absolute and temporal gene expression activity.

  19. A bioluminescence resonance energy transfer (BRET) system: Application to interacting circadian clock proteins

    OpenAIRE

    Xu, Yao; Piston, David W.; Johnson, Carl Hirschie

    1999-01-01

    We describe a method for assaying protein interactions that offers some attractive advantages over previous assays. This method, called bioluminescence resonance energy transfer (BRET), uses a bioluminescent luciferase that is genetically fused to one candidate protein, and a green fluorescent protein mutant fused to another protein of interest. Interactions between the two fusion proteins can bring the luciferase and green fluorescent protein close enough for resonance energy transfer to occ...

  20. An Assessment and Annotated Bibliography of Marine Bioluminescence Research: 1979-1987

    Science.gov (United States)

    1993-01-01

    Deborah W. 108 Bowlby , Mark R. 96, 369 Cormier, Milton J. 97. 110, 111. 227. Arnold, John M. 742, 743 Boyd. S. H. 727 703 Arrio. Bernard 22. 149. 150...Washington, D.C. 14 Johns Hopkins University 13 Laboratoire de Bioluminescence, CNRS, France 13 Marine Biological Laboratory, Woods Hole 13 UniversitAt...free-running circadian rhythm of bioluminescence in individual cells of the 72. Buck. John B. (1978). Functions and Evolutions dinoflagellate Gonyaulax

  1. ATP-Bioluminescence as a method to evaluated microbiological quality of UHT milk

    Directory of Open Access Journals (Sweden)

    A.F. Cunha

    2014-12-01

    Full Text Available New approaches are needed to quickly indicate possible contamination of UHT milk, among them the technique of ATP-Bioluminescence. Therefore, the aim of this study was to compare the results of culture methods with the results of ATP-Bioluminescence technique of 102 UHT whole milk samples incubated at 48, 72, and 168 hours. UHT milk samples were analyzed for the presence of mesophilic and psychrotrophic aerobic microorganisms using Plate Count Agar (PCA, Brain-Heart Infusion (BHI media and PetrifilmTM Aerobic Count (AC plates. The ATP-Bioluminescence technique was applied through the Microbial Luminescent Screening (MLS system. Significant correlations were found between counts of aerobic mesophilic microorganisms on PCA, PetrifilmTM AC, BHI and results of ATP bioluminescence technique (P≤0.05. The ATP-Bioluminescence technique had higher correlation with counting method in PCA than BHI media. At lower pass/fail limits of Relative Light Units (60, 50, 45 and 40 RLU, the number of samples identified as positive increased and statistically agreed with aerobic mesophilic microorganism counts (P>0.05. For the dairy industry, the ATP-Bioluminescence technique may become an important tool that assists the official methods to quickly monitor the microbiological quality of UHT milk though this will likely require a threshold below 150 RLU.

  2. Effects of predator lipids on dinoflagellate defence mechanisms - increased bioluminescence capacity.

    Science.gov (United States)

    Lindström, Jenny; Grebner, Wiebke; Rigby, Kristie; Selander, Erik

    2017-10-12

    Short flashes of blue light (bioluminescence) from dinoflagellates can reduce copepod grazing of light-emitting cells. Other protective strategies against grazing are toxicity, reduced cell chain length and altered swimming patterns in different phytoplankton. Both toxicity and bioluminescence capacity in dinoflagellates decrease in copepod-free cultures, but toxin production can be restored in response to chemical alarm signals from copepods, copepodamides. Here we show that strains of the dinoflagellates Lingulodinium polyedra and Alexandrium tamarense, kept in culture for 14 and 9 years respectively, are capable of increasing their total bioluminescence capacity in response to copepodamides. The luminescence response to mechanical stimulation with air bubbles also increases significantly in L. polyedra after exposure to copepodamides. Effects on size, swimming speed and rate of change of direction in L. polyedra and A. tamarense were not detected, suggesting that post-encounter mechanisms such as bioluminescence and toxin production may constitute the dominating line of defence in these taxa. To our knowledge, this study provides the first evidence of changes in bioluminescence physiology as a response to chemical cues from natural enemies and emphasizes the importance of bioluminescence as an anti-grazing strategy.

  3. Bioluminescent bacteria: lux genes as environmental biosensors Bactérias bioluminescentes: os genes lux como biosensores ambientais

    Directory of Open Access Journals (Sweden)

    Vânia da Silva Nunes-Halldorson

    2003-06-01

    Full Text Available Bioluminescent bacteria are widespread in natural environments. Over the years, many researchers have been studying the physiology, biochemistry and genetic control of bacterial bioluminescence. These discoveries have revolutionized the area of Environmental Microbiology through the use of luminescent genes as biosensors for environmental studies. This paper will review the chronology of scientific discoveries on bacterial bioluminescence and the current applications of bioluminescence in environmental studies, with special emphasis on the Microtox toxicity bioassay. Also, the general ecological significance of bioluminescence will be addressed.Bactérias que emitem bioluminescência são amplamente distribuídas em ambientes naturais. Ao longo dos anos vários pesquisadores vêm estudando a fisiologia, bioquímica e controle genético da bioluminescência. Essas descobertas têm revolucionado a Área de Microbiologia Ambiental através da utilização dos genes lux como biosensores em estudos ambientais. Esta revisão examinará a cronologia de descobertas científicas da bioluminescência bacteriana e as aplicações atuais em estudos ambientais, salientando a utilização do teste de toxicidade Microtox. A significância ecológica da bioluminescência será também examinada.

  4. Computer-aided photometric analysis of dynamic digital bioluminescent images

    Science.gov (United States)

    Gorski, Zbigniew; Bembnista, T.; Floryszak-Wieczorek, J.; Domanski, Marek; Slawinski, Janusz

    2003-04-01

    The paper deals with photometric and morphologic analysis of bioluminescent images obtained by registration of light radiated directly from some plant objects. Registration of images obtained from ultra-weak light sources by the single photon counting (SPC) technique is the subject of this work. The radiation is registered by use of a 16-bit charge coupled device (CCD) camera "Night Owl" together with WinLight EG&G Berthold software. Additional application-specific software has been developed in order to deal with objects that are changing during the exposition time. Advantages of the elaborated set of easy configurable tools named FCT for a computer-aided photometric and morphologic analysis of numerous series of quantitatively imperfect chemiluminescent images are described. Instructions are given how to use these tools and exemplified with several algorithms for the transformation of images library. Using the proposed FCT set, automatic photometric and morphologic analysis of the information hidden within series of chemiluminescent images reflecting defensive processes in poinsettia (Euphorbia pulcherrima Willd) leaves affected by a pathogenic fungus Botrytis cinerea is revealed.

  5. Advancing Molecular Therapies through In Vivo Bioluminescent Imaging

    Directory of Open Access Journals (Sweden)

    Anton McCaffrey

    2003-04-01

    Full Text Available Effective development of therapeutics that target the molecular basis of disease is dependent on testing new therapeutic moieties and delivery strategies in animal models of human disease. Accelerating the analyses of these models and improving their predictive value through whole animal imaging methods, which provide data in real time and are sensitive to the subtle changes, are crucial for rapid advancement of these approaches. Modalities based on optics are rapid, sensitive, and accessible methods for in vivo analyses with relatively low instrumentation costs. In vivo bioluminescent imaging (BLI is one of these optically based imaging methods that enable rapid in vivo analyses of a variety of cellular and molecular events with extreme sensitivity. BLI is based on the use of light-emitting enzymes as internal biological light sources that can be detected externally as biological indicators. BLI has been used to test spatio-temporal expression patterns of both target and therapeutic genes in living laboratory animals where the contextual influences of whole biological systems are preserved. BLI has also been used to analyze gene delivery, immune cell therapies, and the in vivo efficacy of inhibitory RNAs. New tools for BLI are being developed that will offer greater flexibility in detection and analyses. BLI can be used to accelerate the evaluation of experimental therapeutic strategies and whole body imaging offers the opportunity of revealing the effects of novel approaches on key steps in disease processes.

  6. Bioluminescence Imaging of Period1 Gene Expression in Utero

    Directory of Open Access Journals (Sweden)

    Meera T. Saxena

    2007-01-01

    Full Text Available The use of real-time reporters has accelerated our understanding of gene expression in vivo. This study examined the feasibility of a luciferase-based reporter to image spatiotemporal changes in fetal gene expression in utero. We chose to monitor Period1 (Per1 because it is expressed broadly in the body and plays a role in circadian rhythmicity. Using rats carrying a Per1::luc transgene, we repetitively imaged fetuses in utero throughout gestation. We found that bioluminescence was specific to transgenic pups, increased dramatically on embryonic day 10 (10 days after successful mating, and continued to increase logarithmically until birth. Diurnal fluctuations in Per1 expression were apparent several days prior to birth. These results demonstrate the feasibility of in utero imaging of mammalian gene expression, tracking of fetal gene expression from the same litter, and early detection of mammalian clock gene expression. We conclude that luciferase-based reporters can provide a sensitive, noninvasive measure of in utero gene expression.

  7. Rapid drug susceptibility test of mycobacterium tuberculosis by bioluminescence sensor

    Science.gov (United States)

    Lu, Bin; Xu, Shunqing; Chen, Zifei; Zhou, Yikai

    2001-09-01

    With the persisting increase of drug-resistant stains of M. Tuberculosis around the world, rapid and sensitive detection of antibiotic of M. Tuberculosis is becoming more and more important. In the present study, drug susceptibility of M. tuberculosis were detected by recombination mycobacteriophage combined with bioluminescence sensor. It is based on the use of recombination mycobacteriophage which can express firefly luciferase when it infects viable mycobacteria, and can effectively produce quantifiable photon. Meanwhile, in mycobacterium cells treated with active antibiotic, no light is observed. The emitted light is recorded by a bioluminscence sensor, so the result of drug-resistant test can be determined by the naked eye. 159 stains of M. tuberculosis were applied to this test on their resistant to rifampin, streptomycin and isoniazid. It is found that the agreement of this assay with Liewenstein- Jensen slat is: rifampin 95.60 percent, isoniazid 91.82 percent, streptomycin 88.68 percent, which showed that it is a fast and practical method to scene and detect drug resistant of mycobacterium stains.

  8. Spectrally resolved bioluminescence tomography with the third-order simplified spherical harmonics approximation

    Science.gov (United States)

    Lu, Yujie; Douraghy, Ali; Machado, Hidevaldo B.; Stout, David; Tian, Jie; Herschman, Harvey; Chatziioannou, Arion F.

    2009-11-01

    Bioluminescence imaging has been extensively applied to in vivo small animal imaging. Quantitative three-dimensional bioluminescent source information obtained by using bioluminescence tomography can directly and much more accurately reflect biological changes as opposed to planar bioluminescence imaging. Preliminary simulated and experimental reconstruction results demonstrate the feasibility and promise of bioluminescence tomography. However, the use of multiple approximations, particularly the diffusion approximation theory, affects the quality of in vivo small animal-based image reconstructions. In the development of new reconstruction algorithms, high-order approximation models of the radiative transfer equation and spectrally resolved data introduce new challenges to the reconstruction algorithm and speed. In this paper, an SP3-based (the third-order simplified spherical harmonics approximation) spectrally resolved reconstruction algorithm is proposed. The simple linear relationship between the unknown source distribution and the spectrally resolved data is established in this algorithm. A parallel version of this algorithm is realized, making BLT reconstruction feasible for the whole body of small animals especially for fine spatial domain discretization. In simulation validations, the proposed algorithm shows improved reconstruction quality compared with diffusion approximation-based methods when high absorption, superficial sources and detection modes are considered. In addition, comparisons between fine and coarse mesh-based BLT reconstructions show the effects of numerical errors in reconstruction image quality. Finally, BLT reconstructions using in vivo mouse experiments further demonstrate the potential and effectiveness of the SP3-based reconstruction algorithm.

  9. Simultaneous bioluminescent immunoassay of serum total and IgG-bound prolactins.

    Science.gov (United States)

    Kudryavtsev, Alexander N; Krasitskaya, Vasilisa V; Petunin, Alexei I; Burakov, Andrey Y; Frank, Ludmila A

    2012-04-03

    Novel dual-analyte single-well bioluminescence immunoassay (BLIA) for total and IgG-bound prolactins was developed on the base of Ca(2+)-regulated photoprotein obelin mutants with altered color and kinetics of bioluminescence signal as reporters. The mutants W92F-H22E and Y138F were chemically conjugated with monoclonal mouse anti-hPRL and anti-hIgG immunoglobulins and thus displayed signals from total prolactin and IgG-bounded prolactin (macroprolactin) correspondingly. Bioluminescence of the reporters was simultaneously triggered by a single injection of Ca(2+) solution and discriminated via bioluminescent signal spectral and time resolution. The developed microplate-based immunoassay allows detection of two prolactin forms in crude serum without additional manipulations (e.g., gel chromatography or PEG-precipitation). Total prolactin bioluminescence immunoassay in standard, control, and clinical sera offers high sensitivity and reproducibility. The BLIA results show good correlation with those obtained by RIA and immunoassay after gel chromatography.

  10. Autonomous bioluminescent expression of the bacterial luciferase gene cassette (lux in a mammalian cell line.

    Directory of Open Access Journals (Sweden)

    Dan M Close

    Full Text Available The bacterial luciferase (lux gene cassette consists of five genes (luxCDABE whose protein products synergistically generate bioluminescent light signals exclusive of supplementary substrate additions or exogenous manipulations. Historically expressible only in prokaryotes, the lux operon was re-synthesized through a process of multi-bicistronic, codon-optimization to demonstrate for the first time self-directed bioluminescence emission in a mammalian HEK293 cell line in vitro and in vivo.Autonomous in vitro light production was shown to be 12-fold greater than the observable background associated with untransfected control cells. The availability of reduced riboflavin phosphate (FMNH(2 was identified as the limiting bioluminescence substrate in the mammalian cell environment even after the addition of a constitutively expressed flavin reductase gene (frp from Vibrio harveyi. FMNH(2 supplementation led to a 151-fold increase in bioluminescence in cells expressing mammalian codon-optimized luxCDE and frp genes. When injected subcutaneously into nude mice, in vivo optical imaging permitted near instantaneous light detection that persisted independently for the 60 min length of the assay with negligible background.The speed, longevity, and self-sufficiency of lux expression in the mammalian cellular environment provides a viable and powerful alternative for real-time target visualization not currently offered by existing bioluminescent and fluorescent imaging technologies.

  11. Continuous, real-time bioimaging of chemical bioavailability and toxicology using autonomously bioluminescent human cell lines

    Science.gov (United States)

    Xu, Tingting; Close, Dan M.; Webb, James D.; Price, Sarah L.; Ripp, Steven A.; Sayler, Gary S.

    2013-05-01

    Bioluminescent imaging is an emerging biomedical surveillance strategy that uses external cameras to detect in vivo light generated in small animal models of human physiology or in vitro light generated in tissue culture or tissue scaffold mimics of human anatomy. The most widely utilized of reporters is the firefly luciferase (luc) gene; however, it generates light only upon addition of a chemical substrate, thus only generating intermittent single time point data snapshots. To overcome this disadvantage, we have demonstrated substrate-independent bioluminescent imaging using an optimized bacterial bioluminescence (lux) system. The lux reporter produces bioluminescence autonomously using components found naturally within the cell, thereby allowing imaging to occur continuously and in real-time over the lifetime of the host. We have validated this technology in human cells with demonstrated chemical toxicological profiling against exotoxin exposures at signal strengths comparable to existing luc systems (~1.33 × 107 photons/second). As a proof-in-principle demonstration, we have engineered breast carcinoma cells to express bioluminescence for real-time screening of endocrine disrupting chemicals and validated detection of 17β-estradiol (EC50 = ~ 10 pM). These and other applications of this new reporter technology will be discussed as potential new pathways towards improved models of target chemical bioavailability, toxicology, efficacy, and human safety.

  12. Attenuated Bioluminescent Brucella melitensis Mutants GR019 (virB4), GR024 (galE), and GR026 (BMEI1090-BMEI1091) Confer Protection in Mice

    OpenAIRE

    Rajashekara, Gireesh; Glover, David A.; Banai, Menachem; O'Callaghan, David; Splitter, Gary A.

    2006-01-01

    In vivo bioluminescence imaging is a persuasive approach to investigate a number of issues in microbial pathogenesis. Previously, we have applied bioluminescence imaging to gain greater insight into Brucella melitensis pathogenesis. Endowing Brucella with bioluminescence allowed direct visualization of bacterial dissemination, pattern of tissue localization, and the contribution of Brucella genes to virulence. In this report, we describe the pathogenicity of three attenuated bioluminescent B....

  13. Four new bioluminescent taxa of Mycena sect. Calodontes from Peninsular Malaysia.

    Science.gov (United States)

    Chew, Audrey L C; Tan, Yee-Shin; Desjardin, Dennis E; Musa, Md Yusoff; Sabaratnam, Vikineswary

    2014-01-01

    Three new species and one new variety of bioluminescent Mycena collected from Peninsular Malaysia are described herein. All new species belong to Mycena sect. Calodontes in what is known as the Mycena pura complex. Comprehensive descriptions, photographs, illustrations and comparisons with phenetically similar species are provided. Molecular sequences data from the nuclear internal transcribed spacers (ITS-1 and ITS-2, including the 5.8S rRNA) were used to infer relationships within sect. Calodontes. Axenic cultures were obtained to provide data on culture morphology. This is the first published photographic documentation of bioluminescent basidiomes of members of Mycena sect. Calodontes. Also, this addition brings the total known bioluminescent fungi to 77 species. © 2014 by The Mycological Society of America.

  14. Regulated bioluminescence as a tool for bioremediation process monitoring and control of bacterial cultures

    Science.gov (United States)

    Burlage, Robert S.; Heitzer, Armin; Digrazia, Philip M.

    1991-01-01

    An effective on-line monitoring technique for toxic waste bioremediation using bioluminescent microorganisms has shown great potential for the description and optimization of biological processes. The lux genes of the bacterium Vibrio fischeri are used by this species to produce visible light. The lux genes can be genetically fused to the control region of a catabolic gene, with the result that bioluminescence is produced whenever the catabolic gene is induced. Thus the detection of light from a sample indicates that genetic expression from a specific gene is occurring. This technique was used to monitor biodegradation of specific contaminants from waste sites. For these studies, fusions between the lux genes and the operons for naphthalene and toluene/xylene degradation were constructed. Strains carrying one of these fusions respond sensitively and specifically to target substrates. Bioluminescence from these cultures can be rapidly measured in a nondestructive and noninvasive manner. The potential for this technique in this and other biological systems is discussed.

  15. Why does the bioluminescent fungus Armillaria mellea have luminous mycelium but nonluminous fruiting body?

    Science.gov (United States)

    Purtov, K V; Petushkov, V N; Rodionova, N S; Gitelson, J I

    2017-05-01

    By determining the components involved in the bioluminescence process in luminous and nonluminous organs of the honey fungus Armillaria mellea, we have established causes of partial luminescence of this fungus. The complete set of enzymes and substrates required for bioluminescence is formed only in the mycelium and only under the conditions of free oxygen access. Since the synthesis of luciferin precursor (hispidin) and 3-hydroxyhispidin hydroxylase in the fruiting bodies is blocked, the formation of luciferin-the key component of fungal bioluminescent system-was not observed. That is why the fruiting body of Armillaria mellea is nonluminous despite the presence of luciferase, the enzyme that catalyzes the oxidation of luciferin with a photon emission.

  16. An ancestral luciferase in the Malpighi tubules of a non-bioluminescent beetle!

    Science.gov (United States)

    Viviani, V R; Prado, R A; Arnoldi, F C G; Abdalla, F C

    2009-01-01

    The evolutionary origin of beetle bioluminescence is enigmatic. Previously, weak luciferase activity was found in the non-bioluminescent larvae of Tenebrio molitor (Coleoptera: Tenebrionidae), but the detailed tissular origin and identity of the luciferase-like enzyme remained unknown. Using a closely related giant mealworm, Zophobas morio, here we show that the luciferase-like enzyme is located in the Malpighi tubules. cDNA cloning of this luciferase like enzyme, showed that it is a short AMP-ligase with weak luciferase activity which diverged long ago from beetle luciferases. The results indicate that the potential for bioluminescence in AMP-ligases is very ancient and provide a first reasonable protoluciferase model to investigate the origin and evolution of beetle luciferases.

  17. Characterization of an anthraquinone fluor from the bioluminescent, pelagic polychaete Tomopteris

    Science.gov (United States)

    Francis, Warren R; Powers, Meghan L; Haddock, Steven H D

    2014-01-01

    Tomopteris is a cosmopolitan genus of polychaetes. Many species produce yellow luminescence in the parapodia when stimulated. Yellow bioluminescence is rare in the ocean, and the components of this luminescent reaction have not been identified. Only a brief description, half a century ago, noted fluorescence in the parapodia with a remarkably similar spectrum to the bioluminescence, which suggested that it may be the luciferin or terminal light-emitter. Here, we report the isolation of the fluorescent yellow–orange pigment found in the luminous exudate and in the body of the animals. Liquid chromatography-mass spectrometry revealed the mass to be 270 m/z with a molecular formula of C15H10O5, which ultimately was shown to be aloe-emodin, an anthraquinone previously found in plants. We speculate that aloe-emodin could be a factor for resonant-energy transfer or the oxyluciferin for Tomopteris bioluminescence. PMID:24760626

  18. Comparison of Acute Toxicity of Algal Metabolites Using Bioluminescence Inhibition Assay

    Directory of Open Access Journals (Sweden)

    Hansa Jeswani

    2015-01-01

    Full Text Available Microalgae are reported to degrade hazardous compounds. However, algae, especially cyanobacteria are known to produce secondary metabolites which may be toxic to flora, fauna and human beings. The aim of this study was selection of an appropriate algal culture for biological treatment of biomass gasification wastewater based on acute toxicity considerations. The three algae that were selected were Spirulina sp., Scenedesmus abundans and a fresh water algal consortium. Acute toxicity of the metabolites produced by these algal cultures was tested at the end of log phase using the standard bioluminescence inhibition assay based on Vibrio fischeri NRRLB 11174. Scenedesmus abundans and a fresh water algal consortium dominated by cyanobacteria such as Phormidium, Chroococcus and Oscillatoria did not release much toxic metabolites at the end of log phase and caused only about 20% inhibition in bioluminescence. In comparison, Spirulina sp. released toxic metabolites and caused 50% bioluminescence inhibition at 3/5 times dilution of the culture supernatant (EC50.

  19. Photon hunting in the twilight zone: visual features of mesopelagic bioluminescent sharks.

    Science.gov (United States)

    Claes, Julien M; Partridge, Julian C; Hart, Nathan S; Garza-Gisholt, Eduardo; Ho, Hsuan-Ching; Mallefet, Jérôme; Collin, Shaun P

    2014-01-01

    The mesopelagic zone is a visual scene continuum in which organisms have developed various strategies to optimize photon capture. Here, we used light microscopy, stereology-assisted retinal topographic mapping, spectrophotometry and microspectrophotometry to investigate the visual ecology of deep-sea bioluminescent sharks [four etmopterid species (Etmopterus lucifer, E. splendidus, E. spinax and Trigonognathus kabeyai) and one dalatiid species (Squaliolus aliae)]. We highlighted a novel structure, a translucent area present in the upper eye orbit of Etmopteridae, which might be part of a reference system for counterillumination adjustment or acts as a spectral filter for camouflage breaking, as well as several ocular specialisations such as aphakic gaps and semicircular tapeta previously unknown in elasmobranchs. All species showed pure rod hexagonal mosaics with a high topographic diversity. Retinal specialisations, formed by shallow cell density gradients, may aid in prey detection and reflect lifestyle differences; pelagic species display areae centrales while benthopelagic and benthic species display wide and narrow horizontal streaks, respectively. One species (E. lucifer) displays two areae within its horizontal streak that likely allows detection of conspecifics' elongated bioluminescent flank markings. Ganglion cell topography reveals less variation with all species showing a temporal area for acute frontal binocular vision. This area is dorsally extended in T. kabeyai, allowing this species to adjust the strike of its peculiar jaws in the ventro-frontal visual field. Etmopterus lucifer showed an additional nasal area matching a high rod density area. Peak spectral sensitivities of the rod visual pigments (λmax) fall within the range 484-491 nm, allowing these sharks to detect a high proportion of photons present in their habitat. Comparisons with previously published data reveal ocular differences between bioluminescent and non-bioluminescent deep

  20. The application of superweak bioluminescence on freshness degree of chicken egg

    International Nuclear Information System (INIS)

    Zhao Hongxia; Li Guochen; Li Qiangzheng; Li Juan

    2007-01-01

    The luminescence of chicken egg in storage is studied by a detection system of superweak bioluminescence. The results show that egg has the strongest vigour on the third day after it is laid, subsequently the luminescence presents decay with oscillation. These eggs, which have been stored for 3 days, are most suitable for hatching. Different eggs have different luminescence intensities depending on the vigour of the egg. The stronger the vigour of the egg is, the more intensive the luminescence is. Superweak bioluminescence as a comprehensive index of biology and biochemistry response can be used for inspecting the freshness degree of the egg, and the test is nondestructive and sensitive

  1. 'Bioluminescent' reporter phage for the detection of Category A bacterial pathogens.

    Science.gov (United States)

    Schofield, David A; Molineux, Ian J; Westwater, Caroline

    2011-07-08

    Yersinia pestis and Bacillus anthracis are Category A bacterial pathogens that are the causative agents of the plague and anthrax, respectively. Although the natural occurrence of both diseases' is now relatively rare, the possibility of terrorist groups using these pathogens as a bioweapon is real. Because of the disease's inherent communicability, rapid clinical course, and high mortality rate, it is critical that an outbreak be detected quickly. Therefore methodologies that provide rapid detection and diagnosis are essential to ensure immediate implementation of public health measures and activation of crisis management. Recombinant reporter phage may provide a rapid and specific approach for the detection of Y. pestis and B. anthracis. The Centers for Disease Control and Prevention currently use the classical phage lysis assays for the confirmed identification of these bacterial pathogens. These assays take advantage of naturally occurring phage which are specific and lytic for their bacterial hosts. After overnight growth of the cultivated bacterium in the presence of the specific phage, the formation of plaques (bacterial lysis) provides a positive identification of the bacterial target. Although these assays are robust, they suffer from three shortcomings: 1) they are laboratory based; 2) they require bacterial isolation and cultivation from the suspected sample, and 3) they take 24-36 h to complete. To address these issues, recombinant "light-tagged" reporter phage were genetically engineered by integrating the Vibrio harveyi luxAB genes into the genome of Y. pestis and B. anthracis specific phage. The resulting luxAB reporter phage were able to detect their specific target by rapidly (within minutes) and sensitively conferring a bioluminescent phenotype to recipient cells. Importantly, detection was obtained either with cultivated recipient cells or with mock-infected clinical specimens. For demonstration purposes, here we describe the method for the phage

  2. Multimodality imaging of somatostatin receptor-positive tumors with nuclear and bioluminescence imaging

    NARCIS (Netherlands)

    S.E. Pool (Stefan); T.L.M. ten Hagen (Timo); S. Koelewijn (Stuart); M. de Jong (Marcel); G.A. Koning (Gerben)

    2012-01-01

    textabstractMultimodal bioluminescence (BLI) and single-photon emission computed tomography/computed tomography (SPECT/CT) imaging were investigated as means to monitor somatostatin receptor subtype 2 (SST 2)-positive neuroendocrine tumors as both a subcutaneously implanted and a liver metastasis

  3. Bioluminescence ATP Monitoring for the Routine Assessment of Food Contact Surface Cleanliness in a University Canteen

    Directory of Open Access Journals (Sweden)

    Andrea Osimani

    2014-10-01

    Full Text Available ATP bioluminescence monitoring and traditional microbiological analyses (viable counting of total mesophilic aerobes, coliforms and Escherichia coli were used to evaluate the effectiveness of Sanitation Standard Operating Procedures (SSOP at a university canteen which uses a HACCP-based approach. To that end, 10 cleaning control points (CPs, including food contact surfaces at risk of contamination from product residues or microbial growth, were analysed during an 8-month monitoring period. Arbitrary acceptability limits were set for both microbial loads and ATP bioluminescence readings. A highly significant correlation (r = 0.99 between the means of ATP bioluminescence readings and the viable counts of total mesophilic aerobes was seen, thus revealing a strong association of these parameters with the level of surface contamination. Among CPs, the raw meat and multi-purpose chopping boards showed the highest criticalities. Although ATP bioluminescence technology cannot substitute traditional microbiological analyses for the determination of microbial load on food contact surfaces, it has proved to be a powerful tool for the real time monitoring of surface cleanliness at mass catering plants, for verify the correct application of SSOP, and hence for their implementation/revision in the case of poor hygiene.

  4. Comparison of the bioluminescence of Photorhabdus species and subspecies type strains

    Czech Academy of Sciences Publication Activity Database

    Hyršl, P.; Číž, Milan; Lojek, Antonín

    2004-01-01

    Roč. 49, č. 5 (2004), s. 539 ISSN 0015-5632 R&D Projects: GA AV ČR IBS5004009 Institutional research plan: CEZ:AV0Z5004920 Keywords : bioluminescence * Photorhabdus species type strains * Photorhabdus subspecies type strains Subject RIV: BO - Biophysics Impact factor: 1.034, year: 2004

  5. Fast in vivo bioluminescence tomography using a novel pure optical imaging technique

    Directory of Open Access Journals (Sweden)

    Shuang Zhang

    2017-05-01

    Full Text Available Bioluminescence tomography (BLT is a novel optical molecular imaging technique that advanced the conventional planar bioluminescence imaging (BLI into a quantifiable three-dimensional (3D approach in preclinical living animal studies in oncology. In order to solve the inverse problem and reconstruct tumor lesions inside animal body accurately, the prior structural information is commonly obtained from X-ray computed tomography (CT. This strategy requires a complicated hybrid imaging system, extensive post imaging analysis and involvement of ionizing radiation. Moreover, the overall robustness highly depends on the fusion accuracy between the optical and structural information. Here, we present a pure optical bioluminescence tomographic (POBT system and a novel BLT workflow based on multi-view projection acquisition and 3D surface reconstruction. This method can reconstruct the 3D surface of an imaging subject based on a sparse set of planar white-light and bioluminescent images, so that the prior structural information can be offered for 3D tumor lesion reconstruction without the involvement of CT. The performance of this novel technique was evaluated through the comparison with a conventional dual-modality tomographic (DMT system and a commercialized optical imaging system (IVIS Spectrum using three breast cancer xenografts. The results revealed that the new technique offered comparable in vivo tomographic accuracy with the DMT system (P>0.05 in much shorter data analysis time. It also offered significantly better accuracy comparing with the IVIS system (P<0.04 without sacrificing too much time.

  6. Autonomously bioluminescent mammalian cells for continuous and real-time monitoring of cytotoxicity.

    Science.gov (United States)

    Xu, Tingting; Close, Dan M; Webb, James D; Ripp, Steven A; Sayler, Gary S

    2013-10-28

    Mammalian cell-based in vitro assays have been widely employed as alternatives to animal testing for toxicological studies but have been limited due to the high monetary and time costs of parallel sample preparation that are necessitated due to the destructive nature of firefly luciferase-based screening methods. This video describes the utilization of autonomously bioluminescent mammalian cells, which do not require the destructive addition of a luciferin substrate, as an inexpensive and facile method for monitoring the cytotoxic effects of a compound of interest. Mammalian cells stably expressing the full bacterial bioluminescence (luxCDABEfrp) gene cassette autonomously produce an optical signal that peaks at 490 nm without the addition of an expensive and possibly interfering luciferin substrate, excitation by an external energy source, or destruction of the sample that is traditionally performed during optical imaging procedures. This independence from external stimulation places the burden for maintaining the bioluminescent reaction solely on the cell, meaning that the resultant signal is only detected during active metabolism. This characteristic makes the lux-expressing cell line an excellent candidate for use as a biosentinel against cytotoxic effects because changes in bioluminescent production are indicative of adverse effects on cellular growth and metabolism. Similarly, the autonomous nature and lack of required sample destruction permits repeated imaging of the same sample in real-time throughout the period of toxicant exposure and can be performed across multiple samples using existing imaging equipment in an automated fashion.

  7. Bioluminescence resonance energy transfer methods to study G protein-coupled receptor-receptor tyrosine kinase heteroreceptor complexes.

    Science.gov (United States)

    Borroto-Escuela, Dasiel O; Flajolet, Marc; Agnati, Luigi F; Greengard, Paul; Fuxe, Kjell

    2013-01-01

    A large body of evidence indicates that G protein-coupled receptors (GPCRs) and receptor tyrosine kinases (RTKs) can form heteroreceptor complexes. In these complexes, the signaling from each interacting protomer is modulated to produce an integrated and therefore novel response upon agonist(s) activation. In the GPCR-RTK heteroreceptor complexes, GPCRs can activate RTK in the absence of added growth factor through the use of RTK signaling molecules. This integrative phenomenon is reciprocal and can place also RTK signaling downstream of GPCR. Formation of either stable or transient complexes by these two important classes of membrane receptors is involved in regulating all aspects of receptor function, from ligand binding to signal transduction, trafficking, desensitization, and downregulation among others. Functional phenomena can be modulated with conformation-specific inhibitors that stabilize defined GPCR states to abrogate both GPCR agonist- and growth factor-stimulated cell responses or by means of small interfering heteroreceptor complex interface peptides. The bioluminescence resonance energy transfer (BRET) technology has emerged as a powerful method to study the structure of heteroreceptor complexes closely associated with the study of receptor-receptor interactions in such complexes. In this chapter, we provide an overview of different BRET(2) assays that can be used to study the structure of GPCR-RTK heteroreceptor complexes and their functions. Various experimental designs for optimization of these experiments are also described. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Ship track for Islands in the Stream 2002 - Pharmaceutical Discovery, Vision, and Bioluminescence - Office of Ocean Exploration

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Ship track of the R/V Seward Johnson during the 2002 "Islands in the Stream - Pharmaceutical Discovery, Vision, and Bioluminescence" expedition sponsored by the...

  9. Rapid detection of E. Coli O157:H7 by IFAST and ATP bioluminescence assay for water analysis

    CSIR Research Space (South Africa)

    Ngamsom, B

    2016-10-01

    Full Text Available The present investigation reports isolation and detection of E. coli O157:H7 employing a simple and portable microfluidic device based on immiscible filtration assisted by surface tension (IFAST) and adenosine triphosphate (ATP) bioluminescence...

  10. Effects of salinity, pH and temperature on the re-establishment of bioluminescence and copper or SDS toxicity in the marine dinoflagellate Pyrocystis lunula using bioluminescence as an endpoint

    Science.gov (United States)

    Craig, J.M.; Klerks, P.L.; Heimann, K.; Waits, J.L.

    2003-01-01

    Pyrocystis lunula is a unicellular, marine, photoautotrophic, bioluminescent dinoflagellate. This organism is used in the Lumitox ?? bioassay with inhibition of bioluminescence re-establishment as the endpoint. Experiments determined if acute changes in pH, salinity, or temperature had an effect on the organisms' ability to re-establish bioluminescence, or on the bioassay's potential to detect sodium dodecyl sulfate (SDS) and copper toxicity. The re-establishment of bioluminescence itself was not very sensitive to changes in pH within the pH 6-10 range, though reducing pH from 8 to levels below 6 decreased this capacity. Increasing the pH had little effect on Cu or SDS toxicity, but decreasing the pH below 7 virtually eliminated the toxicity of either compound in the bioassay. Lowering the salinity from 33 to 27??? or less resulted in a substantial decrease in re-establishment of bioluminescence, while increasing the salinity to 43 or 48 ??? resulted in a small decline. Salinity had little influence on the bioassay's quantification of Cu toxicity, while the data showed a weak negative relationship between SDS toxicity and salinity. Re-establishment of bioluminescence showed a direct dependence on temperature, but only at 10??C did temperature have an obvious effect on the toxicity of Cu in this bioassay. ?? 2003 Elsevier Science Ltd. All rights reserved.

  11. Photon Counting System for High-Sensitivity Detection of Bioluminescence at Optical Fiber End.

    Science.gov (United States)

    Iinuma, Masataka; Kadoya, Yutaka; Kuroda, Akio

    2016-01-01

    The technique of photon counting is widely used for various fields and also applicable to a high-sensitivity detection of luminescence. Thanks to recent development of single photon detectors with avalanche photodiodes (APDs), the photon counting system with an optical fiber has become powerful for a detection of bioluminescence at an optical fiber end, because it allows us to fully use the merits of compactness, simple operation, highly quantum efficiency of the APD detectors. This optical fiber-based system also has a possibility of improving the sensitivity to a local detection of Adenosine triphosphate (ATP) by high-sensitivity detection of the bioluminescence. In this chapter, we are introducing a basic concept of the optical fiber-based system and explaining how to construct and use this system.

  12. Influence of sediment composition on apparent toxicity in a solid-phase test using bioluminescent bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Benton, M.J. [East Tennessee State Univ., Johnson City, TN (United States). Dept. of Environmental Health; Malott, M.L. [Univ. of Mississippi, University, MS (United States)]|[Dept. of Agriculture, Oxford, MS (United States); Knight, S.S.; Cooper, C.M. [Dept. of Agriculture, Oxford, MS (United States); Benson, W.H. [Univ. of Mississippi, University, MS (United States)

    1995-03-01

    Clean and spiked sediment formulations of various silt:sand and clay:sand ratios were tested for toxicity using a bioassay that utilizes bioluminescent bacteria. Measured toxicities of clean and copper sulfate-spiked sediments were negatively but nonlinearly related with percent silt and percent clay, but no significant relationship existed between measured toxicity and sediment composition for methyl parathion-spiked formulations. Results suggest that solid-phase sediment bioassays using bioluminescence bacteria may be useful for testing the toxicities of single contaminants in formulated artificial sediments of known particle-size composition, and for repeated samples collected from the same site. However, extreme caution must be taken when testing sediments of varying composition or which may be differentially contaminated or contain a suite of contaminants.

  13. Effect of beta-adrenergic antagonists on bioluminescence control in three species of brittlestars (Echinodermata: Ophiuroidea).

    Science.gov (United States)

    Dupont, S; Mallefet, J; Vanderlinden, C

    2004-05-01

    The role of adrenaline in the nervous control of bioluminescence in three brittlestar species, Amphiura filiformis, Amphipholis squamata, and Ophiopsila aranea, was assessed by testing two different beta-adrenergic antagonists (propranolol and labetalol) over a wide concentration range (10(-10)-10(-3)M). We compared the effects of analogues (active vs. inactive) of the same substance (L- and D-enantiomers of propranolol). Propranolol presented both specific and nonspecific effects: (i) nonspecific effects were observed at the higher concentrations tested (10(-4) and 10(-3)M) in all three species; (ii) specific effects were detected only at the lower concentrations tested (10(-6)-10(-5)M). In A. squamata, the involvement of adrenaline in the nervous control of luminescence is supported by propranolol and labetolol specific inhibition. The neuropharmacological implications of nonspecific effects, the involvement of adrenaline and the interspecific differences in the brittlestar nervous control of bioluminescence are discussed.

  14. Investigating real-time activation of adenosine receptors by bioluminescence resonance energy transfer technique

    Science.gov (United States)

    Huang, Yimei; Yang, Hongqin; Zheng, Liqin; Chen, Jiangxu; Wang, Yuhua; Li, Hui; Xie, Shusen

    2013-02-01

    Adenosine receptors play important roles in many physiological and pathological processes, for example regulating myocardial oxygen consumption and the release of neurotransmitters. The activations of adenosine receptors have been studied by some kinds of techniques, such as western blot, immunohistochemistry, etc. However, these techniques cannot reveal the dynamical response of adenosine receptors under stimulation. In this paper, bioluminescence resonance energy transfer technique was introduced to study the real-time activation of adenosine receptors by monitoring the dynamics of cyclic adenosine monophosphate (cAMP) level. The results showed that there were significant differences between adenosine receptors on real-time responses under stimulation. Moreover, the dynamics of cAMP level demonstrated that competition between adenosine receptors existed. Taken together, our study indicates that monitoring the dynamics of cAMP level using bioluminescence resonance energy transfer technique could be one potential approach to investigate the mechanism of competitions between adenosine receptors.

  15. Bioluminescence imaging of β cells and intrahepatic insulin gene activity under normal and pathological conditions.

    Directory of Open Access Journals (Sweden)

    Tokio Katsumata

    Full Text Available In diabetes research, bioluminescence imaging (BLI has been applied in studies of β-cell impairment, development, and islet transplantation. To develop a mouse model that enables noninvasive imaging of β cells, we generated a bacterial artificial chromosome (BAC transgenic mouse in which a mouse 200-kbp genomic fragment comprising the insulin I gene drives luciferase expression (Ins1-luc BAC transgenic mouse. BLI of mice was performed using the IVIS Spectrum system after intraperitoneal injection of luciferin, and the bioluminescence signal from the pancreatic region analyzed. When compared with MIP-Luc-VU mice [FVB/N-Tg(Ins1-lucVUPwrs/J] expressing luciferase under the control of the 9.2-kbp mouse insulin I promoter (MIP, the bioluminescence emission from Ins1-luc BAC transgenic mice was enhanced approximately 4-fold. Streptozotocin-treated Ins1-luc BAC transgenic mice developed severe diabetes concomitant with a sharp decline in the BLI signal intensity in the pancreas. Conversely, mice fed a high-fat diet for 8 weeks showed an increase in the signal, reflecting a decrease or increase in the β-cell mass. Although the bioluminescence intensity of the islets correlated well with the number of isolated islets in vitro, the intensity obtained from a living mouse in vivo did not necessarily reflect an absolute quantification of the β-cell mass under pathological conditions. On the other hand, adenovirus-mediated gene transduction of β-cell-related transcription factors in Ins1-luc BAC transgenic mice generated luminescence from the hepatic region for more than 1 week. These results demonstrate that BLI in Ins1-luc BAC transgenic mice provides a noninvasive method of imaging islet β cells and extrapancreatic activity of the insulin gene in the liver under normal and pathological conditions.

  16. The use of terrestrial bacteria with natural bioluminescence in environmental toxicology

    Czech Academy of Sciences Publication Activity Database

    Lojek, Antonín; Číž, Milan; Kubala, Lukáš; Hyršl, P.; Buňková, Radka

    2007-01-01

    Roč. 101, č. 14 (2007), s116-s118 E-ISSN 1213-7103. [Mezioborová česko-slovenská toxikologická konference /12./. Praha, 11.06.2007-13.06.2007] R&D Projects: GA ČR(CZ) GA525/06/1196 Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : bioluminescence * bacteria * metal ions Subject RIV: BO - Biophysics

  17. Deep-sea bioluminescence blooms after dense water formation at the ocean surface.

    Science.gov (United States)

    Tamburini, Christian; Canals, Miquel; Durrieu de Madron, Xavier; Houpert, Loïc; Lefèvre, Dominique; Martini, Séverine; D'Ortenzio, Fabrizio; Robert, Anne; Testor, Pierre; Aguilar, Juan Antonio; Samarai, Imen Al; Albert, Arnaud; André, Michel; Anghinolfi, Marco; Anton, Gisela; Anvar, Shebli; Ardid, Miguel; Jesus, Ana Carolina Assis; Astraatmadja, Tri L; Aubert, Jean-Jacques; Baret, Bruny; Basa, Stéphane; Bertin, Vincent; Biagi, Simone; Bigi, Armando; Bigongiari, Ciro; Bogazzi, Claudio; Bou-Cabo, Manuel; Bouhou, Boutayeb; Bouwhuis, Mieke C; Brunner, Jurgen; Busto, José; Camarena, Francisco; Capone, Antonio; Cârloganu, Christina; Carminati, Giada; Carr, John; Cecchini, Stefano; Charif, Ziad; Charvis, Philippe; Chiarusi, Tommaso; Circella, Marco; Coniglione, Rosa; Costantini, Heide; Coyle, Paschal; Curtil, Christian; Decowski, Patrick; Dekeyser, Ivan; Deschamps, Anne; Donzaud, Corinne; Dornic, Damien; Dorosti, Hasankiadeh Q; Drouhin, Doriane; Eberl, Thomas; Emanuele, Umberto; Ernenwein, Jean-Pierre; Escoffier, Stéphanie; Fermani, Paolo; Ferri, Marcelino; Flaminio, Vincenzo; Folger, Florian; Fritsch, Ulf; Fuda, Jean-Luc; Galatà, Salvatore; Gay, Pascal; Giacomelli, Giorgio; Giordano, Valentina; Gómez-González, Juan-Pablo; Graf, Kay; Guillard, Goulven; Halladjian, Garadeb; Hallewell, Gregory; van Haren, Hans; Hartman, Joris; Heijboer, Aart J; Hello, Yann; Hernández-Rey, Juan Jose; Herold, Bjoern; Hößl, Jurgen; Hsu, Ching-Cheng; de Jong, Marteen; Kadler, Matthias; Kalekin, Oleg; Kappes, Alexander; Katz, Uli; Kavatsyuk, Oksana; Kooijman, Paul; Kopper, Claudio; Kouchner, Antoine; Kreykenbohm, Ingo; Kulikovskiy, Vladimir; Lahmann, Robert; Lamare, Patrick; Larosa, Giuseppina; Lattuada, Dario; Lim, Gordon; Presti, Domenico Lo; Loehner, Herbert; Loucatos, Sotiris; Mangano, Salvatore; Marcelin, Michel; Margiotta, Annarita; Martinez-Mora, Juan Antonio; Meli, Athina; Montaruli, Teresa; Moscoso, Luciano; Motz, Holger; Neff, Max; Nezri, Emma Nuel; Palioselitis, Dimitris; Păvălaş, Gabriela E; Payet, Kevin; Payre, Patrice; Petrovic, Jelena; Piattelli, Paolo; Picot-Clemente, Nicolas; Popa, Vlad; Pradier, Thierry; Presani, Eleonora; Racca, Chantal; Reed, Corey; Riccobene, Giorgio; Richardt, Carsten; Richter, Roland; Rivière, Colas; Roensch, Kathrin; Rostovtsev, Andrei; Ruiz-Rivas, Joaquin; Rujoiu, Marius; Russo, Valerio G; Salesa, Francisco; Sánchez-Losa, Augustin; Sapienza, Piera; Schöck, Friederike; Schuller, Jean-Pierre; Schussler, Fabian; Shanidze, Rezo; Simeone, Francesco; Spies, Andreas; Spurio, Maurizio; Steijger, Jos J M; Stolarczyk, Thierry; Taiuti, Mauro G F; Toscano, Simona; Vallage, Bertrand; Van Elewyck, Véronique; Vannoni, Giulia; Vecchi, Manuela; Vernin, Pascal; Wijnker, Guus; Wilms, Jorn; de Wolf, Els; Yepes, Harold; Zaborov, Dmitry; De Dios Zornoza, Juan; Zúñiga, Juan

    2013-01-01

    The deep ocean is the largest and least known ecosystem on Earth. It hosts numerous pelagic organisms, most of which are able to emit light. Here we present a unique data set consisting of a 2.5-year long record of light emission by deep-sea pelagic organisms, measured from December 2007 to June 2010 at the ANTARES underwater neutrino telescope in the deep NW Mediterranean Sea, jointly with synchronous hydrological records. This is the longest continuous time-series of deep-sea bioluminescence ever recorded. Our record reveals several weeks long, seasonal bioluminescence blooms with light intensity up to two orders of magnitude higher than background values, which correlate to changes in the properties of deep waters. Such changes are triggered by the winter cooling and evaporation experienced by the upper ocean layer in the Gulf of Lion that leads to the formation and subsequent sinking of dense water through a process known as "open-sea convection". It episodically renews the deep water of the study area and conveys fresh organic matter that fuels the deep ecosystems. Luminous bacteria most likely are the main contributors to the observed deep-sea bioluminescence blooms. Our observations demonstrate a consistent and rapid connection between deep open-sea convection and bathypelagic biological activity, as expressed by bioluminescence. In a setting where dense water formation events are likely to decline under global warming scenarios enhancing ocean stratification, in situ observatories become essential as environmental sentinels for the monitoring and understanding of deep-sea ecosystem shifts.

  18. Deep-sea bioluminescence blooms after dense water formation at the ocean surface.

    Directory of Open Access Journals (Sweden)

    Christian Tamburini

    Full Text Available The deep ocean is the largest and least known ecosystem on Earth. It hosts numerous pelagic organisms, most of which are able to emit light. Here we present a unique data set consisting of a 2.5-year long record of light emission by deep-sea pelagic organisms, measured from December 2007 to June 2010 at the ANTARES underwater neutrino telescope in the deep NW Mediterranean Sea, jointly with synchronous hydrological records. This is the longest continuous time-series of deep-sea bioluminescence ever recorded. Our record reveals several weeks long, seasonal bioluminescence blooms with light intensity up to two orders of magnitude higher than background values, which correlate to changes in the properties of deep waters. Such changes are triggered by the winter cooling and evaporation experienced by the upper ocean layer in the Gulf of Lion that leads to the formation and subsequent sinking of dense water through a process known as "open-sea convection". It episodically renews the deep water of the study area and conveys fresh organic matter that fuels the deep ecosystems. Luminous bacteria most likely are the main contributors to the observed deep-sea bioluminescence blooms. Our observations demonstrate a consistent and rapid connection between deep open-sea convection and bathypelagic biological activity, as expressed by bioluminescence. In a setting where dense water formation events are likely to decline under global warming scenarios enhancing ocean stratification, in situ observatories become essential as environmental sentinels for the monitoring and understanding of deep-sea ecosystem shifts.

  19. Dual monitoring using 124I-FIAU and bioluminescence for HSV1-tk suicide gene therapy

    International Nuclear Information System (INIS)

    Lee, T. S.; Kim, J. H.; Kwon, H. C.

    2007-01-01

    Herpes simplex virus type I thymidine kinase (HSV-tk) is the most common reporter gene and is used in cancer gene therapy with a prodrug nucleoside analog, ganciclovir (GCV). The aim of this study is to evaluate therapeutic efficacy of suicide gene therapy with 2'-fluoro-2'-deoxy-1-D-arabinofuranosyl-5-[ 124 I] iodouracil ( 124 I - FIAU) and bioluminescence in retrovirally HSV -tk and firefly luciferase transduced hepatoma model. The HSV -tk and firefly luciferase (Luc) was retrovirally transduced and expressed in MCA rat Morris hepatoma cells. Nude mice with subcutaneous tumors, MCA and MCA-TK-Luc, were subjected to GCV treatment (50mg/Kg/d intraperitoneally) for 5 day. PET imaging and biodistribution with ( 124 I-FIAU) were performed at before and after initiation of therapy with GCV. Bioluminescent signal was also measured during GCV treatment. Before GCV treatment, no significant difference in tumor volume was found in tumors between MCA and MCA-TK-Luc. After GCV treatment, tumor volume of MCA-TK-Luc markedly reduced compared to that of MCA. In biodistribution study, 124 I-FIAU uptake after GCV therapy significantly decreased compared with pretreatment levels (34.8 13.67 %ID/g vs 7.6 2.59 %ID/g) and bioluminescent signal was also significantly decreased compared with pretreatment levels. In small animal PET imaging, 124 I-FIAU selectively localized in HSV -tk expressing tumor and the therapeutic efficacy of GCV treatment was evaluated by 124 I-FIAU PET imaging. 124 I-FIAU PET and bioluminescence imaging in HSV-tk suicide gene therapy were effective to evaluate the therapeutic response. 124 I-FIAU may serve as an efficient and selective agent for monitoring of transduced HSV1-tk gene expression in vivo in clinical trials

  20. Melatonin Entrains PER2::LUC Bioluminescence Circadian Rhythm in the Mouse Cornea.

    Science.gov (United States)

    Baba, Kenkichi; Davidson, Alec J; Tosini, Gianluca

    2015-07-01

    Previous studies have reported the presence of a circadian rhythm in PERIOD2::LUCIFERASE (PER2::LUC) bioluminescence in mouse photoreceptors, retina, RPE, and cornea. Melatonin (MLT) modulates many physiological functions in the eye and it is believed to be one of the key circadian signals within the eye. The aim of the present study was to investigate the regulation of the PER2::LUC circadian rhythm in mouse cornea and to determine the role played by MLT. Corneas were obtained from PER2::LUC mice and cultured to measure bioluminescence rhythmicity in isolated tissue using a Lumicycle or CCD camera. To determine the time-dependent resetting of the corneal circadian clocks in response to MLT or IIK7 (a melatonin type 2 receptor, MT2, agonist) was added to the cultured corneas at different times of the day. We also defined the location of the MT2 receptor within different corneal layers using immunohistochemistry. A long-lasting bioluminescence rhythm was recorded from cultured PER2::LUC cornea and PER2::LUC signal was localized to the corneal epithelium and endothelium. MLT administration in the early night delayed the cornea rhythm, whereas administration of MLT at late night to early morning advanced the cornea rhythm. Treatment with IIK7 mimicked the MLT phase-shifting effect. Consistent with these results, MT2 immunoreactivity was localized to the corneal epithelium and endothelium. Our work demonstrates that MLT entrains the PER2::LUC bioluminescence rhythm in the cornea. Our data indicate that the cornea may represent a model to study the molecular mechanisms by which MLT affects the circadian clock.

  1. Image Processing for Bioluminescence Resonance Energy Transfer Measurement—BRET-Analyzer

    OpenAIRE

    Chastagnier, Yan; Moutin, Enora; Hemonnot, Anne-Laure; Perroy, Julie

    2018-01-01

    A growing number of tools now allow live recordings of various signaling pathways and protein-protein interaction dynamics in time and space by ratiometric measurements, such as Bioluminescence Resonance Energy Transfer (BRET) Imaging. Accurate and reproducible analysis of ratiometric measurements has thus become mandatory to interpret quantitative imaging. In order to fulfill this necessity, we have developed an open source toolset for Fiji—BRET-Analyzer—allowing a systematic analysis, from ...

  2. Monitoring of gene expression in bacteria during infections using an adaptable set of bioluminescent, fluorescent and colorigenic fusion vectors.

    Directory of Open Access Journals (Sweden)

    Frank Uliczka

    Full Text Available A family of versatile promoter-probe plasmids for gene expression analysis was developed based on a modular expression plasmid system (pZ. The vectors contain different replicons with exchangeable antibiotic cassettes to allow compatibility and expression analysis on a low-, midi- and high-copy number basis. Suicide vector variants also permit chromosomal integration of the reporter fusion and stable vector derivatives can be used for in vivo or in situ expression studies under non-selective conditions. Transcriptional and translational fusions to the reporter genes gfp(mut3.1, amCyan, dsRed2, luxCDABE, phoA or lacZ can be constructed, and presence of identical multiple cloning sites in the vector system facilitates the interchange of promoters or reporter genes between the plasmids of the series. The promoter of the constitutively expressed gapA gene of Escherichia coli was included to obtain fluorescent and bioluminescent expression constructs. A combination of the plasmids allows simultaneous detection and gene expression analysis in individual bacteria, e.g. in bacterial communities or during mouse infections. To test our vector system, we analyzed and quantified expression of Yersinia pseudotuberculosis virulence genes under laboratory conditions, in association with cells and during the infection process.

  3. Monitoring of gene expression in bacteria during infections using an adaptable set of bioluminescent, fluorescent and colorigenic fusion vectors.

    Science.gov (United States)

    Uliczka, Frank; Pisano, Fabio; Kochut, Annika; Opitz, Wiebke; Herbst, Katharina; Stolz, Tatjana; Dersch, Petra

    2011-01-01

    A family of versatile promoter-probe plasmids for gene expression analysis was developed based on a modular expression plasmid system (pZ). The vectors contain different replicons with exchangeable antibiotic cassettes to allow compatibility and expression analysis on a low-, midi- and high-copy number basis. Suicide vector variants also permit chromosomal integration of the reporter fusion and stable vector derivatives can be used for in vivo or in situ expression studies under non-selective conditions. Transcriptional and translational fusions to the reporter genes gfp(mut3.1), amCyan, dsRed2, luxCDABE, phoA or lacZ can be constructed, and presence of identical multiple cloning sites in the vector system facilitates the interchange of promoters or reporter genes between the plasmids of the series. The promoter of the constitutively expressed gapA gene of Escherichia coli was included to obtain fluorescent and bioluminescent expression constructs. A combination of the plasmids allows simultaneous detection and gene expression analysis in individual bacteria, e.g. in bacterial communities or during mouse infections. To test our vector system, we analyzed and quantified expression of Yersinia pseudotuberculosis virulence genes under laboratory conditions, in association with cells and during the infection process.

  4. An ebCMOS camera system for marine bioluminescence observation: The LuSEApher prototype

    Energy Technology Data Exchange (ETDEWEB)

    Dominjon, A., E-mail: a.dominjon@ipnl.in2p3.fr [CNRS/IN2P3, Institut de Physique Nucleaire de Lyon, Villeurbanne F-69622 (France); Ageron, M. [CNRS/IN2P3, Centre de Physique des Particules de Marseille, Marseille, F-13288 (France); Barbier, R. [CNRS/IN2P3, Institut de Physique Nucleaire de Lyon, Villeurbanne F-69622 (France); Universite de Lyon, Universite Lyon 1, Lyon F-69003 (France); Billault, M.; Brunner, J. [CNRS/IN2P3, Centre de Physique des Particules de Marseille, Marseille, F-13288 (France); Cajgfinger, T. [CNRS/IN2P3, Institut de Physique Nucleaire de Lyon, Villeurbanne F-69622 (France); Universite de Lyon, Universite Lyon 1, Lyon F-69003 (France); Calabria, P. [CNRS/IN2P3, Institut de Physique Nucleaire de Lyon, Villeurbanne F-69622 (France); Chabanat, E. [CNRS/IN2P3, Institut de Physique Nucleaire de Lyon, Villeurbanne F-69622 (France); Universite de Lyon, Universite Lyon 1, Lyon F-69003 (France); Chaize, D.; Doan, Q.T.; Guerin, C.; Houles, J.; Vagneron, L. [CNRS/IN2P3, Institut de Physique Nucleaire de Lyon, Villeurbanne F-69622 (France)

    2012-12-11

    The ebCMOS camera, called LuSEApher, is a marine bioluminescence recorder device adapted to extreme low light level. This prototype is based on the skeleton of the LUSIPHER camera system originally developed for fluorescence imaging. It has been installed at 2500 m depth off the Mediterranean shore on the site of the ANTARES neutrino telescope. The LuSEApher camera is mounted on the Instrumented Interface Module connected to the ANTARES network for environmental science purposes (European Seas Observatory Network). The LuSEApher is a self-triggered photo detection system with photon counting ability. The presentation of the device is given and its performances such as the single photon reconstruction, noise performances and trigger strategy are presented. The first recorded movies of bioluminescence are analyzed. To our knowledge, those types of events have never been obtained with such a sensitivity and such a frame rate. We believe that this camera concept could open a new window on bioluminescence studies in the deep sea.

  5. Disposable bioluminescence-based biosensor for detection of bacterial count in food.

    Science.gov (United States)

    Luo, Jinping; Liu, Xiaohong; Tian, Qing; Yue, Weiwei; Zeng, Jing; Chen, Guangquan; Cai, Xinxia

    2009-11-01

    A biosensor for rapid detection of bacterial count based on adenosine 5'-triphosphate (ATP) bioluminescence has been developed. The biosensor is composed of a key sensitive element and a photomultiplier tube used as a detector element. The disposable sensitive element consists of a sampler, a cartridge where intracellular ATP is chemically extracted from bacteria, and a microtube where the extracted ATP reacts with the luciferin-luciferase reagent to produce bioluminescence. The bioluminescence signal is transformed into relevant electrical signal by the detector and further measured with a homemade luminometer. Parameters affecting the amount of the extracted ATP, including the types of ATP extractants, the concentrations of ATP extractant, and the relevant neutralizing reagent, were optimized. Under the optimal experimental conditions, the biosensor showed a linear response to standard bacteria in a concentration range from 10(3) to 10(8) colony-forming units (CFU) per milliliter with a correlation coefficient of 0.925 (n=22) within 5min. Moreover, the bacterial count of real food samples obtained by the biosensor correlated well with those by the conventional plate count method. The proposed biosensor, with characteristics of low cost, easy operation, and fast response, provides potential application to rapid evaluation of bacterial contamination in the food industry, environment monitoring, and other fields.

  6. Distribution of bioluminescent fungi across old-growth and secondary tropical rain forest in Costa Rica.

    Science.gov (United States)

    Seas-Carvajal, Carolina; Avalos, Gerardo

    2013-06-01

    Most research on bioluminescent fungi is concentrated on their taxonomic relationships, while the basics of their natural history and ecological relationships are poorly understood. In this study, we compared the distribution of bioluminescent fungi between old-growth and secondary forest as related to four different soil types at the tropical rainforest of La Selva Biological Station in Costa Rica. The study was conducted during the wet season of 2009. Bioluminescent fungi were sought following eight different transects distributed evenly in old-growth and secondary forests across four different soil types, covering an area of 9 420m2. We found fungi in four different substrates: litter, fallen branches, dead trunks, and roots, for a total of 61 samples. Correspondence analysis showed that the occurrence of fungi and soil types were related (inertia = 0.21, p = 0.071). We found a significant relationship between the presence of fungi and the distribution of soil types (X2 = 18.89, df = 9, p = 0.026). We found only three samples with fruiting bodies, two of which had Mycena and the other had one fungus of the order Xylariales (possibly Hypoxylon sp., Kretzschmariella sp., Xylaria sp.). Future work will concentrate on exploring other aspects of their ecology, such as their dispersal and substrate preference. This information will facilitate field identification and will foster more research on the distribution, seasonality, reproductive phenology and ecological requirements of this group of Fungi.

  7. Detection of Bacillus anthracis spores from environmental water using bioluminescent reporter phage.

    Science.gov (United States)

    Nguyen, C; Makkar, R; Sharp, N J; Page, M A; Molineux, I J; Schofield, D A

    2017-11-01

    We investigated the ability of a temperate Bacillus anthracis reporter phage (Wβ::luxAB-2), which transduces bioluminescence to infected cells, to detect viable spores from deliberately contaminated environmental water samples. Environmental water was inoculated with spores and assayed with Wβ::luxAB-2. Bioluminescent signals directly correlated with input phage and spore concentrations. A limit of detection of 10 1 and 10 2 CFU per ml within 8 h was achieved from pond and lake water, respectively. Detection was greatly simplified by minimizing sample processing steps without spore extraction. The complex endogenous microbial flora and salt content of brackish water challenged the assay, extending the detection time to 12 h for a sensitivity of 10 2 CFU per ml. Phage-mediated bioluminescence was strictly dependent on bacterial physiology, being significantly reduced in mid/late log phase cells. This was shown to be due to an inability of the phage to adsorb. The reporter phage Wβ::luxAB-2 displays potential for simplified detection of viable spores from contaminated water samples within 12 h. A deliberate aerosol release of spores could lead to widespread contamination, leaving large areas uninhabitable until remediation. An essential requirement of this restoration process is the development of simplified detection assays in different environmental matrices. © 2017 The Society for Applied Microbiology.

  8. Development of Optical Molecular Imaging System for the Acquisition of Bioluminescence Signals from Small Animals

    International Nuclear Information System (INIS)

    Lee, Byeong Il; Kim, Hyeon Sik; Jeong, Hye Jin; Lee, Hyung Jae; Moon, Seung Min; Kwon, Seung Young; Jeong, Shin Young; Bom, Hee Seung; Min, Jung Joon; Choi, Eun Seo

    2009-01-01

    Optical imaging is providing great advance and improvement in genetic and molecular imaging of animals and humans. Optical imaging system consists of optical imaging devices, which carry out major function for monitoring, tracing, and imaging in most of molecular in-vivo researches. In bio-luminescent imaging, small animals containing luciferase gene locally irradiate light, and emitted photons transmitted through skin of the small animals are imaged by using a high sensitive charged coupled device (CCD) camera. In this paper, we introduced optical imaging system for the image acquisition of bio-luminescent signals emitted from small animals. In the system, Nikon lens and four LED light sources were mounted at the inside of a dark box. A cooled CCD camera equipped with a control module was used. We tested the performance of the optical imaging system using effendorf tube and light emitting bacteria which injected intravenously into CT26 tumor bearing nude mouse. The performance of implemented optical imaging system for bio-luminescence imaging was demonstrated and the feasibility of the system in small animal imaging application was proved. We anticipate this system could be a useful tool for the molecular imaging of small animals adaptable for various experimental conditions in future

  9. Factors Influencing Quantification of in Vivo Bioluminescence Imaging: Application to Assessment of Pancreatic Islet Transplants

    Directory of Open Access Journals (Sweden)

    John Virostko

    2004-10-01

    Full Text Available The aim of this study is to determine and characterize factors influencing in vivo bioluminescence imaging (BLI and apply them to the specific application of imaging transplanted pancreatic islets. Noninvasive quantitative assessment of transplanted pancreatic islets poses a formidable challenge. Murine pancreatic islets expressing firefly luciferase were transplanted under the renal capsule or into the portal vein of nonobese diabetic–severe combined immunodeficiency mice and the bioluminescence was quantified with a cooled charge coupled device camera and digital photon image analysis. The important, but often neglected, effects of wound healing, mouse positioning, and transplantation site on bioluminescence measurements were investigated by imaging a constant emission, isotropic light-emitting bead (λ = 600 implanted at the renal or hepatic site. The renal beads emitted nearly four times more light than hepatic beads with a smaller spot size, indicating that light absorption and scatter are greatly influenced by the transplant site and must be accounted for in BLI measurements. Detected luminescence decreased with increasing angle between the mouse surface normal and optical axis. By defining imaging parameters such as postsurgical effects, animal positioning, and light attenuation as a function of transplant site, this study develops BLI as a useful imaging modality for quantitative assessment of islets post-transplantation.

  10. Distribution of bioluminescent fungi across old-growth and secondary tropical rain forest in Costa Rica

    Directory of Open Access Journals (Sweden)

    Carolina Seas-Carvajal

    2013-06-01

    Full Text Available Most research on bioluminescent fungi is concentrated on their taxonomic relationships, while the basics of their natural history and ecological relationships are poorly understood. In this study, we compared the distribution of bioluminescent fungi between old-growth and secondary forest as related to four different soil types at the tropical rainforest of La Selva Biological Station in Costa Rica. The study was conducted during the wet season of 2009. Bioluminescent fungi were sought following eight different transects distributed evenly in old-growth and secondary forests across four different soil types, covering an area of 9 420m². We found fungi in four different substrates: litter, fallen branches, dead trunks, and roots, for a total of 61 samples. Correspondence analysis showed that the occurrence of fungi and soil types were related (inertia=0.21, p=0.071. We found a significant relationship between the presence of fungi and the distribution of soil types (X²=18.89, df=9, p=0.026. We found only three samples with fruiting bodies, two of which had Mycena and the other had one fungus of the order Xylariales (possibly Hypoxylon sp., Kretzschmariella sp., Xylaria sp.. Future work will concentrate on exploring other aspects of their ecology, such as their dispersal and substrate preference. This information will facilitate field identification and will foster more research on the distribution, seasonality, reproductive phenology and ecological requirements of this group of Fungi.

  11. Bioluminescent imaging: a critical tool in pre-clinical oncology research.

    LENUS (Irish Health Repository)

    O'Neill, Karen

    2010-02-01

    Bioluminescent imaging (BLI) is a non-invasive imaging modality widely used in the field of pre-clinical oncology research. Imaging of small animal tumour models using BLI involves the generation of light by luciferase-expressing cells in the animal following administration of substrate. This light may be imaged using an external detector. The technique allows a variety of tumour-associated properties to be visualized dynamically in living models. The increasing use of BLI as a small-animal imaging modality has led to advances in the development of xenogeneic, orthotopic, and genetically engineered animal models expressing luciferase genes. This review aims to provide insight into the principles of BLI and its applications in cancer research. Many studies to assess tumour growth and development, as well as efficacy of candidate therapeutics, have been performed using BLI. More recently, advances have also been made using bioluminescent imaging in studies of protein-protein interactions, genetic screening, cell-cycle regulators, and spontaneous cancer development. Such novel studies highlight the versatility and potential of bioluminescent imaging in future oncological research.

  12. A two-hour antibiotic susceptibility test by ATP-bioluminescence.

    Science.gov (United States)

    March Rosselló, Gabriel Alberto; García-Loygorri Jordán de Urries, María Cristina; Gutiérrez Rodríguez, María Purificación; Simarro Grande, María; Orduña Domingo, Antonio; Bratos Pérez, Miguel Ángel

    2016-01-01

    The antibiotic susceptibility test (AST) in Clinical Microbiology laboratories is still time-consuming, and most procedures take 24h to yield results. In this study, a rapid antimicrobial susceptibility test using ATP-bioluminescence has been developed. The design of method was performed using five ATCC collection strains of known susceptibility. This procedure was then validated against standard commercial methods on 10 strains of enterococci, 10 staphylococci, 10 non-fermenting gram negative bacilli, and 13 Enterobacteriaceae from patients. The agreement obtained in the sensitivity between the ATP-bioluminescence method and commercial methods (E-test, MicroScan and VITEK2) was 100%. In summary, the preliminary results obtained in this work show that the ATP-bioluminescence method could provide a fast and reliable AST in two hours. Copyright © 2015 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

  13. Characterization of an anthraquinone fluor from the bioluminescent, pelagic polychaete Tomopteris.

    Science.gov (United States)

    Francis, Warren R; Powers, Meghan L; Haddock, Steven H D

    2014-12-01

    Tomopteris is a cosmopolitan genus of polychaetes. Many species produce yellow luminescence in the parapodia when stimulated. Yellow bioluminescence is rare in the ocean, and the components of this luminescent reaction have not been identified. Only a brief description, half a century ago, noted fluorescence in the parapodia with a remarkably similar spectrum to the bioluminescence, which suggested that it may be the luciferin or terminal light-emitter. Here, we report the isolation of the fluorescent yellow-orange pigment found in the luminous exudate and in the body of the animals. Liquid chromatography-mass spectrometry revealed the mass to be 270 m/z with a molecular formula of C(15)H(10)O(5), which ultimately was shown to be aloe-emodin, an anthraquinone previously found in plants. We speculate that aloe-emodin could be a factor for resonant-energy transfer or the oxyluciferin for Tomopteris bioluminescence. © 2014 The Authors. Luminescence published by John Wiley & Sons Ltd.

  14. A multi-channel bioluminescent bacterial biosensor for the on-line detection of metals and toxicity. Part I: design and optimization of bioluminescent bacterial strains

    Energy Technology Data Exchange (ETDEWEB)

    Charrier, Thomas; Durand, Marie-Jose; Jouanneau, Sulivan; Thouand, Gerald [UMR CNRS 6144 GEPEA, CBAC, Nantes University, PRES UNAM, Campus de la Courtaisiere-IUT, La Roche-sur-Yon cedex (France); Dion, Michel [UMR CNRS 6204, Nantes University, PRES UNAM, Biotechnologie, Biocatalyse, Bioregulation, 2, Rue de la Houssiniere, BP 92208, Nantes cedex 3 (France); Pernetti, Mimma; Poncelet, Denis [ONIRIS-ENITIAA, UMR CNRS GEPEA, Rue de la Geraudiere, BP 82225, Nantes cedex 3 (France)

    2011-05-15

    This study describes the construction of inducible bioluminescent strains via genetic engineering along with their characterization and optimization in the detection of heavy metals. Firstly, a preliminary comparative study enabled us to select a suitable carbon substrate from pyruvate, glucose, citrate, diluted Luria-Bertani, and acetate. The latter carbon source provided the best induction ratios for comparison. Results showed that the three constructed inducible strains, Escherichia coli DH1 pBzntlux, pBarslux, and pBcoplux, were usable when conducting a bioassay after a 14-h overnight culture at 30 C. Utilizing these sensors gave a range of 12 detected heavy metals including several cross-detections. Detection limits for each metal were often close to and sometimes lower than the European standards for water pollution. Finally, in order to maintain sensitive bacteria within the future biosensor-measuring cell, the agarose immobilization matrix was compared to polyvinyl alcohol (PVA). Agarose was selected because the detection limits of the bioluminescent strains were not affected, in contrast to PVA. Specific detection and cross-detection ranges determined in this study will form the basis of a multiple metals detection system by the new multi-channel Lumisens3 biosensor. (orig.)

  15. N-Alkylated 6'-aminoluciferins are bioluminescent substrates for Ultra-Glo and QuantiLum luciferase: new potential scaffolds for bioluminescent assays.

    Science.gov (United States)

    Woodroofe, Carolyn C; Shultz, John W; Wood, Monika G; Osterman, Jean; Cali, James J; Daily, William J; Meisenheimer, Poncho L; Klaubert, Dieter H

    2008-09-30

    A set of 6'-alkylated aminoluciferins are shown to be bioluminescent substrates for Ultra-Glo and QuantiLum luciferases. These studies demonstrate that both the engineered and wild-type firefly luciferases tolerate much greater steric bulk at the 6' position of luciferin than has been previously reported. The nature of the alkyl substituent strongly affects the strength of the bioluminescent signal, which varies widely based on size, shape, and charge. Several compounds were observed to generate more light than the corresponding unsubstituted 6'-aminoluciferin. Determination of Michaelis-Menten constants for the substrates with Ultra-Glo indicated that the variation arises primarily from differences in V max, ranging from 1.33 x 10 (4) to 332 x 10 (4) relative light units, but in some cases K m (0.73-10.8 microM) also plays a role. Molecular modeling results suggest that interactions of the side chain with a hydrogen-bonding network at the base of the luciferin binding pocket may influence substrate-enzyme binding.

  16. Uptake kinetics and biodistribution of 14C-d-luciferin - a radiolabeled substrate for the firefly luciferase catalyzed bioluminescence reaction: impact on bioluminescence based reporter gene imaging

    International Nuclear Information System (INIS)

    Berger, Frank; Bhaumik, Srabani

    2008-01-01

    Firefly luciferase catalyzes the oxidative decarboxylation of d-luciferin to oxyluciferin in the presence of cofactors, producing bioluminescence. This reaction is used in optical bioluminescence-based molecular imaging approaches to detect the expression of the firefly luciferase reporter gene. Biokinetics and distribution of the substrate most likely have a significant impact on levels of light signal and therefore need to be investigated. Benzene ring 14 C(U)-labeled d-luciferin was utilized. Cell uptake and efflux assays, murine biodistribution, autoradiography and CCD-camera based optical bioluminescence imaging were carried out to examine the in vitro and in vivo characteristics of the tracer in cell culture and in living mice respectively. Radiolabeled and unlabeled d-luciferin revealed comparable levels of light emission when incubated with equivalent amounts of the firefly luciferase enzyme. Cell uptake assays in pCMV-luciferase-transfected cells showed slow trapping of the tracer and relatively low uptake values (up to 22.9-fold higher in firefly luciferase gene-transfected vs. nontransfected cells, p=0.0002). Biodistribution studies in living mice after tail-vein injection of 14 C-d-luciferin demonstrated inhomogeneous tracer distribution with early predominant high radioactivity levels in kidneys (10.6% injected dose [ID]/g) and liver (11.9% ID/g), followed at later time points by the bladder (up to 81.3% ID/g) and small intestine (6.5% ID/g), reflecting the elimination routes of the tracer. Kinetics and uptake levels profoundly differed when using alternate injection routes (intravenous versus intraperitoneal). No clear trapping of 14 C-d-luciferin in firefly luciferase-expressing tissues could be observed in vivo. The data obtained with 14 C-d-luciferin provide insights into the dynamics of d-luciferin cell uptake, intracellular accumulation, and efflux. Results of the biodistribution and autoradiographic studies should be useful for optimizing and

  17. Tomographic bioluminescence imaging by use of a combined optical-PET (OPET) system: a computer simulation feasibility study

    Energy Technology Data Exchange (ETDEWEB)

    Alexandrakis, George [David Geffen School of Medicine at UCLA, Crump Institute for Molecular Imaging, University of California, 700 Westwood Plaza, Los Angeles, CA 90095 (United States); Rannou, Fernando R [Departamento de Ingenieria Informatica, Universidad de Santiago de Chile (USACH), Av. Ecuador 3659, Santiago (Chile); Chatziioannou, Arion F [David Geffen School of Medicine at UCLA, Crump Institute for Molecular Imaging, University of California, 700 Westwood Plaza, Los Angeles, CA 90095 (United States)

    2005-09-07

    The feasibility and limits in performing tomographic bioluminescence imaging with a combined optical-PET (OPET) system were explored by simulating its image formation process. A micro-MRI based virtual mouse phantom was assigned appropriate tissue optical properties to each of its segmented internal organs at wavelengths spanning the emission spectrum of the firefly luciferase at 37 deg. C. The TOAST finite-element code was employed to simulate the diffuse transport of photons emitted from bioluminescence sources in the mouse. OPET measurements were simulated for single-point, two-point and distributed bioluminescence sources located in different organs such as the liver, the kidneys and the gut. An expectation maximization code was employed to recover the intensity and location of these simulated sources. It was found that spectrally resolved measurements were necessary in order to perform tomographic bioluminescence imaging. The true location of emission sources could be recovered if the mouse background optical properties were known a priori. The assumption of a homogeneous optical property background proved inadequate for describing photon transport in optically heterogeneous tissues and led to inaccurate source localization in the reconstructed images. The simulation results pointed out specific methodological challenges that need to be addressed before a practical implementation of OPET-based bioluminescence tomography is achieved.

  18. A genetic screen for bioluminescence genes in the fungus Armillaria mellea, through the use of Agrobacterium tumefaciens-mediated random insertional mutagenesis

    Science.gov (United States)

    Bioluminescence is reported from 71 saprobic species of fungi from four, distant lineages in the order Agaricales. Analyses of the fungal luminescent chemistry shows that all four lineages share a functionally conserved substrate and luciferase, indicating that the bioluminescent pathway is likely c...

  19. Bioluminescent detection of peroxynitrite with a boronic acid-caged luciferin.

    Science.gov (United States)

    Sieracki, Nathan A; Gantner, Benjamin N; Mao, Mao; Horner, John H; Ye, Richard D; Malik, Asrar B; Newcomb, Martin E; Bonini, Marcelo G

    2013-08-01

    Peroxynitrite, a highly reactive biological oxidant, is formed under pathophysiologic conditions from the diffusion-limited reaction of nitric oxide and superoxide radical anion. Peroxynitrite has been implicated as the mediator of nitric oxide toxicity in many diseases and as an important signaling disrupting molecule (L. Liaudet et al., Front. Biosci.14, 4809-4814, 2009) [1]. Biosensors effective at capturing peroxynitrite in a specific and fast enough manner for detection, along with readouts compatible with in vivo studies, are lacking. Here we report that the boronic acid-based bioluminescent system PCL-1 (peroxy-caged luciferin-1), previously reported as a chemoselective sensor for hydrogen peroxide (G.C. Van de Bittner et al., Proc. Natl. Acad. Sci. USA107, 21316-21321, 2010) [2], reacts with peroxynitrite stoichiometrically with a rate constant of 9.8±0.3×10(5)M(-1)s(-1) and a bioluminescence detection limit of 16nM, compared to values of 1.2±0.3M(-1)s(-1) and 231nM for hydrogen peroxide. Further, we demonstrate bioluminescent detection of peroxynitrite in the presence of physiological competitors: carbon dioxide, glutathione, albumin, and catalase. We also demonstrate the utility of this method to assess peroxynitrite formation in mammalian cells by measuring peroxynitrite generated under normal culture conditions after stimulation of macrophages with bacterial endotoxin lipopolysaccharide. Thus, the PCL-1 method for measuring peroxynitrite generation shows superior selectivity over other oxidants under in vivo conditions. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. In vivo bioluminescence imaging of Burkholderia mallei respiratory infection and treatment in the mouse model

    Directory of Open Access Journals (Sweden)

    Shane eMassey

    2011-08-01

    Full Text Available Bioluminescent imaging (BLI technology is a powerful tool for monitoring infectious disease progression and treatment approaches. BLI is particularly useful for tracking fastidious intracellular pathogens that might be difficult to recover from certain organs. Burkholderia mallei, the causative agent of glanders, is a facultative intracellular pathogen and has been classified by the CDC as a Category B select agent due to its highly infectious nature and potential use as a biological weapon. Very little is known regarding pathogenesis or treatment of glanders. We investigated the use of bioluminescent reporter constructs to monitor the dynamics of infection as well as the efficacy of therapeutics for B. mallei in real time. A stable luminescent reporter B. mallei strain was created using the pUTmini-Tn5::luxKm2 plasmid and used to monitor glanders in the BALB/c murine model. Mice were infected via the intranasal route with 5x103 bacteria and monitored by BLI at 24, 48 and 72 h. We verified that our reporter construct maintained similar virulence and growth kinetics compared to wild-type B. mallei and confirmed that it maintains luminescent stability in the presence or absence of antibiotic selection. The luminescent signal was initially seen in the lungs, and progressed to the liver and spleen over the course of infection. We demonstrated that antibiotic treatment 24 h post-infection resulted in reduction of bioluminescence that can be attributed to decreased bacterial burden in target organs. These findings suggest that BLI can be used to monitor disease progression and efficacy of therapeutics during glanders infections. Finally, we report an alternative method to mini-Tn5::luxKm2 transposon using mini-Tn7-lux elements that insert site-specifically at known genomic attachment sites and that can also be used to tag bacteria.

  1. Bioluminescence of beetle luciferases with 6'-amino-D-luciferin analogues reveals excited keto-oxyluciferin as the emitter and phenolate/luciferin binding site interactions modulate bioluminescence colors.

    Science.gov (United States)

    Viviani, Vadim R; Neves, Deimison Rodrigues; Amaral, Danilo Trabuco; Prado, Rogilene A; Matsuhashi, Takuto; Hirano, Takashi

    2014-08-19

    Beetle luciferases produce different bioluminescence colors from green to red using the same d-luciferin substrate. Despite many studies of the mechanisms and structural determinants of bioluminescence colors with firefly luciferases, the identity of the emitters and the specific active site interactions responsible for bioluminescence color modulation remain elusive. To address these questions, we analyzed the bioluminescence spectra with 6'-amino-D-luciferin (aminoluciferin) and its 5,5-dimethyl analogue using a set of recombinant beetle luciferases that naturally elicit different colors and different pH sensitivities (pH-sensitive, Amydetes vivianii λmax=538 nm, Macrolampis sp2 λmax=564 nm; pH-insensitive, Phrixotrix hirtus λmax=623 nm, Phrixotrix vivianii λmax=546 nm, and Pyrearinus termitilluminans λmax=534 nm), a luciferase-like enzyme (Tenebrionidae, Zophobas morio λmax=613 nm), and mutants of C311 (S314). The green-yellow-emitting luciferases display red-shifted bioluminescence spectra with aminoluciferin in relation to those with D-luciferin, whereas the red-emitting luciferases displayed blue-shifted spectra. Bioluminescence spectra with 5,5-dimethylaminoluciferin, in which enolization is blocked, were almost identical to those of aminoluciferin. Fluorescence probing using 2-(4-toluidino)naphthalene-6-sulfonate and inference with aminoluciferin confirm that the luciferin binding site of the red-shifted luciferases is more polar than in the case of the green-yellow-emitting luciferases. Altogether, the results show that the keto form of excited oxyluciferin is the emitter in beetle bioluminescence and that bioluminescence colors are essentially modulated by interactions of the 6'-hydroxy group of oxyluciferin and basic moieties under the influence of the microenvironment polarity of the active site: a strong interaction between a base moiety and oxyluciferin phenol in a hydrophobic microenvironment promotes green-yellow emission, whereas a more polar

  2. [Determination of minimal concentrations of biocorrosion inhibitors by a bioluminescence method in relation to bacteria, participating in biocorrosion].

    Science.gov (United States)

    Efremenko, E N; Azizov, R E; Makhlis, T A; Abbasov, V M; Varfolomeev, S D

    2005-01-01

    By using a bioluminescence ATP assay, we have determined the minimal concentrations of some biocorrosion inhibitors (Katon, Khazar, VFIKS-82, Nitro-1, Kaspii-2, and Kaspii-4) suppressing most common microbial corrosion agents: Desulfovibrio desulfuricans, Desulfovibrio vulgaris, Pseudomonas putida, Pseudomonas fluorescens, and Acidithiobacillus ferrooxidans. The cell titers determined by the bioluminescence method, including not only dividing cells but also their dormant living counterparts, are two- to sixfold greater than the values determined microbiologically. It is shown that the bioluminescence method can be applied to determination of cell titers in samples of oil-field waters in the presence of iron ions (up to 260 mM) and iron sulfide (to 186 mg/l) and in the absence or presence of biocidal corrosion inhibitors.

  3. Cancer and glioma : an integrated approach of gene therapy and bioluminescence imaging

    NARCIS (Netherlands)

    Bovenberg, Maria Sarah Sophie; Degeling, Marja Hannah

    2014-01-01

    Glioblastoma multiforme (GBM) is the most malignant variant of glioma. This tumor does not only display an extremely aggressive, invasive growth pattern, but is also very difficult to treat. With a two-year survival rate of 40% and a median survival of 12-18 months after treatment, prognosis is

  4. Integration

    DEFF Research Database (Denmark)

    Emerek, Ruth

    2004-01-01

    Bidraget diskuterer de forskellige intergrationsopfattelse i Danmark - og hvad der kan forstås ved vellykket integration......Bidraget diskuterer de forskellige intergrationsopfattelse i Danmark - og hvad der kan forstås ved vellykket integration...

  5. Hybrid Minimal Core Streptavidin-Obelin as a Versatile Reporter for Bioluminescence-based Bioassay.

    Science.gov (United States)

    Bashmakova, Eugenia E; Krasitskaya, Vasilisa V; Kudryavtsev, Alexander N; Grigorenko, Vitaly G; Frank, Ludmila A

    2017-03-01

    Ca 2+ -regulated photoprotein obelin was genetically fused with a minimum-sized core streptavidin. Hybrid protein (SAV-OL) was produced by bacterial expression and applied as a specific bioluminescent probe in diverse solid-phase assays. The obtained results clearly demonstrate specific activity of each domain indicating its proper folding with favorable space orientation. SAV-OL has been shown to be a much more sensitive label than the chemical conjugate of a full-length streptavidin with obelin. © 2016 The American Society of Photobiology.

  6. Effect of Iron Fe (II and Fe (III in a Binary System Evaluated Bioluminescent Method

    Directory of Open Access Journals (Sweden)

    Elena Sorokina

    2013-01-01

    Full Text Available The effect of iron ions Fe2+ and Fe3+ on the bioluminescent recombinant strain of Escherichia coli in a single-component and binary system. Found that for the bacteria E. coli Fe3+ ions are more toxic than Fe2+. Under the combined effect of iron toxicity increases, the percentage of luminescence quenching increases, but the value is much less than the sum of the indicator for the Fe2+ and Fe3+. The biological effect of insertion of iron is not proportional to their content in the mixture.

  7. Measurement of adenosine triphosphate (ATP) content in single red blood cells using the firefly bioluminescent reaction

    Energy Technology Data Exchange (ETDEWEB)

    Kostuk, R.K.; Muhs, A.G.; Kirkpatrick, F.H.; Gabel, C.W.

    1977-01-01

    A unique optical instrument is described which uses the firefly bioluminscent reaction to measure adenosine triphosphate (ATP) levels in single red blood cells. The method allows chemical content level to be associated with individual cell features. The optical instrument consists of a phase contrast microscope to view cells, a pulsed argon-ion laser to rupture the cell membrane, and a photon counting system to measure the bioluminescent yield. The technique has been calibrated against a standard ATP measurement using bulk analysis methods. The ATP loss mechanism for blood cells in a controlled depletion experiment was also investigated.

  8. Novel peptide chemistry in terrestrial animals: natural luciferin analogues from the bioluminescent earthworm Fridericia heliota.

    Science.gov (United States)

    Dubinnyi, Maxim A; Tsarkova, Aleksandra S; Petushkov, Valentin N; Kaskova, Zinaida M; Rodionova, Natalja S; Kovalchuk, Sergey I; Ziganshin, Rustam H; Baranov, Mikhail S; Mineev, Konstantin S; Yampolsky, Ilia V

    2015-03-02

    We report isolation and structure elucidation of AsLn5, AsLn7, AsLn11 and AsLn12: novel luciferin analogs from the bioluminescent earthworm Fridericia heliota. They were found to be highly unusual modified peptides, comprising either of the two tyrosine-derived chromophores, CompX or CompY and a set of amino acids, including threonine, gamma-aminobutyric acid, homoarginine, and unsymmetrical N,N-dimethylarginine. These natural compounds represent a unique peptide chemistry found in terrestrial animals and rise novel questions concerning their biosynthetic origin. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Evaluation of a bacterial bioluminescence bioassay as a predictive surrogate for mysid chronic estimator tests with produced water

    Energy Technology Data Exchange (ETDEWEB)

    Korenaga, G.L.; Stine, E.R.; Henry, L.R. [and others

    1995-12-01

    Toxicity limits are appearing more frequently in permits for produced water discharges. A bioluminescent bacteria bioassay has been proposed as a screening tool to predict toxicity in higher organisms, which are more expensive and require longer testing times. Before such a surrogate screen can be used, a correlation must be demonstrated between toxicity in the surrogate and the species of interest. This paper describes tests comparing produced water toxicity in the bioluminescent bacteria test and in the myoid shrimp chronic estimator test, which is frequently required in Gulf of Mexico discharge Under these test conditions, the bacteria test was not adequately predictive of produced water chronic toxicity to the myoid shrimp.

  10. GMO detection using a bioluminescent real time reporter (BART of loop mediated isothermal amplification (LAMP suitable for field use

    Directory of Open Access Journals (Sweden)

    Kiddle Guy

    2012-04-01

    Full Text Available Abstract Background There is an increasing need for quantitative technologies suitable for molecular detection in a variety of settings for applications including food traceability and monitoring of genetically modified (GM crops and their products through the food processing chain. Conventional molecular diagnostics utilising real-time polymerase chain reaction (RT-PCR and fluorescence-based determination of amplification require temperature cycling and relatively complex optics. In contrast, isothermal amplification coupled to a bioluminescent output produced in real-time (BART occurs at a constant temperature and only requires a simple light detection and integration device. Results Loop mediated isothermal amplification (LAMP shows robustness to sample-derived inhibitors. Here we show the applicability of coupled LAMP and BART reactions (LAMP-BART for determination of genetically modified (GM maize target DNA at low levels of contamination (0.1-5.0% GM using certified reference material, and compare this to RT-PCR. Results show that conventional DNA extraction methods developed for PCR may not be optimal for LAMP-BART quantification. Additionally, we demonstrate that LAMP is more tolerant to plant sample-derived inhibitors, and show this can be exploited to develop rapid extraction techniques suitable for simple field-based qualitative tests for GM status determination. We also assess the effect of total DNA assay load on LAMP-BART quantitation. Conclusions LAMP-BART is an effective and sensitive technique for GM detection with significant potential for quantification even at low levels of contamination and in samples derived from crops such as maize with a large genome size. The resilience of LAMP-BART to acidic polysaccharides makes it well suited to rapid sample preparation techniques and hence to both high throughput laboratory settings and to portable GM detection applications. The impact of the plant sample matrix and genome loading

  11. Rapid susceptibility testing of Mycobacterium tuberculosis by bioluminescence assay of mycobacterial ATP

    International Nuclear Information System (INIS)

    Nilsson, L.E.; Hoffner, S.E.; Ansehn, S.

    1988-01-01

    Mycobacterial growth was monitored by bioluminescence assay of mycobacterial ATP. Cultures of Mycobacterium tuberculosis H37Rv and of 25 clinical isolates of the same species were exposed to serial dilutions of ethambutol, isoniazid, rifampin, and streptomycin. A suppression of ATP, indicating growth inhibition, occurred for susceptible but not resistant strains within 5 to 7 days of incubation. Breakpoint concentrations between susceptibility and resistance were determined by comparing these results with those obtained by reference techniques. Full agreement was found in 99% of the assays with the resistance ratio method on Lowenstein-Jensen medium, and 98% of the assays were in full agreement with the radiometric system (BACTEC). A main advantage of the bioluminescence method is its rapidity, with results available as fast as with the radiometric system but at a lower cost and without the need for radioactive culture medium. The method provides kinetic data concerning drug effects within available in vivo drug concentrations and has great potential for both rapid routine susceptibility testing and research applications in studies of drug effects on mycobacteria

  12. Fluorescence and Bioluminescence Imaging of Angiogenesis in Flk1-Nano-lantern Transgenic Mice.

    Science.gov (United States)

    Matsushita, Jun; Inagaki, Shigenori; Nishie, Tomomi; Sakasai, Tomoki; Tanaka, Junko; Watanabe, Chisato; Mizutani, Ken-Ichi; Miwa, Yoshihiro; Matsumoto, Ken; Takara, Kazuhiro; Naito, Hisamichi; Kidoya, Hiroyasu; Takakura, Nobuyuki; Nagai, Takeharu; Takahashi, Satoru; Ema, Masatsugu

    2017-04-20

    Angiogenesis is important for normal development as well as for tumour growth. However, the molecular and cellular mechanisms underlying angiogenesis are not fully understood, partly because of the lack of a good animal model for imaging. Here, we report the generation of a novel transgenic (Tg) mouse that expresses a bioluminescent reporter protein, Nano-lantern, under the control of Fetal liver kinase 1 (Flk1). Flk1-Nano-lantern BAC Tg mice recapitulated endogenous Flk1 expression in endothelial cells and lymphatic endothelial cells during development and tumour growth. Importantly, bioluminescence imaging of endothelial cells from the aortic rings of Flk1-Nano-lantern BAC Tg mice enabled us to observe endothelial sprouting for 18 hr without any detectable phototoxicity. Furthermore, Flk1-Nano-lantern BAC Tg mice achieved time-lapse luminescence imaging of tumour angiogenesis in freely moving mice with implanted tumours. Thus, this transgenic mouse line contributes a unique model to study angiogenesis within both physiological and pathological contexts.

  13. Bioluminescence resonance energy transfer (BRET) imaging in plant seedlings and mammalian cells.

    Science.gov (United States)

    Xie, Qiguang; Soutto, Mohammed; Xu, Xiaodong; Zhang, Yunfei; Johnson, Carl Hirschie

    2011-01-01

    Bioluminescence resonance energy transfer (BRET) has become a widely used technique to monitor protein-protein interactions. It involves resonance energy transfer between a bioluminescent donor and a fluorescent acceptor. Because the donor emits photons intrinsically, fluorescence excitation is unnecessary. Therefore, BRET avoids some of the problems inherent in fluorescence resonance energy transfer (FRET) approaches, such as photobleaching, autofluorescence, and undesirable stimulation of photobiological processes. In the past, BRET signals have generally been too dim to be effectively imaged. Newly available cameras that are more sensitive coupled to image splitter now enable BRET imaging in plant and mammalian cells and tissues. In addition, new substrates and enhanced luciferases enable brighter signals that allow even subcellular BRET imaging. Here, we report methods for BRET imaging of (1) localization of COP1 dimerization in plant cells and tissues and (2) subcellular distributions of interactions of the CCAAT/Enhancer Binding Protein α (C/EBPα) in single mammalian cells. We also discuss methods for the correction of BRET images for tissues that absorb light of different spectra. This progress should catalyze further applications of BRET for imaging and high-throughput assays.

  14. Detection of Antibodies in Blood Plasma Using Bioluminescent Sensor Proteins and a Smartphone.

    Science.gov (United States)

    Arts, Remco; den Hartog, Ilona; Zijlema, Stefan E; Thijssen, Vito; van der Beelen, Stan H E; Merkx, Maarten

    2016-04-19

    Antibody detection is of fundamental importance in many diagnostic and bioanalytical assays, yet current detection techniques tend to be laborious and/or expensive. We present a new sensor platform (LUMABS) based on bioluminescence resonance energy transfer (BRET) that allows detection of antibodies directly in solution using a smartphone as the sole piece of equipment. LUMABS are single-protein sensors that consist of the blue-light emitting luciferase NanoLuc connected via a semiflexible linker to the green fluorescent acceptor protein mNeonGreen, which are kept close together using helper domains. Binding of an antibody to epitope sequences flanking the linker disrupts the interaction between the helper domains, resulting in a large decrease in BRET efficiency. The resulting change in color of the emitted light from green-blue to blue can be detected directly in blood plasma, even at picomolar concentrations of antibody. Moreover, the modular architecture of LUMABS allows changing of target specificity by simple exchange of epitope sequences, as demonstrated here for antibodies against HIV1-p17, hemagglutinin (HA), and dengue virus type I. The combination of sensitive ratiometric bioluminescent detection and the intrinsic modularity of the LUMABS design provides an attractive generic platform for point-of-care antibody detection that avoids the complex liquid handling steps associated with conventional immunoassays.

  15. Toxicological study of pesticides in air and precipitations of Paris by means of a bioluminescence method.

    Science.gov (United States)

    Trajkovska, S; Mbaye, M; Gaye Seye, M D; Aaron, J J; Chevreuil, M; Blanchoud, H

    2009-06-01

    A detailed toxicological study on several pesticides, including chlorothalonil, cyprodynil, dichlobénil, pendimethaline, trifluraline, and alpha-endosulfan, present at trace levels in air and total atmospheric precipitations of Paris is presented. The pesticides contained in the atmospheric samples, collected during sampling campaigns in February-March 2007, are identified and quantified by a high-performance liquid chromatographic (HPLC)-UV detection method. The toxicity measurements are performed by means of the Microtox bioluminescence method, based on the evaluation of the bioluminescence inhibition of the Vibrio fischeri marine bacteria at two exposure times to the pesticide solutions. The specific toxicity, corresponding to the particular toxicity of the compound under study and represented by the EC(50) parameter, is determined for these pesticides. Also, the global toxicity, which is the toxicity of all micro-pollutants present in the sample under study, is estimated for the extracts of air and atmospheric precipitation (rainwater) samples. The specific toxicities strongly vary with the nature of the pesticide, the EC(50) parameter values being comprised between 0.17 and 0.83 mg/mL and 0.15 and 0.66 mg/mL, respectively, for exposure times of 5 and 15 min. The importance of the atmospheric samples' global toxicity and the respective contribution of the toxic potency of the various pesticides contained in these samples are discussed.

  16. Association of bioluminescent bacteria from blue swimmer crab Portunus pelagicus (Linneaus, 1758

    Directory of Open Access Journals (Sweden)

    Chinnavenkataraman Govindasamy

    2012-10-01

    Full Text Available Objective: To screen the bioluminescent bacteria from Portunus pelagicus (P. pelagicus at Thondi coast, Palk Strait, Bay of Bengal, India. Methods: Physico-chemical parameter including atmospheric and surfacewater temperature, pH, salinity and dissolved oxygen were analyzed. The population of bioluminescent bacterium was screened in ambient water and blue swimmer crab of P. pelagicus (muscle, gill, hemolymph, shell and colony forming unit (CFU was calculated. Result: Atmospheric and surface water temperatures varied from 26.1 and 27.3 °C to 33.4 and 32.6 °C, respectively; salinity varied from 28.4% to 34.3%, pH varied from 7.6 to 8.6, and dissolved oxygen varied from 4.8 to 6.9 O2 ml/l. In addition, the maximum CFU value was identified (12.63 x104 CFU/ml during postmonsoon season and the minimum level (1.09 x104 CFU/ml identified during summer season. Further, based on the phenotypic characterizations the isolated strain were identified as Vibrio harveyi (V. harveyi. Conclusions: It is concluded from that the incidence of V. harveyi infections was frequently identified with edible crab of P. pelagicus, throughout the study periods in different seasons.

  17. Bayesian sparse-based reconstruction in bioluminescence tomography improves localization accuracy and reduces computational time.

    Science.gov (United States)

    Feng, Jinchao; Jia, Kebin; Li, Zhe; Pogue, Brian W; Yang, Mingjie; Wang, Yaqi

    2017-11-09

    Bioluminescence tomography (BLT) provides fundamental insight into biological processes in vivo. To fully realize its potential, it is important to develop image reconstruction algorithms that accurately visualize and quantify the bioluminescence signals taking advantage of limited boundary measurements. In this study, a new 2-step reconstruction method for BLT is developed by taking advantage of the sparse a priori information of the light emission using multispectral measurements. The first step infers a wavelength-dependent prior by using all multi-wavelength measurements. The second step reconstructs the source distribution based on this developed prior. Simulation, phantom and in vivo results were performed to assess and compare the accuracy and the computational efficiency of this algorithm with conventional sparsity-promoting BLT reconstruction algorithms, and results indicate that the position errors are reduced from a few millimeters down to submillimeter, and reconstruction time is reduced by 3 orders of magnitude in most cases, to just under a few seconds. The recovery of single objects and multiple (2 and 3) small objects is simulated, and the recovery of images of a mouse phantom and an experimental animal with an existing luminescent source in the abdomen is demonstrated. Matlab code is available at https://github.com/jinchaofeng/code/tree/master. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Monitoring and quantitative assessment of tumor burden using in vivo bioluminescence imaging

    Science.gov (United States)

    Chen, Chia-Chi; Hwang, Jeng-Jong; Ting, Gann; Tseng, Yun-Long; Wang, Shyh-Jen; Whang-Peng, Jaqueline

    2007-02-01

    In vivo bioluminescence imaging (BLI) is a sensitive imaging modality that is rapid and accessible, and may comprise an ideal tool for evaluating tumor growth. In this study, the kinetic of tumor growth has been assessed in C26 colon carcinoma bearing BALB/c mouse model. The ability of BLI to noninvasively quantitate the growth of subcutaneous tumors transplanted with C26 cells genetically engineered to stably express firefly luciferase and herpes simplex virus type-1 thymidine kinase (C26/ tk-luc). A good correlation ( R2=0.998) of photon emission to the cell number was found in vitro. Tumor burden and tumor volume were monitored in vivo over time by quantitation of photon emission using Xenogen IVIS 50 and standard external caliper measurement, respectively. At various time intervals, tumor-bearing mice were imaged to determine the correlation of in vivo BLI to tumor volume. However, a correlation of BLI to tumor volume was observed when tumor volume was smaller than 1000 mm 3 ( R2=0.907). γ Scintigraphy combined with [ 131I]FIAU was another imaging modality used for verifying the previous results. In conclusion, this study showed that bioluminescence imaging is a powerful and quantitative tool for the direct assay to monitor tumor growth in vivo. The dual reporter genes transfected tumor-bearing animal model can be applied in the evaluation of the efficacy of new developed anti-cancer drugs.

  19. A miniaturized device for bioluminescence analysis of caspase-3/7 5 activity in a single apoptotic cell

    Czech Academy of Sciences Publication Activity Database

    Adamová, Eva; Lišková, Marcela; Matalová, E.; Klepárník, Karel

    Roč. 406 , č. 22 (2014), s. 5389-5394 ISSN 1618-2642 R&D Projects: GA ČR(CZ) GA14-28254S Institutional support: RVO:68081715 Keywords : apoptosis * bioluminescence * single-cell analysis Subject RIV: CB - Analytical Chemistry, Separation Impact factor: 3.436, year: 2014

  20. Fast monitoring of indoor bioaerosol concentrations with ATP bioluminescence assay using an electrostatic rod-type sampler.

    Directory of Open Access Journals (Sweden)

    Ji-Woon Park

    Full Text Available A culture-based colony counting method is the most widely used analytical technique for monitoring bioaerosols in both indoor and outdoor environments. However, this method requires several days for colony formation. In this study, our goal was fast monitoring (Sampling: 3 min, Detection: < 1 min of indoor bioaerosol concentrations with ATP bioluminescence assay using a bioaerosol sampler. For this purpose, a novel hand-held electrostatic rod-type sampler (110 mm wide, 115 mm long, and 200 mm tall was developed and used with a commercial luminometer, which employs the Adenosine triphosphate (ATP bioluminescence method. The sampler consisted of a wire-rod type charger and a cylindrical collector, and was operated with an applied voltage of 4.5 kV and a sampling flow rate of 150.7 lpm. Its performance was tested using Staphylococcus epidermidis which was aerosolized with an atomizer. Bioaerosol concentrations were measured using ATP bioluminescence method with our sampler and compared with the culture-based method using Andersen cascade impactor under controlled laboratory conditions. Indoor bioaerosol concentrations were also measured using both methods in various indoor environments. A linear correlation was obtained between both methods in lab-tests and field-tests. Our proposed sampler with ATP bioluminescence method may be effective for fast monitoring of indoor bioaerosol concentrations.

  1. Bioluminescence : the potential of a non-invasive bio-optical imaging technique and improvement of animal research

    NARCIS (Netherlands)

    Hesselink, J. W.; van Dam, G. M.

    2007-01-01

    Bioluminescence is an optical imaging technique that exploits the emission of photons at specific wavelengths based on energy-dependent reactions catalysed by luciferases. The technique makes it possible to monitor measure, and track biological processes in living animals. A short review is

  2. Probing intermolecular protein-protein interactions in the calcium-sensing receptor homodimer using bioluminescence resonance energy transfer (BRET)

    DEFF Research Database (Denmark)

    Jensen, Anders A.; Hansen, Jakob L; Sheikh, Søren P

    2002-01-01

    -induced intermolecular movements in the CaR homodimer using the new bioluminescence resonance energy transfer technique, BRET2, which is based on the transference of energy from Renilla luciferase (Rluc) to the green fluorescent protein mutant GFP2. We tagged CaR with Rluc and GFP2 at different intracellular locations...

  3. Bioluminescent bacteria have potential as a marker of drowning in seawater: two immersed cadavers retrieved near estuaries.

    Science.gov (United States)

    Kakizaki, Eiji; Kozawa, Shuji; Sakai, Masahiro; Yukawa, Nobuhiro

    2009-03-01

    We detected numerous bioluminescent bacteria in blood samples from two cadavers that had been immersed in estuarine environments. Autopsy, diatomaceous and toxicological findings indicated death by drowning, which agreed with environmental aspects and the findings of police investigations. Bioluminescent bacteria appeared in blood samples cultured on selective agar containing 2%, 3% and 4% NaCl after about 18h. Blood from the left side of the heart, the right side of the heart and the femoral vein generated 7.0 x 10(2), 2.0 x 10(4) and 8.0 x 10(2) cfu/ml of blood (case 1), and 1.8 x 10(4), 1.1 x 10(3) and 2.5 x 10(1) cfu/ml (case 2) of bioluminescent colonies, respectively, in agar containing 4% NaCl. Homologous analysis based on the 16S rRNA gene also identified the bioluminescent colonies as Vibrio fischeri and V. harveyi, which normally inhabit seawater. This simple assay might serve as an additional indicator to support a conclusion of death by drowning together with the diatom test.

  4. Bioluminescence-Based Method for Measuring Assimilable Organic Carbon in Pretreatment Water for Reverse Osmosis Membrane Desalination ▿

    Science.gov (United States)

    Weinrich, Lauren A.; Schneider, Orren D.; LeChevallier, Mark W.

    2011-01-01

    A bioluminescence-based assimilable organic carbon (AOC) test was developed for determining the biological growth potential of seawater within the reverse osmosis desalination pretreatment process. The test uses Vibrio harveyi, a marine organism that exhibits constitutive luminescence and is nutritionally robust. AOC was measured in both a pilot plant and a full-scale desalination plant pretreatment. PMID:21148685

  5. The in vitro and in vivo effects of constitutive light expression on a bioluminescent strain of the mouse enteropathogen Citrobacter rodentium

    Directory of Open Access Journals (Sweden)

    Hannah M. Read

    2016-06-01

    Full Text Available Bioluminescent reporter genes, such as those from fireflies and bacteria, let researchers use light production as a non-invasive and non-destructive surrogate measure of microbial numbers in a wide variety of environments. As bioluminescence needs microbial metabolites, tagging microorganisms with luciferases means only live metabolically active cells are detected. Despite the wide use of bioluminescent reporter genes, very little is known about the impact of continuous (also called constitutive light expression on tagged bacteria. We have previously made a bioluminescent strain of Citrobacter rodentium, a bacterium which infects laboratory mice in a similar way to how enteropathogenic Escherichia coli (EPEC and enterohaemorrhagic E. coli (EHEC infect humans. In this study, we compared the growth of the bioluminescent C. rodentium strain ICC180 with its non-bioluminescent parent (strain ICC169 in a wide variety of environments. To understand more about the metabolic burden of expressing light, we also compared the growth profiles of the two strains under approximately 2,000 different conditions. We found that constitutive light expression in ICC180 was near-neutral in almost every non-toxic environment tested. However, we also found that the non-bioluminescent parent strain has a competitive advantage over ICC180 during infection of adult mice, although this was not enough for ICC180 to be completely outcompeted. In conclusion, our data suggest that constitutive light expression is not metabolically costly to C. rodentium and supports the view that bioluminescent versions of microbes can be used as a substitute for their non-bioluminescent parents to study bacterial behaviour in a wide variety of environments.

  6. Bioluminescence-based visualization of CD4 T cell dynamics using a T lineage-specific luciferase transgenic model1

    Directory of Open Access Journals (Sweden)

    Zinn Kurt R

    2009-08-01

    Full Text Available Abstract Background Rapid clonal expansion of T cells occurs in response to antigenic challenges. The kinetics of the T cell response has previously been described using tissue-based studies performed at defined time points. Luciferase bioluminescence has recently been utilized for non-invasive analysis of in vivo biologic processes in real-time. Results We have created a novel transgenic mouse model (T-Lux using a human CD2 mini-gene to direct luciferase expression specifically to the T cell compartment. T-Lux T cells demonstrated normal homing patterns within the intact mouse and following adoptive transfer. Bioluminescent signal correlated with T cell numbers in the whole body images as well as within specific organ regions of interest. Following transfer into lymphopenic (RAG2-/- recipients, homeostatic proliferation of T-Lux T cells was visualized using bioluminescent imaging. Real-time bioluminescent analysis of CD4+ T cell antigen-specific responses enabled real-time comparison of the kinetics and magnitude of clonal expansion and contraction in the inductive lymph node and tissue site of antigen injection. T cell expansion was dose-dependent despite the presence of supraphysiologic numbers of OVA-specific OT-II transgenic TCR T-Lux T cells. CD4+ T cells subsequently underwent a rapid (3–4 day contraction phase in the draining lymph node, with a delayed contraction in the antigen delivery site, with bioluminescent signal diminished below initial levels, representing TCR clonal frequency control. Conclusion The T-Lux mouse provides a novel, efficient model for tracking in vivo aspects of the CD4+ T cell response to antigen, providing an attractive approach for studies directed at immunotherapy or vaccine design.

  7. Revealing the ability of a novel polysaccharide bioflocculant in bioremediation of heavy metals sensed in a Vibrio bioluminescence reporter assay.

    Science.gov (United States)

    Sajayan, Arya; Seghal Kiran, G; Priyadharshini, S; Poulose, Navya; Selvin, Joseph

    2017-09-01

    A bioflocculant-producing bacterial strain, designated MSI021, was isolated from the marine sponge Dendrilla nigra and demonstrated 94% flocculation activity in a kaolin clay suspension. MSI021 was identified as Bacillus cereus based on phylogenetic affiliation and biochemical characteristics. The purified extra-cellular bioflocculant was chemically elucidated as a polysaccharide molecule. The polysaccharide bioflocculant was stable under both acidic and alkaline conditions (pH 2.0-10.0) and temperatures up to 100 °C. The purified bioflocculant efficiently nucleated the formation of silver nanoparticles which showed broad spectrum antibacterial activity. The ability of the bioflocculant to remediate heavy metal toxicity was evaluated by measuring the inhibition of bioluminescence expression in Vibrio harveyi. Enrichment of heavy metals such as zinc, mercury and copper at concentrations of 1, 2 and 3 mM in culture media showed significant reduction of bioluminescence in Vibrio, whereas media enriched with heavy metals and bioflocculant showed dose dependent improvement in the expression of bioluminescence. The assay results demonstrated that the polysaccharide bioflocculant effectively mitigates heavy metal toxicity, thereby improving the expression of bioluminescence in Vibrio. This bioluminescence reporter assay can be developed into a high-throughput format to monitor and evaluate of heavy metal toxicity. The findings of this study revealed that a novel polysaccharide bioflocculant produced by a marine B. cereus demonstrated strong flocculating performance and was effective in nucleating the formation antibacterial silver nanoparticles and removing heavy metals. These results suggest that the MSI021 polysaccharide bioflocculant can be used to develop greener waste water treatment systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Comparison of bulb syringe and pulsed lavage irrigation with use of a bioluminescent musculoskeletal wound model.

    Science.gov (United States)

    Svoboda, Steven J; Bice, Terry G; Gooden, Heather A; Brooks, Daniel E; Thomas, Darryl B; Wenke, Joseph C

    2006-10-01

    Despite the fact that wound irrigation is a common surgical procedure, there are many variables, including delivery device, irrigant type, and fluid volume, that have yet to be optimized. The purpose of this study was to compare, with use of transgenic bioluminescent bacteria and standard quantitative microbiological methods, the efficacy of pulsed lavage and bulb syringe irrigation in reducing wound bacterial counts. A caprine model of a complex, contaminated musculoskeletal wound was developed with use of a bioluminescent strain of Pseudomonas aeruginosa that can be quantified. Luminescent activity was recorded as relative luminescent units with use of a photon-counting camera six hours after the wound was created and inoculated. Twelve goats were randomly assigned to either the pulsed lavage group or the bulb syringe irrigation group. Each wound was irrigated with normal saline solution in 3-L increments for a total of 9 L and was imaged after each 3-L increment. In addition, quantitative culture samples were obtained from different tissues within the wound before and after irrigation. Pulsed lavage decreased the amount of relative luminescent units by 52%, 64%, and 70% at 3, 6, and 9 L, respectively. The bulb syringe irrigation reduced the amount of relative luminescent units by 33%, 44%, and 51% at these same time-points. Significant differences in luminescence were noted between the two groups after both 6 and 9 L of irrigation (p irrigation and after irrigation were r = 0.96 and 0.83, respectively. Pulsed lavage was more effective than bulb syringe irrigation in reducing bacterial luminescence after both 6 and 9 L of irrigation. Both device and volume effects can be demonstrated with use of this model. Bioluminescent bacteria provide a method to visualize bacterial distribution and to quantify the bacteria in a wound. Pulsed lavage is a more effective and efficient method of irrigation to remove bacteria in a complex musculoskeletal wound. In the model we

  9. Second-Generation Triple Reporter for Bioluminescence, Micro–Positron Emission Tomography, and Fluorescence Imaging

    Directory of Open Access Journals (Sweden)

    Aparna H. Kesarwala

    2006-10-01

    Full Text Available Bioluminescence, positron emission tomography (PET, and fluorescence modalities are currently available for noninvasive imaging in vivo, each with its own merits. To exploit the combined strengths of each and facilitate multimodality imaging, we engineered a dual-reporter construct in which firefly luciferase (FLuc and a 12–amino acid nonstructural linker were fused in frame to the N-terminus of a mutant herpes simplex virus thymidine kinase (mNLS-SR39TK kinetically enhanced for positron emission tomography (PET. Furthermore, a triple-reporter construct was developed in which monster green fluorescent protein (MGFP, a recently available enhanced fluorescent protein, was introduced into the fusion vector downstream of an internal ribosome entry site (IRES to allow analysis by fluorescence microscopy or flow cytometry without compromising the specific activities of the upstream fusion components. FLuc bioluminescence was measured with a cooled charge-coupled device camera and mNLS-SR39TK activity by 9-[4-[18F]fluoro-3-(hydroxymethyl butyl guanine (18F-FHBG microPET or 3H-penciclovir net accumulation. Importantly, HeLa cells transiently transfected with the FLuc-mNLS-SR39TK-IRES-MGFP triple reporter retained the same specific activities of the FLuc-mNLS-SR39TK heteroenzyme and the individual unfused enzymes with no change in protein half-lives. The presence of the IRES-MGFP modestly decreased upstream heteroprotein expression. In living mice, somatic gene transfer of a ubiquitin promoter-driven FLuc-mNLS-SR39TK-IRES-MGFP plasmid showed a > 1,000-fold increase in liver photon flux and a > 2-fold increase in liver retention of 18F-FHBG by microPET compared with mice treated with control plasmid. Multifocal hepatocellular fluorescence was readily observed by standard confocal microscopy. This second-generation triple reporter incorporating enhanced components enables bioluminescence, PET, and fluorescence imaging of cells and living animals.

  10. Evaluation of biolistic gene transfer methods in vivo using non-invasive bioluminescent imaging techniques

    Directory of Open Access Journals (Sweden)

    Daniell Henry

    2011-06-01

    Full Text Available Abstract Background Gene therapy continues to hold great potential for treating many different types of disease and dysfunction. Safe and efficient techniques for gene transfer and expression in vivo are needed to enable gene therapeutic strategies to be effective in patients. Currently, the most commonly used methods employ replication-defective viral vectors for gene transfer, while physical gene transfer methods such as biolistic-mediated ("gene-gun" delivery to target tissues have not been as extensively explored. In the present study, we evaluated the efficacy of biolistic gene transfer techniques in vivo using non-invasive bioluminescent imaging (BLI methods. Results Plasmid DNA carrying the firefly luciferase (LUC reporter gene under the control of the human Cytomegalovirus (CMV promoter/enhancer was transfected into mouse skin and liver using biolistic methods. The plasmids were coupled to gold microspheres (1 μm diameter using different DNA Loading Ratios (DLRs, and "shot" into target tissues using a helium-driven gene gun. The optimal DLR was found to be in the range of 4-10. Bioluminescence was measured using an In Vivo Imaging System (IVIS-50 at various time-points following transfer. Biolistic gene transfer to mouse skin produced peak reporter gene expression one day after transfer. Expression remained detectable through four days, but declined to undetectable levels by six days following gene transfer. Maximum depth of tissue penetration following biolistic transfer to abdominal skin was 200-300 μm. Similarly, biolistic gene transfer to mouse liver in vivo also produced peak early expression followed by a decline over time. In contrast to skin, however, liver expression of the reporter gene was relatively stable 4-8 days post-biolistic gene transfer, and remained detectable for nearly two weeks. Conclusions The use of bioluminescence imaging techniques enabled efficient evaluation of reporter gene expression in vivo. Our results

  11. Hybrid Multilevel Sparse Reconstruction for a Whole Domain Bioluminescence Tomography Using Adaptive Finite Element

    Directory of Open Access Journals (Sweden)

    Jingjing Yu

    2013-01-01

    Full Text Available Quantitative reconstruction of bioluminescent sources from boundary measurements is a challenging ill-posed inverse problem owing to the high degree of absorption and scattering of light through tissue. We present a hybrid multilevel reconstruction scheme by combining the ability of sparse regularization with the advantage of adaptive finite element method. In view of the characteristics of different discretization levels, two different inversion algorithms are employed on the initial coarse mesh and the succeeding ones to strike a balance between stability and efficiency. Numerical experiment results with a digital mouse model demonstrate that the proposed scheme can accurately localize and quantify source distribution while maintaining reconstruction stability and computational economy. The effectiveness of this hybrid reconstruction scheme is further confirmed with in vivo experiments.

  12. Seasonal variation of deep-sea bioluminescence in the Ionian Sea

    Energy Technology Data Exchange (ETDEWEB)

    Craig, Jessica, E-mail: j.craig@abdn.ac.u [University of Aberdeen, Oceanlab, Main Street, Newburgh, Aberdeenshire, AB41 6AA (United Kingdom); Jamieson, Alan J.; Bagley, Philip M.; Priede, Imants G. [University of Aberdeen, Oceanlab, Main Street, Newburgh, Aberdeenshire, AB41 6AA (United Kingdom)

    2011-01-21

    The ICDeep (Image Intensified Charge Coupled Device for Deep sea research) profiler was used to measure the density of deep bioluminescent animals (BL) through the water column in the east, west and mid-Ionian Sea and in the Algerian Basin. A west to east decrease in BL density was found. Generalized additive modelling was used to investigate seasonal variation in the east and west Ionian Sea (NESTOR and NEMO neutrino telescope sites, respectively) from BL measurements in autumn 2008 and spring 2009. A significant seasonal effect was found in the west Ionian Sea (p<0.001), where a deep autumnal peak in BL density occurred between 500 and 2400 m. No significant seasonal variation in BL density was found in the east Ionian Sea (p=0.07). In both spring and autumn, significant differences in BL density were found through the water column between the east and west Ionian Sea (p<0.001).

  13. Analysis of genetically modified organisms by pyrosequencing on a portable photodiode-based bioluminescence sequencer.

    Science.gov (United States)

    Song, Qinxin; Wei, Guijiang; Zhou, Guohua

    2014-07-01

    A portable bioluminescence analyser for detecting the DNA sequence of genetically modified organisms (GMOs) was developed by using a photodiode (PD) array. Pyrosequencing on eight genes (zSSIIb, Bt11 and Bt176 gene of genetically modified maize; Lectin, 35S-CTP4, CP4EPSPS, CaMV35S promoter and NOS terminator of the genetically modified Roundup ready soya) was successfully detected with this instrument. The corresponding limit of detection (LOD) was 0.01% with 35 PCR cycles. The maize and soya available from three different provenances in China were detected. The results indicate that pyrosequencing using the small size of the detector is a simple, inexpensive, and reliable way in a farm/field test of GMO analysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  14. Application of Bioluminescence Imaging for In Vivo Monitoring of Fungal Infections

    Directory of Open Access Journals (Sweden)

    Matthias Brock

    2012-01-01

    Full Text Available Fungi can cause severe invasive infections especially in the immunocompromised host. Patient populations at risk are increasing due to ongoing developments in cancer treatment and transplantation medicine. Only limited diagnostic tools and few antifungals are available, rendering a significant number of invasive fungal infections life threatening. To reduce mortality rates, a better understanding of the infection processes is urgently required. Bioluminescence imaging (BLI is a powerful tool for such purposes, since it allows visualisation of temporal and spatial progression of infections in real time. BLI has been successfully used to monitor infections caused by various microorganisms, in particular bacteria. However, first studies have also been performed on the fungi Candida albicans and Aspergillus fumigatus. Although BLI was, in principle, suitable to study the infection process, some limitations remained. Here, different luciferase systems are introduced, and current approaches are summarised. Finally, suggestions for further improvements of BLI to monitor fungal infections are provided.

  15. Generation of a new bioluminescent model for visualisation of mammary tumour development in transgenic mice

    LENUS (Irish Health Repository)

    Zagozdzon, Agnieszka M

    2012-05-30

    AbstractBackgroundNumerous transgenic models have been generated to study breast cancer. However, despite many advantages, traditional transgenic models for breast cancer are also burdened with difficulties in early detection and longitudinal observation of transgene-induced tumours, which in most cases are randomly located and occur at various time points. Methods such as palpation followed by mechanical measurement of the tumours are of limited value in transgenic models. There is a crucial need for making these previously generated models suitable for modern methods of tumour visualisation and monitoring, e.g. by bioluminescence-based techniques. This approach was successfully used in the current study.ResultsA new mouse strain (MMTV-Luc2 mice) expressing Luc2 luciferase primarily in mammary tissue in females, with low-level background expression in internal organs, was generated and bred to homozygosity. After these mice were intercrossed with MMTV-PyVT mice, all double transgenic females developed mammary tumours by the age of 10 weeks, the localisation and progression of which could be effectively monitored using the luminescence-based in vivo imaging. Luminescence-based readout allowed for early visualisation of the locally overgrown mammary tissue and for longitudinal evaluation of local progression of the tumours. When sampled ex vivo at the age of 10 weeks, all tumours derived from MMTV-Luc2PyVT females displayed robust bioluminescent signal.ConclusionsWe have created a novel transgenic strain for visualisation and longitudinal monitoring of mammary tumour development in transgenic mice as an addition and\\/or a new and more advanced alternative to manual methods. Generation of this mouse strain is vital for making many of the existing mammary tumour transgenic models applicable for in vivo imaging techniques.

  16. Modular low-light microscope for imaging cellular bioluminescence and radioluminescence.

    Science.gov (United States)

    Kim, Tae Jin; Türkcan, Silvan; Pratx, Guillem

    2017-05-01

    Low-light microscopy methods are receiving increased attention as new applications have emerged. One such application is to allow longitudinal imaging of light-sensitive cells with no phototoxicity and no photobleaching of fluorescent biomarkers. Another application is for imaging signals that are inherently dim and undetectable using standard microscopy techniques, such as bioluminescence, chemiluminescence or radioluminescence. In this protocol, we provide instructions on how to build a modular low-light microscope (1-4 d) by coupling two microscope objective lenses, back to back from each other, using standard optomechanical components. We also provide directions on how to image dim signals such as those of radioluminescence (1-1.5 h), bioluminescence (∼30 min) and low-excitation fluorescence (∼15 min). In particular, radioluminescence microscopy is explained in detail, as it is a newly developed technique that enables the study of small-molecule transport (e.g., radiolabeled drugs, metabolic precursors and nuclear medicine contrast agents) by single cells without perturbing endogenous biochemical processes. In this imaging technique, a scintillator crystal (e.g., CdWO 4 ) is placed in close proximity to the radiolabeled cells, where it converts the radioactive decays into optical flashes detectable using a sensitive camera. Using the image reconstruction toolkit provided in this protocol, the flashes can be reconstructed to yield high-resolution images of the radiotracer distribution. With appropriate timing, the three aforementioned imaging modalities may be performed together on a population of live cells, allowing the user to perform parallel functional studies of cell heterogeneity at the single-cell level.

  17. Generation of a new bioluminescent model for visualisation of mammary tumour development in transgenic mice

    Directory of Open Access Journals (Sweden)

    Zagozdzon Agnieszka M

    2012-05-01

    Full Text Available Abstract Background Numerous transgenic models have been generated to study breast cancer. However, despite many advantages, traditional transgenic models for breast cancer are also burdened with difficulties in early detection and longitudinal observation of transgene-induced tumours, which in most cases are randomly located and occur at various time points. Methods such as palpation followed by mechanical measurement of the tumours are of limited value in transgenic models. There is a crucial need for making these previously generated models suitable for modern methods of tumour visualisation and monitoring, e.g. by bioluminescence-based techniques. This approach was successfully used in the current study. Results A new mouse strain (MMTV-Luc2 mice expressing Luc2 luciferase primarily in mammary tissue in females, with low-level background expression in internal organs, was generated and bred to homozygosity. After these mice were intercrossed with MMTV-PyVT mice, all double transgenic females developed mammary tumours by the age of 10 weeks, the localisation and progression of which could be effectively monitored using the luminescence-based in vivo imaging. Luminescence-based readout allowed for early visualisation of the locally overgrown mammary tissue and for longitudinal evaluation of local progression of the tumours. When sampled ex vivo at the age of 10 weeks, all tumours derived from MMTV-Luc2PyVT females displayed robust bioluminescent signal. Conclusions We have created a novel transgenic strain for visualisation and longitudinal monitoring of mammary tumour development in transgenic mice as an addition and/or a new and more advanced alternative to manual methods. Generation of this mouse strain is vital for making many of the existing mammary tumour transgenic models applicable for in vivo imaging techniques.

  18. Imaging of Bubonic Plague Dynamics by In Vivo Tracking of Bioluminescent Yersinia pestis

    Science.gov (United States)

    Nham, Toan; Filali, Sofia; Danne, Camille; Derbise, Anne; Carniel, Elisabeth

    2012-01-01

    Yersinia pestis dissemination in a host is usually studied by enumerating bacteria in the tissues of animals sacrificed at different times. This laborious methodology gives only snapshots of the infection, as the infectious process is not synchronized. In this work we used in vivo bioluminescence imaging (BLI) to follow Y. pestis dissemination during bubonic plague. We first demonstrated that Y. pestis CO92 transformed with pGEN-luxCDABE stably emitted bioluminescence in vitro and in vivo, while retaining full virulence. The light produced from live animals allowed to delineate the infected organs and correlated with bacterial loads, thus validating the BLI tool. We then showed that the first step of the infectious process is a bacterial multiplication at the injection site (linea alba), followed by a colonization of the draining inguinal lymph node(s), and subsequently of the ipsilateral axillary lymph node through a direct connection between the two nodes. A mild bacteremia and an effective filtering of the blood stream by the liver and spleen probably accounted for the early bacterial blood clearance and the simultaneous development of bacterial foci within these organs. The saturation of the filtering capacity of the spleen and liver subsequently led to terminal septicemia. Our results also indicate that secondary lymphoid tissues are the main targets of Y. pestis multiplication and that colonization of other organs occurs essentially at the terminal phase of the disease. Finally, our analysis reveals that the high variability in the kinetics of infection is attributable to the time the bacteria remain confined at the injection site. However, once Y. pestis has reached the draining lymph nodes, the disease progresses extremely rapidly, leading to the invasion of the entire body within two days and to death of the animals. This highlights the extraordinary capacity of Y. pestis to annihilate the host innate immune response. PMID:22496846

  19. Generation of a new bioluminescent model for visualisation of mammary tumour development in transgenic mice

    International Nuclear Information System (INIS)

    Zagozdzon, Agnieszka M; O’Leary, Patrick; Callanan, John J; Crown, John; Gallagher, William M; Zagozdzon, Radoslaw

    2012-01-01

    Numerous transgenic models have been generated to study breast cancer. However, despite many advantages, traditional transgenic models for breast cancer are also burdened with difficulties in early detection and longitudinal observation of transgene-induced tumours, which in most cases are randomly located and occur at various time points. Methods such as palpation followed by mechanical measurement of the tumours are of limited value in transgenic models. There is a crucial need for making these previously generated models suitable for modern methods of tumour visualisation and monitoring, e.g. by bioluminescence-based techniques. This approach was successfully used in the current study. A new mouse strain (MMTV-Luc2 mice) expressing Luc2 luciferase primarily in mammary tissue in females, with low-level background expression in internal organs, was generated and bred to homozygosity. After these mice were intercrossed with MMTV-PyVT mice, all double transgenic females developed mammary tumours by the age of 10 weeks, the localisation and progression of which could be effectively monitored using the luminescence-based in vivo imaging. Luminescence-based readout allowed for early visualisation of the locally overgrown mammary tissue and for longitudinal evaluation of local progression of the tumours. When sampled ex vivo at the age of 10 weeks, all tumours derived from MMTV-Luc2PyVT females displayed robust bioluminescent signal. We have created a novel transgenic strain for visualisation and longitudinal monitoring of mammary tumour development in transgenic mice as an addition and/or a new and more advanced alternative to manual methods. Generation of this mouse strain is vital for making many of the existing mammary tumour transgenic models applicable for in vivo imaging techniques

  20. Imaging of bubonic plague dynamics by in vivo tracking of bioluminescent Yersinia pestis.

    Directory of Open Access Journals (Sweden)

    Toan Nham

    Full Text Available Yersinia pestis dissemination in a host is usually studied by enumerating bacteria in the tissues of animals sacrificed at different times. This laborious methodology gives only snapshots of the infection, as the infectious process is not synchronized. In this work we used in vivo bioluminescence imaging (BLI to follow Y. pestis dissemination during bubonic plague. We first demonstrated that Y. pestis CO92 transformed with pGEN-luxCDABE stably emitted bioluminescence in vitro and in vivo, while retaining full virulence. The light produced from live animals allowed to delineate the infected organs and correlated with bacterial loads, thus validating the BLI tool. We then showed that the first step of the infectious process is a bacterial multiplication at the injection site (linea alba, followed by a colonization of the draining inguinal lymph node(s, and subsequently of the ipsilateral axillary lymph node through a direct connection between the two nodes. A mild bacteremia and an effective filtering of the blood stream by the liver and spleen probably accounted for the early bacterial blood clearance and the simultaneous development of bacterial foci within these organs. The saturation of the filtering capacity of the spleen and liver subsequently led to terminal septicemia. Our results also indicate that secondary lymphoid tissues are the main targets of Y. pestis multiplication and that colonization of other organs occurs essentially at the terminal phase of the disease. Finally, our analysis reveals that the high variability in the kinetics of infection is attributable to the time the bacteria remain confined at the injection site. However, once Y. pestis has reached the draining lymph nodes, the disease progresses extremely rapidly, leading to the invasion of the entire body within two days and to death of the animals. This highlights the extraordinary capacity of Y. pestis to annihilate the host innate immune response.

  1. Elongation of exogenous fatty acids by the bioluminescent bacterium Vibrio harveyi

    International Nuclear Information System (INIS)

    Byers, D.M.

    1989-01-01

    Bioluminescent bacteria require myristic acid (C14:0) to produce the myristaldehyde substrate of the light-emitting luciferase reaction. Since both endogenous and exogenous C14:0 can be used for this purpose, the metabolism of exogenous fatty acids by luminescent bacteria has been investigated. Both Vibrio harveyi and Vibrio fischeri incorporated label from [1-14C]myristic acid (C14:0) into phospholipid acyl chains as well as into CO2. In contrast, Photobacterium phosphoreum did not exhibit phospholipid acylation or beta-oxidation using exogenous fatty acids. Unlike Escherichia coli, the two Vibrio species can directly elongate fatty acids such as octanoic (C8:0), lauric (C12:0), and myristic acid, as demonstrated by radio-gas liquid chromatography. The induction of bioluminescence in late exponential growth had little effect on the ability of V. harveyi to elongate fatty acids, but it did increase the amount of C14:0 relative to C16:0 labeled from [14C]C8:0. This was not observed in a dark mutant of V. harveyi that is incapable of supplying endogenous C14:0 for luminescence. Cerulenin preferentially decreased the labeling of C16:0 and of unsaturated fatty acids from all 14C-labeled fatty acid precursors as well as from [14C]acetate, suggesting that common mechanisms may be involved in elongation of fatty acids from endogenous and exogenous sources. Fatty acylation of the luminescence-related synthetase and reductase enzymes responsible for aldehyde synthesis exhibited a chain-length preference for C14:0, which also was indicated by reverse-phase thin-layer chromatography of the acyl groups attached to these enzymes. The ability of V. harveyi to activate and elongate exogenous fatty acids may be related to an adaptive requirement to metabolize intracellular C14:0 generated by the luciferase reaction during luminescence development

  2. 'Integration'

    DEFF Research Database (Denmark)

    Olwig, Karen Fog

    2011-01-01

    , while the countries have adopted disparate policies and ideologies, differences in the actual treatment and attitudes towards immigrants and refugees in everyday life are less clear, due to parallel integration programmes based on strong similarities in the welfare systems and in cultural notions...

  3. Turn up the lights: Deep-sea in situ application of a high-speed, high-resolution sCMOS camera to observe marine bioluminescence

    Science.gov (United States)

    Phillips, B. T.; Gruber, D. F.; Sparks, J. S.; Vasan, G.; Roman, C.; Pieribone, V. A.

    2016-02-01

    Observing and measuring marine bioluminescence presents unique challenges in situ. Technology is the greatest limiting factor in this endeavor, with sensitivity, speed and resolution constraining the imaging tools available to researchers. State-of-the-art microscopy cameras offer to bridge this gap. An ultra-low-light, scientific complimentary-metal-oxide-semiconductor (sCMOS) camera was outfitted for in-situ imaging of marine bioluminescence. This system was deployed on multiple deep-sea platforms (manned submersible, remotely operated vehicle, and towed body) in three oceanic regions (Western Tropical Pacific, Eastern Equatorial Pacific, and Northwestern Atlantic) to depths up to 2500m. Using light stimulation, bioluminescent responses were recorded at high frame rates and in high resolution, offering unprecedented low-light imagery of deep-sea bioluminescence in situ. The kinematics and physiology of light production in several zooplankton groups is presented, and luminescent responses at different depths are quantified as intensity vs. time.

  4. The first report of luminescent liver tissue in fishes: evolution and structure of bioluminescent organs in the deep-sea naked barracudinas (Aulopiformes: Lestidiidae).

    Science.gov (United States)

    Ghedotti, Michael J; Barton, Ryan W; Simons, Andrew M; Davis, Matthew P

    2015-03-01

    Bioluminescent organs that provide ventral camouflage are common among fishes in the meso-bathypelagic zones of the deep sea. However, the anatomical structures that have been modified to produce light vary substantially among different groups of fishes. Although the anatomical structure and evolutionary derivation of some of these organs have been well studied, the light organs of the naked barracudinas have received little scientific attention. This study describes the anatomy and evolution of bioluminescent organs in the Lestidiidae (naked barracudinas) in the context of a new phylogeny of barracudinas and closely related alepisauroid fishes. Gross and histological examination of bioluminescent organs or homologous structures from preserved museum specimens indicate that the ventral light organ is derived from hepatopancreatic tissue and that the antorbital spot in Lestrolepis is, in fact, a second dermal light organ. In the context of the phylogeny generated from DNA-sequence data from eight gene fragments (7 nuclear and 1 mitochondrial), a complex liver with a narrow ventral strand running along the ventral midline evolves first in the Lestidiidae. The ventral hepatopancreatic tissue later evolves into a ventral bioluminescent organ in the ancestor of Lestidium and Lestrolepis with the lineage leading to the genus Lestrolepis evolving a dermal antorbital bioluminescent organ, likely for light-intensity matching. This is the first described hepatopancreatic bioluminescent organ in fishes. © 2014 Wiley Periodicals, Inc.

  5. Evaluation of a bioluminescence method, contact angle measurements and topography for testing the cleanability of plastic surfaces under laboratory conditions

    Science.gov (United States)

    Redsven, I.; Kymäläinen, H.-R.; Pesonen-Leinonen, E.; Kuisma, R.; Ojala-Paloposki, T.; Hautala, M.; Sjöberg, A.-M.

    2007-04-01

    Detection of adenosine triphosphate (ATP) by bioluminescence is used, for instance, in the food industry and in hospitals to assess the hygiene status of surfaces. The aim of this laboratory study was to investigate the feasibility of the ATP method for estimating the cleanability of resilient floor coverings from biological soil. The surfaces were worn using a Soiling and Wearing Drum Tester, and soiled and cleaned with an Erichsen Washability and Scrubbing Resistance Tester. In the laboratory test carried out with the bioluminescence method, most of the new and worn floor coverings that were biologically soiled were cleaned efficiently. According to this study, the semiquantitative ATP screening method can be used for hygiene monitoring of flooring materials. No correlation was found between cleanability and contact angles or surface topography measured using a profilometer. However, by revealing local irregularities and damage on surfaces, scanning electron micrographs appeared useful in explaining differences in cleanability.

  6. Description of the larva of Alampoides alychnus (Kirsch, 1873, the first known species with bioluminescent immatures in Euplinthini (Elateridae, Agrypninae

    Directory of Open Access Journals (Sweden)

    Simone Policena Rosa

    2013-01-01

    Full Text Available Mature larva of Alampoides alychnus (Kirsch is described and compared to known Pyrophorini immatures. Larvae were collected live in the soil of a region dominated by sugarcane plantation and gallery forest in Campo Novo dos Parecis, Mato Grosso, Brazil. They were maintained in laboratory and the pupal period lasted 14 days. This larva differs from other Pyrophorini larvae mainly by bioluminescent pattern: one pair of luminous spots on the mesonotum, and a longitudinal series of median spots on the metanotum and all abdominal segments. The morphology of larva and the bioluminescent pattern of larva and pupa are described for the first time to the genus and the tribe. The fact that adults show no trace of luminescence is emphasized.

  7. Dual monitoring using {sup 124}I-FIAU and bioluminescence for HSV1-tk suicide gene therapy

    Energy Technology Data Exchange (ETDEWEB)

    Lee, T. S.; Kim, J. H.; Kwon, H. C. [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)] (and others)

    2007-07-01

    Herpes simplex virus type I thymidine kinase (HSV-tk) is the most common reporter gene and is used in cancer gene therapy with a prodrug nucleoside analog, ganciclovir (GCV). The aim of this study is to evaluate therapeutic efficacy of suicide gene therapy with 2'-fluoro-2'-deoxy-1-D-arabinofuranosyl-5-[{sup 124}I] iodouracil ({sup 124}I - FIAU) and bioluminescence in retrovirally HSV -tk and firefly luciferase transduced hepatoma model. The HSV -tk and firefly luciferase (Luc) was retrovirally transduced and expressed in MCA rat Morris hepatoma cells. Nude mice with subcutaneous tumors, MCA and MCA-TK-Luc, were subjected to GCV treatment (50mg/Kg/d intraperitoneally) for 5 day. PET imaging and biodistribution with ({sup 124}I-FIAU) were performed at before and after initiation of therapy with GCV. Bioluminescent signal was also measured during GCV treatment. Before GCV treatment, no significant difference in tumor volume was found in tumors between MCA and MCA-TK-Luc. After GCV treatment, tumor volume of MCA-TK-Luc markedly reduced compared to that of MCA. In biodistribution study, {sup 124}I-FIAU uptake after GCV therapy significantly decreased compared with pretreatment levels (34.8 13.67 %ID/g vs 7.6 2.59 %ID/g) and bioluminescent signal was also significantly decreased compared with pretreatment levels. In small animal PET imaging, {sup 124}I-FIAU selectively localized in HSV -tk expressing tumor and the therapeutic efficacy of GCV treatment was evaluated by {sup 124}I-FIAU PET imaging. {sup 124}I-FIAU PET and bioluminescence imaging in HSV-tk suicide gene therapy were effective to evaluate the therapeutic response. {sup 124}I-FIAU may serve as an efficient and selective agent for monitoring of transduced HSV1-tk gene expression in vivo in clinical trials.

  8. Standard procedure for testing acute toxic effects on bioluminescent bacteria; Saggio di tossicita` acuta con batteri bioluminescenti

    Energy Technology Data Exchange (ETDEWEB)

    Guzzella, L. [CNR, Brugherio, Milan (Italy). Istituto di Ricerca Sulle Acque

    1996-06-01

    A standardized method for the determination of 15-30 min toxicity of `Vibrio fisheri` bioluminescent bacteria is evaluated. The proposed method can be applied for the analysis of liquid (superficial and drinking waters, eluates and wastes) and solid (sediments and muds) samples and permits the quantification of the EC50 and EC20 values and of the no-effective sample dilution. The results of a interlaboratory ring tests conducted with reference substances are illustrated.

  9. Distribution of bioluminescent fungi across old-growth and secondary tropical rain forest in Costa Rica

    Directory of Open Access Journals (Sweden)

    Carolina Seas-Carvajal

    2013-06-01

    Full Text Available Most research on bioluminescent fungi is concentrated on their taxonomic relationships, while the basics of their natural history and ecological relationships are poorly understood. In this study, we compared the distribution of bioluminescent fungi between old-growth and secondary forest as related to four different soil types at the tropical rainforest of La Selva Biological Station in Costa Rica. The study was conducted during the wet season of 2009. Bioluminescent fungi were sought following eight different transects distributed evenly in old-growth and secondary forests across four different soil types, covering an area of 9 420m². We found fungi in four different substrates: litter, fallen branches, dead trunks, and roots, for a total of 61 samples. Correspondence analysis showed that the occurrence of fungi and soil types were related (inertia=0.21, p=0.071. We found a significant relationship between the presence of fungi and the distribution of soil types (X²=18.89, df=9, p=0.026. We found only three samples with fruiting bodies, two of which had Mycena and the other had one fungus of the order Xylariales (possibly Hypoxylon sp., Kretzschmariella sp., Xylaria sp.. Future work will concentrate on exploring other aspects of their ecology, such as their dispersal and substrate preference. This information will facilitate field identification and will foster more research on the distribution, seasonality, reproductive phenology and ecological requirements of this group of Fungi.La mayoría de las investigaciones sobre los hongos bioluminiscentes se ha centrado en relaciones taxonómicas. Los aspectos básicos de la historia natural y relaciones ecológicas de este grupo son poco conocidos. En este estudio, comparamos la distribución de hongos bioluminiscentes entre el bosque primario y el secundario en la Estación Biológica La Selva, Costa Rica en relación con cuatro tipos de suelo. El estudio se realizó durante la estación lluviosa

  10. An advanced preclinical mouse model for acute myeloid leukemia using patients' cells of various genetic subgroups and in vivo bioluminescence imaging.

    Directory of Open Access Journals (Sweden)

    Binje Vick

    Full Text Available Acute myeloid leukemia (AML is a clinically and molecularly heterogeneous disease with poor outcome. Adequate model systems are required for preclinical studies to improve understanding of AML biology and to develop novel, rational treatment approaches. Xenografts in immunodeficient mice allow performing functional studies on patient-derived AML cells. We have established an improved model system that integrates serial retransplantation of patient-derived xenograft (PDX cells in mice, genetic manipulation by lentiviral transduction, and essential quality controls by immunophenotyping and targeted resequencing of driver genes. 17/29 samples showed primary engraftment, 10/17 samples could be retransplanted and some of them allowed virtually indefinite serial transplantation. 5/6 samples were successfully transduced using lentiviruses. Neither serial transplantation nor genetic engineering markedly altered sample characteristics analyzed. Transgene expression was stable in PDX AML cells. Example given, recombinant luciferase enabled bioluminescence in vivo imaging and highly sensitive and reliable disease monitoring; imaging visualized minimal disease at 1 PDX cell in 10000 mouse bone marrow cells and facilitated quantifying leukemia initiating cells. We conclude that serial expansion, genetic engineering and imaging represent valuable tools to improve the individualized xenograft mouse model of AML. Prospectively, these advancements enable repetitive, clinically relevant studies on AML biology and preclinical treatment trials on genetically defined and heterogeneous subgroups.

  11. Interaction of Protease-Activated Receptor 2 with G Proteins and Beta-Arrestin 1 Studied by Bioluminescence Resonance Energy Transfer

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    Mohammed Akli eAyoub

    2013-12-01

    Full Text Available G protein-coupled receptors (GPCRs are well recognized as being able to activate several signaling pathways through the activation of different G proteins as well as other signaling proteins such as beta-arrestins. Therefore, understanding how such multiple GPCR-mediated signaling can be integrated constitute an important aspect. Here, we applied bioluminescence resonance energy transfer (BRET to shed more light on the G protein coupling profile of trypsin receptor, or protease-activated receptor 2 (PAR2, and its interaction with beta-arrestin1. Using YFP and Rluc fusion constructs expressed in COS-7 cells, BRET data revealed a pre-assembly of PAR2 with both Galphai1 and Galphao and a rapid and transient activation of these G proteins upon receptor activation. In contrast, no preassembly of PAR2 with Galpha12 could be detected and their physical association can be measured with a very slow and sustained kinetics similar to that of beta-arrestin1 recruitment. These data demonstrate the coupling of PAR2 with Galphai1, Galphao and Galpha12 in COS-7 cells with differences in the kinetics of GPCR-G protein coupling, a parameter that very likely influences the cellular response. Moreover, this further illustrates that preassembly or agonist-induced G protein interaction depends on receptor-G protein pairs indicating another level of complexity and regulation of the signaling of GPCR-G protein complexes and its multiplicity.

  12. Assessment of chitosan-affected metabolic response by peroxisome proliferator-activated receptor bioluminescent imaging-guided transcriptomic analysis.

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    Chia-Hung Kao

    Full Text Available Chitosan has been widely used in food industry as a weight-loss aid and a cholesterol-lowering agent. Previous studies have shown that chitosan affects metabolic responses and contributes to anti-diabetic, hypocholesteremic, and blood glucose-lowering effects; however, the in vivo targeting sites and mechanisms of chitosan remain to be clarified. In this study, we constructed transgenic mice, which carried the luciferase genes driven by peroxisome proliferator-activated receptor (PPAR, a key regulator of fatty acid and glucose metabolism. Bioluminescent imaging of PPAR transgenic mice was applied to report the organs that chitosan acted on, and gene expression profiles of chitosan-targeted organs were further analyzed to elucidate the mechanisms of chitosan. Bioluminescent imaging showed that constitutive PPAR activities were detected in brain and gastrointestinal tract. Administration of chitosan significantly activated the PPAR activities in brain and stomach. Microarray analysis of brain and stomach showed that several pathways involved in lipid and glucose metabolism were regulated by chitosan. Moreover, the expression levels of metabolism-associated genes like apolipoprotein B (apoB and ghrelin genes were down-regulated by chitosan. In conclusion, these findings suggested the feasibility of PPAR bioluminescent imaging-guided transcriptomic analysis on the evaluation of chitosan-affected metabolic responses in vivo. Moreover, we newly identified that downregulated expression of apoB and ghrelin genes were novel mechanisms for chitosan-affected metabolic responses in vivo.

  13. Multimodal Imaging Reveals Improvement of Blood Supply to an Artificial Cell Transplant Site Induced by Bioluminescent Mesenchymal Stem Cells.

    Science.gov (United States)

    Gálisová, Andrea; Fábryová, Eva; Jirák, Daniel; Sticová, Eva; Lodererová, Alena; Herynek, Vít; Kříž, Jan; Hájek, Milan

    2017-02-01

    An artificial site for cell or pancreatic islet transplantation can be created using a polymeric scaffold, even though it suffers subcutaneously from improper vascularisation. A sufficient blood supply is crucial for graft survival and function and can be enhanced by transplantation of mesenchymal stem cells (MSCs). The purpose of this study was to assess the effect of syngeneic MSCs on neoangiogenesis and cell engraftment in an artificial site by multimodal imaging. MSCs expressing a gene for luciferase were injected into the artificial subcutaneous site 7 days after scaffold implantation. MRI experiments (anatomical and dynamic contrast-enhanced images) were performed on a 4.7-T scanner using gradient echo sequences. Bioluminescent images were acquired on an IVIS Lumina optical imager. Longitudinal examination was performed for 2 months, and one animal was monitored for 16 months. We confirmed the long-term presence (lasting more than 16 months) of viable donor cells inside the scaffolds using bioluminescence imaging with an optical signal peak appearing on day 3 after MSC implantation. When compared to controls, the tissue perfusion and vessel permeability in the scaffolds were significantly improved at the site with MSCs with a maximal peak on day 9 after MSC transplantation. Our data suggest that the maximal signal obtained by bioluminescence and magnetic resonance imaging from an artificially created site between 3 and 9 days after MSC transplantation can predict the optimal time range for subsequent cellular or tissue transplantation, including pancreatic islets.

  14. Detection of Actaea racemosa Adulteration by Thin-Layer Chromatography and Combined Thin-Layer Chromatography-Bioluminescence

    Science.gov (United States)

    Verbitski, Sheryl M.; Gourdin, Gerald T.; Ikenouye, Larissa M.; McChesney, James D.; Hildreth, Jana

    2014-01-01

    Actaea racemosa L. (black cohosh; syn. Cimicifuga racemosa L. Nutt.) is a native North American perennial whose root and rhizome preparations are commercially available as phytomedicines and dietary supplements, primarily for management of menopausal symptoms. Despite its wide use, methods that accurately identify processed A. racemosa are not well established; product adulteration remains a concern. Because of its similar appearance and growing locales, A. racemosa has been unintentionally mixed with other species of the genus, such as Actaea pachypoda Ell. (white cohosh) and more commonly Actaea podocarpa DC. (yellow cohosh). The genus Actaea also has 23 temperate species with numerous common names, which can also contribute to the misidentification of plant material. Consequently, a variety of Actaea spp. are common adulterants of commercially available black cohosh preparations. Thin-layer chromatography (TLC) and combined TLC-bioluminescence (Bioluminex™) are efficient, economical, and effective techniques which provide characteristic patterns and toxicity profiles for each plant species. These data indicate that common black cohosh adulterants, such as yellow cohosh, can be differentiated from black cohosh by TLC and TLC-bioluminescence. This study also showed that unknown contaminants that were not detected using standard A. racemosa identity techniques were readily detected by TLC and TLC-bioluminescence. PMID:18476337

  15. BarTeL, a Genetically Versatile, Bioluminescent and Granule Neuron Precursor-Targeted Mouse Model for Medulloblastoma.

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    Gregory M Shackleford

    Full Text Available Medulloblastomas are the most common malignant pediatric brain tumor and have been divided into four major molecular subgroups. Animal models that mimic the principal molecular aberrations of these subgroups will be important tools for preclinical studies and allow greater understanding of medulloblastoma biology. We report a new transgenic model of medulloblastoma that possesses a unique combination of desirable characteristics including, among others, the ability to incorporate multiple and variable genes of choice and to produce bioluminescent tumors from a limited number of somatic cells within a normal cellular environment. This model, termed BarTeL, utilizes a Barhl1 homeobox gene promoter to target expression of a bicistronic transgene encoding both the avian retroviral receptor TVA and an eGFP-Luciferase fusion protein to neonatal cerebellar granule neuron precursor (cGNP cells, which are cells of origin for the sonic hedgehog (SHH subgroup of human medulloblastomas. The Barhl1 promoter-driven transgene is expressed strongly in mammalian cGNPs and weakly or not at all in mature granule neurons. We efficiently induced bioluminescent medulloblastomas expressing eGFP-luciferase in BarTeL mice by infection of a limited number of somatic cGNPs with avian retroviral vectors encoding the active N-terminal fragment of SHH and a stabilized MYCN mutant. Detection and quantification of the increasing bioluminescence of growing tumors in young BarTeL mice was facilitated by the declining bioluminescence of their uninfected maturing cGNPs. Inclusion of eGFP in the transgene allowed enriched sorting of cGNPs from neonatal cerebella. Use of a single bicistronic avian vector simultaneously expressing both Shh and Mycn oncogenes increased the medulloblastoma incidence and aggressiveness compared to mixed virus infections. Bioluminescent tumors could also be produced by ex vivo transduction of neonatal BarTeL cerebellar cells by avian retroviruses and

  16. Dual-Color Bioluminescent Sensor Proteins for Therapeutic Drug Monitoring of Antitumor Antibodies.

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    van Rosmalen, Martijn; Ni, Yan; Vervoort, Daan F M; Arts, Remco; Ludwig, Susann K J; Merkx, Maarten

    2018-03-06

    Monitoring the levels of therapeutic antibodies in individual patients would allow patient-specific dose optimization, with the potential for major therapeutic and financial benefits. Our group recently developed a new platform of bioluminescent sensor proteins (LUMABS; LUMinescent AntiBody Sensor) that allow antibody detection directly in blood plasma. In this study, we targeted four clinically important therapeutic antibodies, the Her2-receptor targeting trastuzumab, the anti-CD20 antibodies rituximab and obinutuzumab, and the EGFR-blocking cetuximab. A strong correlation was found between the affinity of the antibody binding peptide and sensor performance. LUMABS sensors with physiologically relevant affinities and decent sensor responses were obtained for trastuzumab and cetuximab using mimotope and meditope peptides, respectively, with affinities in the 10 -7 M range. The lower affinity of the CD20-derived cyclic peptide employed in the anti-CD20 LUMABS sensor ( K d = 10 -5 M), translated in a LUMABS sensor with a strongly attenuated sensor response. The trastuzumab and cetuximab sensors were further characterized with respect to binding kinetics and their performance in undiluted blood plasma. For both antibodies, LUMABS-based detection directly in plasma compared well to the analytical performance of commercial ELISA kits. Besides identifying important design parameters for the development of new LUMABS sensors, this work demonstrates the potential of the LUMABS platform for point-of-care detection of therapeutic antibodies.

  17. Rapid and Quantitative Assessment of Cancer Treatment Response Using In Vivo Bioluminescence Imaging

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    Alnawaz Rehemtulla

    2000-01-01

    Full Text Available Current assessment of orthotopic tumor models in animals utilizes survival as the primary therapeutic end point. In vivo bioluminescence imaging (BLI is a sensitive imaging modality that is rapid and accessible, and may comprise an ideal tool for evaluating antineoplastic therapies [1 ]. Using human tumor cell lines constitutively expressing luciferase, the kinetics of tumor growth and response to therapy have been assessed in intraperitoneal [2], subcutaneous, and intravascular [3] cancer models. However, use of this approach for evaluating orthotopic tumor models has not been demonstrated. In this report, the ability of BLI to noninvasively quantitate the growth and therapeuticinduced cell kill of orthotopic rat brain tumors derived from 9L gliosarcoma cells genetically engineered to stably express firefly luciferase (9LLuc was investigated. Intracerebral tumor burden was monitored over time by quantitation of photon emission and tumor volume using a cryogenically cooled CCD camera and magnetic resonance imaging (MRI, respectively. There was excellent correlation (r=0.91 between detected photons and tumor volume. A quantitative comparison of tumor cell kill determined from serial MRI volume measurements and BLI photon counts following 1,3-bis(2-chloroethyl-1-nitrosourea (BCNU treatment revealed that both imaging modalities yielded statistically similar cell kill values (P=.951. These results provide direct validation of BLI imaging as a powerful and quantitative tool for the assessment of antineoplastic therapies in living animals.

  18. Combining intracellular and secreted bioluminescent reporter proteins for multicolor cell-based assays.

    Science.gov (United States)

    Michelini, Elisa; Cevenini, Luca; Mezzanotte, Laura; Ablamsky, Danielle; Southworth, Tara; Branchini, Bruce R; Roda, Aldo

    2008-02-01

    Bioluminescent (BL) proteins are a promising tool for diverse applications based on reporter gene technology thanks to their high sensitivity and range of linear response. Due to their widespread use in the environmental, medical and agro-food fields, there is a great need for new BL reporter proteins with improved characteristics to provide researches a wide range of suitable reporters. Few efforts have been made in this direction and further improvement of BL reporter features (e.g., thermostability, narrower emission bandwidth, emission at different wavelengths) tailored for specific applications would be a remarkable progress toward the development of ultrasensitive multiplexed assays either in vitro or in vivo. The suitability of using red- and green-emitting thermostable mutants of Photinus pyralis firefly luciferase and two click beetle luciferases in combination with a secreted luciferase from Gaussia princeps was evaluated to develop a triple-color mammalian assay. Two triple-reporter model mammalian systems were developed in a human hepatoblastoma cell line to monitor the transcriptional regulation of cholesterol 7-alpha hydroxylase (cyp7a1), the enzyme that catalyzes the first and rate-limiting step of the main pathway responsible for cholesterol degradation in humans. These model systems allowed us to evaluate the feasibility of using two intracellular BL reporters and a secreted one in the same cell-based assay. The selection of reporter proteins characterized by similar expression levels was identified as a critical point for the development of a multicolor assay.

  19. CD4+ T cell effects on CD8+ T cell location defined using bioluminescence.

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    Mitra Azadniv

    2011-01-01

    Full Text Available T lymphocytes of the CD8+ class are critical in delivering cytotoxic function and in controlling viral and intracellular infections. These cells are "helped" by T lymphocytes of the CD4+ class, which facilitate their activation, clonal expansion, full differentiation and the persistence of memory. In this study we investigated the impact of CD4+ T cells on the location of CD8+ T cells, using antibody-mediated CD4+ T cell depletion and imaging the antigen-driven redistribution of bioluminescent CD8+ T cells in living mice. We documented that CD4+ T cells influence the biodistribution of CD8+ T cells, favoring their localization to abdominal lymph nodes. Flow cytometric analysis revealed that this was associated with an increase in the expression of specific integrins. The presence of CD4+ T cells at the time of initial CD8+ T cell activation also influences their biodistribution in the memory phase. Based on these results, we propose the model that one of the functions of CD4+ T cell "help" is to program the homing potential of CD8+ T cells.

  20. Functional imaging of interleukin 1 beta expression in inflammatory process using bioluminescence imaging in transgenic mice

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    Liu Zhihui

    2008-08-01

    Full Text Available Abstract Background Interleukin 1 beta (IL-1β plays an important role in a number of chronic and acute inflammatory diseases. To understand the role of IL-1β in disease processes and develop an in vivo screening system for anti-inflammatory drugs, a transgenic mouse line was generated which incorporated the transgene firefly luciferase gene driven by a 4.5-kb fragment of the human IL-1β gene promoter. Luciferase gene expression was monitored in live mice under anesthesia using bioluminescence imaging in a number of inflammatory disease models. Results In a LPS-induced sepsis model, dramatic increase in luciferase activity was observed in the mice. This transgene induction was time dependent and correlated with an increase of endogenous IL-1β mRNA and pro-IL-1β protein levels in the mice. In a zymosan-induced arthritis model and an oxazolone-induced skin hypersensitivity reaction model, luciferase expression was locally induced in the zymosan injected knee joint and in the ear with oxazolone application, respectively. Dexamethasone suppressed the expression of luciferase gene both in the acute sepsis model and in the acute arthritis model. Conclusion Our data suggest that the transgenic mice model could be used to study transcriptional regulation of the IL-1β gene expression in the inflammatory process and evaluation the effect of anti-inflammatory drug in vivo.

  1. Light Emission Requires Exposure to the Atmosphere in Ex Vivo Bioluminescence Imaging

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    Yusuke Inoue

    2006-04-01

    Full Text Available The identification of organs bearing luciferase activity by in vivo bioluminescence imaging (BLI is often difficult, and ex vivo imaging of excised organs plays a complementary role. This study investigated the importance of exposure to the atmosphere in ex vivo BLI. Mice were inoculated with murine pro-B cell line Ba/F3 transduced with firefly luciferase and p190 BCR-ABL. They were killed following in vivo BLI, and whole-body imaging was done after death and then after intraperitoneal air injection. In addition, the right knee was exposed and imaged before and after the adjacent bones were cut. Extensive light signals were seen on in vivo imaging. The luminescence disappeared after the animal was killed, and air injection restored the light emission from the abdomen only, suggesting a critical role of atmospheric oxygen in luminescence after death. Although no substantial light signal at the right knee was seen before bone cutting, light emission was evident after cutting. In conclusion, in ex vivo BLI, light emission requires exposure to the atmosphere. Bone destruction is required to demonstrate luciferase activity in the bone marrow after death.

  2. L{sub 1/2} regularization based numerical method for effective reconstruction of bioluminescence tomography

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xueli, E-mail: xlchen@xidian.edu.cn, E-mail: jimleung@mail.xidian.edu.cn; Yang, Defu; Zhang, Qitan; Liang, Jimin, E-mail: xlchen@xidian.edu.cn, E-mail: jimleung@mail.xidian.edu.cn [School of Life Science and Technology, Xidian University, Xi' an 710071 (China); Engineering Research Center of Molecular and Neuro Imaging, Ministry of Education (China)

    2014-05-14

    Even though bioluminescence tomography (BLT) exhibits significant potential and wide applications in macroscopic imaging of small animals in vivo, the inverse reconstruction is still a tough problem that has plagued researchers in a related area. The ill-posedness of inverse reconstruction arises from insufficient measurements and modeling errors, so that the inverse reconstruction cannot be solved directly. In this study, an l{sub 1/2} regularization based numerical method was developed for effective reconstruction of BLT. In the method, the inverse reconstruction of BLT was constrained into an l{sub 1/2} regularization problem, and then the weighted interior-point algorithm (WIPA) was applied to solve the problem through transforming it into obtaining the solution of a series of l{sub 1} regularizers. The feasibility and effectiveness of the proposed method were demonstrated with numerical simulations on a digital mouse. Stability verification experiments further illustrated the robustness of the proposed method for different levels of Gaussian noise.

  3. Bioluminescent avian pathogenic Escherichia coli for monitoring colibacillosis in experimentally infected chickens.

    Science.gov (United States)

    Oosterik, Leon H; Tuntufye, Huruma N; Tsonos, Jessica; Luyten, Tom; Noppen, Sam; Liekens, Sandra; Lavigne, Rob; Butaye, Patrick; Goddeeris, Bruno M

    2016-10-01

    Avian pathogenic Escherichia coli (APEC) are responsible for significant economic losses in the poultry industry. In this study, a model for investigating the pathogenesis of APEC infections was established. APEC strain CH2 (O78) was marked with the luciferase operon (luxCDABE) using a Tn7 transposon and tissues of experimentally infected chickens were analysed for a correlation between the bioluminescent signal and the number of bacteria. Transposition of the lux operon into the chromosome of the APEC isolate did not affect sensitivity to lytic bacteriophages and there was no effect on virulence in an intratracheal infection model in 1-day-old chicks, although results with a subcutaneous infection model were inconclusive. A correlation between the number of bacteria and the luminescent signal was found in liquid medium, as well as in homogenised heart, liver, spleen and lung of 4-week-old experimentally infected chickens. This study showed that lux could be used for identification of the infecting strain after experimental infection with APEC in poultry. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. Surface-Tunable Bioluminescence Resonance Energy Transfer via Geometry-Controlled ZnO Nanorod Coordination.

    Science.gov (United States)

    Lim, Jun Hyung; Park, Geun Chul; Lee, Seung Muk; Lee, Jung Heon; Lim, Butaek; Hwang, Soo Min; Kim, Jung Ho; Park, Hansoo; Joo, Jinho; Kim, Young-Pil

    2015-07-01

    The use of ZnO nanorods (NRs) as an effective coordinator and biosensing platform to create bioluminescence resonance energy transfer (BRET) is reported. Herein, a hydrothermal approach is applied to obtain morphologically controlled ZnO NRs, which are directly bound to luciferase (Luc) and carboxy-modified quantum dot (QD) acting as a donor-acceptor pair for BRET. BRET efficiency varies significantly with the geometry of ZnO NRs, which modulates the coordination between hexahistidine-tagged Luc (Luc-His6 ) and QD, owing to the combined effect of the total surface area consisting of (001) and (100) planes and their surface polarities. Unlike typical QD-BRET reactions with metal ions (e.g., zinc ions), a geometry-controlled ZnO NR platform can facilitate the design of surface-initiated BRET sensors without being supplemented by copious metal ions: the geometry-controlled ZnO NR platform can therefore pave the way for nanostructure-based biosensors with enhanced analytical performance. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Comparative bioluminescence dynamics among multiple Armillaria gallica, A. mellea, and A. tabescens genets.

    Science.gov (United States)

    Mihail, Jeanne D

    2013-03-01

    Bioluminescence is well known among white-spored species of Basidiomycota including several species of the white-rot wood decay genus Armillaria. Previous work demonstrated consistent differences among A. gallica, A. mellea, and A. tabescens in luminescence magnitude and in luminescence expression relative to environmental stimuli. In the present studies, temporal fluctuations in mycelial luminescence were quantitatively characterized using genets matched for geographical location. All genets derived from rhizomorphs or basdiomata were constitutively luminescent while six of 13 genets originating from mycelial fans were inconsistently luminescent. Using time series of 1000 consecutive measurements over 800 ms intervals, fluctuation patterns had significantly quantifiable structure and were not simply 'white noise'. Fluctuation patterns were qualitatively similar with alternating periods of rapid fluctuation and relative stability, regardless of luminescence magnitude. Anomalous spikes or shifts in luminescence were recorded for several genets suggesting further work to identify the transient stimuli which elicited these altered luminescence patterns. Copyright © 2013 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  6. Quick, portable toxicity testing of marine or terrigenous fluids, sediments, or chemicals with bioluminescent organism

    Energy Technology Data Exchange (ETDEWEB)

    Sabate, R.W.; Stiffey, A.V.; Dewailly, E.L. [Lumitox Gulf L.C., New Orleans, LA (United States)

    1995-12-31

    A hand-held, battery-operated instrument, which measures bioluminescence inhibition of the microscopic marine dinoflagellate Pyrocystis lunula, is capable of field-testing substances for toxicity. The organism is sensitive to ppb of strong toxicants. It tolerates some solvents in concentrations necessary for testing lipophylic samples. A test consumes only micrograms of sample. This method requires no adjustments for salinity, pH, color, or turbidity. It has been used successfully to test oil-well drilling fluids, brines produced with oil, waters and sediments from streams and lakes and petroleum-plant effluents containing contaminants such as benzene. The test is non-specific; however, if the substance is known, the end-point effects a direct measurement of its concentration. One-hour toxicity screening tests in the field produce results comparable to the standard four-hour laboratory test. Keeping the sample in the dark during incubation and testing, together with shortness of the overall procedure, eliminates anomalies from light-sensitive substances. Day-to-day variation, as well as among test replicates, is less than 10%. This quick method yields results comparable with a quick test that uses Photobacterium phosphoria, and with 96-hour tests that use Mysidopsis bahia, Artemia salina, Gonyaulax polyedra, Pimephales promelas, Ceriodaphnia dubia, and Cyprinodon variegatus.

  7. Hunting in bioluminescent light: Vision in the nocturnal box jellyfish Copula sivickisi

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    Anders eGarm

    2016-03-01

    Full Text Available Cubomedusae all have a similar set of six eyes on each of their four rhopalia. Still, there is a great variation in activity patterns with some species being strictly day active while others are strictly night active. Here we have examined the visual ecology of the medusa of the night active Copula sivickisi from Okinawa using optics, morphology, electrophysiology, and behavioural experiments. We found the lenses of both the upper and the lower lens eyes to be image forming but under-focused, resulting in low spatial resolution in the order of 10 – 15 degrees. The photoreceptor physiology is similar in the two lens eyes and they have a single opsin peaking around 460 nm and low temporal resolution with a flicker fusion frequency (fff of 2.5 Hz indicating adaptions to vision in low light intensities. Further, the outer segments have fluid filled swellings, which may concentrate the light in the photoreceptor membrane by total internal reflections, and thus enhance the signal to noise ratio in the eyes. Finally our behavioural experiments confirmed that the animals use vision when hunting. When they are active at night they seek out high prey-concentration by visual attraction to areas with abundant bioluminescent flashes triggered by their prey.

  8. Evaluation of monkeypox virus infection of prairie dogs (Cynomys ludovicianus) using in vivo bioluminescent imaging

    Science.gov (United States)

    Falendysz, Elizabeth A.; Londoño-Navas, Angela M.; Meteyer, Carol U.; Pussini, Nicola; Lopera, Juan G.; Osorio, Jorge E.; Rocke, Tonie E.

    2014-01-01

    Monkeypox (MPX) is a re-emerging zoonotic disease that is endemic in Central and West Africa, where it can cause a smallpox-like disease in humans. Despite many epidemiologic and field investigations of MPX, no definitive reservoir species has been identified. Using recombinant viruses expressing the firefly luciferase (luc) gene, we previously demonstrated the suitability of in vivo bioluminescent imaging (BLI) to study the pathogenesis of MPX in animal models. Here, we evaluated BLI as a novel approach for tracking MPX virus infection in black-tailed prairie dogs (Cynomys ludovicianus). Prairie dogs were affected during a multistate outbreak of MPX in the US in 2003 and have since been used as an animal model of this disease. Our BLI results were compared with PCR and virus isolation from tissues collected postmortem. Virus was easily detected and quantified in skin and superficial tissues by BLI before and during clinical phases, as well as in subclinical secondary cases, but was not reliably detected in deep tissues such as the lung. Although there are limitations to viral detection in larger wild rodent species, BLI can enhance the use of prairie dogs as an animal model of MPX and can be used for the study of infection, disease progression, and transmission in potential wild rodent reservoirs.

  9. DNA Nanoparticles: Detection of Long-Term Transgene Activity in Brain using Bioluminescence Imaging

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    David M. Yurek

    2011-09-01

    Full Text Available In this study, we used bioluminescence imaging (BLI to track long-term transgene activity following the transfection of brain cells using a nonviral gene therapy technique. Formulations of deoxyribonucleic acid (DNA combined with 30-mer lysine polymers (substituted with 10 kDa polyethylene glycol form nanoparticles that transfect brain cells in vivo and produce transgene activity. Here we show that a single intracerebral injection of these DNA nanoparticles (DNPs into the rat cortex, striatum, or substantia nigra results in long-term and persistent luciferase transgene activity over an 8- to 11-week period as evaluated by in vivo BLI analysis, and single injections of DNPs into the mouse striatum showed stable luciferase transgene activity for 1 year. Compacted DNPs produced in vivo signals 7- to 34-fold higher than DNA alone. In contrast, ex vivo BLI analysis, which is subject to less signal quenching from surrounding tissues, demonstrated a DNP to DNA alone ratio of 76- to 280-fold. Moreover, the ex vivo BLI analysis confirmed that signals originated from the targeted brain structures. In summary, BLI permits serial analysis of luciferase transgene activity at multiple brain locations following gene transfer with DNPs. Ex vivo analysis may permit more accurate determination of relative activities of gene transfer vectors.

  10. Screening inhibitors of P. berghei blood stages using bioluminescent reporter parasites.

    Science.gov (United States)

    Lin, Jing-Wen; Sajid, Mohammed; Ramesar, Jai; Khan, Shahid M; Janse, Chris J; Franke-Fayard, Blandine

    2013-01-01

    We describe two improved assays for in vitro and in vivo screening of inhibitors and chemicals for antimalarial activity against blood stages of the rodent malaria parasite, Plasmodium berghei. These assays are based on the determination of bioluminescence in small blood samples that is produced by reporter parasites expressing luciferase. Luciferase production increases as the parasite develops in a red blood cell and as the numbers of parasites increase during an infection. In the first assay, in vitro drug luminescence (ITDL) assay, the in vitro development of ring-stage parasites into mature schizonts in the presence and absence of candidate inhibitor(s) is quantified by measuring luciferase activity after the parasites have been allowed to mature into schizonts in culture. In the second assay, the in vivo drug luminescence (IVDL) assay, in vivo parasite growth (using a standard 4-day suppressive drug test) is quantified by measuring the luciferase activity of circulating parasites in samples of tail blood of drug-treated mice.

  11. Measuring Cytotoxicity by Bioluminescence Imaging Outperforms the Standard Chromium-51 Release Assay

    Science.gov (United States)

    Karimi, Mobin A.; Lee, Eric; Bachmann, Michael H.; Salicioni, Ana Maria; Behrens, Edward M.; Kambayashi, Taku; Baldwin, Cynthia L.

    2014-01-01

    The chromium-release assay developed in 1968 is still the most commonly used method to measure cytotoxicity by T cells and by natural killer cells. Target cells are loaded in vitro with radioactive chromium and lysis is determined by measuring chromium in the supernatant released by dying cells. Since then, alternative methods have been developed using different markers of target cell viability that do not involve radioactivity. Here, we compared and contrasted a bioluminescence (BLI)-based cytotoxicity assay to the standard radioactive chromium-release assay using an identical set of effector cells and tumor target cells. For this, we stably transduced several human and murine tumor cell lines to express luciferase. When co-cultured with cytotoxic effector cells, highly reproducible decreases in BLI were seen in an effector to target cell dose-dependent manner. When compared to results obtained from the chromium release assay, the performance of the BLI-based assay was superior, because of its robustness, increased signal-to-noise ratio, and faster kinetics. The reduced/delayed detection of cytotoxicity by the chromium release method was attributable to the association of chromium with structural components of the cell, which are released quickly by detergent solubilization but not by hypotonic lysis. We conclude that the (BLI)-based measurement of cytotoxicity offers a superior non-radioactive alternative to the chromium-release assay that is more robust and quicker to perform. PMID:24586714

  12. Oculogryphus chenghoiyanae sp. n. (Coleoptera, Lampyridae: a new ototretine firefly from Hong Kong with descriptions of its bioluminescent behavior and ultraviolet-induced fluorescence in females

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    Vor Yiu

    2018-02-01

    Full Text Available The first Oculogryphus species with associated males and female was found in Hong Kong and is described as new: O. chenghoiyanae sp. n. Adults of both sexes were collected live in the field and their bioluminescent behavior is reported for the first time in the genus. The captive males emit weak and continuous light from a pair of light spots on abdominal ventrite 6 or do so when disturbed. The larviform (highly paedomorphic females can glow brightly from a pair of light-emitting organs on the abdomen. The females of Oculogryphus and Stenocladius are to date the only documented representatives of paedomorphism in ototretine fireflies. The finding is consistent with the evidence from male morphology and bioluminescent behavior, supporting the close relationship between the two genera. A key to the Oculogryphus species is provided. The Oculogryphus females can fluoresce with a blue-green light through the whole body under ultraviolet illumination, a phenomenon reported in the Lampyridae for the first time. The co-occurrence of bioluminescence and fluorescence is rare in terrestrial ecosystems, previously known only in some millipedes (Diplopoda. The fluorescence and bioluminescence abilities of Oculogryphus females are functionally independent: abdominal light-emitting organs producing bright yellowish green light while the body wall fluoresces with blue-green light. In contrast, fluorescence and bioluminescence in millipedes are biochemically linked, like in some jellyfish (Cnidaria: Medusozoa.

  13. Systematic study of target localization for bioluminescence tomography guided radiation therapy

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Jingjing [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University, Baltimore, Maryland 21231 and School of Physics and Information Technology, Shaanxi Normal University, Shaanxi 710119 (China); Zhang, Bin; Reyes, Juvenal; Wong, John W.; Wang, Ken Kang-Hsin, E-mail: kwang27@jhmi.edu [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University, Baltimore, Maryland 21231 (United States); Iordachita, Iulian I. [Laboratory for Computational Sensing and Robotics, Johns Hopkins University, Baltimore, Maryland 21218 (United States); Lu, Zhihao [Department of Oncology and Department of Surgery, Johns Hopkins University, Baltimore, Maryland 21231 and Key laboratory of Carcinogenesis and Translational Research, Department of GI Oncology, Peking University, Beijing Cancer Hospital and Institute, Beijing 100142 (China); Brock, Malcolm V. [Department of Oncology and Department of Surgery, Johns Hopkins University, Baltimore, Maryland 21231 (United States); Patterson, Michael S. [Department of Medical Physics and Applied Radiation Sciences, McMaster University, Hamilton, Ontario L8S 4L8 (Canada)

    2016-05-15

    Purpose: To overcome the limitation of CT/cone-beam CT (CBCT) in guiding radiation for soft tissue targets, the authors developed a spectrally resolved bioluminescence tomography (BLT) system for the small animal radiation research platform. The authors systematically assessed the performance of the BLT system in terms of target localization and the ability to resolve two neighboring sources in simulations, tissue-mimicking phantom, and in vivo environments. Methods: Multispectral measurements acquired in a single projection were used for the BLT reconstruction. The incomplete variables truncated conjugate gradient algorithm with an iterative permissible region shrinking strategy was employed as the optimization scheme to reconstruct source distributions. Simulation studies were conducted for single spherical sources with sizes from 0.5 to 3 mm radius at depth of 3–12 mm. The same configuration was also applied for the double source simulation with source separations varying from 3 to 9 mm. Experiments were performed in a standalone BLT/CBCT system. Two self-illuminated sources with 3 and 4.7 mm separations placed inside a tissue-mimicking phantom were chosen as the test cases. Live mice implanted with single-source at 6 and 9 mm depth, two sources at 3 and 5 mm separation at depth of 5 mm, or three sources in the abdomen were also used to illustrate the localization capability of the BLT system for multiple targets in vivo. Results: For simulation study, approximate 1 mm accuracy can be achieved at localizing center of mass (CoM) for single-source and grouped CoM for double source cases. For the case of 1.5 mm radius source, a common tumor size used in preclinical study, their simulation shows that for all the source separations considered, except for the 3 mm separation at 9 and 12 mm depth, the two neighboring sources can be resolved at depths from 3 to 12 mm. Phantom experiments illustrated that 2D bioluminescence imaging failed to distinguish two sources

  14. Propagation and perception of bioluminescence: factors affecting counterillumination as a cryptic strategy.

    Science.gov (United States)

    Johnsen, Sönke; Widder, Edith A; Mobley, Curtis D

    2004-08-01

    Many deep-sea species, particularly crustaceans, cephalopods, and fish, use photophores to illuminate their ventral surfaces and thus disguise their silhouettes from predators viewing them from below. This strategy has several potential limitations, two of which are examined here. First, a predator with acute vision may be able to detect the individual photophores on the ventral surface. Second, a predator may be able to detect any mismatch between the spectrum of the bioluminescence and that of the background light. The first limitation was examined by modeling the perceived images of the counterillumination of the squid Abralia veranyi and the myctophid fish Ceratoscopelus maderensis as a function of the distance and visual acuity of the viewer. The second limitation was addressed by measuring downwelling irradiance under moonlight and starlight and then modeling underwater spectra. Four water types were examined: coastal water at a depth of 5 m and oceanic water at 5, 210, and 800 m. The appearance of the counterillumination was more affected by the visual acuity of the viewer than by the clarity of the water, even at relatively large distances. Species with high visual acuity (0.11 degrees resolution) were able to distinguish the individual photophores of some counterilluminating signals at distances of several meters, thus breaking the camouflage. Depth and the presence or absence of moonlight strongly affected the spectrum of the background light, particularly near the surface. The increased variability near the surface was partially offset by the higher contrast attenuation at shallow depths, which reduced the sighting distance of mismatches. This research has implications for the study of spatial resolution, contrast sensitivity, and color discrimination in deep-sea visual systems.

  15. Comparison of in vivo bioluminescence imaging and lavage biomarkers to assess pulmonary inflammation.

    Science.gov (United States)

    Hadina, Suzana; Wohlford-Lenane, Christine L; Thorne, Peter S

    2012-01-27

    Gram-negative bacterial endotoxin triggers innate immunity via TLR-4 and NF-kB signal activation. The aim of this study was to evaluate the use of transgenic mice expressing luciferase as a marker of NF-kB activation for exploring innate immune responses to pulmonary endotoxin exposure over time thus obviating the need for serial necropsies. Transgenic rNF-kB-Luc BALB/c mice were exposed to two different types of endotoxin (Neisseria meningitidis lipooligosaccharide, and Escherichia coli lipopolysaccharide) at multiple doses by nasal instillation. Bioluminescence was quantified in vivo at five time points in three separate experiments. In the fourth experiment lungs were imaged ex vivo 8h post exposure and tissue was analyzed for luciferase activity. Non-transgenic BALB/c mice were similarly exposed to lipooligosaccharide and bronchoalveolar lavage was assessed for neutrophil recruitment and IL-6. Non-transgenic BALB/c mice exhibited highly significant increases of IL-6 and neutrophils in bronchoalveolar lavage 4h after the exposure to instilled doses as low as 30EU/mouse. In contrast, luciferase imaging of NF-kB signal activation in vivo in transgenic rNF-kB-Luc mice did not show significant changes over time or over doses from 30EU to 300,000EU/mouse of nasally-instilled endotoxin. Ex vivo lung imaging 8h after endotoxin exposure to 3000EU demonstrated a strong signal. An intravenous LPS dose of 300,000EU/mouse produced a measurable luminescence signal in vivo. This non-terminal assessment method is useful only with extremely high doses of endotoxin that induce systemic injury and cannot be applied to research of occupational and environmental exposures at relevant levels of endotoxin. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  16. Comparison of static and microfluidic protease assays using modified bioluminescence resonance energy transfer chemistry.

    Directory of Open Access Journals (Sweden)

    Nan Wu

    Full Text Available BACKGROUND: Fluorescence and bioluminescence resonance energy transfer (F/BRET are two forms of Förster resonance energy transfer, which can be used for optical transduction of biosensors. BRET has several advantages over fluorescence-based technologies because it does not require an external light source. There would be benefits in combining BRET transduction with microfluidics but the low luminance of BRET has made this challenging until now. METHODOLOGY: We used a thrombin bioprobe based on a form of BRET (BRET(H, which uses the BRET(1 substrate, native coelenterazine, with the typical BRET(2 donor and acceptor proteins linked by a thrombin target peptide. The microfluidic assay was carried out in a Y-shaped microfluidic network. The dependence of the BRET(H ratio on the measurement location, flow rate and bioprobe concentration was quantified. Results were compared with the same bioprobe in a static microwell plate assay. PRINCIPAL FINDINGS: The BRET(H thrombin bioprobe has a lower limit of detection (LOD than previously reported for the equivalent BRET(1-based version but it is substantially brighter than the BRET(2 version. The normalised BRET(H ratio of the bioprobe changed 32% following complete cleavage by thrombin and 31% in the microfluidic format. The LOD for thrombin in the microfluidic format was 27 pM, compared with an LOD of 310 pM, using the same bioprobe in a static microwell assay, and two orders of magnitude lower than reported for other microfluidic chip-based protease assays. CONCLUSIONS: These data demonstrate that BRET based microfluidic assays are feasible and that BRET(H provides a useful test bed for optimising BRET-based microfluidics. This approach may be convenient for a wide range of applications requiring sensitive detection and/or quantification of chemical or biological analytes.

  17. Image Processing for Bioluminescence Resonance Energy Transfer Measurement-BRET-Analyzer.

    Science.gov (United States)

    Chastagnier, Yan; Moutin, Enora; Hemonnot, Anne-Laure; Perroy, Julie

    2017-01-01

    A growing number of tools now allow live recordings of various signaling pathways and protein-protein interaction dynamics in time and space by ratiometric measurements, such as Bioluminescence Resonance Energy Transfer (BRET) Imaging. Accurate and reproducible analysis of ratiometric measurements has thus become mandatory to interpret quantitative imaging. In order to fulfill this necessity, we have developed an open source toolset for Fiji- BRET-Analyzer -allowing a systematic analysis, from image processing to ratio quantification. We share this open source solution and a step-by-step tutorial at https://github.com/ychastagnier/BRET-Analyzer. This toolset proposes (1) image background subtraction, (2) image alignment over time, (3) a composite thresholding method of the image used as the denominator of the ratio to refine the precise limits of the sample, (4) pixel by pixel division of the images and efficient distribution of the ratio intensity on a pseudocolor scale, and (5) quantification of the ratio mean intensity and standard variation among pixels in chosen areas. In addition to systematize the analysis process, we show that the BRET-Analyzer allows proper reconstitution and quantification of the ratiometric image in time and space, even from heterogeneous subcellular volumes. Indeed, analyzing twice the same images, we demonstrate that compared to standard analysis BRET-Analyzer precisely define the luminescent specimen limits, enlightening proficient strengths from small and big ensembles over time. For example, we followed and quantified, in live, scaffold proteins interaction dynamics in neuronal sub-cellular compartments including dendritic spines, for half an hour. In conclusion, BRET-Analyzer provides a complete, versatile and efficient toolset for automated reproducible and meaningful image ratio analysis.

  18. Image Processing for Bioluminescence Resonance Energy Transfer Measurement—BRET-Analyzer

    Directory of Open Access Journals (Sweden)

    Yan Chastagnier

    2018-01-01

    Full Text Available A growing number of tools now allow live recordings of various signaling pathways and protein-protein interaction dynamics in time and space by ratiometric measurements, such as Bioluminescence Resonance Energy Transfer (BRET Imaging. Accurate and reproducible analysis of ratiometric measurements has thus become mandatory to interpret quantitative imaging. In order to fulfill this necessity, we have developed an open source toolset for Fiji—BRET-Analyzer—allowing a systematic analysis, from image processing to ratio quantification. We share this open source solution and a step-by-step tutorial at https://github.com/ychastagnier/BRET-Analyzer. This toolset proposes (1 image background subtraction, (2 image alignment over time, (3 a composite thresholding method of the image used as the denominator of the ratio to refine the precise limits of the sample, (4 pixel by pixel division of the images and efficient distribution of the ratio intensity on a pseudocolor scale, and (5 quantification of the ratio mean intensity and standard variation among pixels in chosen areas. In addition to systematize the analysis process, we show that the BRET-Analyzer allows proper reconstitution and quantification of the ratiometric image in time and space, even from heterogeneous subcellular volumes. Indeed, analyzing twice the same images, we demonstrate that compared to standard analysis BRET-Analyzer precisely define the luminescent specimen limits, enlightening proficient strengths from small and big ensembles over time. For example, we followed and quantified, in live, scaffold proteins interaction dynamics in neuronal sub-cellular compartments including dendritic spines, for half an hour. In conclusion, BRET-Analyzer provides a complete, versatile and efficient toolset for automated reproducible and meaningful image ratio analysis.

  19. Specific expression of bioluminescence reporter gene in cardiomyocyte regulated by tissue specific promoter

    Energy Technology Data Exchange (ETDEWEB)

    Nguyen, Vu Hong; Tae, Seong Ho; Le, Nguyen Uyen Chi; Min, Jung Joon [Chonnam National University Medical School, Gwangju (Korea, Republic of)

    2007-07-01

    As the human heart is not capable of regenerating the great numbers of cardiac cells that are lost after myocardial infarction, impaired cardiac function is the inevitable result of ischemic disease. Recently, human embryonic stem cells (hESCs) have gained popularity as a potentially ideal cell candidate for tissue regeneration. In particular, hESCs are capable of cardiac lineage-specific differentiation and confer improvement of cardiac function following transplantation into animal models. Although such data are encouraging, the specific strategy for in vivo and non-invasive detection of differentiated cardiac lineage is still limited. Therefore, in the present study, we established the gene construction in which the optical reporter gene Firefly luciferase was controlled by Myosin Heavy Chain promoter for specific expressing in heart cells. The vector consisting of - MHC promoter and a firefly luciferase coding sequence flanked by full-length bovine growth hormone (BGH) 3'-polyadenylation sequence based on pcDNA3.1- vector backbone. To test the specific transcription of this promoter in g of MHC-Fluc or CMV-Flue (for control) plasmid DNA in myocardial tissue, 20 phosphate-buffered saline was directly injected into mouse myocardium through a midline sternotomy and liver. After 1 week of injection, MHC-Fluc expression was detected from heart region which was observed under cooled CCD camera of in vivo imaging system but not from liver. In control group injected with CMV-Flue, the bioluminescence was detected from all these organs. The expression of Flue under control of Myosin Heavy Chain promoter may become a suitable optical reporter gene for stem cell-derived cardiac lineage differentiation study.

  20. Bioluminescence inhibition assays for toxicity screening of wood extractives and biocides in paper mill process waters.

    Science.gov (United States)

    Rigol, Anna; Latorre, Anna; Lacorte, Sílvia; Barceló, Damià

    2004-02-01

    The risk associated with wood extractives, biocides, and other additives in pulp and paper mill effluents was evaluated by performing a characterization of process waters and effluents in terms of toxicity and chemical analysis. The individual toxicity of 10 resin acids, two unsaturated fatty acids, and three biocides was estimated by measuring the bioluminescence inhibition with a ToxAlert 100 system. Median effective concentration values (EC50) of 4.3 to 17.9, 1.2 to 1.5, and 0.022 to 0.50 mg/L were obtained, respectively. Mixtures of these three families of compounds showed antagonistic effects. Chemical analysis of process waters was performed by liquid chromatography- and gas chromatography-mass spectrometry. Biocides such as 2-(thiocyanomethylthio)-benzotiazole (TCMTB) (EC50 = 0.022 mg/L) and 2,2-dibromo-3-nitrilpropionamide (DBNPA) (EC50 = 0.50 mg/L) were the most toxic compounds tested and were detected at concentrations of 16 and 59 microg/L, respectively, in a closed-circuit recycling paper mill. Process waters from kraft pulp mills, printing paper mills, and packing board paper mills showed the highest concentration of resin acids (up to 400 microg/L) and accounted for inhibition percentages up to 100%. Detergent degradation products such as nonylphenol (NP) and octylphenol (OP) and the plasticizer bisphenol A (BPA) were also detected in the waters at levels of 0.6 to 10.6, 0.3 to 1.4, and 0.7 to 187 microg/L, respectively. However, once these waters were biologically treated, the concentration of detected organic compounds diminished and the toxicity decreased in most cases to values of inhibition lower than 20%.

  1. Continuous long-term cytotoxicity monitoring in 3D spheroids of beetle luciferase-expressing hepatocytes by nondestructive bioluminescence measurement.

    Science.gov (United States)

    Yasunaga, Mayu; Fujita, Yasuko; Saito, Rumiko; Oshimura, Mitsuo; Nakajima, Yoshihiro

    2017-06-20

    Three-dimensional (3D) spheroids are frequently used in toxicological study because their morphology and function closely resemble those of tissue. As these properties are maintained over a long term, repeated treatment of the spheroids with a test object is possible. Generally, in the repeated treatment test to assess cytotoxicity in the spheroids, ATP assay, colorimetric measurement using pigments or high-content imaging analysis is performed. However, continuous assessment of cytotoxicity in the same spheroids using the above assays or analysis is impossible because the spheroids must be disrupted or killed. To overcome this technical limitation, we constructed a simple monitoring system in which cytotoxicity in the spheroids can be continuously monitored by nondestructive bioluminescence measurement. Mouse primary hepatocytes were isolated from transchromosomic (Tc) mice harboring a mouse artificial chromosome (MAC) vector expressing beetle luciferase Emerald Luc (ELuc) under the control of cytomegalovirus immediate early enhancer/chicken β-actin promoter/rabbit β-globin intron II (CAG) promoter, and used in 3D cultures. We confirmed that both luminescence and albumin secretion from the spheroids seeded in the 96-well format Cell-able TM were maintained for approximately 1 month. Finally, we repetitively treated the luminescent 3D spheroids with representative hepatotoxicants for approximately 1 month, and continuously and nondestructively measured bioluminescence every day. We successfully obtained daily changes of the dose-response bioluminescence curves for the respective toxicants. In this study, we constructed a monitoring system in which cytotoxicity in the same 3D spheroids was continuously and sensitively monitored over a long term. Because this system can be easily applied to other cells, such as human primary cells or stem cells, it is expected to serve as the preferred platform for simple and cost-effective long-term monitoring of cellular events

  2. Phylogenetic relationships of click beetles (Coleoptera: Elateridae) inferred from 28S ribosomal DNA: insights into the evolution of bioluminescence in Elateridae.

    Science.gov (United States)

    Sagegami-Oba, Reiko; Oba, Yuichi; Ohira, Hitoo

    2007-02-01

    Although the taxonomy of click beetles (family Elateridae) has been studied extensively, inconsistencies remain. We examine here the relationships between species of Elateridae based on partial sequences of nuclear 28S ribosomal DNA. Specimens were collected primarily from Japan, while luminous click beetles were also sampled from Central and South America to investigate the origins of bioluminescence in Elateridae. Neighbor-joining, maximum-parsimony, and maximum-likelihood analyses produced a consistent basal topology with high statistical support that is partially congruent with the results of previous investigations based on the morphological characteristics of larvae and adults. The most parsimonious reconstruction of the "luminous" and "nonluminous" states, based on the present molecular phylogeny, indicates that the ancestral state of Elateridae was nonluminous. This suggests that the bioluminescence in click beetle evolved independent of that of other luminous beetles, such as Lampyridae, despite their common mechanisms of bioluminescence.

  3. Ultra-thin titanium nanolayers for plasmon-assisted enhancement of bioluminescence of chloroplast in biological light emitting devices

    Energy Technology Data Exchange (ETDEWEB)

    Hsun Su, Yen [Department of Materials Science and Engineering, National Cheng Kung University, Tainan 70101, Taiwan (China); Advanced Optoelectronic Technology Center, National Cheng Kung University, Tainan 70101, Taiwan (China); Hsu, Chia-Yun; Chang, Chung-Chien [Science and Technology of Accelerator Light Source, Hsinchu 300, Taiwan (China); Department of Materials Science and Engineering, National Chiao Tung University, Hsinchu 300, Taiwan (China); Tu, Sheng-Lung; Shen, Yun-Hwei [Department of Resource Engineering, National Cheng Kung University, Tainan 70101, Taiwan (China)

    2013-08-05

    Ultra-thin titanium films were deposited via ultra-high vacuum ion beam sputter deposition. Since the asymmetric electric field of the metal foil plane matches the B-band absorption of chlorophyll a, the ultra-thin titanium nanolayers were able to generate surface plasmon resonance, thus enhancing the photoluminescence of chlorophyll a. Because the density of the states of plasmon resonance increases, the enhancement of photoluminescence also rises. Due to the biocompatibility and inexpensiveness of titanium, it can be utilized to enhance the bioluminescence of chloroplast in biological light emitting devices, bio-laser, and biophotonics.

  4. Chronic toxicity evaluation of wastewater treatment plant effluents with bioluminescent bacteria: A comparison with invertebrates and fish

    Energy Technology Data Exchange (ETDEWEB)

    Sweet, L.I.; Travers, D.F.; Meier, P.G. [Univ. of Michigan, Ann Arbor, MI (United States)

    1997-10-01

    The use of bioluminescent bacteria in chronic toxicity testing is a potentially useful yet unexplored tool in whole effluent biomonitoring. The purpose of this study was to determine the chronic toxicity of 14 different wastewater treatment plant effluents to Chronic Microtox{reg_sign} bacteria (Vibrio fischeri), a cladoceran (Ceriodaphnia dubia), and a minnow (Pimephales promelas). The invertebrate and fish have been utilized extensively for the evaluation of effluents and in establishing water quality criteria. The results of this study suggest that the 22-h Microtox Chronic Toxicity Test may correlate well with the most sensitive chronic no-observed-effect concentration value of the three-brood C. dubia test.

  5. Light and vision in the deep-sea benthos: I. Bioluminescence at 500-1000 m depth in the Bahamian islands.

    Science.gov (United States)

    Johnsen, Sönke; Frank, Tamara M; Haddock, Steven H D; Widder, Edith A; Messing, Charles G

    2012-10-01

    Bioluminescence is common and well studied in mesopelagic species. However, the extent of bioluminescence in benthic sites of similar depths is far less studied, although the relatively large eyes of benthic fish, crustaceans and cephalopods at bathyal depths suggest the presence of significant biogenic light. Using the Johnson-Sea-Link submersible, we collected numerous species of cnidarians, echinoderms, crustaceans, cephalopods and sponges, as well as one annelid from three sites in the northern Bahamas (500-1000 m depth). Using mechanical and chemical stimulation, we tested the collected species for light emission, and photographed and measured the spectra of the emitted light. In addition, in situ intensified video and still photos were taken of different benthic habitats. Surprisingly, bioluminescence in benthic animals at these sites was far less common than in mesopelagic animals from similar depths, with less than 20% of the collected species emitting light. Bioluminescent taxa comprised two species of anemone (Actinaria), a new genus and species of flabellate Parazoanthidae (formerly Gerardia sp.) (Zoanthidea), three sea pens (Pennatulacea), three bamboo corals (Alcyonacea), the chrysogorgiid coral Chrysogorgia desbonni (Alcyonacea), the caridean shrimp Parapandalus sp. and Heterocarpus ensifer (Decapoda), two holothuroids (Elasipodida and Aspidochirota) and the ophiuroid Ophiochiton ternispinus (Ophiurida). Except for the ophiuroid and the two shrimp, which emitted blue light (peak wavelengths 470 and 455 nm), all the species produced greener light than that measured in most mesopelagic taxa, with the emissions of the pennatulaceans being strongly shifted towards longer wavelengths. In situ observations suggested that bioluminescence associated with these sites was due primarily to light emitted by bioluminescent planktonic species as they struck filter feeders that extended into the water column.

  6. Transfection efficiency of normal and cancer cell lines and monitoring of promoter activity by single-cell bioluminescence imaging.

    Science.gov (United States)

    Horibe, Tomohisa; Torisawa, Aya; Akiyoshi, Ryutaro; Hatta-Ohashi, Yoko; Suzuki, Hirobumi; Kawakami, Koji

    2014-02-01

    The bioluminescence system (luciferase reporter assay system) is widely used to study gene expression, signal transduction and other cellular activities. Although transfection of reporter plasmid DNA to mammalian cell lines is an indispensable experimental step, the transfection efficiency of DNA varies among cell lines, and several cell lines are not suitable for this type of assay because of the low transfection efficiency. In this study, we confirm the transfection efficiency of reporter DNA to several cancer and normal cell lines after transient transfection by single-cell imaging. Luminescence images could be obtained from living single cells after transient transfection, and the calculated transfection efficiency of this method was similar to that of the conventional reporter assay using a luminometer. We attempted to measure the activity of the Bip promoter under endoplasmic reticulum stress conditions using both high and low transfection efficiency cells for plasmid DNA at the single-cell level, and observed activation of this promoter even in cells with the lowest transfection efficiency. These results show that bioluminescence imaging of single cells is a powerful tool for the analysis of gene expression based on a reporter assay using limited samples such as clinical specimens or cells from primary culture, and could provide additional information compared with the conventional assay. Copyright © 2013 John Wiley & Sons, Ltd.

  7. Proteomic analysis and bioluminescent reporter gene assays to investigate effects of simulated microgravity on Caco-2 cells.

    Science.gov (United States)

    La Barbera, Giorgia; Capriotti, Anna Laura; Michelini, Elisa; Piovesana, Susy; Calabretta, Maria Maddalena; Zenezini Chiozzi, Riccardo; Roda, Aldo; Laganà, Aldo

    2017-08-01

    Microgravity is one of the most important features in spaceflight. Previous evidence from in-vitro studies has shown that significant changes occur under simulated microgravity. For this reason, human colon adenocarcinoma Caco-2 cells were selected as cell model of intestinal epithelial barrier and their response to altered gravity conditions was investigated, especially on the protein level. In this study, we combined label-free shotgun proteomics and bioluminescent reporter gene assays to identify key proteins and pathways involved in the response of Caco-2 cells under reference and microgravity conditions. A two-dimensional clinostat was modified with 3D-printed adaptors to hold conventional T25 culture flasks. The comparative proteome analysis led to identify 38 and 26 proteins differently regulated by simulated microgravity after 48 and 72 h, respectively. Substantial fractions of these proteins are involved in regulation, cellular and metabolic processes and localization. Bioluminescent reporter gene assays were carried out to investigate microgavity-induced alterations on the transcriptional regulation of key targets, such as NF-kB pathway and CYP27A1. While no significant difference was found in the basal transcription, a lower NF-kB basal activation in simulated microgravity conditions was reported, corroborating the hypothesis of reduced immunity in microgravity conditions. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Effect of optical property estimation accuracy on tomographic bioluminescence imaging: simulation of a combined optical-PET (OPET) system

    Energy Technology Data Exchange (ETDEWEB)

    Alexandrakis, George [Crump Institute for Molecular Imaging, David Geffen School of Medicine at UCLA, University of California, 700 Westwood Plaza, Los Angeles, CA 90095 (United States); Rannou, Fernando R [Departamento de Ingenieria Informatica, Universidad de Santiago de Chile (USACH), Av. Ecuador 3659, Santiago (Chile); Chatziioannou, Arion F [Crump Institute for Molecular Imaging, David Geffen School of Medicine at UCLA, University of California, 700 Westwood Plaza, Los Angeles, CA 90095 (United States)

    2006-04-21

    Inevitable discrepancies between the mouse tissue optical properties assumed by an experimenter and the actual physiological values may affect the tomographic localization of bioluminescent sources. In a previous work, the simplifying assumption of optically homogeneous tissues led to inaccurate localization of deep sources. Improved results may be obtained if a mouse anatomical map is provided by a high-resolution imaging modality and optical properties are assigned to segmented tissues. In this work, the feasibility of this approach was explored by simulating the effect of different magnitude optical property errors on the image formation process of a combined optical-PET system. Some comparisons were made with corresponding simulations using higher spatial resolution data that are typically attainable by CCD cameras. In addition, simulation results provided insights on some of the experimental conditions that could lead to poor localization of bioluminescent sources. They also provided a rough guide on how accurately tissue optical properties need to be known in order to achieve correct localization of point sources with increasing tissue depth under low background noise conditions.

  9. Development and application of the lux gene for environmental bioremediation

    Energy Technology Data Exchange (ETDEWEB)

    Burlage, R.S.; Yang, Z. [Oak Ridge National Lab., TN (United States). Environmental Sciences Div.; Palmer, R.J. [Univ. of Tennessee, Knoxville, TN (United States). Center for Environmental Biotechnology; Khang, Y. [Yeungnam Univ., Kyongsan (Korea, Republic of)

    1996-09-01

    Bioremediation is the use of living systems, usually microorganisms, to treat a quantity of soil or water for the presence of hazardous wastes. Bioremediation has many advantages over other remediation approaches, including cost savings, versatility, and the ability to treat the wastes in situ. In order to study the processes of microbial bioremediation, the authors have constructed bacterial strains that incorporate genetically engineered bioreporter genes. These bioreporter genes allow the bacteria to be detected during in situ processes, as manifested by their ability to bioluminescence or to fluoresce. This bioreporter microorganisms are described, along with the technology for detecting them and the projects which are benefiting from their application.

  10. Noninvasive monitoring of placenta-specific transgene expression by bioluminescence imaging.

    Directory of Open Access Journals (Sweden)

    Xiujun Fan

    Full Text Available BACKGROUND: Placental dysfunction underlies numerous complications of pregnancy. A major obstacle to understanding the roles of potential mediators of placental pathology has been the absence of suitable methods for tissue-specific gene manipulation and sensitive assays for studying gene functions in the placentas of intact animals. We describe a sensitive and noninvasive method of repetitively tracking placenta-specific gene expression throughout pregnancy using lentivirus-mediated transduction of optical reporter genes in mouse blastocysts. METHODOLOGY/PRINCIPAL FINDINGS: Zona-free blastocysts were incubated with lentivirus expressing firefly luciferase (Fluc and Tomato fluorescent fusion protein for trophectoderm-specific infection and transplanted into day 3 pseudopregnant recipients (GD3. Animals were examined for Fluc expression by live bioluminescence imaging (BLI at different points during pregnancy, and the placentas were examined for tomato expression in different cell types on GD18. In another set of experiments, blastocysts with maximum photon fluxes in the range of 2.0E+4 to 6.0E+4 p/s/cm(2/sr were transferred. Fluc expression was detectable in all surrogate dams by day 5 of pregnancy by live imaging, and the signal increased dramatically thereafter each day until GD12, reaching a peak at GD16 and maintaining that level through GD18. All of the placentas, but none of the fetuses, analyzed on GD18 by BLI showed different degrees of Fluc expression. However, only placentas of dams transferred with selected blastocysts showed uniform photon distribution with no significant variability of photon intensity among placentas of the same litter. Tomato expression in the placentas was limited to only trophoblast cell lineages. CONCLUSIONS/SIGNIFICANCE: These results, for the first time, demonstrate the feasibility of selecting lentivirally-transduced blastocysts for uniform gene expression in all placentas of the same litter and early

  11. Strong electron correlation in the decomposition reaction of dioxetanone with implications for firefly bioluminescence.

    Science.gov (United States)

    Greenman, Loren; Mazziotti, David A

    2010-10-28

    Dioxetanone, a key component of the bioluminescence of firefly luciferin, is itself a chemiluminescent molecule due to two conical intersections on its decomposition reaction surface. While recent calculations of firefly luciferin have employed four electrons in four active orbitals [(4,4)] for the dioxetanone moiety, a study of dioxetanone [F. Liu et al., J. Am. Chem. Soc. 131, 6181 (2009)] indicates that a much larger active space is required. Using a variational calculation of the two-electron reduced-density-matrix (2-RDM) [D. A. Mazziotti, Acc. Chem. Res. 39, 207 (2006)], we present the ground-state potential energy surface as a function of active spaces from (4,4) to (20,17) to determine the number of molecular orbitals required for a correct treatment of the strong electron correlation near the conical intersections. Because the 2-RDM method replaces exponentially scaling diagonalizations with polynomially scaling semidefinite optimizations, we readily computed large (18,15) and (20,17) active spaces that are inaccessible to traditional wave function methods. Convergence of the electron correlation with active-space size was measured with complementary RDM-based metrics, the von Neumann entropy of the one-electron RDM as well as the Frobenius and infinity norms of the cumulant 2-RDM. Results show that the electron correlation is not correctly described until the (14,12) active space with small variations present through the (20,17) space. Specifically, for active spaces smaller than (14,12), we demonstrate that at the first conical intersection, the electron in the σ(∗) orbital of the oxygen-oxygen bond is substantially undercorrelated with the electron of the σ orbital and overcorrelated with the electron of the carbonyl oxygen's p orbital. Based on these results, we estimate that in contrast to previous treatments, an accurate calculation of the strong electron correlation in firefly luciferin requires an active space of 28 electrons in 25 orbitals

  12. In vivo bioluminescence imaging for leptomeningeal dissemination of medulloblastoma in mouse models

    International Nuclear Information System (INIS)

    Choi, Seung Ah; Kwak, Pil Ae; Kim, Seung-Ki; Park, Sung-Hye; Lee, Ji Yeoun; Wang, Kyu-Chang; Oh, Hyun Jeong; Kim, Kyuwan; Lee, Dong Soo; Hwang, Do Won; Phi, Ji Hoon

    2016-01-01

    The primary cause of treatment failure in medulloblastomas (MB) is the development of leptomeningeal dissemination (seeding). For translational research on MB seeding, one of the major challenges is the development of reliable experimental models that simulate the seeding and growth characteristics of MBs. To overcome this obstacle, we improved an experimental mouse model by intracisternal inoculation of human MB cells and monitoring with in vivo live images. Human MB cells (UW426, D283 and MED8A) were transfected with a firefly luciferase gene and a Thy1.1 (CD90.1) marker linked with IRES under the control of the CMV promoter in a retroviral DNA backbone (effLuc). The MB-effLuc cells were injected into the cisterna magna using an intrathecal catheter, and bioluminescence images were captured. We performed histopathological analysis to confirm the extent of tumor seeding. The luciferase activity of MB-effLuc cells displayed a gradually increasing pattern, which correlated with a quantitative luminometric assay. Live imaging showed that the MB-effLuc cells were diffusely distributed in the cervical spinal cord and the lumbosacral area. All mice injected with UW426-effLuc, D283-effLuc and MED8A-effLuc died within 51 days. The median survival was 22, 41 and 12 days after injection of 1.2 × 10 6 UW426-effLuc, D283-effLuc and MED8A-effLuc cells, respectively. The histopathological studies revealed that the MB-effLuc cells spread extensively and diffusely along the leptomeninges of the brain and spinal cord, forming tumor cell-coated layers. The tumor cells in the subarachnoid space expressed a human nuclei marker and Ki-67. Compared with the intracerebellar injection method in which the subfrontal area and distal spinal cord were spared by tumor cell seeding in some mice, the intracisternal injection model more closely resembled the widespread leptomeningeal seeding observed in MB patients. The results and described method are valuable resources for further

  13. A Bone Metastasis Nude Mouse Model Created by Ultrasound Guided Intracardiac Injection of Breast Cancer Cells: the Micro-CT, MRI and Bioluminescence Imaging Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Park, Young Jin; Song, Eun Hye; Kim, Seol Hwa; Song, Ho Taek; Suh, Jin Suck [Yonsei University College of Medicine, Seoul (Korea, Republic of); Choi, Sang Hyun [Korean Minjok Leadership Academy, Heongsung (Korea, Republic of)

    2011-01-15

    The purpose of this study was to develop a nude mouse model of bone metastasis by performing intracardiac injection of breast cancer cells under ultrasonography guidance and we wanted to evaluate the development and the distribution of metastasis in vivo using micro-CT, MRI and bioluminescence imaging. Animal experiments were performed in 6-week-old female nude mice. The animals underwent left ventricular injection of 2x105 MDA-MB-231Bo-Luc cells. After injection of the tumor cells, serial bioluminescence imaging was performed for 7 weeks. The findings of micro-CT, MRI and the histology were correlated with the 'hot' lesions seen on the bioluminescence imaging. Metastasis was found in 62.3% of the animals. Two weeks after intracardiac injection, metastasis to the brain, spine and femur was detected with bioluminescence imaging with an increasing intensity by week 7. Micro-CT scan confirmed multiple osteolytic lesions at the femur, spine and skull. MRI and the histology were able to show metastasis in the brain and extraskeletal metastasis around the femur. The intracardiac injection of cancer cells under ultrasonography guidance is a safe and highly reproducible method to produce bone metastasis in nude mice. This bone metastasis nude mouse model will be useful to study the mechanism of bone metastasis and to validate new therapeutics

  14. Stably Integrated luxCDABE for Assessment of Salmonella Invasion Kinetics

    Directory of Open Access Journals (Sweden)

    Kelly N. Flentie

    2008-09-01

    Full Text Available Salmonella Typhimurium is a common cause of gastroenteritis in humans and also localizes to neoplastic tumors in animals. Invasion of specific eukaryotic cells is a key mechanism of Salmonella interactions with host tissues. Early stages of gastrointestinal cell invasion are mediated by a Salmonella type III secretion system, powered by the adenosine triphosphatase invC. The aim of this work was to characterize the invC dependence of invasion kinetics into disparate eukaryotic cells traditionally used as models of gut epithelium or neoplasms. Thus, a nondestructive real-time assay was developed to report eukaryotic cell invasion kinetics using lux+ Salmonella that contain chromosomally integrated luxCDABE genes. Bioluminescence-based invasion assays using lux+ Salmonella exhibited inoculum dose-response correlation, distinguished invasion-competent from invasion-incompetent Salmonella, and discriminated relative Salmonella invasiveness in accordance with environmental conditions that induce invasion gene expression. In standard gentamicin protection assays, bioluminescence from lux+ Salmonella correlated with recovery of colony-forming units of internalized bacteria and could be visualized by bioluminescence microscopy. Furthermore, this assay distinguished invasion-competent from invasion-incompetent bacteria independent of gentamicin treatment in real time. Bioluminescence reported Salmonella invasion of disparate eukaryotic cell lines, including neoplastic melanoma, colon adenocarcinoma, and glioma cell lines used in animal models of malignancy. In each case, Salmonella invasion of eukaryotic cells was invC dependent.

  15. Live imaging of bioluminescent leptospira interrogans in mice reveals renal colonization as a stealth escape from the blood defenses and antibiotics.

    Directory of Open Access Journals (Sweden)

    Gwenn Ratet

    2014-12-01

    Full Text Available Leptospira (L. interrogans are bacteria responsible for a worldwide reemerging zoonosis. Some animals asymptomatically carry L. interrogans in their kidneys and excrete bacteria in their urine, which contaminates the environment. Humans are infected through skin contact with leptospires and develop mild to severe leptospirosis. Previous attempts to construct fluorescent or bioluminescent leptospires, which would permit in vivo visualization and investigation of host defense mechanisms during infection, have been unsuccessful. Using a firefly luciferase cassette and random transposition tools, we constructed bioluminescent chromosomal transformants in saprophytic and pathogenic leptospires. The kinetics of leptospiral dissemination in mice, after intraperitoneal inoculation with a pathogenic transformant, was tracked by bioluminescence using live imaging. For infective doses of 106 to 107 bacteria, we observed dissemination and exponential growth of leptospires in the blood, followed by apparent clearance of bacteria. However, with 2×108 bacteria, the septicemia led to the death of mice within 3 days post-infection. In surviving mice, one week after infection, pathogenic leptospires reemerged only in the kidneys, where they multiplied and reached a steady state, leading to a sustained chronic renal infection. These experiments reveal that a fraction of the leptospiral population escapes the potent blood defense, and colonizes a defined number of niches in the kidneys, proportional to the infective dose. Antibiotic treatments failed to eradicate leptospires that colonized the kidneys, although they were effective against L. interrogans if administered before or early after infection. To conclude, mice infected with bioluminescent L. interrogans proved to be a novel model to study both acute and chronic leptospirosis, and revealed that, in the kidneys, leptospires are protected from antibiotics. These bioluminescent leptospires represent a

  16. Mitrocomin from the jellyfish Mitrocoma cellularia with deleted C-terminal tyrosine reveals a higher bioluminescence activity compared to wild type photoprotein

    Energy Technology Data Exchange (ETDEWEB)

    Burakova, Ludmila P.; Natashin, Pavel V.; Markova, Svetlana V.; Eremeeva, Elena V.; Malikova, Natalia P.; Cheng, Chongyun; Liu, Zhi-Jie; Vysotski, Eugene S.

    2016-09-01

    The full-length cDNA genes encoding five new isoforms of Ca2 +-regulated photoprotein mitrocomin from a small tissue sample of the outer bell margin containing photocytes of only one specimen of the luminous jellyfish Mitrocoma cellularia were cloned, sequenced, and characterized after their expression in Escherichia coli and subsequent purification. The analysis of cDNA nucleotide sequences encoding mitrocomin isoforms allowed suggestion that two isoforms might be the products of two allelic genes differing in one amino acid residue (64R/Q) whereas other isotypes appear as a result of transcriptional mutations. In addition, the crystal structure of mitrocomin was determined at 1.30 Å resolution which expectedly revealed a high similarity with the structures of other hydromedusan photoproteins. Although mitrocomin isoforms reveal a high degree of identity of amino acid sequences, they vary in specific bioluminescence activities. At that, all isotypes displayed the identical bioluminescence spectra (473–474 nm with no shoulder at 400 nm). Fluorescence spectra of Ca2 +-discharged mitrocomins were almost identical to their light emission spectra similar to the case of Ca2 +-discharged aequorin, but different from Ca2 +-discharged obelins and clytin which fluorescence is red-shifted by 25–30 nm from bioluminescence spectra. The main distinction of mitrocomin from other hydromedusan photoproteins is an additional Tyr at the C-terminus. Using site-directed mutagenesis, we showed that this Tyr is not important for bioluminescence because its deletion even increases specific activity and efficiency of apo-mitrocomin conversion into active photoprotein, in contrast to C-terminal Pro of other photoproteins. Since genes in a population generally exist as different isoforms, it makes us anticipate the cloning of even more isoforms of mitrocomin and other hydromedusan photoproteins with different bioluminescence properties.

  17. Attenuated bioluminescent Brucella melitensis mutants GR019 (virB4), GR024 (galE), and GR026 (BMEI1090-BMEI1091) confer protection in mice.

    Science.gov (United States)

    Rajashekara, Gireesh; Glover, David A; Banai, Menachem; O'Callaghan, David; Splitter, Gary A

    2006-05-01

    In vivo bioluminescence imaging is a persuasive approach to investigate a number of issues in microbial pathogenesis. Previously, we have applied bioluminescence imaging to gain greater insight into Brucella melitensis pathogenesis. Endowing Brucella with bioluminescence allowed direct visualization of bacterial dissemination, pattern of tissue localization, and the contribution of Brucella genes to virulence. In this report, we describe the pathogenicity of three attenuated bioluminescent B. melitensis mutants, GR019 (virB4), GR024 (galE), and GR026 (BMEI1090-BMEI1091), and the dynamics of bioluminescent virulent bacterial infection following vaccination with these mutants. The virB4, galE, and BMEI1090-BMEI1091 mutants were attenuated in interferon regulatory factor 1-deficient (IRF-1(-/-)) mice; however, only the GR019 (virB4) mutant was attenuated in cultured macrophages. Therefore, in vivo imaging provides a comprehensive approach to identify virulence genes that are relevant to in vivo pathogenesis. Our results provide greater insights into the role of galE in virulence and also suggest that BMEI1090 and downstream genes constitute a novel set of genes involved in Brucella virulence. Survival of the vaccine strain in the host for a critical period is important for effective Brucella vaccines. The galE mutant induced no changes in liver and spleen but localized chronically in the tail and protected IRF-1(-/-) and wild-type mice from virulent challenge, implying that this mutant may serve as a potential vaccine candidate in future studies and that the direct visualization of Brucella may provide insight into selection of improved vaccine candidates.

  18. The cAMP-dependent protein kinase inhibitor H-89 attenuates the bioluminescence signal produced by Renilla Luciferase.

    Directory of Open Access Journals (Sweden)

    Katie J Herbst

    2009-05-01

    Full Text Available Investigations into the regulation and functional roles of kinases such as cAMP-dependent protein kinase (PKA increasingly rely on cellular assays. Currently, there are a number of bioluminescence-based assays, for example reporter gene assays, that allow the study of the regulation, activity, and functional effects of PKA in the cellular context. Additionally there are continuing efforts to engineer improved biosensors that are capable of detecting real-time PKA signaling dynamics in cells. These cell-based assays are often utilized to test the involvement of PKA-dependent processes by using H-89, a reversible competitive inhibitor of PKA.We present here data to show that H-89, in addition to being a competitive PKA inhibitor, attenuates the bioluminescence signal produced by Renilla luciferase (RLuc variants in a population of cells and also in single cells. Using 10 microM of luciferase substrate and 10 microM H-89, we observed that the signal from RLuc and RLuc8, an eight-point mutation variant of RLuc, in cells was reduced to 50% (+/-15% and 54% (+/-14% of controls exposed to the vehicle alone, respectively. In vitro, we showed that H-89 decreased the RLuc8 bioluminescence signal but did not compete with coelenterazine-h for the RLuc8 active site, and also did not affect the activity of Firefly luciferase. By contrast, another competitive inhibitor of PKA, KT5720, did not affect the activity of RLuc8.The identification and characterization of the adverse effect of H-89 on RLuc signal will help deconvolute data previously generated from RLuc-based assays looking at the functional effects of PKA signaling. In addition, for the current application and future development of bioluminscence assays, KT5720 is identified as a more suitable PKA inhibitor to be used in conjunction with RLuc-based assays. These principal findings also provide an important lesson to fully consider all of the potential effects of experimental conditions on a cell

  19. LuxCDABE—Transformed Constitutively Bioluminescent Escherichia coli for Toxicity Screening: Comparison with Naturally Luminous Vibrio fischeri

    Directory of Open Access Journals (Sweden)

    Anne Kahru

    2011-08-01

    Full Text Available We show that in vitro toxicity assay based on inhibition of the bioluminescence of recombinant Escherichia coli encoding thermostable luciferase from Photorhabdus luminescens is a versatile alternative to Vibrio fischeri MicrotoxTM test. Performance of two luxCDABE-transformed E. coli MC1061 constructs (pDNlux and (pSLlux otherwise identical, but having 100-fold different background luminescence was compared with the performance of V. fischeri. The microplate luminometer and a kinetic Flash-Assay test format was used that differently from Microtox test is also applicable for high throughput analysis. Toxic effects (30-s till 30-min EC50 of four heavy metals (Zn, Cd, Hg, Cu and three organic chemicals (aniline, 3,5-dichloroaniline and 3,5-dichlorophenol were studied. Both E. coli strains had comparable sensitivity and the respective 30-min EC50 values highly correlated (log-log R2 = 0.99; p < 0.01 showing that the sensitivity of the recombinant bacteria towards chemicals analyzed did not depend on the bioluminescence level of the recombinant cells. The most toxic chemical for all used bacterial strains (E. coli, V. fischeri was mercury whereas the lowest EC50 values for Hg (0.04–0.05 mg/L and highest EC50 values for aniline (1,300–1,700 mg/L were observed for E. coli strains. Despite of that, toxicity results obtained with both E. coli strains (pSLlux and pDNlux significantly correlated with V. fischeri results (log-log R2 = 0.70/0.75; p < 0.05/0.01. The use of amino acids (0.25% and glucose (0.05%-supplemented M9 medium instead of leucine-supplemented saline significantly (p < 0.05 reduced the apparent toxicity of heavy metals to both E. coli strains up to three orders of magnitude, but had little or no complexing effect on organic compounds. Thus, P. luminescens luxCDABE-transformed E. coli strains can be successfully used for the acute toxicity screening of various types of organic chemicals and heavy metals and can replace V. fischeri in

  20. How do marine bacteria produce light, why are they luminescent, and can we employ bacterial bioluminescence in aquatic biotechnology?

    Directory of Open Access Journals (Sweden)

    Grzegorz Wêgrzyn

    2002-09-01

    Full Text Available Bioluminescence, the phenomenon of light production by living organisms, occurs in forms of life as various as bacteria, fungi and animals. Nevertheless, light-emitting bacteria are the most abundant and widespread of luminescent organisms. Interestingly, most species of such bacteria live in marine environments. In this article, the biochemical mechanism of bacterial luminescence and its genetic regulation are summarized. Although the biochemistry and genetics of light emission by cells have been investigated in detail, the biological role of bacterial luminescence has remained obscure. Here, we discuss recent discoveries that shed new light on this problem. Finally, we provide examples of how bacterial luminescence can be employed in marine biotechnology, especially in the detection of toxic and mutagenic pollution in aquatic environments.

  1. Promising Biological Indicator of Heavy Metal Pollution: Bioluminescent Bacterial Strains Isolated and Characterized from Marine Niches of Goa, India.

    Science.gov (United States)

    Thakre, Neha A; Shanware, Arti S

    2015-09-01

    In present study, several marine water samples collected from the North Goa Beaches, India for isolation of luminescent bacterial species. Isolates obtained labelled as DP1-5 and AB1-6. Molecular characterization including identification of a microbial culture using 16S rRNA gene based molecular technique and phylogenetic analysis confirmed that DP3 & AB1 isolates were Vibrio harveyi. All of the isolates demonstrated multiple metal resistances in terms of growth, with altered luminescence with variable metal concentration. Present investigations were an attempt towards exploring and reporting an updated diversity of bioluminescent bacterial species from various sites around the Goa, India which would be explored in future for constructing luminescence based biosensor for efficiently monitoring the level of hazardous metals in the environment.

  2. Probing intermolecular protein-protein interactions in the calcium-sensing receptor homodimer using bioluminescence resonance energy transfer (BRET)

    DEFF Research Database (Denmark)

    Jensen, Anders A.; Hansen, Jakob L; Sheikh, Søren P

    2002-01-01

    The calcium-sensing receptor (CaR) belongs to family C of the G-protein coupled receptor superfamily. The receptor is believed to exist as a homodimer due to covalent and non-covalent interactions between the two amino terminal domains (ATDs). It is well established that agonist binding to family C...... receptors takes place at the ATD and that this causes the ATD dimer to twist. However, very little is known about the translation of the ATD dimer twist into G-protein coupling to the 7 transmembrane moieties (7TMs) of these receptor dimers. In this study we have attempted to delineate the agonist......-induced intermolecular movements in the CaR homodimer using the new bioluminescence resonance energy transfer technique, BRET2, which is based on the transference of energy from Renilla luciferase (Rluc) to the green fluorescent protein mutant GFP2. We tagged CaR with Rluc and GFP2 at different intracellular locations...

  3. Avaliação da qualidade microbiológica de bebida láctea e creme de leite UAT por ATP-Bioluminescência Evaluation of microbiological quality of UHT milk drink and UHT milk cream by ATP-Bioluminescence

    Directory of Open Access Journals (Sweden)

    A.F. Cunha

    2013-04-01

    Full Text Available Embora métodos tradicionais sejam utilizados na avaliação microbiológica de produtos UAT, metodologias rápidas, baseadas em ATP-Bioluminescência, têm sido desenvolvidas. Os resultados da aplicação dessa técnica em 54 amostras de bebida láctea UAT achocolatada e 12 de creme de leite UAT foram comparados com os resultados de métodos microbiológicos, utilizando-se diferentes meios de cultura e tempos de incubação das referidas amostras. A técnica de ATP-Bioluminescência foi aplicada por meio do sistema MLS, e os resultados foram expressos em unidades relativas de luz (RLU. Em todos os tempos de incubação - 48, 72 e 168 horas - , as amostras apresentaram contagens baixas de microrganismos mesófilos e psicrotróficos aeróbios quando analisadas em meio PCA, BHI, PetrifilmTM AC e por ATP-Bioluminescência (Although traditional methods are used for the microbiological evaluation of UHT products, rapid methodologies based on ATP-Bioluminescence have been developed. The results of applying this technique in 54 samples of chocolate UHT milk drink and 12 of UHT milk cream were compared with the results of microbiological methods, using different culture media and incubation times for the referred samples. The ATP-Bioluminescence technique was applied through the MLS system and the results were expressed as relative light units (RLU. In all incubation times - 48, 72, and 168 hours - , the samples showed lower counts of mesophilic and psychrotrophic aerobic microorganisms when analyzed using PCA, BHI, PetrifilmTM AC and ATP-Bioluminescence (<150RLU, demonstrating the technique's high specificity. Only one sample of UHT milk cream showed a mesophilic aerobic count above the standard established by Brazilian legislation (<100CFU/mL when analyzed in PCA (260 CFU/mL and PetrifilmTM AC (108CFU/mL at 168 hours. This high count of aerobic mesophilic microorganisms was also detected by the ATP-Bioluminescence (416 RLU technique. The results of

  4. Quantitative imaging of D-2-hydroxyglutarate (D2HG in selected histological tissue areas by a novel bioluminescence technique

    Directory of Open Access Journals (Sweden)

    Nadine Fabienne Voelxen

    2016-03-01

    Full Text Available AbstractPatients with malignant gliomas have a poor prognosis with average survival of less than one year. Whereas in other tumor entities the characteristics of tumor metabolism are successfully used for therapeutic approaches, such developments are very rare in brain tumors, notably in gliomas. One metabolic feature characteristic of gliomas, in particular diffuse astrocytomas and oligodendroglial tumors, is the variable content of D-2-hydroxyglutarate (D2HG, a metabolite, which was discovered first in this tumor entity. D2HG is generated in large amounts due to various gain-of–function mutations in the isocitrate dehydrogenases IDH-1 and IDH-2. Meanwhile, D2HG has been detected in several other tumor entities including intrahepatic bile-duct cancer, chondrosarcoma, acute myeloid leukemia, and angioimmunoblastic T-cell lymphoma. D2HG is barely detectable in healthy tissue (< 0.1 mM, but its concentration increases up to 35 mM in malignant tumor tissues. Consequently, the oncometabolite D2HG has gained increasing interest in the field of tumor metabolism. To facilitate its quantitative measurement without loss of spatial resolution at a microscopical level, we have developed a novel bioluminescence assay for determining D2HG in sections of snap-frozen tissue. The assay was verified independently by photometric tests and liquid chromatography / mass spectrometry (LC/MS. The novel technique allows the microscopically resolved determination of D2HG in a concentration range of 0 – 10 µmol/g tissue (wet weight. In combination with the already established bioluminescence imaging techniques for ATP, glucose, pyruvate, and lactate, the novel D2HG assay enables a comparative characterization of the metabolic profile of individual tumors in a further dimension.

  5. SU-E-T-20: A Novel Hybrid CBCT, Bioluminescence and Fluorescence Tomography System for Preclinical Radiation Research

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, B; Eslami, S; Iordachita, I [Johns Hopkins University, Baltimore, Maryland (United States); Yang, Y [University of Miami School of Medicine, Miami, FL (United States); Patterson, M [Hamilton Regional Cancer Ctr., Hamilton, ON (Canada); Wong, J [Johns Hopkins University, Baltimore, MD (United States); Wang, K [Johns Hopkins Hospital, Baltimore, MD (United States)

    2014-06-01

    Purpose: A novel standalone bioluminescence and fluorescence tomography (BLT and FT) system equipped with high resolution CBCT has been built in our group. In this work, we present the system calibration method and validate our system in both phantom and in vivo environment. Methods: The CBCT is acquired by rotating the animal stage while keeping the x-ray source and detector panel static. The optical signal is reflected by the 3-mirror system to a multispectral filter set and then delivered to the CCD camera with f/1.4 lens mounted. Nine fibers passing through the stage and in contact with the mouse skin serve as the light sources for diffuse optical tomography (DOT) and FT. The anatomical information and optical properties acquired from the CBCT and DOT, respectively, are used as the priori information to improve the BLT/FT reconstruction accuracy. Flat field correction for the optical system was acquired at multiple wavelengths. A home-built phantom is used to register the optical and CBCT coordinates. An absolute calibration relating the CCD photon counts rate to the light fluence rate emitted at animal surface was developed to quantify the bioluminescence power or fluorophore concentration. Results: An optical inhomogeneous phantom with 2 light sources (3mm separation) imbedded is used to test the system. The optical signal is mapped onto the mesh generated from CBCT for optical reconstruction. Our preliminary results show that the center of mass can be reconstructed within 2.8mm accuracy. A live mouse with the light source imbedded is also used to validate our system. Liver or lung metastatic luminescence tumor model will be used for further testing. Conclusion: This hybrid system transforms preclinical research to a level that even sub-palpable volume of cells can be imaged rapidly and non-invasively, which largely extends the scope of radiobiological research. The research is supported by the NCI grant R01CA158100-01.

  6. Hybrid radiosity-SP{sub 3} equation based bioluminescence tomography reconstruction for turbid medium with low- and non-scattering regions

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xueli, E-mail: xlchen@xidian.edu.cn, E-mail: jimleung@mail.xidian.edu.cn; Zhang, Qitan; Yang, Defu; Liang, Jimin, E-mail: xlchen@xidian.edu.cn, E-mail: jimleung@mail.xidian.edu.cn [School of Life Science and Technology, Xidian University, Xi' an, Shaanxi 710071 (China)

    2014-01-14

    To provide an ideal solution for a specific problem of gastric cancer detection in which low-scattering regions simultaneously existed with both the non- and high-scattering regions, a novel hybrid radiosity-SP{sub 3} equation based reconstruction algorithm for bioluminescence tomography was proposed in this paper. In the algorithm, the third-order simplified spherical harmonics approximation (SP{sub 3}) was combined with the radiosity equation to describe the bioluminescent light propagation in tissues, which provided acceptable accuracy for the turbid medium with both low- and non-scattering regions. The performance of the algorithm was evaluated with digital mouse based simulations and a gastric cancer-bearing mouse based in situ experiment. Primary results demonstrated the feasibility and superiority of the proposed algorithm for the turbid medium with low- and non-scattering regions.

  7. A multi-channel bioluminescent bacterial biosensor for the on-line detection of metals and toxicity. Part II: technical development and proof of concept of the biosensor

    Energy Technology Data Exchange (ETDEWEB)

    Charrier, Thomas; Thouand, Gerald [UMR CNRS 6144 GEPEA, CBAC, Nantes University, PRES UNAM, Campus de la Courtaisiere-IUT, La Roche-sur-Yon cedex (France); Chapeau, Cyrille [Biolumine, Biokar Diagnostic, Rue des Quarante Mines ZAC de Ther-Allonne, Beauvais Cedex (France); Bendria, Loubna; Daniel, Philippe [UMR CNRS 6087 LPEC, Universite du Maine, Av Olivier Messiaen, Le Mans cedex 9 (France); Picart, Pascal [UMR CNRS 6613 IAM-LAUM, Ecole Nationale des Ingenieurs du Mans, Universite du Maine, Le Mans Cedex 9 (France)

    2011-05-15

    This research study deals with the on-line detection of heavy metals and toxicity within the context of environmental pollution monitoring. It describes the construction and the proof of concept of a multi-channel bioluminescent bacterial biosensor in immobilized phase: Lumisens3. This new versatile device, designed for the non-stop analysis of water pollution, enables the insertion of any bioluminescent strains (inducible or constitutive), immobilized in a multi-well removable card. The technical design of Lumisens3 has benefited from both a classical and a robust approach and includes four main parts: (1) a dedicated removable card contains 64 wells, 3 mm in depth, arranged in eight grooves within which bacteria are immobilized, (2) this card is incubated on a Pelletier block with a CCD cooled camera on top for bioluminescence monitoring, (3) a fluidic network feeds the card with the sample to be analyzed and finally (4) a dedicated computer interface, BIOLUX 1.0, controls all the elements of the biosensor, allowing it to operate autonomously. The proof of concept of this biosensor was performed using a set of four bioluminescent bacteria (Escherichia coli DH1 pBzntlux, pBarslux, pBcoplux, and E. coli XL1 pBfiluxCDABE) in the on-line detection of CdCl{sub 2} 0.5 {mu}M and As{sub 2}O{sub 3} 5 {mu}M from an influent. When considering metals individually, the ''fingerprints'' from the biosensor were as expected. However, when metals were mixed together, cross reaction and synergistic effects were detected. This biosensor allowed us to demonstrate the simultaneous on-line cross detection of one or several heavy metals as well as the measurement of the overall toxicity of the sample. (orig.)

  8. Development of a Novel Preclinical Pancreatic Cancer Research Model: Bioluminescence Image-Guided Focal Irradiation and Tumor Monitoring of Orthotopic Xenografts1

    OpenAIRE

    Tuli, Richard; Surmak, Andrew; Reyes, Juvenal; Hacker-Prietz, Amy; Armour, Michael; Leubner, Ashley; Blackford, Amanda; Tryggestad, Erik; Jaffee, Elizabeth M; Wong, John; DeWeese, Theodore L; Herman, Joseph M

    2012-01-01

    PURPOSE: We report on a novel preclinical pancreatic cancer research model that uses bioluminescence imaging (BLI)-guided irradiation of orthotopic xenograft tumors, sparing of surrounding normal tissues, and quantitative, noninvasive longitudinal assessment of treatment response. MATERIALS AND METHODS: Luciferase-expressing MiaPaCa-2 pancreatic carcinoma cells were orthotopically injected in nude mice. BLI was compared to pathologic tumor volume, and photon emission was assessed over time. B...

  9. In vivo visualization and monitoring of viable neural stem cells using noninvasive bioluminescence imaging in the 6-hydroxydopamine-induced mouse model of Parkinson disease.

    Science.gov (United States)

    Im, Hyung-Jun; Hwang, Do Won; Lee, Han Kyu; Jang, Jaeho; Lee, Song; Youn, Hyewon; Jin, Yeona; Kim, Seung U; Kim, E Edmund; Kim, Yong Sik; Lee, Dong Soo

    2013-06-01

    Transplantation of neural stem cells (NSCs) has been proposed as a treatment for Parkinson disease (PD). The aim of this study was to monitor the viability of transplanted NSCs expressing the enhanced luciferase gene in a mouse model of PD in vivo. The PD animal model was induced by unilateral injection of 6-hydroxydopamine (6-OHDA). The behavioral test using apomorphine-induced rotation and positron emission tomography with [18F]N-(3-fluoropropyl)-2'-carbomethoxy-3'-(4-iodophenyl)nortropane ([18F]FP-CIT) were conducted. HB1.F3 cells transduced with an enhanced firefly luciferase retroviral vector (F3-effLuc cells) were transplanted into the right striatum. In vivo bioluminescence imaging was repeated for 2 weeks. Four weeks after transplantation, [18F]FP-CIT PET and the rotation test were repeated. All 6-OHDA-injected mice showed markedly decreased [18F]FP-CIT uptake in the right striatum. Transplanted F3-effLuc cells were visualized on the right side of the brain in all mice by bioluminescence imaging. The bioluminescence intensity of the transplanted F3-effLuc cells gradually decreased until it was undetectable by 10 days. The behavioral test showed that stem cell transplantation attenuated the motor symptoms of PD. No significant change was found in [18F]FP-CIT imaging after cell transplantation. We successfully established an in vivo bioluminescence imaging system for the detection of transplanted NSCs in a mouse model of PD. NSC transplantation induced behavioral improvement in PD model mice.

  10. Effect of concentrating and exposing the bioluminescent bacteria to the non-luminescent allo-bacterial extracellular products on their luminescence

    Digital Repository Service at National Institute of Oceanography (India)

    Ravindran, J.; Priya, G.G.; Kannapiran, E.

    and the other bacteria with the help of selective receptors sense the same (2). The low molecular weight molecules secreted are called autoinducer that act as “Molecular Messengers” (3). The auto inducers increase with increase in bacterial population... of bioluminescent bacteria causes significant economic loss. It cannot be predicted when they will appear. The present solution is frequent exchange of water to dilute the risk. It is not known the kind response of the luminescent bacteria in presence of various...

  11. Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications

    Science.gov (United States)

    Herfst, Sander; Bestebroer, Theo M.; Vaes, Vincent P.; van der Hoeven, Barbara; Koster, Abraham J.; Kremers, Gert-Jan; Scott, Dana P.; Gultyaev, Alexander P.; Sorell, Erin M.; de Graaf, Miranda; Bárcena, Montserrat; Rimmelzwaan, Guus F.; Fouchier, Ron A.

    2015-01-01

    Bioluminescent and fluorescent influenza A viruses offer new opportunities to study influenza virus replication, tropism and pathogenesis. To date, several influenza A reporter viruses have been described. These strategies typically focused on a single reporter gene (either bioluminescent or fluorescent) in a single virus backbone. However, whilst bioluminescence is suited to in vivo imaging, fluorescent viruses are more appropriate for microscopy. Therefore, the idea l reporter virus varies depending on the experiment in question, and it is important that any reporter virus strategy can be adapted accordingly. Herein, a strategy was developed to create five different reporter viruses in a single virus backbone. Specifically, enhanced green fluorescent protein (eGFP), far-red fluorescent protein (fRFP), near-infrared fluorescent protein (iRFP), Gaussia luciferase (gLUC) and firefly luciferase (fLUC) were inserted into the PA gene segment of A/PR/8/34 (H1N1). This study provides a comprehensive characterisation of the effects of different reporter genes on influenza virus replication and reporter activity. In vivo reporter gene expression, in lung tissues, was only detected for eGFP, fRFP and gLUC expressing viruses. In vitro, the eGFP-expressing virus displayed the best reporter stability and could be used for correlative light electron microscopy (CLEM). This strategy was then used to create eGFP-expressing viruses consisting entirely of pandemic H1N1, highly pathogenic avian influenza (HPAI) H5N1 and H7N9. The HPAI H5N1 eGFP-expressing virus infected mice and reporter gene expression was detected, in lung tissues, in vivo. Thus, this study provides new tools and insights for the creation of bioluminescent and fluorescent influenza A reporter viruses. PMID:26241861

  12. Effects of Photodynamic Therapy on Gram-Positive and Gram-Negative Bacterial Biofilms by Bioluminescence Imaging and Scanning Electron Microscopic Analysis

    Science.gov (United States)

    Núñez, Silvia C.; Azambuja, Nilton; Fregnani, Eduardo R.; Rodriguez, Helena M.H.; Hamblin, Michael R.; Suzuki, Hideo; Ribeiro, Martha S.

    2013-01-01

    Abstract Objective: The aim of this study was to test photodynamic therapy (PDT) as an alternative approach to biofilm disruption on dental hard tissue, We evaluated the effect of methylene blue and a 660 nm diode laser on the viability and architecture of Gram-positive and Gram-negative bacterial biofilms. Materials and methods: Ten human teeth were inoculated with bioluminescent Pseudomonas aeruginosa or Enterococcus faecalis to form 3 day biofilms in prepared root canals. Bioluminescence imaging was used to serially quantify and evaluate the bacterial viability, and scanning electron microscopic (SEM) imaging was used to assess architecture and morphology of bacterial biofilm before and after PDT employing methylene blue and 40 mW, 660 nm diode laser light delivered into the root canal via a 300 μm fiber for 240 sec, resulting in a total energy of 9.6 J. The data were statistically analyzed with analysis of variance (ANOVA) followed by Tukey test. Results: The bacterial reduction showed a dose dependence; as the light energy increased, the bioluminescence decreased in both planktonic suspension and in biofilms. The SEM analysis showed a significant reduction of biofilm on the surface. PDT promoted disruption of the biofilm and the number of adherent bacteria was reduced. Conclusions: The photodynamic effect seems to disrupt the biofilm by acting both on bacterial cells and on the extracellular matrix. PMID:23822168

  13. An endogenous green fluorescent protein-photoprotein pair in Clytia hemisphaerica eggs shows co-targeting to mitochondria and efficient bioluminescence energy transfer.

    Science.gov (United States)

    Fourrage, Cécile; Swann, Karl; Gonzalez Garcia, Jose Raul; Campbell, Anthony K; Houliston, Evelyn

    2014-04-09

    Green fluorescent proteins (GFPs) and calcium-activated photoproteins of the aequorin/clytin family, now widely used as research tools, were originally isolated from the hydrozoan jellyfish Aequora victoria. It is known that bioluminescence resonance energy transfer (BRET) is possible between these proteins to generate flashes of green light, but the native function and significance of this phenomenon is unclear. Using the hydrozoan Clytia hemisphaerica, we characterized differential expression of three clytin and four GFP genes in distinct tissues at larva, medusa and polyp stages, corresponding to the major in vivo sites of bioluminescence (medusa tentacles and eggs) and fluorescence (these sites plus medusa manubrium, gonad and larval ectoderms). Potential physiological functions at these sites include UV protection of stem cells for fluorescence alone, and prey attraction and camouflaging counter-illumination for bioluminescence. Remarkably, the clytin2 and GFP2 proteins, co-expressed in eggs, show particularly efficient BRET and co-localize to mitochondria, owing to parallel acquisition by the two genes of mitochondrial targeting sequences during hydrozoan evolution. Overall, our results indicate that endogenous GFPs and photoproteins can play diverse roles even within one species and provide a striking and novel example of protein coevolution, which could have facilitated efficient or brighter BRET flashes through mitochondrial compartmentalization.

  14. In Vivo Imaging with Bioluminescent Enterovirus 71 Allows for Real-Time Visualization of Tissue Tropism and Viral Spread.

    Science.gov (United States)

    Caine, Elizabeth A; Osorio, Jorge E

    2017-03-01

    Hand, foot, and mouth disease (HFMD) is a reemerging illness caused by a variety of enteroviruses. The main causative agents are enterovirus 71 (EV71), coxsackievirus A16 (CVA16), and, most recently, coxsackievirus A6 (CVA6). Enterovirus infections can vary from asymptomatic infections to those with a mild fever and blisters on infected individuals' hands, feet, and throats to infections with severe neurological complications. Viral persistence for weeks postinfection (wpi) has also been documented by the demonstration of virus in children's stools. However, little is known about disease progression, viral spread, and tissue tropism of these viruses. These types of studies are limited because many recently developed mouse models mimic the severe neurological complications that occur in a small percentage of enterovirus infections. In the present study, we documented real-time EV71 infection in two different mouse strains by the use of in vivo imaging. Infection of BALB/c mice with a bioluminescent mouse-adapted EV71 construct (mEV71-NLuc) resulted in a lack of clinical signs of disease but in relatively high viral replication, as visualized by luminescence, for 2 wpi. In contrast, mEV71-NLuc infection of AG129 mice (alpha/beta and gamma interferon receptor deficient) showed rapid spread and long-term persistence of the virus in the brain. Interestingly, AG129 mice that survived infection maintained luminescence in the brain for up to 8 wpi. The results we present here will allow future studies on EV71 antiviral drug susceptibility, vaccine efficacy, transmissibility, and pathogenesis. IMPORTANCE We report here that a stable full-length enterovirus 71 (EV71) reporter construct was used to visualize real-time viral spread in AG129 and BALB/c mice. To our knowledge, this is the first report of in vivo imaging of infection with any member of the Picornaviridae family. The nanoluciferase (NLuc) gene, one of the smallest luciferase genes currently available, was shown to

  15. Bioluminescent human breast cancer cell lines that permit rapid and sensitive in vivo detection of mammary tumors and multiple metastases in immune deficient mice

    International Nuclear Information System (INIS)

    Jenkins, Darlene E; Hornig, Yvette S; Oei, Yoko; Dusich, Joan; Purchio, Tony

    2005-01-01

    Our goal was to generate xenograft mouse models of human breast cancer based on luciferase-expressing MDA-MB-231 tumor cells that would provide rapid mammary tumor growth; produce metastasis to clinically relevant tissues such as lymph nodes, lung, and bone; and permit sensitive in vivo detection of both primary and secondary tumor sites by bioluminescent imaging. Two clonal cell sublines of human MDA-MB-231 cells that stably expressed firefly luciferase were isolated following transfection of the parental cells with luciferase cDNA. Each subline was passaged once or twice in vivo to enhance primary tumor growth and to increase metastasis. The resulting luciferase-expressing D3H1 and D3H2LN cells were analyzed for long-term bioluminescent stability, primary tumor growth, and distal metastasis to lymph nodes, lungs, bone and soft tissues by bioluminescent imaging. Cells were injected into the mammary fat pad of nude and nude-beige mice or were delivered systemically via intracardiac injection. Metastasis was also evaluated by ex vivo imaging and histologic analysis postmortem. The D3H1 and D3H2LN cell lines exhibited long-term stable luciferase expression for up to 4–6 months of accumulative tumor growth time in vivo. Bioluminescent imaging quantified primary mammary fat pad tumor development and detected early spontaneous lymph node metastasis in vivo. Increased frequency of spontaneous lymph node metastasis was observed with D3H2LN tumors as compared with D3H1 tumors. With postmortem ex vivo imaging, we detected additional lung micrometastasis in mice with D3H2LN mammary tumors. Subsequent histologic evaluation of tissue sections from lymph nodes and lung lobes confirmed spontaneous tumor metastasis at these sites. Following intracardiac injection of the MDA-MB-231-luc tumor cells, early metastasis to skeletal tissues, lymph nodes, brain and various visceral organs was detected. Weekly in vivo imaging data permitted longitudinal analysis of metastasis at

  16. Following in real time the impact of pneumococcal virulence factors in an acute mouse pneumonia model using bioluminescent bacteria.

    Science.gov (United States)

    Saleh, Malek; Abdullah, Mohammed R; Schulz, Christian; Kohler, Thomas; Pribyl, Thomas; Jensch, Inga; Hammerschmidt, Sven

    2014-02-23

    Pneumonia is one of the major health care problems in developing and industrialized countries and is associated with considerable morbidity and mortality. Despite advances in knowledge of this illness, the availability of intensive care units (ICU), and the use of potent antimicrobial agents and effective vaccines, the mortality rates remain high(1). Streptococcus pneumoniae is the leading pathogen of community-acquired pneumonia (CAP) and one of the most common causes of bacteremia in humans. This pathogen is equipped with an armamentarium of surface-exposed adhesins and virulence factors contributing to pneumonia and invasive pneumococcal disease (IPD). The assessment of the in vivo role of bacterial fitness or virulence factors is of utmost importance to unravel S. pneumoniae pathogenicity mechanisms. Murine models of pneumonia, bacteremia, and meningitis are being used to determine the impact of pneumococcal factors at different stages of the infection. Here we describe a protocol to monitor in real-time pneumococcal dissemination in mice after intranasal or intraperitoneal infections with bioluminescent bacteria. The results show the multiplication and dissemination of pneumococci in the lower respiratory tract and blood, which can be visualized and evaluated using an imaging system and the accompanying analysis software.

  17. Quick bacteriophage-mediated bioluminescence assay for detecting Staphylococcus spp. in sonicate fluid of orthopaedic artificial joints.

    Science.gov (United States)

    Šuster, Katja; Podgornik, Aleš; Cör, Andrej

    2017-07-01

    Staphylococcus spp. accounts for up to two thirds of all microorganisms causing prosthetic joint infections, with Staphylococcus aureus and Staphylococcus epidermidis being the major cause. The present study describes a diagnostic model to detect staphylococci using a specific bacteriophage and bioluminescence detection, exploring the possibility of its use on sonicate fluid of orthopaedic artificial joints. Intracellular adenosine-5'-triphosphate release by bacteriophage mediated lysis of staphylococci was assessed to determine optimal parameters for detection. With the optimized method, a limit of detection of around 103 CFU/mL was obtained after incubation with bacteriophage for 2 h. Importantly, sonicate fluid did not prevent the ability of bacteriophage to infect bacteria and all simulated infected sonicate fluid as well as 6 clinical samples with microbiologically proven staphylococcal infection were detected as positive. The total assay took approximately 4 h. Collectively, the results indicate that the developed method promises a rapid, inexpensive and specific diagnostic detection of staphylococci in sonicate fluid of infected prosthetic joints. In addition, the unlimited pool of different existing bacteriophages, with different specificity for all kind of bacteria gives the opportunity for further investigations, improvements of the current model and implementation in other medical fields for the purpose of the establishment of a rapid diagnosis.

  18. Dual Monitoring of Secretion and ATP Levels during Chondrogenesis Using Perfusion Culture-Combined Bioluminescence Monitoring System

    Directory of Open Access Journals (Sweden)

    Hyuck Joon Kwon

    2015-01-01

    Full Text Available Skeletal pattern formation in limb development depends on prechondrogenic condensation which prefigures the cartilage template. However, although morphogens such as TGF-βs and BMPs have been known to play essential roles in skeletal patterning, how the morphogens induce prechondrogenic cells to aggregate and determine patterns of cartilage elements has remained unclear. Our previous study reported that ATP oscillations are induced during chondrogenesis. This result suggests the possibility that ATP oscillations lead to the oscillatory secretion of morphogens, due to the fact that secretion process requires ATP. To examine the correlation between ATP oscillations and secretion levels of morphogens, we have developed perfusion culture-combined bioluminescence monitoring system to simultaneously monitor intracellular ATP levels and secretion levels. Using this system, we found that secretory activity oscillates in phase with ATP oscillations and that secretion levels of TGF-β1 and BMP2 oscillate during chondrogenesis. The oscillatory secretion of the morphogens would contribute to amplifying the fluctuation of the morphogens, underlie the spatial patterning of morphogens, and consequently lead to skeletal pattern formation.

  19. Susceptibility of the wild-derived inbred CAST/Ei mouse to infection by orthopoxviruses analyzed by live bioluminescence imaging

    Energy Technology Data Exchange (ETDEWEB)

    Americo, Jeffrey L.; Sood, Cindy L.; Cotter, Catherine A.; Vogel, Jodi L.; Kristie, Thomas M.; Moss, Bernard, E-mail: bmoss@nih.gov; Earl, Patricia L., E-mail: pearl@nih.gov

    2014-01-20

    Classical inbred mice are extensively used for virus research. However, we recently found that some wild-derived inbred mouse strains are more susceptible than classical strains to monkeypox virus. Experiments described here indicated that the 50% lethal dose of vaccinia virus (VACV) and cowpox virus (CPXV) were two logs lower in wild-derived inbred CAST/Ei mice than classical inbred BALB/c mice, whereas there was little difference in the susceptibility of the mouse strains to herpes simplex virus. Live bioluminescence imaging was used to follow spread of pathogenic and attenuated VACV strains and CPXV virus from nasal passages to organs in the chest and abdomen of CAST/Ei mice. Luminescence increased first in the head and then simultaneously in the chest and abdomen in a dose-dependent manner. The spreading kinetics was more rapid with VACV than CPXV although the peak photon flux was similar. These data suggest advantages of CAST/Ei mice for orthopoxvirus studies. - Highlights: • Wild-derived inbred CAST/Ei mice are susceptible to vaccinia virus and cowpox virus. • Morbidity and mortality from orthopoxviruses are greater in CAST/Ei than BALB/c mice. • Morbidity and mortality from herpes simplex virus type 1 are similar in both mice. • Imaging shows virus spread from nose to lungs, abdominal organs and brain. • Vaccinia virus spreads more rapidly than cowpox virus.

  20. Evaluation of bioluminescent imaging for noninvasive monitoring of colorectal cancer progression in the liver and its response to immunogene therapy

    Directory of Open Access Journals (Sweden)

    Gonzalez-Aparicio Manuela

    2009-01-01

    Full Text Available Abstract Background Bioluminescent imaging (BLI is based on the detection of light emitted by living cells expressing a luciferase gene. Stable transfection of luciferase in cancer cells and their inoculation into permissive animals allows the noninvasive monitorization of tumor progression inside internal organs. We have applied this technology for the development of a murine model of colorectal cancer involving the liver, with the aim of improving the pre-clinical evaluation of new anticancer therapies. Results A murine colon cancer cell line stably transfected with the luciferase gene (MC38Luc1 retains tumorigenicity in immunocompetent C57BL/6 animals. Intrahepatic inoculation of MC38Luc1 causes progressive liver infiltration that can be monitored by BLI. Compared with ultrasonography (US, BLI is more sensitive, but accurate estimation of tumor mass is impaired in advanced stages. We applied BLI to evaluate the efficacy of an immunogene therapy approach based on the liver-specific expression of the proinflammatory cytokine interleukin-12 (IL-12. Individualized quantification of light emission was able to determine the extent and duration of antitumor responses and to predict long-term disease-free survival. Conclusion We show that BLI is a rapid, convenient and safe technique for the individual monitorization of tumor progression in the liver. Evaluation of experimental treatments with complex mechanisms of action such as immunotherapy is possible using this technology.

  1. Subject-Author Index 989..1009

    Indian Academy of Sciences (India)

    Between molecular biology and molecular cell biology. 225. Bacterial identification. Development of rapid phenotypic system for the identification of Gram-negative oxidase-positive bacilli in resource-limited settings. 329. Bacterial sensor. Bioluminescent bioreporter for assessment of arsenic contamination in water samples.

  2. Journal of Biosciences | Indian Academy of Sciences

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Biosciences. Shahzada Asad. Articles written in Journal of Biosciences. Volume 38 Issue 2 June 2013 pp 251-258 Articles. Bioluminescent bioreporter for assessment of arsenic contamination in water samples of India · Pratima Sharma Shahzada Asad Arif Ali · More Details Abstract Fulltext PDF.

  3. Whole-Cell Optical Biosensor for Mercury – Operational Conditions in Saline Water

    Czech Academy of Sciences Publication Activity Database

    Solovyev, Andrey; Kuncová, Gabriela; Demnerová, K.

    2015-01-01

    Roč. 69, č. 1 (2015), s. 183-191 ISSN 0366-6352 R&D Projects: GA TA ČR TA03010544 Institutional support: RVO:67985858 Keywords : mercury detection assay * bioluminescent bioreporter * sea water Subject RIV: CA - Inorganic Chemistry Impact factor: 1.326, year: 2015

  4. Further assessment of Monkeypox Virus infection in Gambian pouched rats (Cricetomys gambianus) using in vivo bioluminescent imaging

    Science.gov (United States)

    Falendysz, Elizabeth; Lopera, Juan G.; Faye Lorenzsonn,; Salzer, Johanna S.; Hutson, Christina L.; Doty, Jeffrey; Gallardo-Romero, Nadia; Carroll, Darin S.; Osorio, Jorge E.; Rocke, Tonie E.

    2015-01-01

    Monkeypox is a zoonosis clinically similar to smallpox in humans. Recent evidence has shown a potential risk of increased incidence in central Africa. Despite attempts to isolate the virus from wild rodents and other small mammals, no reservoir host has been identified. In 2003,Monkeypox virus (MPXV) was accidentally introduced into the U.S. via the pet trade and was associated with the Gambian pouched rat (Cricetomys gambianus). Therefore, we investigated the potential reservoir competence of the Gambian pouched rat for MPXV by utilizing a combination of in vivo and in vitro methods. We inoculated three animals by the intradermal route and three animals by the intranasal route, with one mock-infected control for each route. Bioluminescent imaging (BLI) was used to track replicating virus in infected animals and virological assays (e.g. real time PCR, cell culture) were used to determine viral load in blood, urine, ocular, nasal, oral, and rectal swabs. Intradermal inoculation resulted in clinical signs of monkeypox infection in two of three animals. One severely ill animal was euthanized and the other affected animal recovered. In contrast, intranasal inoculation resulted in subclinical infection in all three animals. All animals, regardless of apparent or inapparent infection, shed virus in oral and nasal secretions. Additionally, BLI identified viral replication in the skin without grossly visible lesions. These results suggest that Gambian pouched rats may play an important role in transmission of the virus to humans, as they are hunted for consumption and it is possible for MPXV-infected pouched rats to shed infectious virus without displaying overt clinical signs.

  5. Effect of enhanced Renilla luciferase and fluorescent protein variants on the Foerster distance of Bioluminescence resonance energy transfer (BRET)

    Energy Technology Data Exchange (ETDEWEB)

    Dacres, Helen, E-mail: helen.dacres@csiro.au [CSIRO Food Futures Flagship and Ecosystem Sciences, Canberra (Australia); Michie, Michelle; Wang, Jian [CSIRO Food Futures Flagship and Ecosystem Sciences, Canberra (Australia); Pfleger, Kevin D.G. [Laboratory for Molecular Endocrinology-GPCRs, Western Australian Institute for Medical Research (WAIMR) and Centre for Medical Research, The University of Western Australia, Perth (Australia); Trowell, Stephen C. [CSIRO Food Futures Flagship and Ecosystem Sciences, Canberra (Australia)

    2012-08-31

    Highlights: Black-Right-Pointing-Pointer First experimental determination of Foerster distance (R{sub 0}) for enhanced BRET systems. Black-Right-Pointing-Pointer Effect of brighter BRET components RLuc2, RLuc8 and Venus was assessed. Black-Right-Pointing-Pointer Using brighter BRET components substantially increased (25%) R{sub 0} of the BRET{sup 1} system. Black-Right-Pointing-Pointer Using brighter BRET components marginally increased (2-9%) R{sub 0} of the BRET{sup 2} system. Black-Right-Pointing-Pointer Brighter BRET components improve the different weaknesses of BRET{sup 1} and BRET{sup 2} systems. -- Abstract: Bioluminescence resonance energy transfer (BRET) is an important tool for monitoring macromolecular interactions and is useful as a transduction technique for biosensor development. Foerster distance (R{sub 0}), the intermolecular separation characterized by 50% of the maximum possible energy transfer, is a critical BRET parameter. R{sub 0} provides a means of linking measured changes in BRET ratio to a physical dimension scale and allows estimation of the range of distances that can be measured by any donor-acceptor pair. The sensitivity of BRET assays has recently been improved by introduction of new BRET components, RLuc2, RLuc8 and Venus with improved quantum yields, stability and brightness. We determined R{sub 0} for BRET{sup 1} systems incorporating novel RLuc variants RLuc2 or RLuc8, in combination with Venus, as 5.68 or 5.55 nm respectively. These values were approximately 25% higher than the R{sub 0} of the original BRET{sup 1} system. R{sub 0} for BRET{sup 2} systems combining green fluorescent proteins (GFP{sup 2}) with RLuc2 or RLuc8 variants was 7.67 or 8.15 nm, i.e. only 2-9% greater than the original BRET{sup 2} system despite being {approx}30-fold brighter.

  6. Bioavailability of Cd in 110 polluted topsoils to recombinant bioluminescent sensor bacteria. Effect of soil particulate matter

    Energy Technology Data Exchange (ETDEWEB)

    Ivask, Angela; Pollumaa, Lee; Kahru, Anne [National Inst. of Chemical Physics and Biophysics, Lab. of Molecular Genetics, Tallin (Estonia); Dubourguier, Henri-Charles [National Inst. of Chemical Physics and Biophysics, Lab. of Molecular Genetics, Tallin (Estonia); Estonian Univ. of Life Sciences, Tartu (Estonia); Inst. Superieur d' Agriculture, Lille (France)

    2011-02-15

    In this study, bioavailability and water extractability of Cd in a panel of 110 natural aged heavy metal-polluted soils from northern France containing up to 20.1 mg of Cd per kilogramme was evaluated. Materials and methods Particulate matter was removed by differential centrifugation of soil-water suspensions (liquid to solid ratio 10) resulting in soil-water extracts containing different size of particles. Chemical analysis of Cd and analysis of bioavailable Cd with recombinant bioluminescent Cd-sensing bacteria were applied in parallel to these fractionated soil solutions. Results and Discussion Extractability of Cd from soil to the aqueous phase was low-only 0.13% of the soil total Cd as a mean; however, Cd-sensing recombinant luminescent bacteria Bacillus subtilis incubated in soil-water suspensions for 2 h showed that in the conditions of contact exposure, the bioavailable fraction of Cd increased about 30-fold being 3.74% of the soil total Cd as a mean value. The total Cd content of soils was not a good predictor of either bioavailable or water-extracted fraction of Cd, but these fractions were rather determined by the combination of soil total Cd and physico-chemical properties-texture and organic matter content. Analysis of two selected ''model'' soils with Cd sensor bacteria showed that about 90% of the bioavailable Cd was associated with larger soil particles that were removed from the soil suspensions by centrifugation at 4,500 x g, and even settling of the soil suspensions for 2 h removed already 65% of bioavailable Cd. Conclusions Thus, our results indicate a potential for remarkably higher environmental hazard for soil-associated heavy metals than just aqueous exposure. (orig.)

  7. Further Assessment of Monkeypox Virus Infection in Gambian Pouched Rats (Cricetomys gambianus Using In Vivo Bioluminescent Imaging.

    Directory of Open Access Journals (Sweden)

    Elizabeth A Falendysz

    Full Text Available Monkeypox is a zoonosis clinically similar to smallpox in humans. Recent evidence has shown a potential risk of increased incidence in central Africa. Despite attempts to isolate the virus from wild rodents and other small mammals, no reservoir host has been identified. In 2003, Monkeypox virus (MPXV was accidentally introduced into the U.S. via the pet trade and was associated with the Gambian pouched rat (Cricetomys gambianus. Therefore, we investigated the potential reservoir competence of the Gambian pouched rat for MPXV by utilizing a combination of in vivo and in vitro methods. We inoculated three animals by the intradermal route and three animals by the intranasal route, with one mock-infected control for each route. Bioluminescent imaging (BLI was used to track replicating virus in infected animals and virological assays (e.g. real time PCR, cell culture were used to determine viral load in blood, urine, ocular, nasal, oral, and rectal swabs. Intradermal inoculation resulted in clinical signs of monkeypox infection in two of three animals. One severely ill animal was euthanized and the other affected animal recovered. In contrast, intranasal inoculation resulted in subclinical infection in all three animals. All animals, regardless of apparent or inapparent infection, shed virus in oral and nasal secretions. Additionally, BLI identified viral replication in the skin without grossly visible lesions. These results suggest that Gambian pouched rats may play an important role in transmission of the virus to humans, as they are hunted for consumption and it is possible for MPXV-infected pouched rats to shed infectious virus without displaying overt clinical signs.

  8. In Vivo Bioluminescence Imaging To Evaluate Systemic and Topical Antibiotics against Community-Acquired Methicillin-Resistant Staphylococcus aureus-Infected Skin Wounds in Mice

    Science.gov (United States)

    Guo, Yi; Ramos, Romela Irene; Cho, John S.; Donegan, Niles P.; Cheung, Ambrose L.

    2013-01-01

    Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) frequently causes skin and soft tissue infections, including impetigo, cellulitis, folliculitis, and infected wounds and ulcers. Uncomplicated CA-MRSA skin infections are typically managed in an outpatient setting with oral and topical antibiotics and/or incision and drainage, whereas complicated skin infections often require hospitalization, intravenous antibiotics, and sometimes surgery. The aim of this study was to develop a mouse model of CA-MRSA wound infection to compare the efficacy of commonly used systemic and topical antibiotics. A bioluminescent USA300 CA-MRSA strain was inoculated into full-thickness scalpel wounds on the backs of mice and digital photography/image analysis and in vivo bioluminescence imaging were used to measure wound healing and the bacterial burden. Subcutaneous vancomycin, daptomycin, and linezolid similarly reduced the lesion sizes and bacterial burden. Oral linezolid, clindamycin, and doxycycline all decreased the lesion sizes and bacterial burden. Oral trimethoprim-sulfamethoxazole decreased the bacterial burden but did not decrease the lesion size. Topical mupirocin and retapamulin ointments both reduced the bacterial burden. However, the petrolatum vehicle ointment for retapamulin, but not the polyethylene glycol vehicle ointment for mupirocin, promoted wound healing and initially increased the bacterial burden. Finally, in type 2 diabetic mice, subcutaneous linezolid and daptomycin had the most rapid therapeutic effect compared with vancomycin. Taken together, this mouse model of CA-MRSA wound infection, which utilizes in vivo bioluminescence imaging to monitor the bacterial burden, represents an alternative method to evaluate the preclinical in vivo efficacy of systemic and topical antimicrobial agents. PMID:23208713

  9. Adenosina trifosfato bioluminescência para avaliação da limpeza de superfícies: uma revisão integrativa

    Directory of Open Access Journals (Sweden)

    Adriana Cristina de Oliveira

    2014-12-01

    Full Text Available Objetivo: Identificar na literatura indicações e controvérsias do ATP bioluminescência para avaliação da efetividade da limpeza de superfícies em estabelecimentos de saúde. Método: Revisão integrativa da literatura, entre 2000 e 2012, nas bases de dados MEDLINE, LILACS, Science Direct, SCOPUS e Isi Web of Knowledge. Resultados: Selecionou-se para esta revisão 15 artigos. O ATP bioluminescência foi apontado como importante recurso educacional e método complementar à inspeção visual e às análises microbiológicas na avaliação da efetividade da limpeza. A impossibilidade de indicar a contaminação da superfície por micro-organismos viáveis, a interferência por substâncias químicas e a dificuldade de interpretação dos resultados constituem as principais controvérsias para o uso deste nos serviços de saúde. Conclusão: Apesar de constituir importante recurso na avaliação da limpeza de superfícies, mais estudos são necessários para incorporação efetiva do método nos serviços de saúde.

  10. Bioluminescence-based cytotoxicity assay for simultaneous evaluation of cell viability and membrane damage in human hepatoma HepG2 cells.

    Science.gov (United States)

    Uno, Katsuhiro; Murotomi, Kazutoshi; Kazuki, Yasuhiro; Oshimura, Mitsuo; Nakajima, Yoshihiro

    2018-05-01

    We have developed a bioluminescence-based non-destructive cytotoxicity assay in which cell viability and membrane damage are simultaneously evaluated using Emerald luciferase (ELuc) and endoplasmic reticulum (ER)-targeted copepod luciferase (GLuc-KDEL), respectively, by using multi-integrase mouse artificial chromosome (MI-MAC) vector. We have demonstrated that the time-dependent concentration response curves of ELuc luminescence intensity and WST-1 assay, and GLuc-KDEL luminescence intensity and lactate dehydrogenase (LDH) activity in the culture medium accompanied by cytotoxicity show good agreement in toxicant-treated ELuc- and GLuc-KDEL-expressing HepG2 stable cell lines. We have clarified that the increase of GLuc-KDEL luminescence intensity in the culture medium reflects the type of cell death, including necrosis and late apoptosis, but not early apoptosis. We have also uncovered a strong correlation between GLuc-KDEL luminescence intensity in the culture medium and the extracellular release of high mobility group box 1 (HMGB1), a representative damage-associated molecular pattern (DAMP) molecule. The bioluminescence measurement assay using ELuc and GLuc-KDEL developed in this study can simultaneously monitor cell viability and membrane damage, respectively, and the increase of GLuc-KDEL luminescence intensity in the culture medium accompanied by the increase of cytotoxicity is an index of necrosis and late apoptosis associated with the extracellular release of DAMP molecules. Copyright © 2018 John Wiley & Sons, Ltd.

  11. A Trp53fl/flPtenfl/fl mouse model of undifferentiated pleomorphic sarcoma mediated by adeno-Cre injection and in vivo bioluminescence imaging.

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    Marisa R Buchakjian

    Full Text Available Genetic mouse models of soft tissue sarcoma provide critical insights into disease pathophysiology, which are oftentimes unable to be extracted from human tumor samples or xenograft models. In this study we describe a mouse model of soft tissue sarcoma mediated by adenoviral-Cre recombinase injection into Trp53fl/fl/Ptenfl/fl lox-stop-lox luciferase mice. Injection of adenovirus expressing Cre recombinase, either subcutaneously or intramuscularly in two experimental groups, results in viral infection and gene recombination with 100% penetrance within the first 24 hours following injection. Luciferase expression measured by real-time bioluminescence imaging increases over time, with an initial robust increase following viral injection, followed by a steady rise over the next several weeks as primary tumors develop and grow. Intramuscular injections were more commonly associated with evidence of systemic viral distribution than subcutaneous injections. All mice developed soft tissue sarcomas at the primary injection site, with histological examination identifying 93% of tumors as invasive pleomorphic sarcomas based on microscopic morphology and immunohistochemical expression of sarcoma markers. A lymphocytic infiltrate was present in 64% of the sarcomas in this immunocompetent model and 71% of tumors expressed PD-L1. This is the first report of a viral-Cre mediated Trp53/Pten mouse model of undifferentiated pleomorphic sarcoma. The bioluminescence imaging feature, along with high penetrance of the model and its immunological characteristics, makes it suited for pre-clinical studies of soft tissue sarcoma.

  12. Revisiting Coleoptera a + T-rich region: structural conservation, phylogenetic and phylogeographic approaches in mitochondrial control region of bioluminescent Elateridae species (Coleoptera).

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    Amaral, Danilo T; Mitani, Yasuo; Oliveira, Gabriela; Ohmiya, Yoshihiro; Viviani, Vadim R

    2017-09-01

    The control region (CR) or A + T-rich region in Coleoptera mt genome is poorly characterized, including the Elateroidea bioluminescent species. Here, we provided the first attempt to characterize and compare the structure and organization of the CR of different species within Elateridae. We also revisited some sequenced Coleoptera CR and observed consensus T-stretches, non-conserved sequences near the stem-loop and unusual inner tRNAs-like sequences. All these features are probably involved in the replication start of the mt genome. The phylogenetic relationships in Elateridae bioluminescent groups using partial sequence of CR showed the monophyly of Pyrearinus pumilus group and Pyrearinus as a polyphyletic genus, corroborating our previous results. The wider genetic variation obtained by CR analysis could separate two different lineages that occur within P. termitilluminans populations. In Elateridae, the CR exhibited high polymorphism within and between populations, which was also observed in other Coleoptera species, suggesting that the CR could be described as a suitable molecular marker to be applied in phylogenetic and phylogeographic studies.

  13. SU-C-303-04: Evaluation of On- and Off-Line Bioluminescence Tomography System for Focal Irradiation Guidance

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    Zhang, B; Wang, K; Reyes, J; Tran, P; Wong, J [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University, Baltimore, MD (United States); Iordachita, I [Laboratory for Computational Sensing and Robotics, Johns Hopkins University, Baltimore, MD (United States)

    2015-06-15

    Purpose: We have developed offline and on-board bioluminescence tomography(BLT) systems for the small animal radiation research platform(SARRP) for radiation guidance of soft tissue targets. We investigated the effectiveness of offline BLT guidance. Methods: CBCT is equipped on both the offline BLT system and SARRP that are 10 ft. apart. To evaluate the setup error during animal transport between the two systems, we implanted a luminescence source in the abdomen of anesthetized mice. Five mice were studied. After CBCT was acquired on both systems, source centers and correlation coefficients were calculated. CBCT was also used to generate object mesh for BLT reconstruction. To assess target localization, we compared the localization of the luminescence source based on (1)on-board SARRP BLT and CBCT, (2)offline BLT and CBCT, and (3)offline BLT and SARRP CBCT. The 3rd comparison examines if an offline BLT system can be used to guide radiation when there is minimal target contrast in CBCT. Results: Our CBCT results show the offset of the light source center can be maintained within 0.2 mm during animal transport. The center of mass(CoM) of the light source reconstructed by the offline BLT has an offset of 1.0 ± 0.4 mm from the ‘true’ CoM as derived from the SARRP CBCT. The results compare well with the offset of 1.0 ± 0.2 mm using on-line BLT. Conclusion: With CBCT information provided by the SARRP and effective animal immobilization during transport, these findings support the use of offline BLT in close vicinity for accurate soft tissue target localization for irradiation. However, the disadvantage of the off-line system is reduced efficiency as care is required to maintain stable animal transport. We envisage a dual use system where the on-board arrangement allows convenient access to CBCT and avoids disturbance of animal setup. The off-line capability would support standalone longitudinal imaging studies. The work is supported by NIH R01CA158100 and Xstrahl

  14. Rapid obtention of stable, bioluminescent tumor cell lines using a tCD2-luciferase chimeric construct

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    Gourzones Claire

    2011-03-01

    Full Text Available Abstract Background Bioluminescent tumor cell lines are experimental tools of major importance for cancer investigation, especially imaging of tumors in xenografted animals. Stable expression of exogenous luciferase in tumor cells combined to systemic injection of luciferin provides an excellent signal/background ratio for external optical imaging. Therefore, there is a need to rationalize and speed up the production of luciferase-positive tumor cell lines representative of multiple tumor phenotypes. For this aim we have designed a fusion gene linking the luciferase 2 protein to the c-terminus of a truncated form of the rat CD2 protein (tCD2-luc2. To allow simultaneous assessment of the wild-type luciferase 2 in a context of tCD2 co-expression, we have made a bicistronic construct for concomitant but separate expression of these two proteins (luc2-IRES-tCD2. Both the mono- and bi-cistronic constructs were transduced in lymphoid and epithelial cells using lentiviral vectors. Results The tCD2-luc2 chimera behaves as a type I membrane protein with surface presentation of CD2 epitopes. One of these epitopes reacts with the OX34, a widely spread, high affinity monoclonal antibody. Stably transfected cells are sorted by flow cytometry on the basis of OX34 staining. In vitro and, moreover, in xenografted tumors, the tCD2-luc2 chimera retains a substantial and stable luciferase activity, although not as high as the wild-type luciferase expressed from the luc2-IRES-tCD2 construct. Expression of the tCD2-luc2 chimera does not harm cell and tumor growth. Conclusion Lentiviral transduction of the chimeric tCD2-luc2 fusion gene allows selection of cell clones with stable luciferase expression in less than seven days without antibiotic selection. We believe that it will be helpful to increase the number of tumor cell lines available for in vivo imaging and assessment of novel therapeutic modalities. On a longer term, the tCD2-luc2 chimera has the potential to

  15. Developmental and Microbiological Analysis of the Inception of Bioluminescent Symbiosis in the Marine Fish Nuchequula nuchalis (Perciformes: Leiognathidae)▿

    Science.gov (United States)

    Dunlap, Paul V.; Davis, Kimberly M.; Tomiyama, Shinichi; Fujino, Misato; Fukui, Atsushi

    2008-01-01

    Many marine fish harbor luminous bacteria as bioluminescent symbionts. Despite the diversity, abundance, and ecological importance of these fish and their apparent dependence on luminous bacteria for survival and reproduction, little is known about developmental and microbiological events surrounding the inception of their symbioses. To gain insight on these issues, we examined wild-caught larvae of the leiognathid fish Nuchequula nuchalis, a species that harbors Photobacterium leiognathi as its symbiont, for the presence, developmental state, and microbiological status of the fish's internal, supraesophageal light organ. Nascent light organs were evident in the smallest specimens obtained, flexion larvae of 6.0 to 6.5 mm in notochord length (NL), a developmental stage at which the stomach had not yet differentiated and the nascent gasbladder had not established an interface with the light organ. Light organs of certain of the specimens in this size range apparently lacked bacteria, whereas light organs of other specimens of 6.5 mm in NL and of all larger specimens harbored large populations of bacteria, representatives of which were identified as P. leiognathi. Bacteria identified as Vibrio harveyi were also present in the light organ of one larval specimen. Light organ populations were composed typically of two or three genetically distinct strain types of P. leiognathi, similar to the situation in adult fish, and the same strain type was only rarely found in light organs of different larval, juvenile, or adult specimens. Light organs of larvae carried a smaller proportion of strains merodiploid for the lux-rib operon, 79 of 249 strains, than those of adults (75 of 91 strains). These results indicate that light organs of N. nuchalis flexion and postflexion larvae of 6.0 to 6.7 mm in NL are at an early stage of development and that inception of the symbiosis apparently occurs in flexion larvae of 6.0 to 6.5 mm in NL. Ontogeny of the light organ therefore

  16. ATP-bioluminescence assay as an alternative for hygiene-monitoring procedures of stainless steel milk contact surfaces Técnica do ATP-bioliminescência como uma alternativa para o monitoramento do procedimento de higienização de superfícies de aço inoxidável na indústria de laticínios

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    Patrícia Dolabela Costa

    2006-09-01

    Full Text Available ATP-bioluminescence assay was used to evaluate the cleaning and sanitizing procedures of stainless steel milk contact surfaces. Surfaces of a raw milk transportation tank, a raw milk cooling storage tank, an equalization pasteurizing tank, a pasteurized milk storage tank and a pasteurized milk packaging tank, as well as a milk centrifuge were evaluated. The Relative Light Unit (RLU using a commercial luminometer and mesophilic aerobic counts (CFU.cm-2 on surfaces were determined. There were no significant differences (p>0.05 in RLU on the assessed surfaces before and after the cleaning and sanitizing procedures. Also, there were no significant differences (p>0.05 in CFU.cm-2 on the surfaces before the cleaning and sanitizing procedures, whereas different bacterial numbers were found on surfaces after the hygiene procedures. The surfaces of the milk centrifuge, raw milk cooling tank and milk packaging tank showed the largest mesophilic aerobic numbers after the hygiene, reaching 2.21, 2.18, and 1.21 log10, respectively. The results demonstrated that there is no agreement among the techniques for classification of hygienic conditions of the evaluated surfaces, showing a non-direct relation between RLU and CFU.cm-2. The ATP-bioluminescence method showed that 100% of the surfaces were under inadequate hygiene conditions, while the plate count method detected only 50%, based on the APHA's recommendation, and 33%, based on the WHO's recommendation. High variations in RLU measurements were observed, suggesting the need for more than only one surface analysis. The ATP-bioluminescence is not a direct replacement for microbiological testing, but provides complementary information and should be used as part of an integrated strategy.A técnica do ATP-bioluminescência avaliou os procedimentos de higienização de superfícies em aço inoxidável dos tanques de transporte de leite cru, de estocagem de leite cru resfriado, de equilíbrio de um pasteurizador

  17. STUDIES ON BIOLUMINESCENCE

    Science.gov (United States)

    Harvey, E. Newton

    1919-01-01

    There seems to be very little doubt but that luciferase is a protein or so closely associated with proteins that their removal destroys its characteristic properties. The particular group of proteins to which it belongs may be arrived at by a process of exclusion, and only the group of albumins has properties which agree completely with those of luciferase. Dubois believes Pholas luciferase to be an oxidizing enzyme similar to the oxydones of Battelli and Stern because it is readily destroyed by fat solvents such as chloroform, strong alcohol, etc. He has detected iron in a luciferase solution which has dialyzed against running water for a long time, and believes it to be made up of protein in combination with iron and to act as an "oxyzymase ferrique." Cypridina luciferase, on the other hand, is not readily destroyed by fat solvents. Toluene and chloroform are good preservatives, and I often make use of them for this purpose, keeping the luciferase solutions for many months. Professor A. H. Phillips of Princeton University has very kindly analyzed some whole dried Cypridinœ for me and finds iron, copper, and manganese but no zinc or vanadium to be present. Whether these metals are connected with the action of Cypridina luciferase is uncertain, but it is significant that all three of the metals thought to be concerned in organic oxidations are present. Although a large amount of luciferin mixed with a small amount of luciferase will use up all the latter, I agree with Dubois that luciferase has sufficient properties in common with the enzymes as a class to be considered an enzyme. The peroxidases are well known to be used up in the reactions they accelerate. All workers on enzymes agree that the more enzymes are purified the less active they become. The chemical procedures necessary to remove foreign material bring about irreversible changes in the enzyme itself, a characteristic also of many protein groups and of the colloidal state in general. This is true in the case of luciferase, for the crude luciferase solution is the most active preparation that can be obtained. I believe that Cypridina luciferase should be placed in a class of oxidizing enzymes by itself—a group having the chemical reactions of an albumin, possibly in combination with some heavy metal, and which as far as we know, acts specifically on only one substance, Cypridina luciferin. It resembles the plant peroxidases in resisting the action of chloroform, toluene, etc., but will not oxidize any of the hydroxyphenol or aminophenol compounds so readily oxidized by the peroxidases, nor will the peroxidases oxidize luciferin with light production. Dubois' researches show that Pholas luciferase differs in some properties from Cypridina luciferase, and my own work indicates that firefly luciferase is more like that of Pholas. A comparative study of other species of luminous animals is needed in order to delimit more accurately the class of luciferases as a whole. Luciferin presents many characteristics in common with the proteins, but two, which, to say the least, throw doubt on its protein nature: (1) its peculiar solubility (in alcohols, esters, and glacial acetic acid), (2) and its resistance to digestion by proteases, even by trypsin which has almost universal digestive action. These two peculiarities have been discussed above. We can only say that if a protein, luciferin must belong to a new group differing from known natural proteins in these respects. In general characteristics this new group would fall somewhere on the border-line between the proteoses and peptones. It would not be surprising to find in nature proteoses or peptones soluble in absolute alcohol. We know also that some NH-CO linkages of proteins are broken down with great difficulty by trypsin as it is difficult to obtain a tryptic digest of protein which does not give the biuret reaction, and the work of Fischer and Abderhalden has shown that certain artificial polypeptides are not digested by pure activated pancreatic juice. We have, then, three possibilities. Luciferin is (1) either a natural proteose not attacked by trypsin, or (2) if attacked by trypsin, its decomposition products (presumably amino-acids) still contain the group oxidizable with light production, or (3) it is not a protein at all. I believe that luciferin has too many protein characteristics to conform to the last possibility. I have been unable to oxidize with light production various mixtures of amino-acids (from beef and casein) by means of luciferase and consequently am led to believe that luciferin is a new natural proteose, soluble in absolute alcohol and not digested by trypsin. Dubois believes Pholas luciferin to be a natural albumin with acid properties. Cypridina luciferin could not possibly be regarded as an albumin, but it is very likely that the luciferins of different species of luminous animals differ in certain characteristics. As in the case of the luciferases, we know that the luciferins are not identical substances, and only future work can determine in what particulars they differ. A summary of the properties of Cypridina luciferin and Cypridina luciferase will be found in the tables accompanying this paper. PMID:19871744

  18. Synthetic strategies for controlling inter- and intramolecular interactions: Applications in single-molecule fluorescence imaging, bioluminescence imaging, and palladium catalysis

    Science.gov (United States)

    Conley, Nicholas R.

    proximity of the Cy3 and Cy5 fluorophores, behaves as an optical photoswitch in the presence of a thiol reagent. This unique property was employed to achieve sub-diffraction-limited imaging of the stalks of Caulobacter crescentus cells with 30-nm resolution using STORM (stochastic optical reconstruction microscopy). Lastly, the synthesis of the first selenium analogue of firefly luciferin is described, and this analogue is shown to be a competent substrate for firefly luciferase (fLuc). Remarkably, it exhibits red-shifted bioluminescence emission relative to the native sulfur analogue. The in vivo performance of the selenium and sulfur analogues in imaging are compared by tail-vein injection into nude mice bearing subcutaneous tumor xenografts of a human breast cancer cell line that was stably transduced to express fLuc. Part II of this thesis begins by addressing design considerations in the development of palladium catalysts that effect oxidative transformations under mild conditions (i.e., 1 atm air, room temperature) using molecular oxygen as the terminal oxidant. A newly synthesized cationic palladium complex, [(2,9-dimethylphenanthroline)Pd(OAc)]2[OTf]2, is shown to catalyze aerobic alcohol oxidation under such conditions with an unprecedented initial turnover frequency, but the presence of partially reduced oxygen species results in competitive ligand oxidation with concomitant decrease in catalyst activity. To remedy this, oxidatively resistant ligands, which are essential for the development of next-generation, high-turnover-frequency palladium catalysts that utilize oxygen as a terminal oxidant, have been prepared and effectively employed. In addition, the first general palladium-catalyzed route to the carbonylation of diols is reported. In this system, carbon monoxide (1 atm) serves the carbonyl source, (2,9-dimethylphenanthroline)Pd(OAc) 2 acts as the catalyst, and N-chlorosuccinimide and iodosobenzene are the oxidants for 1,2- and 1,3-diols, respectively. This

  19. [ Bioluminescent assay to detect melanocortin-1 receptor (MC1R) polymorphisms (R160W, R151C, and D294H)].

    Science.gov (United States)

    Bashmakova, E E; Krasitskaya, V V; Bondar, A A; Kozlova, A V; Ruksha, T G; Frank, L A

    2015-01-01

    Several polymorphisms in melanocortin-1 receptor (MC1R) gene are shown to have associations with melanoma risk. In particular, rs1805007, rs1805008, and rs1805009 mutations causing the corresponding R151C, R160W, and D294H changes and associated with the phenotype ("red-hair mutations") are connected with melanoma and non-melanoma skin cancer risks. The work describes the approach to detect these polymorphisms based on primer extension reaction with the following dual bioluminescent assay. Model plasmids with polymorphic MC1R fragments as well as several clinical DNA samples were tested using the developed technique. The results were in good correlation with those obtained by Sanger sequencing.

  20. Horse species symposium: a novel approach to monitoring pathogen progression during uterine and placental infection in the mare using bioluminescence imaging technology and lux-modified bacteria.

    Science.gov (United States)

    Ryan, P L; Christiansen, D L; Hopper, R M; Walters, F K; Moulton, K; Curbelo, J; Greene, J M; Willard, S T

    2011-05-01

    Uterine and placental infections are the leading cause of abortion, stillbirth, and preterm delivery in the mare. Whereas uterine and placental infections in women have been studied extensively, a comprehensive examination of the pathogenic processes leading to this unsatisfactory pregnancy outcome in the mare has yet to be completed. Most information in the literature relating to late-term pregnancy loss in mares is based on retrospective studies of clinical cases submitted for necropsy. Here we report the development and application of a novel approach, whereby transgenically modified bacteria transformed with lux genes of Xenorhabdus luminescens or Photorhabdus luminescens origin and biophotonic imaging are utilized to better understand pathogen-induced preterm birth in late-term pregnant mares. This technology uses highly sensitive bioluminescence imaging camera systems to localize and monitor pathogen progression during tissue invasion by measuring the bioluminescent signatures emitted by the lux-modified pathogens. This method has an important advantage in that it allows for the potential tracking of pathogens in vivo in real time and over time, which was hitherto impossible. Although the application of this technology in domestic animals is in its infancy, investigators were successful in identifying the fetal lungs, sinuses, nares, urinary, and gastrointestinal systems as primary tissues for pathogen invasion after experimental infection of pregnant mares with lux-modified Escherichia coli. It is important that pathogens were not detected in other vital organs, such as the liver, brain, and cardiac system. Such precision in localizing sites of pathogen invasion provides potential application for this novel approach in the development of more targeted therapeutic interventions for pathogen-related diseases in the equine and other domestic species.

  1. In vivo bioluminescence imaging of Escherichia coli O104:H4 and role of aerobactin during colonization of a mouse model of infection

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    Torres Alfredo G

    2012-06-01

    Full Text Available Abstract Background A major outbreak of bloody diarrhea associated with Shiga toxin-producing Escherichia coli O104:H4 occurred early in 2011, to which an unusual number of hemolytic uremic syndrome cases were linked. Due to limited information regarding pathogenesis and/or virulence properties of this particular serotype, we investigated the contribution of the aerobactin iron transport system during in vitro and in vivo conditions. Results A bioluminescent reporter construct was used to perform real-time monitoring of E. coli O104:H4 in a mouse model of infection. We verified that our reporter strain maintained characteristics and growth kinetics that were similar to those of the wild-type E. coli strain. We found that the intestinal cecum of ICR (CD-1 mice was colonized by O104:H4, with bacteria persisting for up to 7 days after intragastric inoculation. MALDI-TOF analysis of heat-extracted proteins was performed to identify putative surface-exposed virulence determinants. A protein with a high similarity to the aerobactin iron receptor was identified and further demonstrated to be up-regulated in E. coli O104:H4 when grown on MacConkey agar or during iron-depleted conditions. Because the aerobactin iron acquisition system is a key virulence factor in Enterobacteriaceae, an isogenic aerobactin receptor (iutA mutant was created and its intestinal fitness assessed in the murine model. We demonstrated that the aerobactin mutant was out-competed by the wild-type E. coli O104:H4 during in vivo competition experiments, and the mutant was unable to persist in the cecum. Conclusion Our findings demonstrate that bioluminescent imaging is a useful tool to monitor E. coli O104:H4 colonization properties, and the murine model can become a rapid way to evaluate bacterial factors associated with fitness and/or colonization during E. coli O104:H4 infections.

  2. In vivo bioluminescence imaging of Escherichia coli O104:H4 and role of aerobactin during colonization of a mouse model of infection

    Science.gov (United States)

    2012-01-01

    Background A major outbreak of bloody diarrhea associated with Shiga toxin-producing Escherichia coli O104:H4 occurred early in 2011, to which an unusual number of hemolytic uremic syndrome cases were linked. Due to limited information regarding pathogenesis and/or virulence properties of this particular serotype, we investigated the contribution of the aerobactin iron transport system during in vitro and in vivo conditions. Results A bioluminescent reporter construct was used to perform real-time monitoring of E. coli O104:H4 in a mouse model of infection. We verified that our reporter strain maintained characteristics and growth kinetics that were similar to those of the wild-type E. coli strain. We found that the intestinal cecum of ICR (CD-1) mice was colonized by O104:H4, with bacteria persisting for up to 7 days after intragastric inoculation. MALDI-TOF analysis of heat-extracted proteins was performed to identify putative surface-exposed virulence determinants. A protein with a high similarity to the aerobactin iron receptor was identified and further demonstrated to be up-regulated in E. coli O104:H4 when grown on MacConkey agar or during iron-depleted conditions. Because the aerobactin iron acquisition system is a key virulence factor in Enterobacteriaceae, an isogenic aerobactin receptor (iutA) mutant was created and its intestinal fitness assessed in the murine model. We demonstrated that the aerobactin mutant was out-competed by the wild-type E. coli O104:H4 during in vivo competition experiments, and the mutant was unable to persist in the cecum. Conclusion Our findings demonstrate that bioluminescent imaging is a useful tool to monitor E. coli O104:H4 colonization properties, and the murine model can become a rapid way to evaluate bacterial factors associated with fitness and/or colonization during E. coli O104:H4 infections. PMID:22716772

  3. Using an Adenosine Triphosphate Bioluminescent Assay to Determine Effective Antibiotic Combinations against Carbapenem-Resistant Gram Negative Bacteria within 24 Hours.

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    Yiying Cai

    Full Text Available Current in vitro combination testing methods involve enumeration by bacterial plating, which is labor-intensive and time-consuming. Measurement of bioluminescence, released when bacterial adenosine triphosphate binds to firefly luciferin-luciferase, has been proposed as a surrogate for bacterial counts. We developed an ATP bioluminescent combination testing assay with a rapid turnaround time of 24h to determine effective antibiotic combinations.100 strains of carbapenem-resistant (CR GNB [30 Acinetobacter baumannii (AB, 30 Pseudomonas aeruginosa (PA and 40 Klebsiella pneumoniae (KP] were used. Bacterial suspensions (105 CFU/ml were added to 96-well plates containing clinically achievable concentrations of multiple single and two-antibiotic combinations. At 24h, the luminescence intensity of each well was measured. Receiver operator characteristic curves were plotted to determine optimal luminescence threshold (TRLU to discriminate between inhibitory/non-inhibitory combinations when compared to viable plating. The unweighted accuracy (UA [(sensitivity + specificity/2] of TRLU values was determined. External validation was further done using 50 additional CR-GNB.Predictive accuracies of TRLU were high for when all antibiotic combinations and species were collectively analyzed (TRLU = 0.81, UA = 89%. When individual thresholds for each species were determined, UA remained high. Predictive accuracy was highest for KP (TRLU = 0.81, UA = 91%, and lowest for AB (TRLU = 0.83, UA = 87%. Upon external validation, high overall accuracy (91% was observed. The assay distinguished inhibitory/non-inhibitory combinations with UA of 80%, 94% and 93% for AB, PA and KP respectively.We developed an assay that is robust at identifying useful combinations with a rapid turn-around time of 24h, and may be employed to guide the timely selection of effective antibiotic combinations.

  4. A non-invasive in vivo imaging system to study dissemination of bioluminescent Yersinia pestis CO92 in a mouse model of pneumonic plague.

    Science.gov (United States)

    Sha, Jian; Rosenzweig, Jason A; Kirtley, Michelle L; van Lier, Christina J; Fitts, Eric C; Kozlova, Elena V; Erova, Tatiana E; Tiner, Bethany L; Chopra, Ashok K

    2013-02-01

    The gold standard in microbiology for monitoring bacterial dissemination in infected animals has always been viable plate counts. This method, despite being quantitative, requires sacrificing the infected animals. Recently, however, an alternative method of in vivo imaging of bioluminescent bacteria (IVIBB) for monitoring microbial dissemination within the host has been employed. Yersinia pestis is a Gram-negative bacterium capable of causing bubonic, septicemic, and pneumonic plague. In this study, we compared the conventional counting of bacterial colony forming units (cfu) in the various infected tissues to IVIBB in monitoring Y. pestis dissemination in a mouse model of pneumonic plague. By using a transposon mutagenesis system harboring the luciferase (luc) gene, we screened approximately 4000 clones and obtained a fully virulent, luc-positive Y. pestis CO92 (Y. pestis-luc2) reporter strain in which transposition occurred within the largest pMT1 plasmid which possesses murine toxin and capsular antigen encoding genes. The aforementioned reporter strain and the wild-type CO92 exhibited similar growth curves, formed capsule based on immunofluorescence microscopy and flow cytometry, and had a similar LD(50). Intranasal infection of mice with 15 LD(50) of CO92-luc2 resulted in animal mortality by 72 h, and an increasing number of bioluminescent bacteria were observed in various mouse organs over a 24-72 h period when whole animals were imaged. However, following levofloxacin treatment (10 mg/kg/day) for 6 days 24 h post infection, no luminescence was observed after 72 h of infection, indicating that the tested antimicrobial killed bacteria preventing their detection in host peripheral tissues. Overall, we demonstrated that IVIBB is an effective and non-invasive way of monitoring bacterial dissemination in animals following pneumonic plague having strong correlation with cfu, and our reporter CO92-luc2 strain can be employed as a useful tool to monitor the efficacy

  5. Measuring ligand-dependent and ligand-independent interactions between nuclear receptors and associated proteins using Bioluminescence Resonance Energy Transfer (BRET).

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    Koterba, Kristen L; Rowan, Brian G

    2006-07-26

    Bioluminescent resonance energy transfer (BRET2) is a recently developed technology for the measurement of protein-protein interactions in a live, cell-based system. BRET2 is characterized by the efficient transfer of excited energy between a bioluminescent donor molecule (Renilla luciferase) and a fluorescent acceptor molecule (a mutant of Green Fluorescent Protein (GFP2). The BRET2 assay offers advantages over fluorescence resonance energy transfer (FRET) because it does not require an external light source thereby eliminating problems of photobleaching and autoflourescence. The absence of contamination by light results in low background that permits detection of very small changes in the BRET2 signal. BRET2 is dependent on the orientation and distance between two fusion proteins and therefore requires extensive preliminary standardization experiments to conclude a positive BRET2 signal independent of variations in protein titrations and arrangement in tertiary structures. Estrogen receptor (ER) signaling is modulated by steroid receptor coactivator 1 (SRC-1). To establish BRET2 in a ligand inducible system we used SRC-1 as the donor moiety and ER as the acceptor moiety. Expression and functionality of the fusion proteins were assessed by transient transfection in HEK-293 cells followed by Western blot analysis and measurement of ER-dependent reporter gene activity. These preliminary determinations are required prior to measuring nuclear receptor protein-protein interactions by BRET2. This article describes in detail the BRET2 methodology for measuring interaction between full-length ER and coregulator proteins in real-time, in an in vivo environment.

  6. Enhanced Healing of Diabetic Wounds by Topical Administration of Adipose Tissue-Derived Stromal Cells Overexpressing Stromal-Derived Factor-1: Biodistribution and Engraftment Analysis by Bioluminescent Imaging

    Directory of Open Access Journals (Sweden)

    Giuliana Di Rocco

    2011-01-01

    Full Text Available Chronic ulcers represent a major health problem in diabetic patients resulting in pain and discomfort. Conventional therapy does not guarantee adequate wound repair. In diabetes, impaired healing is partly due to poor endothelial progenitor cells mobilisation and homing, with altered levels of the chemokine stromal-derived factor-1 (SDF-1 at the wound site. Adipose tissue-associated stromal cells (AT-SCs can provide an accessible source of progenitor cells secreting proangiogenic factors and differentiating into endothelial-like cells. We demonstrated that topical administration of AT-SCs genetically modified ex vivo to overexpress SDF-1, promotes wound healing into diabetic mice. In particular, by in vivo bioluminescent imaging analysis, we monitored biodistribution and survival after transplantation of luciferase-expressing cells. In conclusion, this study indicates the therapeutic potential of AT-SCs administration in wound healing, through cell differentiation, enhanced cellular recruitment at the wound site, and paracrine effects associated with local growth-factors production.

  7. High-performance thin-layer chromatography screening of multi class antibiotics in animal food by bioluminescent bioautography and electrospray ionization mass spectrometry.

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    Chen, Yisheng; Schwack, Wolfgang

    2014-08-22

    The world-wide usage and partly abuse of veterinary antibiotics resulted in a pressing need to control residues in animal-derived foods. Large-scale screening for residues of antibiotics is typically performed by microbial agar diffusion tests. This work employing high-performance thin-layer chromatography (HPTLC) combined with bioautography and electrospray ionization mass spectrometry introduces a rapid and efficient method for a multi-class screening of antibiotic residues. The viability of the bioluminescent bacterium Aliivibrio fischeri to the studied antibiotics (16 species of 5 groups) was optimized on amino plates, enabling detection sensitivity down to the strictest maximum residue limits. The HPTLC method was developed not to separate the individual antibiotics, but for cleanup of sample extracts. The studied antibiotics either remained at the start zones (tetracyclines, aminoglycosides, fluoroquinolones, and macrolides) or migrated into the front (amphenicols), while interfering co-extracted matrix compounds were dispersed at hRf 20-80. Only after a few hours, the multi-sample plate image clearly revealed the presence or absence of antibiotic residues. Moreover, molecular information as to the suspected findings was rapidly achieved by HPTLC-mass spectrometry. Showing remarkable sensitivity and matrix-tolerance, the established method was successfully applied to milk and kidney samples. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. Rapid identification and susceptibility testing of uropathogenic microbes via immunosorbent ATP-bioluminescence assay on a microfluidic simulator for antibiotic therapy.

    Science.gov (United States)

    Dong, Tao; Zhao, Xinyan

    2015-02-17

    The incorporation of pathogen identification with antimicrobial susceptibility testing (AST) was implemented on a concept microfluidic simulator, which is well suited for personalizing antibiotic treatment of urinary tract infections (UTIs). The microfluidic device employs a fiberglass membrane sandwiched between two polypropylene components, with capture antibodies immobilized on the membrane. The chambers in the microfluidic device share the same geometric distribution as the wells in a standard 384-well microplate, resulting in compatibility with common microplate readers. Thirteen types of common uropathogenic microbes were selected as the analytes in this study. The microbes can be specifically captured by various capture antibodies and then quantified via an ATP bioluminescence assay (ATP-BLA) either directly or after a variety of follow-up tests, including urine culture, antibiotic treatment, and personalized antibiotic therapy simulation. Owing to the design of the microfluidic device, as well as the antibody specificity and the ATP-BLA sensitivity, the simulator was proven to be able to identify UTI pathogen species in artificial urine samples within 20 min and to reliably and simultaneously verify the antiseptic effects of eight antibiotic drugs within 3-6 h. The measurement range of the device spreads from 1 × 10(3) to 1 × 10(5) cells/mL in urine samples. We envision that the medical simulator might be broadly employed in UTI treatment and could serve as a model for the diagnosis and treatment of other diseases.

  9. Integral or integrated marketing

    Directory of Open Access Journals (Sweden)

    Davčik Nebojša

    2006-01-01

    Full Text Available Marketing theorists and experts try to develop business efficient organization and to get marketing performance at higher, business integrated level since its earliest beginnings. The core issue in this paperwork is the dialectic and practical approach dilemma should we develop integrated or integral marketing approach in the organization. The presented company cases as well as dialectic and functional explanations of this dilemma clearly shows that integrated marketing is narrower approach than integral marketing if we take as focal point new, unique and completed entity. In the integration the essence is in getting different parts together, which do not have to make necessary the new entity. The key elements in the definition of the integral marketing are necessity and holistic, e.g. necessity to develop new, holistic entity.

  10. Monitoring of Gene Expression in Bacteria during Infections Using an Adaptable Set of Bioluminescent, Fluorescent and Colorigenic Fusion Vectors

    OpenAIRE

    Uliczka, Frank; Pisano, Fabio; Kochut, Annika; Opitz, Wiebke; Herbst, Katharina; Stolz, Tatjana; Dersch, Petra

    2011-01-01

    A family of versatile promoter-probe plasmids for gene expression analysis was developed based on a modular expression plasmid system (pZ). The vectors contain different replicons with exchangeable antibiotic cassettes to allow compatibility and expression analysis on a low-, midi- and high-copy number basis. Suicide vector variants also permit chromosomal integration of the reporter fusion and stable vector derivatives can be used for in vivo or in situ expression studies under non-selective...

  11. Rapid detection of Salmonella in food and feed by coupling loop-mediated isothermal amplification with bioluminescent assay in real-time.

    Science.gov (United States)

    Yang, Qianru; Domesle, Kelly J; Wang, Fei; Ge, Beilei

    2016-06-17

    Salmonella is among the most significant pathogens causing food and feed safety concerns. This study examined the rapid detection of Salmonella in various types of food and feed samples by coupling loop-mediated isothermal amplification (LAMP) with a novel reporter, bioluminescent assay in real-time (BART). Performance of the LAMP-BART assay was compared to a conventional LAMP and the commercially available 3M Molecular Detection Assay (MDA) Salmonella. The LAMP-BART assay was 100 % specific among 178 strains (151 Salmonella and 27 non-Salmonella) tested. The detection limits were 36 cells per reaction in pure culture and 10(4) to 10(6) CFU per 25 g in spiked food and feed samples without enrichment, which were comparable to those of the conventional LAMP and 3M MDA Salmonella but 5-10 min faster. Ground turkey showed a strong inhibition on 3M MDA Salmonella, requiring at least 10(8) CFU per 25 g for detection. The correlation between Salmonella cell numbers and LAMP-BART signals was high (R (2) = 0.941-0.962), suggesting good quantification capability. After 24 h enrichment, all three assays accurately detected 1 to 3 CFU per 25 g of Salmonella among five types of food (cantaloupe, ground beef, ground turkey, shell eggs, and tomato) and three types of feed (cattle feed, chicken feed, and dry dog food) examined. However, 10(1) CFU per 25 g was required for cattle feed when tested by 3M MDA Salmonella. The Salmonella LAMP-BART assay was rapid, specific, sensitive, quantitative, and robust. Upon further validation, it may become a valuable tool for routine screening of Salmonella in various types of food and feed samples.

  12. Use of L-Glutamic Acid in a New Enrichment Broth (R-TATP Broth) for Detecting the Presence or Absence of Molds in Raw Ingredients/Personal Care Product Formulations by Using an ATP Bioluminescence Assay.

    Science.gov (United States)

    Yang, Youjun; English, Donald J

    The present study reports the effects of adding L-glutamic acid to a new enrichment broth designated as R-TATP broth, to promote the growth of slow-growing mold microorganisms such as Aspergillus brasiliensis and Aspergillus oryzae , without interfering in the growth of other types of microorganisms. This L-glutamic acid containing enrichment broth would be particularly valuable in a rapid microbial detection assay such as an adenosine triphosphate (ATP) bioluminescence assay. By using this new enrichment broth, the amount of ATP (represented as relative light unit ratio after normalized with the negative test control) from mold growth was significantly increased by reducing the time of detection of microbial contamination in a raw ingredient or personal care product formulation from an incubation period of 48-18 h. By using L-glutamic acid in this enrichment broth, the lag phase of the mold growth cycle was shortened. In response to various concentrations of L-glutamic acid in R-TATP broth, there was an increased amount of ATP that had been produced by mold metabolism in an ATP bioluminescence assay. By using L-glutamic acid in R-TATP broth in an ATP bioluminescence assay, the presence of mold could be detected in 18 h as well as other types of microorganisms that may or may not be present in a test sample. By detecting the presence or absence of microbial contamination in 18 h, it is superior in comparison to a 48-96 h incubation period by using either a standard or rapid detection method.

  13. Detection of Heteromers Formed by Cannabinoid CB1, Dopamine D2, and Adenosine A2A G-Protein-Coupled Receptors by Combining Bimolecular Fluorescence Complementation and Bioluminescence Energy Transfer

    Directory of Open Access Journals (Sweden)

    Gemma Navarro

    2008-01-01

    Full Text Available Functional interactions in signaling occur between dopamine D2 (D2R and cannabinoid CB1 (CB1R receptors, between CB1R and adenosine A2A (A2AR receptors, and between D2R and A2AR. Furthermore, direct molecular interactions have been reported for the pairs CB1R-D2R, A2AR-D2R, and CB1R-A2AR. Here a combination of bimolecular fluorescence complementation and bioluminescence energy transfer techniques was used to identify the occurrence of D2R-CB1R-A2AR hetero-oligomers in living cells.

  14. Systematic calibration of an integrated x-ray and optical tomography system for preclinical radiation research

    International Nuclear Information System (INIS)

    Yang, Yidong; Wang, Ken Kang-Hsin; Wong, John W.; Eslami, Sohrab; Iordachita, Iulian I.; Patterson, Michael S.

    2015-01-01

    Purpose: The cone beam computed tomography (CBCT) guided small animal radiation research platform (SARRP) has been developed for focal tumor irradiation, allowing laboratory researchers to test basic biological hypotheses that can modify radiotherapy outcomes in ways that were not feasible previously. CBCT provides excellent bone to soft tissue contrast, but is incapable of differentiating tumors from surrounding soft tissue. Bioluminescence tomography (BLT), in contrast, allows direct visualization of even subpalpable tumors and quantitative evaluation of tumor response. Integration of BLT with CBCT offers complementary image information, with CBCT delineating anatomic structures and BLT differentiating luminescent tumors. This study is to develop a systematic method to calibrate an integrated CBCT and BLT imaging system which can be adopted onboard the SARRP to guide focal tumor irradiation. Methods: The integrated imaging system consists of CBCT, diffuse optical tomography (DOT), and BLT. The anatomy acquired from CBCT and optical properties acquired from DOT serve as a priori information for the subsequent BLT reconstruction. Phantoms were designed and procedures were developed to calibrate the CBCT, DOT/BLT, and the entire integrated system. Geometrical calibration was performed to calibrate the CBCT system. Flat field correction was performed to correct the nonuniform response of the optical imaging system. Absolute emittance calibration was performed to convert the camera readout to the emittance at the phantom or animal surface, which enabled the direct reconstruction of the bioluminescence source strength. Phantom and mouse imaging were performed to validate the calibration. Results: All calibration procedures were successfully performed. Both CBCT of a thin wire and a euthanized mouse revealed no spatial artifact, validating the accuracy of the CBCT calibration. The absolute emittance calibration was validated with a 650 nm laser source, resulting in a 3

  15. Systematic calibration of an integrated x-ray and optical tomography system for preclinical radiation research

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Yidong, E-mail: yidongyang@med.miami.edu [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University School of Medicine, Baltimore, Maryland 21231 and Department of Radiation Oncology, University of Miami School of Medicine, Miami, Florida 33136 (United States); Wang, Ken Kang-Hsin; Wong, John W. [Department of Radiation Oncology and Molecular Radiation Sciences, Johns Hopkins University School of Medicine, Baltimore, Maryland 21231 (United States); Eslami, Sohrab; Iordachita, Iulian I. [Laboratory for Computational Sensing and Robotics, Johns Hopkins University, Baltimore, Maryland 21218 (United States); Patterson, Michael S. [Juravinski Cancer Centre and Department of Medical Physics and Applied Radiation Sciences, McMaster University, Hamilton, Ontario L8S4K1 (Canada)

    2015-04-15

    Purpose: The cone beam computed tomography (CBCT) guided small animal radiation research platform (SARRP) has been developed for focal tumor irradiation, allowing laboratory researchers to test basic biological hypotheses that can modify radiotherapy outcomes in ways that were not feasible previously. CBCT provides excellent bone to soft tissue contrast, but is incapable of differentiating tumors from surrounding soft tissue. Bioluminescence tomography (BLT), in contrast, allows direct visualization of even subpalpable tumors and quantitative evaluation of tumor response. Integration of BLT with CBCT offers complementary image information, with CBCT delineating anatomic structures and BLT differentiating luminescent tumors. This study is to develop a systematic method to calibrate an integrated CBCT and BLT imaging system which can be adopted onboard the SARRP to guide focal tumor irradiation. Methods: The integrated imaging system consists of CBCT, diffuse optical tomography (DOT), and BLT. The anatomy acquired from CBCT and optical properties acquired from DOT serve as a priori information for the subsequent BLT reconstruction. Phantoms were designed and procedures were developed to calibrate the CBCT, DOT/BLT, and the entire integrated system. Geometrical calibration was performed to calibrate the CBCT system. Flat field correction was performed to correct the nonuniform response of the optical imaging system. Absolute emittance calibration was performed to convert the camera readout to the emittance at the phantom or animal surface, which enabled the direct reconstruction of the bioluminescence source strength. Phantom and mouse imaging were performed to validate the calibration. Results: All calibration procedures were successfully performed. Both CBCT of a thin wire and a euthanized mouse revealed no spatial artifact, validating the accuracy of the CBCT calibration. The absolute emittance calibration was validated with a 650 nm laser source, resulting in a 3

  16. Preclinical evaluation of destruxin B as a novel Wnt signaling target suppressing proliferation and metastasis of colorectal cancer using non-invasive bioluminescence imaging

    Energy Technology Data Exchange (ETDEWEB)

    Yeh, Chi-Tai [Graduate Institute of Clinical Medicine, Taipei Medical University, Taipei, Taiwan (China); Center of Excellence for Cancer Research, Taipei Medical University, Taipei, Taiwan (China); Department of Surgery, Taipei Medical University-Shuang Ho Hospital, Taipei, Taiwan (China); Rao, Yerra Koteswara [Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, Taichung, Taiwan (China); Ye, Min [Department of Natural Medicine, School of Pharmaceutical Sciences, Peking University, Beijing (China); Wu, Wen-Shi [Department of Horticulture and Biotechnology, Chinese Culture University, Taipei, Taiwan (China); Chang, Tung-Chen [Department of Surgery, Taipei Medical University-Shuang Ho Hospital, Taipei, Taiwan (China); Wang, Liang-Shun [Graduate Institute of Clinical Medicine, Taipei Medical University, Taipei, Taiwan (China); Division of Thoracic Surgery, Department of Surgery, Shuang Ho Hospital, Taipei Medical University, Taipei, Taiwan (China); Wu, Chih-Hsiung [Center of Excellence for Cancer Research, Taipei Medical University, Taipei, Taiwan (China); Department of Surgery, Taipei Medical University-Shuang Ho Hospital, Taipei, Taiwan (China); Wu, Alexander T.H., E-mail: chaw1211@tmu.edu.tw [Ph.D. Program for Translational Medicine, College of Medical Science and Technology, Taipei Medical University, Taipei, Taiwan (China); Department of Radiation Oncology, Taipei Medical University Hospital, Taipei, Taiwan (China); Tzeng, Yew-Min, E-mail: ymtzeng@cyut.edu.tw [Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, Taichung, Taiwan (China)

    2012-05-15

    In continuation to our studies toward the identification of direct anti-cancer targets, here we showed that destruxin B (DB) from Metarhizium anisopliae suppressed the proliferation and induced cell cycle arrest in human colorectal cancer (CRC) HT29, SW480 and HCT116 cells. Additionally, DB induced apoptosis in HT29 cells by decreased expression level of anti-apoptotic proteins Bcl-2 and Bcl-xL while increased pro-apoptotic Bax. On the other hand, DB attenuated Wnt-signaling by downregulation of β-catenin, Tcf4 and β-catenin/Tcf4 transcriptional activity, concomitantly with decreased expression of β-catenin target genes cyclin D1, c-myc and survivin. Furthermore, DB affected the migratory and invasive ability of HT29 cells through suppressed MMPs-2 and -9 enzymatic activities. We also found that DB targeted the MAPK and/or PI3K/Akt pathway by reduced expression of Akt, IKK-α, JNK, NF-κB, c-Jun and c-Fos while increased that of IκBα. Finally, we demonstrated that DB inhibited tumorigenesis in HT29 xenograft mice using non-invasive bioluminescence technique. Consistently, tumor samples from DB-treated mice demonstrated suppressed expression of β-catenin, cyclin D1, survivin, and endothelial marker CD31 while increased caspase-3 expression. Collectively, our data supports DB as an inhibitor of Wnt/β-catenin/Tcf signaling pathway that may be beneficial in the CRC management. Highlights: ► Destruxin B (DB) inhibited colorectal cancer cells growth and induced apoptosis. ► MAPK and/or PI3K/Akt cascade cooperates in DB induced apoptosis. ► DB affected the migratory and invasive ability of HT29 cells through MMP-9. ► DB attenuated Wnt-signaling components β-catenin, Tcf4. ► DB attenuated cyclin D1, c-myc, survivin and tumorigenesis in HT29 xenograft mice.

  17. Cavalieri integration

    CSIR Research Space (South Africa)

    Ackermann, ER

    2012-09-01

    Full Text Available We use Cavalieri’s principle to develop a novel integration technique which we call Cavalieri integration. Cavalieri integrals differ from Riemann integrals in that non-rectangular integration strips are used. In this way we can use single Cavalieri...

  18. Integral trees and integral graphs

    NARCIS (Netherlands)

    Wang, Ligong

    2005-01-01

    This monograph deals with integral graphs, Laplacian integral regular graphs, cospectral graphs and cospectral integral graphs. The organization of this work, which consists of eight chapters, is as follows.

  19. Multimodal Imaging of Integrin Receptor-Positive Tumors by Bioluminescence, Fluorescence, Gamma Scintigraphy, and Single-Photon Emission Computed Tomography Using a Cyclic RGD Peptide Labeled with a Near-Infrared Fluorescent Dye and a Radionuclide

    Directory of Open Access Journals (Sweden)

    W. Barry Edwards

    2009-03-01

    Full Text Available Integrins, particularly the αvβ3 heterodimers, play important roles in tumor-induced angiogenesis and invasiveness. To image the expression pattern of the αvβ3 integrin in tumors through a multimodality imaging paradigm, we prepared a cyclic RGDyK peptide analogue (LS308 bearing a tetraazamacrocycle 1,4,7,10-tetraazacyclododecane-N, N′, N″, N‴-tetraacetic acid (DOTA and a lipophilic near-infrared (NIR fluorescent dye cypate. The αvβ3 integrin binding affinity and the internalization properties of LS308 mediated by the αvβ3 integrin in 4t1luc cells were investigated by receptor binding assay and fluorescence microscopy, respectively. The in vivo distribution of 111In-labeled LS308 in a 4t1luc tumor-bearing mouse model was studied by fluorescence, bioluminescence, planar gamma, and single-photon emission computed tomography (SPECT. The results show that LS308 has high affinity for αvβ3 integrin and internalized preferentially via the αvβ3 integrin-mediated endocytosis in 4t1luc cells. We also found that LS308 selectively accumulated in αvβ3-positve tumors in a receptor-specific manner and was visualized by the four imaging methods. Whereas the endogenous bioluminescence imaging identified the ensemble of the tumor tissue, the fluorescence and SPECT methods with the exogenous contrast agent LS308 reported the local expression of αvβ3 integrin. Thus, the multimodal imaging approach could provide important complementary diagnostic information for monitoring the efficacy of new antiangiogenic drugs.

  20. Superdecomposition integrals

    Czech Academy of Sciences Publication Activity Database

    Mesiar, Radko; Li, J.; Pap, E.

    2015-01-01

    Roč. 259, č. 1 (2015), s. 3-11 ISSN 0165-0114 R&D Projects: GA ČR GAP402/11/0378 Institutional support: RVO:67985556 Keywords : Choquet integral * Decomposition integral * Superdecomposition integral * Convex integral Subject RIV: BA - General Mathematics Impact factor: 2.098, year: 2015 http://library.utia.cas.cz/separaty/2015/E/mesiar-0442006.pdf

  1. Riemann Integration

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 11; Issue 11. Riemann Integration. Dilip P Patil. General Article Volume 11 Issue 11 November 2006 pp 61-80 ... Keywords. Area; lower and upper sums; Riemann sums; integrable functions; definite integral; derivative; primitive; anti-derivative.

  2. VECTOR INTEGRATION

    NARCIS (Netherlands)

    Thomas, E. G. F.

    2012-01-01

    This paper deals with the theory of integration of scalar functions with respect to a measure with values in a, not necessarily locally convex, topological vector space. It focuses on the extension of such integrals from bounded measurable functions to the class of integrable functions, proving

  3. Interorganisational Integration

    DEFF Research Database (Denmark)

    Lyngsø, Anne Marie; Godtfredsen, Nina Skavlan; Frølich, Anne

    2016-01-01

    INTRODUCTION: Despite many initiatives to improve coordination of patient pathways and intersectoral cooperation, Danish health care is still fragmented, lacking intra- and interorganisational integration. This study explores barriers to and facilitators of interorganisational integration...... at a university hospital in the Capital Region of Denmark. RESULTS AND DISCUSSION: Our results can be grouped into five influencing areas for interorganisational integration: communication/information transfer, committed leadership, patient engagement, the role and competencies of the general practitioner...... and organisational culture. Proposed solutions to barriers in each area hold the potential to improve care integration as experienced by individuals responsible for supporting and facilitating it. Barriers and facilitators to integrating care relate to clinical, professional, functional and normative integration...

  4. Integrated Science.

    Science.gov (United States)

    Rainey, Larry; Miller, Roxanne Greitz

    1997-01-01

    Describes the Integrated Science program that integrates biology, earth/space science, chemistry, and physics over a three-year, spiraling sequence arranged around broad themes such as cycles, changes, patterns, and waves. Includes weekly telecasts via public television and satellite, teacher manuals, student handbooks, e-mail connections, staff…

  5. Integrated Design

    DEFF Research Database (Denmark)

    Lenau, Torben Anker

    1999-01-01

    A homepage on the internet with course material, lecture plan, student exercises, etc. Continuesly updated during the course Integrated Design (80402, 80403)......A homepage on the internet with course material, lecture plan, student exercises, etc. Continuesly updated during the course Integrated Design (80402, 80403)...

  6. The integration of immigrants

    OpenAIRE

    Bauböck, Rainer

    1995-01-01

    from the Table of Contents: Migration and integration - Basic concepts and definitions; Immigration and Integration policies; The legal framework for integration; Dimension of social integration; Cultural integration; Conclusions;

  7. Integration of Chandrasekhar's integral equation

    International Nuclear Information System (INIS)

    Tanaka, Tasuku

    2003-01-01

    We solve Chandrasekhar's integration equation for radiative transfer in the plane-parallel atmosphere by iterative integration. The primary thrust in radiative transfer has been to solve the forward problem, i.e., to evaluate the radiance, given the optical thickness and the scattering phase function. In the area of satellite remote sensing, our problem is the inverse problem: to retrieve the surface reflectance and the optical thickness of the atmosphere from the radiance measured by satellites. In order to retrieve the optical thickness and the surface reflectance from the radiance at the top-of-the atmosphere (TOA), we should express the radiance at TOA 'explicitly' in the optical thickness and the surface reflectance. Chandrasekhar formalized radiative transfer in the plane-parallel atmosphere in a simultaneous integral equation, and he obtained the second approximation. Since then no higher approximation has been reported. In this paper, we obtain the third approximation of the scattering function. We integrate functions derived from the second approximation in the integral interval from 1 to ∞ of the inverse of the cos of zenith angles. We can obtain the indefinite integral rather easily in the form of a series expansion. However, the integrals at the upper limit, ∞, are not yet known to us. We can assess the converged values of those series expansions at ∞ through calculus. For integration, we choose coupling pairs to avoid unnecessary terms in the outcome of integral and discover that the simultaneous integral equation can be deduced to the mere integral equation. Through algebraic calculation, we obtain the third approximation as a polynomial of the third degree in the atmospheric optical thickness

  8. Strobed integrator

    International Nuclear Information System (INIS)

    Krasnokutskij, R.N.; Kurchaninov, L.L.; Fedyakin, N.N.; Shuvalov, R.S.

    1989-01-01

    Two measuring techniques for signal amplitude of particle detector without the in-side amplification: with peak detector and with strobed integrator are compared. Simple formulae to calculate noise value at the output of strobed integrator are derived. Usage of strobed integrator is shown to enable to increase essentially counting rate as compared to peak detector at the same noise level. Optimal positioning strobe-pulse with regard to the input signal is determined. Equivalent noise charge ar signal shaping sifferent schemes is calculated

  9. Stochastic integrals

    CERN Document Server

    McKean, Henry P

    2005-01-01

    This little book is a brilliant introduction to an important boundary field between the theory of probability and differential equations. -E. B. Dynkin, Mathematical Reviews This well-written book has been used for many years to learn about stochastic integrals. The book starts with the presentation of Brownian motion, then deals with stochastic integrals and differentials, including the famous Itô lemma. The rest of the book is devoted to various topics of stochastic integral equations, including those on smooth manifolds. Originally published in 1969, this classic book is ideal for supplemen

  10. Estudo comparativo da eficácia de sanitizantes por meio da técnica de ATP - bioluminescência em sistemas de higienização do tipo clean-in-place (CIP

    Directory of Open Access Journals (Sweden)

    Adbeel de Lima Santos

    2009-10-01

    Full Text Available Utilizou-se o método de ATP-bioluminescência para avaliação de procedimentos de higienização CIP, com diferentes sanitizantes, de um homogeneizador em uma indústria de laticínios de Juiz de Fora - MG. A eficácia do processo foi comparada com base na utilização de dióxido de cloro estabilizado em solução aquosa, hipoclorito de sódio, ácido peracético e iodóforo. Os resultados obtidos permitiram concluir que o dióxido de cloro e o ácido peracético possuem a mesma eficácia. O mesmo ocorre com o hipoclorito de sódio e o iodóforo, porém, estes últimos não foram eficazes nas condições analisadas.

  11. Discrimination of methicillin-resistant Staphylococcus aureus from methicillin-susceptible Staphylococcus aureus or coagulase-negative staphylococci by detection of penicillin-binding protein 2 and penicillin-binding protein 2' using a bioluminescent enzyme immunoassay.

    Science.gov (United States)

    Shiga, Kazuki; Gomi, Keiko; Nishimura, Motoi; Watanabe, Masaharu; Nomura, Fumio; Kajiyama, Naoki

    2013-02-28

    For the discrimination of methicillin-resistant S. aureus (MRSA) from methicillin-susceptible S. aureus (MSSA) or coagulase-negative staphylococci (CNS), we developed a bioluminescent enzyme immunoassay (BLEIA) for detecting penicillin-binding protein 2 (PBP2) and penicillin-binding protein 2' (PBP2') using biotinylated firefly luciferase. The BLEIA was able to detect recombinant PBP2 at 50 pg/ml and recombinant PBP2' at 500 pg/ml. PBP2 and PBP2' present in the membranes of S. aureus were extracted by acid and detergent treatment. The method was able to detect PBP2 or PBP2' extracted from 10(6) colony forming units of S. aureus because of efficient extraction and the high sensitivity of luciferase. In a study of clinical isolates previously characterized as either MRSA or MSSA by antibiotic susceptibility testing, all 34 specimens identified as MRSA were both PBP2 and PBP2' positive. The 34 MSSA specimens were PBP2 positive and PBP2' negative. Moreover, the BLEIA could detect PBP2' extracted from four species of methicillin-resistant CNS, but not PBP2 extracted from four species of methicillin-resistant and methicillin-susceptible CNS. This result suggested that PBP2 could be a unique marker for discrimination of S. aureus from CNS. A BLEIA that is able to detect PBP2 and PBP2' may be useful in clinical diagnostics. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. Integrated optics

    International Nuclear Information System (INIS)

    Jekowski, J.; Aeby, I.

    1979-01-01

    The purpose of the R and D effort is to establish a baseline technology in integrated optic techniques within the EMG. Significant investment presently being made by industry in this area indicates that this technology may offer an alternative solution for high speed digital data acquisition and transmission. The objectives of this year's integrated optics program were three-fold: (1) to establish a literature search and maintain industry contacts in an attempt to make the needs of the Weapons Test Program known; (2) to assist the Las Vegas Hybrid facility in establising integrated optic fabrication techniques for their new Thin Film Laboratory; and (3) to conduct basic experiments at Los Alamos in the construction of photolithograhicaly etched plastic film devices. The report describes the successful completion of preliminary investigations into photolithographic techniques and continuing investigations into newly announced integrated optic techniques and devices that would be applicable to the weapons test program

  13. Integral cryptanalysis

    DEFF Research Database (Denmark)

    Knudsen, Lars Ramkilde; Wagner, David

    2002-01-01

    This paper considers a cryptanalytic approach called integral cryptanalysis. It can be seen as a dual to differential cryptanalysis and applies to ciphers not vulnerable to differential attacks. The method is particularly applicable to block ciphers which use bijective components only.......This paper considers a cryptanalytic approach called integral cryptanalysis. It can be seen as a dual to differential cryptanalysis and applies to ciphers not vulnerable to differential attacks. The method is particularly applicable to block ciphers which use bijective components only....

  14. Organising integration

    DEFF Research Database (Denmark)

    Axelsson, Runo

    2013-01-01

    to large-scale bureaucratic structures of centralised control. In the 1980’s there was a period of decentralisation to smaller organisational units, which was followed in the 1990’s by an introduction of market mechanisms in accordance with the New Public Management. During this period, there has been......Background: In Sweden, as in many other countries, there has been a succession of trends in the organisation of health care and other welfare services. These trends have had different implications for the integration of services in the health and welfare system. Aims: One aim is to discuss...... the implications of different organisational trends for the integration of health and welfare services. Another aim is to introduce a Swedish model of financial coordination as a flexible way to organise integration. Organisational trends: In the 1960’s there was an expansion of health and welfare services leading...

  15. Lebesgue integration

    CERN Document Server

    Williamson, JH

    2014-01-01

    This concise introduction to Lebesgue integration is geared toward advanced undergraduate math majors and may be read by any student possessing some familiarity with real variable theory and elementary calculus. The self-contained treatment features exercises at the end of each chapter that range from simple to difficult. The approach begins with sets and functions and advances to Lebesgue measure, including considerations of measurable sets, sets of measure zero, and Borel sets and nonmeasurable sets. A two-part exploration of the integral covers measurable functions, convergence theorems, co

  16. Integrated Design

    DEFF Research Database (Denmark)

    Jørgensen, Michael; Nielsen, M. W.; Strømann-Andersen, Jakob Bjørn

    2011-01-01

    This paper presents a case study of the implementation of integrated design in an actual architectural competition. The design process was carried out at a highly esteemed architectural office and attended by both engineers and architects working towards mutual goals of architectural excellence......, low-energy consumption, and high-quality indoor environment. We use this case study to investigate how technical knowledge about building performance can be integrated into the conceptual design stage. We have selected certain points during the design process that represented design challenges......'s performance. This article illustrates how a continuous implementation of technical knowledge early in the design process for an actual architectural competition resulted in a building design with an energy demand approximately 30% lower than Danish building regulations, yet which still maintains a high...

  17. Integrated Toys

    DEFF Research Database (Denmark)

    Petersson, Eva

    2005-01-01

    Toys play a crucial role in supporting children’s learning and creation of meaning in their everyday life. Children also play with toys out of an interest to interact with others e.g. peers and adults. Tendencies of digital technology in toys have led to greater opportunities for manipulation...... changed our everyday lives. However, its contribution in development of new suitable material for play and learning has not been appropriate. We argue that play is a fruitful base for learning and training and that the design of physical and virtual artifacts advantageously should rest upon...... and interaction with the integrated toy as a virtual experience without attachment or wearable on the body, which in context would not be appropriate. In the field of rehabilitation integrated toys are still at an embryonic stage and as such an under used resource for rehabilitation. Qualitative method...

  18. Scientific integrity

    DEFF Research Database (Denmark)

    Merlo, Domenico Franco; Vahakangas, Kirsi; Knudsen, Lisbeth E.

    2008-01-01

    Environmental health research is a relatively new scientific area with much interdisciplinary collaboration. Regardless of which human population is included in field studies (e.g., general population, working population, children, elderly, vulnerable sub-groups, etc.) their conduct must guarantee...... consent was obtained.Integrity is central to environmental health research searching for causal relations. It requires open communication and trust and any violation (i.e., research misconduct, including fabrication or falsification of data, plagiarism, conflicting interests, etc.) may endanger...

  19. Bio-testing integral toxicity of corrosion inhibitors, biocides and oil hydrocarbons in oil-and gas-processing industry

    Energy Technology Data Exchange (ETDEWEB)

    Chugunov, V.A.; Kholodenko, V.P.; Irkhina, I.A.; Fomchenkov, V.M.; Novikov, I.A. [State Research Center for Applied Microbiology, Obolensk, Moscow (Russian Federation)

    2004-07-01

    In recent years bioassays have been widely used for assessing levels of contamination of the environment. This is due to the fact that test-organisms provide a general response to toxicants present in samples. Based on microorganisms as test objects, it is possible to develop cheap, sensitive and rapid assays to identify environmental xenobiotics and toxicants. The objective of the research was to develop different microbiological assays for assessing integral toxicity of water environments polluted with corrosion inhibitors, biocides and hydrocarbons in oil- and gas-processing industry. Bio-luminescent, electro-orientational, osmo-optic and microorganism reducing activity assays were used for express evaluation of integral toxicity. They are found to determine promptly integral toxicity of water environments containing various pollutants (oil, oil products, corrosion inhibitors, biocides). Results conclude that the assays may be used for analyzing integral toxicity of water polluted with hydrocarbons, as well as for monitoring of water changes as a result of biodegradation of pollutants by microorganisms and their associations. Using a kit of different assays, it is also possible to evaluate ecological safety of biocides, corrosion inhibitors, and their compositions. Bioassays used as a kit are more effective than each assay individually, allowing one to get complete characterization of a reaction of bacterial test organisms to different environments. (authors)

  20. TH-C-17A-12: Integrated CBCT and Optical Tomography System On-Board a Small Animal Radiation Research Platform (SARRP)

    Energy Technology Data Exchange (ETDEWEB)

    Wang, K; Zhang, B; Eslami, S; Iordachita, I; Wong, J [Johns Hopkins University, Baltimore, MD (United States); Patterson, M [Hamilton Regional Cancer Ctr., Hamilton, ON (Canada)

    2014-06-15

    Purpose: We present a newly developed on-board optical tomography system for SARRP. Innovative features include the compact design and fast acquisition optical method to perform 3D soft tissue radiation guidance. Because of the on-board feature and the combination of the CBCT, diffusive optical tomography (DOT), bioluminescence and fluorescence tomography (BLT and FT), this integrated system is expected to provide more accurate soft tissue guidance than an off-line system as well as highly sensitive functional imaging in preclinical research. Methods: Images are acquired in the order of CBCT, DOT and then BLT/FT, where the SARRP CBCT and DOT are used to provide the anatomical and optical properties information to enhance the subsequent BLT/FT optical reconstruction. The SARRP stage is redesigned to include 9 imbedded optical fibers in contact with the animal's skin. These fibers, connected to a white light lamp or laser, serve as the light sources for the DOT or FT, respectively. A CCD camera with f/1.4 lens and multi-spectral filter set is used as the optical detector and is mounted on a portable cart ready to dock into the SARRP. No radiation is delivered during optical image acquisition. A 3-way mirror system capable of 180 degree rotation around the animal reflects the optical signal to the camera at multiple projection angles. A special black-painted dome covers the stage and provides the light shielding. Results: Spontaneous metastatic bioluminescent liver and lung tumor models will be used to validate the 3D BLT reconstruction. To demonstrate the capability of our FT system, GastroSense750 fluorescence agent will be used to imaging the mouse stomach and intestinal region in 3D. Conclusion: We expect that this integrated CBCT and optical tomography on-board a SARRP will present new research opportunities for pre-clinical radiation research. Supported by NCI RO1-CA 158100.

  1. Homeland Integration

    Directory of Open Access Journals (Sweden)

    Bin Ai

    2017-03-01

    Full Text Available In this study, the first author narrates his experiences of the challenges of integration into several Chinese universities as a PhD graduate after returning from Australia. His patterns of communication and psychological changes are examined in terms of identity construction and transformation. His insider position as a Chinese native and academic returnee enables him to see the realities of practice in average Chinese universities at close range, yet with the altered vision gained from his overseas experience. This study highlights the challenges for academic returnees in Chinese higher education institutions and may also have resonances for academic returnees in other countries. Wider questions about the assessment of English research writing and the attitudes to academic returnees in Chinese universities are raised, contributing to debate over the future development of Chinese higher education institutions in a globalizing world.

  2. Numerical Contour Integration for Loop Integrals

    OpenAIRE

    Kurihara, Y.; Kaneko, T.

    2005-01-01

    A fully numerical method to calculate loop integrals, a numerical contour-integration method, is proposed. Loop integrals can be interpreted as a contour integral in a complex plane for an integrand with multi-poles in the plane. Stable and efficient numerical integrations an along appropriate contour can be performed for tensor integrals as well as for scalar ones appearing in loop calculations of the standard model. Examples of 3- and 4-point diagrams in 1-loop integrals and 2- and 3-point ...

  3. Safe genetic modification of cardiac stem cells using a site-specific integration technique.

    Science.gov (United States)

    Lan, Feng; Liu, Junwei; Narsinh, Kazim H; Hu, Shijun; Han, Leng; Lee, Andrew S; Karow, Marisa; Nguyen, Patricia K; Nag, Divya; Calos, Michele P; Robbins, Robert C; Wu, Joseph C

    2012-09-11

    Human cardiac progenitor cells (hCPCs) are a promising cell source for regenerative repair after myocardial infarction. Exploitation of their full therapeutic potential may require stable genetic modification of the cells ex vivo. Safe genetic engineering of stem cells, using facile methods for site-specific integration of transgenes into known genomic contexts, would significantly enhance the overall safety and efficacy of cellular therapy in a variety of clinical contexts. We used the phiC31 site-specific recombinase to achieve targeted integration of a triple fusion reporter gene into a known chromosomal context in hCPCs and human endothelial cells. Stable expression of the reporter gene from its unique chromosomal integration site resulted in no discernible genomic instability or adverse changes in cell phenotype. Namely, phiC31-modified hCPCs were unchanged in their differentiation propensity, cellular proliferative rate, and global gene expression profile when compared with unaltered control hCPCs. Expression of the triple fusion reporter gene enabled multimodal assessment of cell fate in vitro and in vivo using fluorescence microscopy, bioluminescence imaging, and positron emission tomography. Intramyocardial transplantation of genetically modified hCPCs resulted in significant improvement in myocardial function 2 weeks after cell delivery, as assessed by echocardiography (P=0.002) and MRI (P=0.001). We also demonstrated the feasibility and therapeutic efficacy of genetically modifying differentiated human endothelial cells, which enhanced hind limb perfusion (P<0.05 at day 7 and 14 after transplantation) on laser Doppler imaging. The phiC31 integrase genomic modification system is a safe, efficient tool to enable site-specific integration of reporter transgenes in progenitor and differentiated cell types.

  4. Bioluminescence for Detection of Trace Compounds

    Science.gov (United States)

    1990-03-08

    to EtBr. 13 ’ 4 ,1 I lI I I C€ Omw . 006W .i- - OONG 6.5 1.tt I’ 5TIM 4 0| *lwn 30M a 40 Anw000 0.0 CoiM1.- .mAn 0060 6 lollIn 48 go * 40I ’ 1ramtraO...of distilled water at 4"C, and then 1ilowed to stand on ice for one hour. Following this, a small amount of the concentrated cells was placed in 20

  5. Sensory Control and Function of Bacterial Bioluminescence

    National Research Council Canada - National Science Library

    Silverman, Michael

    1998-01-01

    ...) and for the synthesis of signal receptors (sensor kinases and response regulators). The genes and proteins defined by this study resemble elements of the phosphorelay paradigm known as two-component signal transduction...

  6. Bacterial bioluminescence in marine pollution assessment

    Digital Repository Service at National Institute of Oceanography (India)

    Ramaiah, N.; Chandramohan, D.

    of these species was monitored upon additions of micro and nanomolar concentrations of numerous toxic (heavy metals, pesticides, phenolic compounds, amines, aldehydes, organic solvents, commercial detergents, crude petroleum extracts) and nontoxic (essential salts...

  7. Selectivity of Whole Cell Optical Biosensor with Immobilized Bioreporter Pseudomonas Fluorescens HK44

    Czech Academy of Sciences Publication Activity Database

    Trögl, Josef; Ripp, S.; Kuncová, Gabriela; Sayler, G. S.; Churavá, Alena; Pařík, P.; Demnerová, K.; Hálová, Jaroslava; Kubicová, L.

    2005-01-01

    Roč. 107, č. 1 (2005), s. 98-103 ISSN 0925-4005. [European Conference on Optical Chemical Sensors and Biosensors EUROPT(R)ODE /7./. Madrid, 04.04.2004-07.04.2004] R&D Projects: GA ČR(CZ) GA104/01/0461 Institutional research plan: CEZ:AV0Z40720504 Keywords : biosensor * Pseudomonas fluorescens HK44 * naphthalene Subject RIV: CE - Biochemistry Impact factor: 2.646, year: 2005

  8. The mechanics of bacterial cluster formation on plant leaf surfaces as revealed by bioreporter technology

    NARCIS (Netherlands)

    Tecon, R.; Leveau, J.H.J.

    2012-01-01

    Bacteria that colonize the leaves of terrestrial plants often occur in clusters whose size varies from a few to thousands of cells. For the formation of such bacterial clusters, two non-mutually exclusive but very different mechanisms may be proposed: aggregation of multiple cells or clonal

  9. Development of a multistrain bacterial bioreporter platform for the monitoring of hydrocarbon contaminants in marine environments

    NARCIS (Netherlands)

    Tecon, R.; Beggah, S.; Czechowska, K.; Sentchilo, V.; Chronopoulou, P.M.; McGenity, T.J.; van der Meer, J.R.

    2010-01-01

    Petroleum hydrocarbons are common contaminants in marine and freshwater aquatic habitats, often occurring as a result of oil spillage. Rapid and reliable on-site tools for measuring the bioavailable hydrocarbon fractions, i.e., those that are most likely to cause toxic effects or are available for

  10. Definite Integrals using Orthogonality and Integral Transforms

    Directory of Open Access Journals (Sweden)

    Howard S. Cohl

    2012-10-01

    Full Text Available We obtain definite integrals for products of associated Legendre functions with Bessel functions, associated Legendre functions, and Chebyshev polynomials of the first kind using orthogonality and integral transforms.

  11. Sesquilinear uniform vector integral

    Indian Academy of Sciences (India)

    absolute integral which generalizes the Lebesgue integral and is more easy to manipulate. We speak so far about scalar integrals (scalar functions integrated with respect to positive measures-speaking approximately). The idea of integrating vector functions with respect to scalar measures came naturally on the scene of ...

  12. Determine Arbitrary Feynman Integrals by Vacuum Integrals

    OpenAIRE

    Liu, Xiao; Ma, Yan-Qing

    2018-01-01

    By introducing an auxiliary parameter, we find a series representation for Feynman integral, which is defined as analytical continuation of a calculable series. To obtain the series representation, one only needs to deal with some much simpler vacuum integrals. The series representation therefore translates the problem of computing Feynman integrals to the problem of performing analytical continuations. As a Feynman integral is fully determined by its series representation, its reduction rela...

  13. Wealth Through Integration: Regional Integration and Poverty ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    Wealth Through Integration: Regional Integration and Poverty-Reduction Strategies in West Africa. Book cover Wealth Through Integration. Directeur(s) : Elias T. Ayuk and Samuel T. Kaboré. Maison(s) d'édition : Springer, IDRC. 27 février 2013. ISBN : 9781461444145. 297 pages. e-ISBN : 9781552505441. Téléchargez le ...

  14. Simplification of integrity constraints for data integration

    DEFF Research Database (Denmark)

    Christiansen, Henning; Martinenghi, Davide

    2004-01-01

    When two or more databases are combined into a global one, integrity may be violated even when each database is consistent with its own local integrity constraints. Efficient methods for checking global integrity in data integration systems are called for: answers to queries can then be trusted......, because either the global database is known to be consistent or suitable actions have been taken to provide consistent views. The present work generalizes simplification techniques for integrity checking in traditional databases to the combined case. Knowledge of local consistency is employed, perhaps...... together with given a priori constraints on the combination, so that only a minimal number of tuples needs to be considered. Combination from scratch, integration of a new source, and absorption of local updates are dealt with for both the local-as-view and global-as-view approaches to data integration....

  15. Tactical Systems Integration Laboratory

    Data.gov (United States)

    Federal Laboratory Consortium — The Tactical Systems Integration Laboratory is used to design and integrate computer hardware and software and related electronic subsystems for tactical vehicles....

  16. Pro Spring Integration

    CERN Document Server

    Lui, M; Chan, Andy; Long, Josh

    2011-01-01

    Pro Spring Integration is an authoritative book from the experts that guides you through the vast world of enterprise application integration (EAI) and application of the Spring Integration framework towards solving integration problems. The book is:. * An introduction to the concepts of enterprise application integration * A reference on building event-driven applications using Spring Integration * A guide to solving common integration problems using Spring Integration What makes this book unique is its coverage of contemporary technologies and real-world information, with a focus on common p

  17. SAP crm integration testing

    OpenAIRE

    Černiavskaitė, Marija

    2017-01-01

    This Bachelor's thesis presents SAP CRM and integration systems testing analysis: investigation in SAP CRM and SAP PO systems, presentation of relationship between systems, introduction to third-party system (non-SAP) – Network Informational System (NIS) which has integration with SAP, presentation of best CRM testing practises, analysis and recommendation of integration testing. Practical integration testing is done in accordance to recommendations.

  18. Probabilistic Data Integration

    NARCIS (Netherlands)

    van Keulen, Maurice

    2017-01-01

    Probabilistic data integration is a specific kind of data integration where integration problems such as inconsistency and uncertainty are handled by means of a probabilistic data representation. The approach is based on the view that data quality problems (as they occur in an integration process)

  19. How integrated are immigrants?

    Directory of Open Access Journals (Sweden)

    Rickard Sandell

    2015-12-01

    Full Text Available Background: The successful integration of immigrants is seen as a principal means to secure economic growth and welfare in many countries. Success in this task depends on the capacity to formulate effective integration policies, which in turn is based on research capable of describing and explaining the integration process properly. Objective: Our objective is to define a conceptual and quantifiable measure of full immigrant integration.This enables a quantitative evaluation of how integrated immigrants are in a specific context in an immigrant - native system - a question poorly addressed by past research. Methods: Our approach consists of looking at the functional dependency of different integrationquantifiers on immigrant density. The empirical analysis uses register data from Spain. We focus on social integration and labour market integration in formal employment. Results: In our empirical analysis we find dramatic differences in immigrant integration levels across integration contexts. While labour market integration approaches the level of full integration, social integration quickly declines as immigration levels surge. It is shown that these differences are primarily due to the presence of social network effects in the social integration process, absent in the labour market integration process. Conclusions: Proper identification of integration deficits and its causes is likely to improve the efficiency of integration policy making, and the capacity to reach integration targets. Our framework has this quality. The research presented here shows that full labour market integration of immigrants is a realistic target. However, it also shows that, if left unattended, the segregation forces contained in social networks, quite dramatically obstruct the social integration process. Lack of social integration undermines the strategy of reliance on increasing immigration to secure future economic well-being that many governments andinternational

  20. Development and validation of non-integrative, self-limited, and replicating minicircles for safe reporter gene imaging of cell-based therapies.

    Directory of Open Access Journals (Sweden)

    John A Ronald

    Full Text Available Reporter gene (RG imaging of cell-based therapies provides a direct readout of therapeutic efficacy by assessing the fate of implanted cells. To permit long-term cellular imaging, RGs are traditionally required to be integrated into the cellular genome. This poses a potential safety risk and regulatory bottleneck for clinical translation as integration can lead to cellular transformation. To address this issue, we have developed non-integrative, replicating minicircles (MCs as an alternative platform for safer monitoring of cells in living subjects. We developed both plasmids and minicircles containing the scaffold/matrix attachment regions (S/MAR of the human interferon-beta gene, driven by the CMV promoter, and expressing the bioluminescence RG firefly luciferase. Constructs were transfected into breast cancer cells, and expanded S/MAR minicircle clones showed luciferase signal for greater than 3 months in culture and minicircles remained as episomes. Importantly, luciferase activity in clonal populations was slowly lost over time and this corresponded to a loss of episome, providing a way to reversibly label cells. To monitor cell proliferation in vivo, 1.5 × 10(6 cells carrying the S/MAR minicircle were implanted subcutaneously into mice (n = 5 and as tumors developed significantly more bioluminescence signal was noted at day 35 and 43 compared to day 7 post-implant (p<0.05. To our knowledge, this is the first work examining the use of episomal, self-limited, replicating minicircles to track the proliferation of cells using non-invasive imaging in living subjects. Continued development of S/MAR minicircles will provide a broadly applicable vector platform amenable with any of the numerous RG technologies available to allow therapeutic cell fate to be assessed in individual patients, and to achieve this without the need to manipulate the cell's genome so that safety concerns are minimized. This will lead to safe tools to assess treatment

  1. Buried waste integrated demonstration technology integration process

    International Nuclear Information System (INIS)

    Ferguson, J.S.; Ferguson, J.E.

    1992-04-01

    A Technology integration Process was developed for the Idaho National Energy Laboratories (INEL) Buried Waste Integrated Demonstration (BWID) Program to facilitate the transfer of technology and knowledge from industry, universities, and other Federal agencies into the BWID; to successfully transfer demonstrated technology and knowledge from the BWID to industry, universities, and other Federal agencies; and to share demonstrated technologies and knowledge between Integrated Demonstrations and other Department of Energy (DOE) spread throughout the DOE Complex. This document also details specific methods and tools for integrating and transferring technologies into or out of the BWID program. The document provides background on the BWID program and technology development needs, demonstrates the direction of technology transfer, illustrates current processes for this transfer, and lists points of contact for prospective participants in the BWID technology transfer efforts. The Technology Integration Process was prepared to ensure compliance with the requirements of DOE's Office of Technology Development (OTD)

  2. Cavalieri Integration - CSIR Technical report

    CSIR Research Space (South Africa)

    Grobler, TL

    2011-10-01

    Full Text Available The authors use Cavalieri's principle to develop a novel integration technique which they call Cavalieri integration. Cavalieri integrals differ from Riemann integrals in that non-rectangular integration strips are used. In this way they can use...

  3. Communicating European Integration – Information vs. Integration?

    Directory of Open Access Journals (Sweden)

    Alexander Reinfeldt

    2014-02-01

    Full Text Available This article deals with the apparent contradiction between elite-driven supranational European integration and public information efforts by supranational political actors from the early 1950s. Supranational European integration relied on rational governance by independent experts which provoked a structural democratic deficit. Until the early 1970s public participation had never been the main preoccupation of supranational political actors who instead considered the ‘permissive consensus’ to be a precondition for progress towards further European integration. And yet, from the very beginning European supranational political actors pursued information policies. At first, the article reconsiders the rationale of supranational European integration and outlines the basic features of supranational information policies. Then, the article empirically analyses the purposes that guided supranational political actors like the High Authority of the European Coal and Steel Community (ECSC and the Commissions of the European Economic Community (EEC and the European Atomic Community (EURATOM in pursuing information policies at all by discussing two significant examples: the EC information efforts directed at young audiences and the participation of the EC in fairs and exhibitions. It is argued that information policies were only in part pursued to communicate European integration to the public or to influence public opinion on European integration issues. In fact, the main intention and impact of supranational information efforts was to foster transnational European integration and co-operation, for example between member state governments and administrations, non-governmental actors or associations.

  4. One loop integrals reduction

    International Nuclear Information System (INIS)

    Sun Yi; Chang Haoran

    2012-01-01

    By further examining the symmetry of external momenta and masses in Feynman integrals, we fulfilled the method proposed by Battistel and Dallabona, and showed that recursion relations in this method can be applied to simplify Feynman integrals directly. (authors)

  5. The Integrated Renovation Process

    DEFF Research Database (Denmark)

    Galiotto, Nicolas; Heiselberg, Per; Knudstrup, Mary-Ann

    The Integrated Renovation Process (IRP) is a user customized methodology based on judiciously selected constructivist and interactive multi-criteria decision making methods (Galiotto, Heiselberg, & Knudstrup, 2014 (expected)). When applied for home renovation, the Integrated Renovation Process...

  6. Semantic Integration Technonolgies Survey

    OpenAIRE

    Kalfoglou, Yannis; Hu, Bo; Reynolds, Dave; Shadbolt, Nigel

    2005-01-01

    The \\underline{University of Southampton} and \\underline{Hewlett Packard Laboratories at Bristol} are collaborating in a joint project, CROSI, to investigate semantic integration. \\textbf{CROSI}, which stands for Capturing, Representing, and Operationalising Semantic Integration, aims to advance the state-of-the-art for semantic integration technologies. Semantic integration has become a much debated topic in today's research agenda, especially with the advent of the Semantic Web. Its roots, ...

  7. Integrated optics technology study

    Science.gov (United States)

    Chen, B.

    1982-01-01

    The materials and processes available for the fabrication of single mode integrated electrooptical components are described. Issues included in the study are: (1) host material and orientation, (2) waveguide formation, (3) optical loss mechanisms, (4) wavelength selection, (5) polarization effects and control, (6) laser to integrated optics coupling,(7) fiber optic waveguides to integrated optics coupling, (8) souces, (9) detectors. The best materials, technology and processes for fabrication of integrated optical components for communications and fiber gyro applications are recommended.

  8. Integrated vs. Federated Search

    DEFF Research Database (Denmark)

    Løvschall, Kasper

    2009-01-01

    Oplæg om forskelle og ligheder mellem integrated og federated search i bibliotekskontekst. Holdt ved temadag om "Integrated Search - samsøgning i alle kilder" på Danmarks Biblioteksskole den 22. januar 2009.......Oplæg om forskelle og ligheder mellem integrated og federated search i bibliotekskontekst. Holdt ved temadag om "Integrated Search - samsøgning i alle kilder" på Danmarks Biblioteksskole den 22. januar 2009....

  9. Boolean integral calculus

    Science.gov (United States)

    Tucker, Jerry H.; Tapia, Moiez A.; Bennett, A. Wayne

    1988-01-01

    The concept of Boolean integration is developed, and different Boolean integral operators are introduced. Given the changes in a desired function in terms of the changes in its arguments, the ways of 'integrating' (i.e. realizing) such a function, if it exists, are presented. The necessary and sufficient conditions for integrating, in different senses, the expression specifying the changes are obtained. Boolean calculus has applications in the design of logic circuits and in fault analysis.

  10. Integration phenomena Paralympic cyclists

    OpenAIRE

    Diepoldová, Tereza

    2017-01-01

    Title: Integration phenomena Paralympic cyclists. Objectives of work: Try to find integration phenomena in relation to sport training and its impact on selected cyclists with disabilities. Methods: Case report structured interview, data collection method - the method of interrogation. Results: Based on case studies developed a structured interview, we found differences in the integration, which we have divided into phases - before obtaining disability, acclimatization, sports integration. Fur...

  11. Spring integration essentials

    CERN Document Server

    Pandey, Chandan

    2015-01-01

    This book is intended for developers who are either already involved with enterprise integration or planning to venture into the domain. Basic knowledge of Java and Spring is expected. For newer users, this book can be used to understand an integration scenario, what the challenges are, and how Spring Integration can be used to solve it. Prior experience of Spring Integration is not expected as this book will walk you through all the code examples.

  12. Optimizing Computer Technology Integration

    Science.gov (United States)

    Dillon-Marable, Elizabeth; Valentine, Thomas

    2006-01-01

    The purpose of this study was to better understand what optimal computer technology integration looks like in adult basic skills education (ABSE). One question guided the research: How is computer technology integration best conceptualized and measured? The study used the Delphi method to map the construct of computer technology integration and…

  13. Foundations for Psychotherapy Integration

    Directory of Open Access Journals (Sweden)

    António Branco Vasco

    2014-10-01

    Full Text Available The movement for integration in psychotheray is clearly one of the main trends that can be observed in the field. The author stresses three main reasons for this state of affairs and as a way of justifying the importance of integration: historical and psychosocial, empirical and philosophical. A specific way of thinking in integrative terms is also outlined - "paradigmatic complementarity."

  14. Discrete bipolar universal integrals

    Czech Academy of Sciences Publication Activity Database

    Greco, S.; Mesiar, Radko; Rindone, F.

    2014-01-01

    Roč. 252, č. 1 (2014), s. 55-65 ISSN 0165-0114 R&D Projects: GA ČR GAP402/11/0378 Institutional support: RVO:67985556 Keywords : bipolar integral * universal integral * Choquet integral Subject RIV: BA - General Mathematics Impact factor: 1.986, year: 2014 http://library.utia.cas.cz/separaty/2014/E/mesiar-0432224.pdf

  15. Sledge-Hammer Integration

    Science.gov (United States)

    Ahner, Henry

    2009-01-01

    Integration (here visualized as a pounding process) is mathematically realized by simple transformations, successively smoothing the bounding curve into a straight line and the region-to-be-integrated into an area-equivalent rectangle. The relationship to Riemann sums, and to the trapezoid and midpoint methods of numerical integration, is…

  16. Four integration patterns

    DEFF Research Database (Denmark)

    Bygstad, Bendik; Nielsen, Peter Axel; Munkvold, Bjørn Erik

    2010-01-01

    . Expanding the perspective of traditional project management research, we draw extensively on central insights from IS research. Building on socio-technical IS research and Software Engineering research we suggest four generic patterns of integration: Big Bang, Stakeholder Integration, Technical Integration...

  17. Curriculum Integration: An Overview

    Science.gov (United States)

    Wall, Amanda; Leckie, Alisa

    2017-01-01

    Curriculum integration is a tenet of middle level education. "This We Believe," the position paper of the Association for Middle Level Education, advocates for curriculum that is exploratory, relevant, integrative, and meaningful for young adolescents. Teachers can integrate curriculum across content areas by anchoring units of study in…

  18. Steffensen's integral inequality for conformable fractional integrals

    Directory of Open Access Journals (Sweden)

    Mehmet Zeki Sarikaya

    2017-09-01

    Full Text Available The aim of this paper is to establish some Steffensen’s type inequalities for conformable fractional integral. The results presented here would provide generalizations of those given in earlier works.

  19. Photonic Integrated Circuits

    Science.gov (United States)

    Krainak, Michael; Merritt, Scott

    2016-01-01

    Integrated photonics generally is the integration of multiple lithographically defined photonic and electronic components and devices (e.g. lasers, detectors, waveguides passive structures, modulators, electronic control and optical interconnects) on a single platform with nanometer-scale feature sizes. The development of photonic integrated circuits permits size, weight, power and cost reductions for spacecraft microprocessors, optical communication, processor buses, advanced data processing, and integrated optic science instrument optical systems, subsystems and components. This is particularly critical for small spacecraft platforms. We will give an overview of some NASA applications for integrated photonics.

  20. Integration of generic issues

    International Nuclear Information System (INIS)

    Thatcher, D.

    1989-01-01

    The NRC has recognized the need to integrate generic issues (GIs). The GI process includes a number of phases, all of which should recognize the potential for overlap and conflict among related issues. In addition to the issues themselves, other related NRC and industry programs and activities need to be factored into the GI process. Integration has taken place, or is taking place, for a number of GIs. Each case of integration involves a specific set of circumstances and, as a result, the way in which integration proceeds can vary. This paper discusses the integration of issues in the generic issue process and provides a number of examples

  1. Nature-based integration

    DEFF Research Database (Denmark)

    Pitkänen, Kati; Oratuomi, Joose; Hellgren, Daniela

    Increased attention to, and careful planning of the integration of migrants into Nordic societies is ever more important. Nature based integration is a new solution to respond to this need. This report presents the results of a Nordic survey and workshop and illustrates current practices of nature...... based integration by case study descriptions from Denmark, Sweden Norway and Finland. Across Nordic countries several practical projects and initiatives have been launched to promote the benefits of nature in integration and there is also growing academic interest in the topic. Nordic countries have...... the potential of becoming real forerunners in nature based integration even at the global scale....

  2. Entrepreneurial Integration Skills

    DEFF Research Database (Denmark)

    Bauer, Florian; Schriber, Svante; King, David R.

    2016-01-01

    Acquisition integration is important to realize synergies and to achieve acquisition success. However, there is a lack of clarity on pertinent integration approaches suggesting that integration is more complex and dynamic than traditionally assumed. In this chapter, we shed light on ambiguous cause...... effect relationships by investigating the effect of integration related decisions on intermediate goals. Additionally, we argue that entrepreneurial integration skills, or proactivity under ambiguity, are needed to keep pace with the dynamism inherent in acquisition integration. Based on primary data...... on 116 acquisitions, we find that entrepreneurial integration skills can display both advantages and disadvantages. While it helps to realize expected and serendipitous synergies, it can also trigger employee uncertainty due to decreased transparency. In supplementary analysis, we show measures...

  3. Searching for integrable systems

    International Nuclear Information System (INIS)

    Cary, J.R.

    1984-01-01

    Lack of integrability leads to undesirable consequences in a number of physical systems. The lack of integrability of the magnetic field leads to enhanced particle transport in stellarators and tokamaks with tearing-mode turbulence. Limitations of the luminosity of colliding beams may be due to the onset of stochasticity. Enhanced radial transport in mirror machines caused by the lack of integrability and/or the presence of resonances may be a significant problem in future devices. To improve such systems one needs a systematic method for finding integrable systems. Of course, it is easy to find integrable systems if no restrictions are imposed; textbooks are full of such examples. The problem is to find integrable systems given a set of constraints. An example of this type of problem is that of finding integrable vacuum magnetic fields with rotational transform. The solution to this problem is relevant to the magnetic-confinement program

  4. Immigrant Integration: Acculturation and Social Integration

    Directory of Open Access Journals (Sweden)

    Astrid HAMBERGER

    2009-11-01

    Full Text Available This article tackles the concept of “immigrant integration” as it is analyzed by different authors in the international migration field. In this article, I will use the terms “refugee” and “immigrant” as equivalent to each other due to the interchangeable character of these concepts throughout the integration literature. First, the article brings into discussion the definitional and conceptual battle around the concept of immigrant “integration”, and second, it will describe and analyze cultural and social integration with their presupposing processes.

  5. Integrated management systems

    CERN Document Server

    Bugdol, Marek

    2015-01-01

    Examining the challenges of integrated management, this book explores the importance and potential benefits of using an integrated approach as a cross-functional concept of management. It covers not only standardized management systems (e.g. International Organization for Standardization), but also models of self-assessment, as well as different types of integration. Furthermore, it demonstrates how processes and systems can be integrated, and how management efficiency can be increased. The major part of this book focuses on management concepts which use integration as a key tool of management processes (e.g. the systematic approach, supply chain management, virtual and network organizations, processes management and total quality management). Case studies, illustrations, and tables are also provided to exemplify and illuminate the content, as well as examples of successful and failed integrations. Providing a particularly useful resource to managers and specialists involved in the improvement of organization...

  6. Integration of reusable systems

    CERN Document Server

    Rubin, Stuart

    2014-01-01

    Software reuse and integration has been described as the process of creating software systems from existing software rather than building software systems from scratch. Whereas reuse solely deals with the artifacts creation, integration focuses on how reusable artifacts interact with the already existing parts of the specified transformation. Currently, most reuse research focuses on creating and integrating adaptable components at development or at compile time. However, with the emergence of ubiquitous computing, reuse technologies that can support adaptation and reconfiguration of architectures and components at runtime are in demand. This edited book includes 15 high quality research papers written by experts in information reuse and integration to cover the most recent advances in the field. These papers are extended versions of the best papers which were presented at IEEE International Conference on Information Reuse and Integration and IEEE International Workshop on Formal Methods Integration, which wa...

  7. What constitutes information integrity?

    Directory of Open Access Journals (Sweden)

    S. Flowerday

    2007-12-01

    Full Text Available This research focused on what constitutes information integrity as this is a problem facing companies today. Moreover, information integrity is a pillar of information security and is required in order to have a sound security management programme. However, it is acknowledged that 100% information integrity is not currently achievable due to various limitations and therefore the auditing concept of reasonable assurance is adopted. This is in line with the concept that 100% information security is not achievable and the notion that adequate security is the goal, using appropriate countermeasures. The main contribution of this article is to illustrate the importance of and provide a macro view of what constitutes information integrity. The findings are in harmony with Samuel Johnson's words (1751: 'Integrity without knowledge is weak and useless, and knowledge without integrity is dangerous and dreadful.'

  8. What constitutes information integrity?

    Directory of Open Access Journals (Sweden)

    S. Flowerday

    2008-01-01

    Full Text Available This research focused on what constitutes information integrity as this is a problem facing companies today. Moreover, information integrity is a pillar of information security and is required in order to have a sound security management programme. However, it is acknowledged that 100% information integrity is not currently achievable due to various limitations and therefore the auditing concept of reasonable assurance is adopted. This is in line with the concept that 100% information security is not achievable and the notion that adequate security is the goal, using appropriate countermeasures. The main contribution of this article is to illustrate the importance of and provide a macro view of what constitutes information integrity. The findings are in harmony with Samuel Johnson's words (1751: 'Integrity without knowledge is weak and useless, and knowledge without integrity is dangerous and dreadful.'

  9. Integrated management systems

    DEFF Research Database (Denmark)

    Jørgensen, Tine Herreborg; Remmen, Arne; Mellado, M. Dolores

    2006-01-01

    of integration. The focus of the article is primarily to discuss three ambition levels of integration: from increased compatibility of system elements over coordination of generic processes to an embeddedness of an integrated management system (IMS) in a culture of learning and continuous improvements......Different approaches to integration of management systems (ISO 9001, ISO 14001, OHSAS 18001 and SA 8000) with various levels of ambition have emerged. The tendency of increased compatibility between these standards has paved the road for discussions of, how to understand the different aspects....... At present, national IMS standards are being developed, and the IMS standards in Denmark and Spain are being analysed regarding the ambition level for integration. Should the ISO organisation decide to make a standard for IMS, then it would be necessary to consider the different levels of integration...

  10. Classical Mechanics and Symplectic Integration

    DEFF Research Database (Denmark)

    Nordkvist, Nikolaj; Hjorth, Poul G.

    2005-01-01

    Content: Classical mechanics: Calculus of variations, Lagrange’s equations, Symmetries and Noether’s theorem, Hamilton’s equations, cannonical transformations, integrable systems, pertubation theory. Symplectic integration: Numerical integrators, symplectic integrators, main theorem on symplectic...

  11. Integration i Danmark

    OpenAIRE

    Mikkelsen, Caroline; Olsen, Kristine Falk; Weng, Millie Cammilla; Andersen, Stine Bjørnstrup

    2012-01-01

    We found interest in this project, when we through the media discovered that there is a lot of prejudicesregardingimmigrants in Denmark. We saw a problem in these prejudices and wanted to get an insight in howthese prejudicesaffect the integration in Denmark, which led us to following question of reach: What are the problems about the Danish integration in relation to stigma and what part do the conquests play in the debate, regarding assimilation or integration in the Danish society. Our ...

  12. Integrated Sensitivity Analysis Workflow

    Energy Technology Data Exchange (ETDEWEB)

    Friedman-Hill, Ernest J. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Hoffman, Edward L. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Gibson, Marcus J. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Clay, Robert L. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States)

    2014-08-01

    Sensitivity analysis is a crucial element of rigorous engineering analysis, but performing such an analysis on a complex model is difficult and time consuming. The mission of the DART Workbench team at Sandia National Laboratories is to lower the barriers to adoption of advanced analysis tools through software integration. The integrated environment guides the engineer in the use of these integrated tools and greatly reduces the cycle time for engineering analysis.

  13. Branching trajectory continual integral

    International Nuclear Information System (INIS)

    Maslov, V.P.; Chebotarev, A.M.

    1980-01-01

    Heuristic definition of the Feynman continual integral over branching trajectories is suggested which makes it possible to obtain in the closed form the solution of the Cauchy problem for the model Hartree equation. A number of properties of the solution is derived from an integral representation. In particular, the quasiclassical asymptotics, exact solution in the gaussian case and perturbation theory series are described. The existence theorem for the simpliest continual integral over branching trajectories is proved [ru

  14. An integral heuristic

    International Nuclear Information System (INIS)

    Kauffman, L.H.

    1990-01-01

    This paper gives a heuristic derivation of the skein relation for the Homfly polynomial in an integral formalism. The derivation is formally correct but highly simplified. In the light of Witten's proposal for invariants of links via functional integrals, it is useful to have a formal pattern to compare with the complexities of the full approach. The formalism is a heuristic. However, it is closely related to the actual structure of the Witten functional integral

  15. The integrated PWR

    International Nuclear Information System (INIS)

    Gautier, G.M.

    2002-01-01

    This document presents the integrated reactors concepts by a presentation of four reactors: PIUS, SIR, IRIS and CAREM. The core conception, the operating, the safety, the economical aspects and the possible users are detailed. From the performance of the classical integrated PWR, the necessity of new innovative fuels utilization, the research of a simplified design to make easier the safety and the KWh cost decrease, a new integrated reactor is presented: SCAR 600. (A.L.B.)

  16. Modularly Integrated MEMS Technology

    National Research Council Canada - National Science Library

    Eyoum, Marie-Angie N

    2006-01-01

    Process design, development and integration to fabricate reliable MEMS devices on top of VLSI-CMOS electronics without damaging the underlying circuitry have been investigated throughout this dissertation...

  17. Handbook of integral equations

    CERN Document Server

    Polyanin, Andrei D

    2008-01-01

    This handbook contains over 2,500 integral equations with solutions as well as analytical and numerical methods for solving linear and nonlinear equations. It explores Volterra, Fredholm, WienerHopf, Hammerstein, Uryson, and other equations that arise in mathematics, physics, engineering, the sciences, and economics. This second edition includes new chapters on mixed multidimensional equations and methods of integral equations for ODEs and PDEs, along with over 400 new equations with exact solutions. With many examples added for illustrative purposes, it presents new material on Volterra, Fredholm, singular, hypersingular, dual, and nonlinear integral equations, integral transforms, and special functions.

  18. Avionics systems integration technology

    Science.gov (United States)

    Stech, George; Williams, James R.

    1988-01-01

    A very dramatic and continuing explosion in digital electronics technology has been taking place in the last decade. The prudent and timely application of this technology will provide Army aviation the capability to prevail against a numerically superior enemy threat. The Army and NASA have exploited this technology explosion in the development and application of avionics systems integration technology for new and future aviation systems. A few selected Army avionics integration technology base efforts are discussed. Also discussed is the Avionics Integration Research Laboratory (AIRLAB) that NASA has established at Langley for research into the integration and validation of avionics systems, and evaluation of advanced technology in a total systems context.

  19. Extending Database Integration Technology

    National Research Council Canada - National Science Library

    Buneman, Peter

    1999-01-01

    Formal approaches to the semantics of databases and database languages can have immediate and practical consequences in extending database integration technologies to include a vastly greater range...

  20. Integrated Information Management (IIM)

    National Research Council Canada - National Science Library

    McIlvain, Jason

    2007-01-01

    Information Technology is the core capability required to align our resources and increase our effectiveness on the battlefield by integrating and coordinating our preventative measures and responses...

  1. Building integrated photovoltaic; Photovaltaique integre aux batiments

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2004-01-01

    Durable, modular and flexible in use, as demonstrated by the different case studies in this publication, photovoltaic can replace diverse building elements, from glass facades to weather proof roofs. This leaflet towards architects describes aesthetic, technical, economic and environmental interest of building integrated photovoltaic. (author)

  2. Wealth Through Integration: Regional Integration and Poverty ...

    International Development Research Centre (IDRC) Digital Library (Canada)

    2013-02-27

    Feb 27, 2013 ... Could regional integration be a first step toward joining the global market? Since trade liberalization reduces revenues from duties and taxes, the available resources for public investments are negatively affected. The proposed solution encourages less-developed countries to emphasize regional ...

  3. Feynman integrals and hyperlogarithms

    International Nuclear Information System (INIS)

    Panzer, Erik

    2015-01-01

    We study Feynman integrals in the representation with Schwinger parameters and derive recursive integral formulas for massless 3- and 4-point functions. Properties of analytic (including dimensional) regularization are summarized and we prove that in the Euclidean region, each Feynman integral can be written as a linear combination of convergent Feynman integrals. This means that one can choose a basis of convergent master integrals and need not evaluate any divergent Feynman graph directly. Secondly we give a self-contained account of hyperlogarithms and explain in detail the algorithms needed for their application to the evaluation of multivariate integrals. We define a new method to track singularities of such integrals and present a computer program that implements the integration method. As our main result, we prove the existence of infinite families of massless 3- and 4-point graphs (including the ladder box graphs with arbitrary loop number and their minors) whose Feynman integrals can be expressed in terms of multiple polylogarithms, to all orders in the ε-expansion. These integrals can be computed effectively with the presented program. We include interesting examples of explicit results for Feynman integrals with up to 6 loops. In particular we present the first exactly computed counterterm in massless φ 4 theory which is not a multiple zeta value, but a linear combination of multiple polylogarithms at primitive sixth roots of unity (and divided by the √(3)). To this end we derive a parity result on the reducibility of the real- and imaginary parts of such numbers into products and terms of lower depth.

  4. Factorization of Behavioral Integrity

    DEFF Research Database (Denmark)

    Li, Ximeng; Nielson, Flemming; Nielson, Hanne Riis

    2015-01-01

    We develop a bisimulation-based nonintereference property that describes the allowed dependencies between communication behaviors of different integrity levels. The property is able to capture all possible combinations of integrity levels for the “presence” and “content” of actual communications...

  5. Fuzzy measures and integrals

    Czech Academy of Sciences Publication Activity Database

    Mesiar, Radko

    2005-01-01

    Roč. 28, č. 156 (2005), s. 365-370 ISSN 0165-0114 R&D Projects: GA ČR(CZ) GA402/04/1026 Institutional research plan: CEZ:AV0Z10750506 Keywords : fuzzy measures * fuzzy integral * regular fuzzy integral Subject RIV: BA - General Mathematics Impact factor: 1.039, year: 2005

  6. Hermeneutics of Integrative Knowledge.

    Science.gov (United States)

    Shin, Un-chol

    This paper examines and compares the formation processes and structures of three types of integrative knowledge that in general represent natural sciences, social sciences, and humanities. These three types can be observed, respectively, in the philosophies of Michael Polanyi, Jurgen Habermas, and Paul Ricoeur. These types of integrative knowledge…

  7. Ramjets: Airframe Integration

    Science.gov (United States)

    2010-09-01

    Thermal Management (Propulsion a vitesse elevee : Conception du moteur - integration et gestion thermique ) 14. ABSTRACT These notes deal with the...analogy for heat transfer from the outer flow to the conical intake shell via the Ramjets: Airframe Integration 6 - 24 RTO-EN-AVT-185 boundary

  8. Integrated Marketing Communications

    Science.gov (United States)

    Black, Jim

    2004-01-01

    Integration has become a cliche in enrollment management and student services circles. The term is used to describe everything from integrated marketing to seamless services. Often, it defines organizational structures, processes, student information systems, and even communities. In Robert Sevier's article in this issue of "College and…

  9. Integration and Polar Coordinates

    Indian Academy of Sciences (India)

    Jacobian matrix, Riemann- integrable function, co-area for- mula, Lipschitzfunction, levelset. S Kesavan. A simple proof of the formula for the integration of radial functions on R", N > 2, is given. As an application, the volume of the N-dimensional unit ball is computed. 1. Introduction. The method of substitution, also known as ...

  10. Integration Report 2007

    NARCIS (Netherlands)

    Jaco Dagevos; Mérove Gijsberts

    2007-01-01

    Original title: Jaarrapport integratie 2007. The Annual Report on Integration 2007 describes the integration of non-Western ethnic minorities in the Netherlands using a wide range of indicators. It focuses among other things on the situation of ethnic minority pupils in education, the

  11. On an integral transform

    Directory of Open Access Journals (Sweden)

    D. Naylor

    1986-01-01

    Full Text Available This paper establishes properties of a convolution type integral transform whose kernel is a Macdonald type Bessel function of zero order. An inversion formula is developed and the transform is applied to obtain the solution of some related integral equations.

  12. Integral fast reactor concept

    International Nuclear Information System (INIS)

    Chang, Y.I.; Marchaterre, J.F.; Sevy, R.H.

    1984-01-01

    Key features of the IFR consist of a pool-type plant arrangement, a metal fuel-based core design, and an integral fuel cycle with colocated fuel cycle facility. Both the basic concept and the technology base have been demonstrated through actual integral cycle operation in EBR-II. This paper discusses the inherent safety characteristics of the IFR concept

  13. The Fresnel Integrals Revisited

    Science.gov (United States)

    Chen, Hongwei

    2009-01-01

    This note presents another elementary method to evaluate the Fresnel integrals. It is interesting to see that this technique is also strong enough to capture a number of pairs of parameter integrals. The main ingredients of the method are the consideration of some related derivatives and linear differential equations.

  14. Hamiltonian path integrals

    International Nuclear Information System (INIS)

    Prokhorov, L.V.

    1982-01-01

    The properties of path integrals associated with the allowance for nonstandard terms reflecting the operator nature of the canonical variables are considered. Rules for treating such terms (''equivalence rules'') are formulated. Problems with a boundary, the behavior of path integrals under canonical transformations, and the problem of quantization of dynamical systems with constraints are considered in the framework of the method

  15. Integrity, Concept of

    DEFF Research Database (Denmark)

    Rendtorff, Jacob Dahl

    2015-01-01

    Integrity can be defined in the following sense: Integrity accounts for the inviolability of the human being. Although originally a virtue of uncorrupted character, expressing uprightness, honesty, and good intentions, it has, like dignity, been universalized as a quality of the person as such. T...

  16. Integral transformational coaching

    NARCIS (Netherlands)

    Keizer, W.A.J.; Nandram, S.S.

    2009-01-01

    In Chap. 12, Keizer and Nandram present the concept of Integral Transformational Coaching based on the concept of Flow and its effects on work performance. Integral Transformational Coaching is a method that prevents and cures unhealthy stress and burnout. They draw on some tried and tested

  17. Integration i flere variable

    DEFF Research Database (Denmark)

    Markvorsen, Steen

    2010-01-01

    -integralerne. Undervejs introduceres \\texttt{Integrator8}. Det er en pakke med Maple procedurer, som er udviklet specielt med henblik på eksempelbaseret visuel læring af de indledende integrationsbegreber og deres mangfoldige anvendelser. Vi giver eksempler på, hvordan integration i flere variable anvendes til beregning...

  18. Musik, Sprog og Integration

    DEFF Research Database (Denmark)

    Holst, Finn

    2010-01-01

    Evaluering af projektet Musik, Sprog og Integration, Søndermarkskolen, Horsens. Andenårsevaluering af et treårigt projekt.......Evaluering af projektet Musik, Sprog og Integration, Søndermarkskolen, Horsens. Andenårsevaluering af et treårigt projekt....

  19. Kanonisk sikkerhed og integration

    DEFF Research Database (Denmark)

    Larsen, Vibe; Øland, Trine

    2006-01-01

    Artiklen diskuterer, med indragelse af den lignende debat i andre europæiske lande, kulturkanonen i forhold til integration......Artiklen diskuterer, med indragelse af den lignende debat i andre europæiske lande, kulturkanonen i forhold til integration...

  20. Does energy integrate?

    Energy Technology Data Exchange (ETDEWEB)

    Hira, A.; Amaya, L. [Simon Fraser University, Burnaby, BC (Canada). Dept. of Political Science

    2003-01-01

    Amidst the international movement to privatize and deregulate electricity and gas sectors of economics, the question of the integration of those sectors has been somewhat underestimated. In fact, the integration of energy markets across boundaries is occurring. We examine this process in three regions: Europe, Central America, and South America. We analyze the forces driving integration in each area, and estimate the prospects for progress. We take a close look at Nordpool, which is now the most integrated market in the world, to see if it can serve as a model for other regions. We close with a set of conditions that we suggest are necessary for a successful international integration of energy markets. (author)

  1. Does energy integrate?

    International Nuclear Information System (INIS)

    Hira, Anil; Amaya, Libardo

    2003-01-01

    Amidst the international movement to privatize and deregulate electricity and gas sectors of economies, the question of the integration of those sectors has been somewhat underestimated. In fact, the integration of energy markets across boundaries is occurring. We examine this process in three regions: Europe, Central America, and South America. We analyze the forces driving integration in each area, and estimate the prospects for progress. We take a close look at Nordpool, which is now the most integrated market in the world, to see if it can serve as a model for other regions. We close with a set of conditions that we suggest are necessary for a successful international integration of energy markets

  2. Laboring for integration

    DEFF Research Database (Denmark)

    Fogelman, Tatiana

    Over the past fifteen years the district of Berlin-Marzahn became home to the largest concentration of post-Soviet migrants of German ancestry in the former East Germany. Drawing on my research in the district, including two of its integration projects for middle-aged, unemployed migrants, I...... examine these projects’ failed attempts to create spaces of encounter between the migrants and district’s long-term residents. In this presentation I focus specifically on the structural position of integration work (Integrationsarbeit) that creates first and foremost conditions of possibility for its...... temporal imaginaries underpinning the ideas of integration through encounter, as well as integration more broadly, I argue for reflexivity-centric public reformulations of both integration and encounter....

  3. Personal Integrity and Leadership

    Directory of Open Access Journals (Sweden)

    Antonius Gea

    2016-07-01

    Full Text Available Generally, integrity is associated with leadership, especially in the organization or company. Integrity in leadership becomes a growing concern in business and organizations. The aim of this study was to find out the connection that could be shaped between personal integrity and the skill of leadership especially in the performance of work. This study used a library research, a literature study that done by using the available resources and relevant literature, related to the topic being discussed by the author. Books or journal articles related to topics were used as background reading to understand well about the problems of integrity and leadership, especially in the organization or company. This study finds out that the leader integrity is related to follower work role performance and that this effect is fully mediated through follower affective organizational commitment. 

  4. Neuroeconomics and Integrated Care

    DEFF Research Database (Denmark)

    Larsen, Torben

    2012-01-01

    Background: Fragmented specialized care for the frail elderly as claimed by WHO needs horizontal integration across settings. The home of the patient seems to be a promising place to integrate hospital care, primary care and social services for high-risk discharges where the quality...... of rehabilitation makes a difference. Objective: The study aims to reveal how integrated home care may be organised to improve quality of care as compared to usual hospital care. Method: A qualitative case study of the use of a neuroeconomic model in relation to multidisciplianry collaboration on a RCT...... of integrated home care for stroke patients. Results: (1) The classical understanding of CNS is that of a dual system of ANS and Cortex. The new neuroeconomic understanding is that of a reciprocal balance of Limbic System (LS) and Neocortex (NC). This applies directly in favour of integrated homecare compared...

  5. Retroviral integration: Site matters

    Science.gov (United States)

    Demeulemeester, Jonas; De Rijck, Jan

    2015-01-01

    Here, we review genomic target site selection during retroviral integration as a multistep process in which specific biases are introduced at each level. The first asymmetries are introduced when the virus takes a specific route into the nucleus. Next, by co‐opting distinct host cofactors, the integration machinery is guided to particular chromatin contexts. As the viral integrase captures a local target nucleosome, specific contacts introduce fine‐grained biases in the integration site distribution. In vivo, the established population of proviruses is subject to both positive and negative selection, thereby continuously reshaping the integration site distribution. By affecting stochastic proviral expression as well as the mutagenic potential of the virus, integration site choice may be an inherent part of the evolutionary strategies used by different retroviruses to maximise reproductive success. PMID:26293289

  6. Vertcal integration: origins of oil industry integration

    International Nuclear Information System (INIS)

    Rainbow, R.

    2001-01-01

    This article examines the structure of the US oil industry in relation to the history of the industry, capital market assumptions, and advice on the enhancement of 'shareholder value'. The driving forces behind the attempts to establish cartels and vertical integration are considered, and the emergence of US companies resulting from the break-up of the Standard Oil Trust cartel, the influence of costs and logistics on the structure of the industry outside the USA , and the structure of the natural gas industry are discussed along with the discovery of large Middle East oil reserves, the enormous growth in demand for oil products in Europe and Japan, and the formation of the OPEC cartel. Details are given of the breaking down of vertical integration in the big oil companies, the theory of conglomerates, the success of big companies, the importance of scale to balance exploration risks, and the need to adjust in time to shifts in the business environment

  7. Long Pulse Integrator of Variable Integral Time Constant

    International Nuclear Information System (INIS)

    Wang Yong; Ji Zhenshan; Du Xiaoying; Wu Yichun; Li Shi; Luo Jiarong

    2010-01-01

    A kind of new long pulse integrator was designed based on the method of variable integral time constant and deducting integral drift by drift slope. The integral time constant can be changed by choosing different integral resistors, in order to improve the signal-to-noise ratio, and avoid output saturation; the slope of integral drift of a certain period of time can be calculated by digital signal processing, which can be used to deduct the drift of original integral signal in real time to reduce the integral drift. The tests show that this kind of long pulse integrator is good at reducing integral drift, which also can eliminate the effects of changing integral time constant. According to experiments, the integral time constant can be changed by remote control and manual adjustment of integral drift is avoided, which can improve the experiment efficiency greatly and can be used for electromagnetic measurement in Tokamak experiment. (authors)

  8. Integrated assessment briefs

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1995-04-01

    Integrated assessment can be used to evaluate and clarify resource management policy options and outcomes for decision makers. The defining characteristics of integrated assessment are (1) focus on providing information and analysis that can be understood and used by decision makers rather than for merely advancing understanding and (2) its multidisciplinary approach, using methods, styles of study, and considerations from a broader variety of technical areas than would typically characterize studies produced from a single disciplinary standpoint. Integrated assessment may combine scientific, social, economic, health, and environmental data and models. Integrated assessment requires bridging the gap between science and policy considerations. Because not everything can be valued using a single metric, such as a dollar value, the integrated assessment process also involves evaluating trade-offs among dissimilar attributes. Scientists at Oak Ridge National Laboratory (ORNL) recognized the importance and value of multidisciplinary approaches to solving environmental problems early on and have pioneered the development of tools and methods for integrated assessment over the past three decades. Major examples of ORNL`s experience in the development of its capabilities for integrated assessment are given.

  9. Three dimensional system integration

    CERN Document Server

    Papanikolaou, Antonis; Radojcic, Riko

    2010-01-01

    Three-dimensional (3D) integrated circuit (IC) stacking is the next big step in electronic system integration. It enables packing more functionality, as well as integration of heterogeneous materials, devices, and signals, in the same space (volume). This results in consumer electronics (e.g., mobile, handheld devices) which can run more powerful applications, such as full-length movies and 3D games, with longer battery life. This technology is so promising that it is expected to be a mainstream technology a few years from now, less than 10-15 years from its original conception. To achieve thi

  10. In times of Integration

    DEFF Research Database (Denmark)

    Val, Maria Rosa Rovira; Lehmann, Martin; Zinenko, Anna

    dramatically with many of the world’s economies facing downturn and a looming possible recession; and the global economic and political balance changing; (ii) most larger companies and quite a few SMEs now have a mature knowledge of these standards; and (iii) some standards are advocating for integration...... procedures, such as cases of for example ISO integrated management systems, mutual equivalences recognition of Global Compact-GRI-ISO26000, or the case of IIRC initiative to develop integrated reporting on an organization’s Financial, Environmental, Social and Governance performance. This paper focuses...

  11. SQL Server Integration Services

    CERN Document Server

    Hamilton, Bill

    2007-01-01

    SQL Server 2005 Integration Services (SSIS) lets you build high-performance data integration solutions. SSIS solutions wrap sophisticated workflows around tasks that extract, transform, and load (ETL) data from and to a wide variety of data sources. This Short Cut begins with an overview of key SSIS concepts, capabilities, standard workflow and ETL elements, the development environment, execution, deployment, and migration from Data Transformation Services (DTS). Next, you'll see how to apply the concepts you've learned through hands-on examples of common integration scenarios. Once you've

  12. Social integration i arbejdslivet

    DEFF Research Database (Denmark)

    Nielsen, Kjeld; Christensen, Allan

    2002-01-01

    of the study that social integration depends on working environment conditions as well as cultural conditions. The social integration again is crucial for the way in which employees perceive the daily work situation and indirectly also for the well-being of the clients of the institution.......The study reported in this article is based on a combination of methods related to working environment and to organisational culture. The combination is aimed at studying the extent and meaning of the social integration of the employees in an institution for handicapped adults. It is the conclusion...

  13. Improper Riemann integrals

    CERN Document Server

    Roussos, Ioannis Markos

    2013-01-01

    Improper Riemann Integrals is the first book to collect classical and modern material on the subject for undergraduate students. The book gives students the prerequisites and tools to understand the convergence, principal value, and evaluation of the improper/generalized Riemann integral. It also illustrates applications to science and engineering problems.The book contains the necessary background, theorems, and tools, along with two lists of the most important integrals and sums computed in the text. Numerous examples at various levels of difficulty illustrate the concepts and theorems. The

  14. The Structural Integrity Centre

    International Nuclear Information System (INIS)

    Tomkins, B.

    1987-01-01

    The paper concerns the development and work of the Structural Integrity Centre (SIC) at Risley Nuclear Laboratories, United Kingdom. The centre was set up to provide authoritative advice to plant designers and operators on the integrity and life assessment of structures and components across the reactor projects in the United Kingdom. A description is given of the structure and role of the SIC, as well as the Structural Integrity Assessment work. The assessment methods are described for thermally loaded structures and welded structures. Finally, defect significance assessment and environmental effects are outlined. (U.K.)

  15. Principles of data integration

    CERN Document Server

    Doan, AnHai; Ives, Zachary

    2012-01-01

    How do you approach answering queries when your data is stored in multiple databases that were designed independently by different people? This is first comprehensive book on data integration and is written by three of the most respected experts in the field. This book provides an extensive introduction to the theory and concepts underlying today's data integration techniques, with detailed, instruction for their application using concrete examples throughout to explain the concepts. Data integration is the problem of answering queries that span multiple data sources (e.g., databases, web

  16. MOS integrated circuit design

    CERN Document Server

    Wolfendale, E

    2013-01-01

    MOS Integral Circuit Design aims to help in the design of integrated circuits, especially large-scale ones, using MOS Technology through teaching of techniques, practical applications, and examples. The book covers topics such as design equation and process parameters; MOS static and dynamic circuits; logic design techniques, system partitioning, and layout techniques. Also featured are computer aids such as logic simulation and mask layout, as well as examples on simple MOS design. The text is recommended for electrical engineers who would like to know how to use MOS for integral circuit desi

  17. Path integration quantization

    International Nuclear Information System (INIS)

    DeWitt-Morette, C.

    1983-01-01

    Much is expected of path integration as a quantization procedure. Much more is possible if one recognizes that path integration is at the crossroad of stochastic and differential calculus and uses the full power of both stochastic and differential calculus in setting up and computing path integrals. In contrast to differential calculus, stochastic calculus has only comparatively recently become an instrument of thought. It has nevertheless already been used in a variety of challenging problems, for instance in the quantization problem. The author presents some applications of the stochastic scheme. (Auth.)

  18. GMRES and integral operators

    Energy Technology Data Exchange (ETDEWEB)

    Kelley, C.T.; Xue, Z.Q. [North Carolina State Univ., Raleigh, NC (United States)

    1994-12-31

    Many discretizations of integral equations and compact fixed point problems are collectively compact and strongly convergent in spaces of continuous functions. These properties not only lead to stable and convergent approximations but also can be used in the construction of fast multilevel algorithms. Recently the GMRES algorithm has become a standard coarse mesh solver. The purpose of this paper is to show how the special properties of integral operators and their approximations are reflected in the performance of the GMRES iteration and how these properties can be used to strengthen the norm in which convergence takes place. The authors illustrate these ideas with composite Gauss rules for integral equations on the unit interval.

  19. Hamiltonian path integrals

    International Nuclear Information System (INIS)

    Prokhorov, L.V.

    1982-01-01

    Problems related to consideration of operator nonpermutability in Hamiltonian path integral (HPI) are considered in the review. Integrals are investigated using trajectories in configuration space (nonrelativistic quantum mechanics). Problems related to trajectory integrals in HPI phase space are discussed: the problem of operator nonpermutability consideration (extra terms problem) and corresponding equivalence rules; ambiguity of HPI usual recording; transition to curvilinear coordinates. Problem of quantization of dynamical systems with couplings has been studied. As in the case of canonical transformations, quantization of the systems with couplings of the first kind requires the consideration of extra terms

  20. Introduction to integral discriminants

    International Nuclear Information System (INIS)

    Morozov, A.; Shakirov, Sh.

    2009-01-01

    The simplest partition function, associated with homogeneous symmetric forms S of degree r in n variables, is integral discriminant J n|r (S) = ∫e -S(x 1 ,...,x n ) dx 1 ...dx n . Actually, S-dependence remains the same if e -S in the integrand is substituted by arbitrary function f(S), i.e. integral discriminant is a characteristic of the form S itself, and not of the averaging procedure. The aim of the present paper is to calculate J n|r in a number of non-Gaussian cases. Using Ward identities - linear differential equations, satisfied by integral discriminants - we calculate J 2|3 ,J 2|4 ,J 2|5 and J 3|3 . In all these examples, integral discriminant appears to be a generalized hypergeometric function. It depends on several SL(n) invariants of S, with essential singularities controlled by the ordinary algebraic discriminant of S.

  1. Financial Integrity Benchmarks

    Data.gov (United States)

    City of Jackson, Mississippi — This data compiles standard financial integrity benchmarks that allow the City to measure its financial standing. It measure the City's debt ratio and bond ratings....

  2. Immigration und Integration

    OpenAIRE

    Rauscher, Anton

    2003-01-01

    Immigration und Integration : eine Herausforderung für Kirche, Gesellschaft und Politik in Deutschland und den USA / hrsg. von Anton Rauscher. - Berlin : Duncker & Humblot, 2003. - 174 S. - (Soziale Orientierung ; 15)

  3. Integrated Health Management Definitions

    Data.gov (United States)

    National Aeronautics and Space Administration — The Joint Army Navy NASA Air Force Modeling and Simulation Subcommittee's Integrated Health Management panel was started about 6 years ago to help foster...

  4. Integrated Disposal Facility

    Data.gov (United States)

    Federal Laboratory Consortium — Located near the center of the 586-square-mile Hanford Site is the Integrated Disposal Facility, also known as the IDF.This facility is a landfill similar in concept...

  5. Measuring integrated care

    DEFF Research Database (Denmark)

    Strandberg-Larsen, Martin

    2011-01-01

    differences were found in the perception of clinical integration in the two settings. More primary care clinicians in the Northern California region of Kaiser Permanente reported being part of a clinical integrated environment than did Danish general practitioners. By measuring the level of clinical...... respond to these needs, patients and providers face the multiple challenges of today's healthcare environment. Decision makers, planners and managers need evidence based policy options and information on the scope of the integrated care challenges they are facing. The US managed care organization Kaiser...... Permanente has been put forward as an example for European healthcare systems to follow, although the evidence base is far from conclusive. The thesis has five objectives: 1) To contribute to the understanding of the concept of integration in healthcare systems and to identify measurement methods to capture...

  6. Integrating Forensic Science.

    Science.gov (United States)

    Funkhouser, John; Deslich, Barbara J.

    2000-01-01

    Explains the implementation of forensic science in an integrated curriculum and discusses the advantages of this approach. Lists the forensic science course syllabi studied in three high schools. Discusses the unit on polymers in detail. (YDS)

  7. State Program Integrity Reviews

    Data.gov (United States)

    U.S. Department of Health & Human Services — State program integrity reviews play a critical role in how CMS provides effective support and assistance to states in their efforts to combat provider fraud and...

  8. Advances in integrated optics

    CERN Document Server

    Chester, A; Bertolotti, M

    1994-01-01

    This volwne contains the Proceedings of a two-week summer conference titled "Advances in Integrated Optics" held June 1-9, 1993, in Erice, Sicily. This was the 18th annual course organized by the International School of Quantum Electronics, under the auspices of the "Ettore Majorana" Centre for Scientific Culture. The term Integrated Optics signifies guided-wave optical circuits consisting of two or more devices on a single substrate. Since its inception in the late 1960's, Integrated Optics has evolved from a specialized research topic into a broad field of work, ranging from basic research through commercial applications. Today many devices are available on market while a big effort is devolved to research on integrated nonlinear optical devices. This conference was organized to provide a comprehensive survey of the frontiers of this technology, including fundamental concepts, nonlinear optical materials, devices both in the linear and nonlinear regimes, and selected applications. These Proceedings update a...

  9. Integrated Surface Dataset (Global)

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — The Integrated Surface (ISD) Dataset (ISD) is composed of worldwide surface weather observations from over 35,000 stations, though the best spatial coverage is...

  10. Integration, measure and probability

    CERN Document Server

    Pitt, H R

    2012-01-01

    Introductory treatment develops the theory of integration in a general context, making it applicable to other branches of analysis. More specialized topics include convergence theorems and random sequences and functions. 1963 edition.

  11. European Economic Integration

    Science.gov (United States)

    Huston, James A.

    1971-01-01

    Recounts the history and problems of European Economic Integration from the first post World War II organization, the OEEC, to the EEC (Common Market) and the EFTA. Suggestions for further reading are included. (JB)

  12. Integrals of Bessel functions

    OpenAIRE

    Babusci, D.; Dattoli, G.; Germano, B.; Martinelli, M. R.; Ricci, P. E.

    2011-01-01

    We use the operator method to evaluate a class of integrals involving Bessel or Bessel-type functions. The technique we propose is based on the formal reduction of these family of functions to Gaussians.

  13. Integrated inventory information system

    Digital Repository Service at National Institute of Oceanography (India)

    Sarupria, J.S.; Kunte, P.D.

    The nature of oceanographic data and the management of inventory level information are described in Integrated Inventory Information System (IIIS). It is shown how a ROSCOPO (report on observations/samples collected during oceanographic programme...

  14. Measuring integrated care

    DEFF Research Database (Denmark)

    Strandberg-Larsen, Martin

    2011-01-01

    respond to these needs, patients and providers face the multiple challenges of today's healthcare environment. Decision makers, planners and managers need evidence based policy options and information on the scope of the integrated care challenges they are facing. The US managed care organization Kaiser...... Permanente has been put forward as an example for European healthcare systems to follow, although the evidence base is far from conclusive. The thesis has five objectives: 1) To contribute to the understanding of the concept of integration in healthcare systems and to identify measurement methods to capture...... the multi-dimensional aspects of integrated healthcare delivery. 2) To assess the level of integration of the Danish healthcare system. 3) To assess the use of joint health plans as a tool for coordination between the regional and local level in the Danish healthcare system. 4) To compare the inputs...

  15. Integration a functional approach

    CERN Document Server

    Bichteler, Klaus

    1998-01-01

    This book covers Lebesgue integration and its generalizations from Daniell's point of view, modified by the use of seminorms. Integrating functions rather than measuring sets is posited as the main purpose of measure theory. From this point of view Lebesgue's integral can be had as a rather straightforward, even simplistic, extension of Riemann's integral; and its aims, definitions, and procedures can be motivated at an elementary level. The notion of measurability, for example, is suggested by Littlewood's observations rather than being conveyed authoritatively through definitions of (sigma)-algebras and good-cut-conditions, the latter of which are hard to justify and thus appear mysterious, even nettlesome, to the beginner. The approach taken provides the additional benefit of cutting the labor in half. The use of seminorms, ubiquitous in modern analysis, speeds things up even further. The book is intended for the reader who has some experience with proofs, a beginning graduate student for example. It might...

  16. Systems Integration Fact Sheet

    Energy Technology Data Exchange (ETDEWEB)

    None

    2016-06-01

    This fact sheet is an overview of the Systems Integration subprogram at the U.S. Department of Energy SunShot Initiative. The Systems Integration subprogram enables the widespread deployment of safe, reliable, and cost-effective solar energy technologies by addressing the associated technical and non-technical challenges. These include timely and cost-effective interconnection procedures, optimal system planning, accurate prediction of solar resources, monitoring and control of solar power, maintaining grid reliability and stability, and many more. To address the challenges associated with interconnecting and integrating hundreds of gigawatts of solar power onto the electricity grid, the Systems Integration program funds research, development, and demonstration projects in four broad, interrelated focus areas: grid performance and reliability, dispatchability, power electronics, and communications.

  17. The Executive as Integrator.

    Science.gov (United States)

    Cohn, Hans M.

    1983-01-01

    Argues that although the executive has many tasks, he or she must view internal organizational integration as a primary task, making use of organizational charts, job descriptions, statements of goals and objectives, evaluations, and feedback devices. (RH)

  18. On Kontsevich integrals

    International Nuclear Information System (INIS)

    Itzykson, C.

    1992-01-01

    Some mathematical results are described on a remarkable application of matrix interaction due to Witten and Kontsevich, to study topological invariants. Some interesting properties of the Kontsevich integrals are identified. (K.A.) 4 refs

  19. Pair Correlation Function Integrals

    DEFF Research Database (Denmark)

    Wedberg, Nils Hejle Rasmus Ingemar; O'Connell, John P.; Peters, Günther H.J.

    2011-01-01

    We describe a method for extending radial distribution functions obtained from molecular simulations of pure and mixed molecular fluids to arbitrary distances. The method allows total correlation function integrals to be reliably calculated from simulations of relatively small systems. The long-d...... distribution function has structure beyond the sampling limit imposed by the system size, the integration is more reliable, and usually more accurate, than simple integral truncation.......We describe a method for extending radial distribution functions obtained from molecular simulations of pure and mixed molecular fluids to arbitrary distances. The method allows total correlation function integrals to be reliably calculated from simulations of relatively small systems. The long......, and J. Abildskov, Mol. Simul. 36, 1243 (2010); Fluid Phase Equilib. 302, 32 (2011)], but describe here its theoretical basis more thoroughly and derive long-distance approximations for the direct correlation functions. We describe the numerical implementation of the method in detail, and report...

  20. Introduction to integral discriminants

    Science.gov (United States)

    Morozov, A.; Shakirov, Sh.

    2009-12-01

    The simplest partition function, associated with homogeneous symmetric forms S of degree r in n variables, is integral discriminant Jn|r(S) = ∫e-S(x1,...,xn)dx1...dxn. Actually, S-dependence remains the same if e-S in the integrand is substituted by arbitrary function f(S), i.e. integral discriminant is a characteristic of the form S itself, and not of the averaging procedure. The aim of the present paper is to calculate Jn|r in a number of non-Gaussian cases. Using Ward identities — linear differential equations, satisfied by integral discriminants — we calculate J2|3,J2|4,J2|5 and J3|3. In all these examples, integral discriminant appears to be a generalized hypergeometric function. It depends on several SL(n) invariants of S, with essential singularities controlled by the ordinary algebraic discriminant of S.

  1. Retroviral DNA Integration

    Science.gov (United States)

    2016-01-01

    The integration of a DNA copy of the viral RNA genome into host chromatin is the defining step of retroviral replication. This enzymatic process is catalyzed by the virus-encoded integrase protein, which is conserved among retroviruses and LTR-retrotransposons. Retroviral integration proceeds via two integrase activities: 3′-processing of the viral DNA ends, followed by the strand transfer of the processed ends into host cell chromosomal DNA. Herein we review the molecular mechanism of retroviral DNA integration, with an emphasis on reaction chemistries and architectures of the nucleoprotein complexes involved. We additionally discuss the latest advances on anti-integrase drug development for the treatment of AIDS and the utility of integrating retroviral vectors in gene therapy applications. PMID:27198982

  2. Complementary and Integrative Medicine

    Science.gov (United States)

    ... medical treatments that are not part of mainstream medicine. When you are using these types of care, it may be called complementary, integrative, or alternative medicine. Complementary medicine is used together with mainstream medical ...

  3. Integrated Reporting Information System -

    Data.gov (United States)

    Department of Transportation — The Integrated Reporting Information System (IRIS) is a flexible and scalable web-based system that supports post operational analysis and evaluation of the National...

  4. Integrating corpora and dictionaries

    DEFF Research Database (Denmark)

    Kristoffersen, Jette Hedegaard; Troelsgård, Thomas

    2012-01-01

    In this paper, we will discuss different possibilities for integration of corpus data with dictionary data, mainly seen from a lexicographic point of view and in a sign language context. For about 25 years a text corpus has been considered a useful, if not necessary tool for editing dictionaries...... of written and spoken languages. Corpora are equally useful to sign language lexicographers, but sign language corpora have not become accessible until recent years. Nowadays corpora exist, or are being developed, for several sign languages, and we have the possibility of editing new, truly corpus-based sign...... language dictionaries, and of developing interfaces that integrate corpus and dictionary data. After a brief look at three existing integrated interfaces, one for German, one for Danish, and one for Danish Sign Language, we point out some of the problems that should be considered when making an integrated...

  5. Penitentiary integral defense

    OpenAIRE

    Francisco Javier Rodríguez Román

    2012-01-01

    Penitentiary integral defense is a set of techniques designed to solve the breach of the peace that may appear in the penitentiary world. This paper studies the scope and legal framework of penitentiary self-defense. This kind of self-defense is defined by the particular characteristics of a confinement context and a very specific regulation aimed at preserving the integrity of prisoners and penitentiary staff as well as prison facilities. A technical section illustrating the main penitentiar...

  6. Integrated parasite management

    DEFF Research Database (Denmark)

    Clausen, Jesper Hedegaard; Madsen, Henry; Van, Phan Thi

    2015-01-01

    communities at risk through mass drug administration. However, we argue that treatment alone will not reduce the risk from eating infected fish and that sustainable effective control must adopt an integrated FZT control approach based on education, infrastructure improvements, and management practices...... that target critical control points in the aquaculture production cycle identified from a thorough understanding of FZT and host biology and epidemiology. We present recommendations for an integrated parasite management (IPM) program for aquaculture farms....

  7. IDC Integrated Master Plan.

    Energy Technology Data Exchange (ETDEWEB)

    Clifford, David J.; Harris, James M.

    2014-12-01

    This is the IDC Re-Engineering Phase 2 project Integrated Master Plan (IMP). The IMP presents the major accomplishments planned over time to re-engineer the IDC system. The IMP and the associate Integrated Master Schedule (IMS) are used for planning, scheduling, executing, and tracking the project technical work efforts. REVISIONS Version Date Author/Team Revision Description Authorized by V1.0 12/2014 IDC Re- engineering Project Team Initial delivery M. Harris

  8. Function integrated track system

    OpenAIRE

    Hohnecker, Eberhard

    2010-01-01

    The paper discusses a function integrated track system that focuses on the reduction of acoustic emissions from railway lines. It is shown that the combination of an embedded rail system (ERS), a sound absorbing track surface, and an integrated mini sound barrier has significant acoustic advantages compared to a standard ballast superstructure. The acoustic advantages of an embedded rail system are particularly pronounced in the case of railway bridges. Finally, it is shown that a...

  9. Writing against integration

    DEFF Research Database (Denmark)

    Rytter, Mikkel

    2018-01-01

    The article addresses some of the problems related to the concept of integration, which has been used (and abused) in Denmark since the 1980s to discuss socio-economic, cultural and religious challenges related to the everyday life of ethnic minorities. The concept of integration is not innocent,......-Lughod’s seminal article “writing against culture”, the article suggests strategies of “writing against integration” in order to regain the critical potential of academic analysis....

  10. Holomorphic matrix integrals

    International Nuclear Information System (INIS)

    Felder, Giovanni; Riser, Roman

    2004-01-01

    We study a class of holomorphic matrix models. The integrals are taken over middle-dimensional cycles in the space of complex square matrices. As the size of the matrices tends to infinity, the distribution of eigenvalues is given by a measure with support on a collection of arcs in the complex planes. We show that the arcs are level sets of the imaginary part of a hyperelliptic integral connecting branch points

  11. Integrated Electric Gas Turbine

    OpenAIRE

    Millsaps, Knox T.

    2010-01-01

    Patent An integrated electric gas turbine comprises a compressor that includes a plurality of airfoils. An electric motor is arranged to drive the compressor, and a combustor is arranged to receive compressed air from the compressor and further arranged to receive a fuel input. A turbine is arranged to receive the combustion gases from the combustor. A generator is integrated with the turbine and arranged to provide a power output. A controller is connected between the...

  12. Discrete pseudo-integrals

    Czech Academy of Sciences Publication Activity Database

    Mesiar, Radko; Li, J.; Pap, E.

    2013-01-01

    Roč. 54, č. 3 (2013), s. 357-364 ISSN 0888-613X R&D Projects: GA ČR GAP402/11/0378 Institutional support: RVO:67985556 Keywords : concave integral * pseudo-addition * pseudo- multiplication Subject RIV: BA - General Mathematics Impact factor: 1.977, year: 2013 http://library.utia.cas.cz/separaty/2013/E/mesiar-discrete pseudo-integrals. pdf

  13. Experimental integrated photovoltaic systems

    International Nuclear Information System (INIS)

    Pop-Jordanov, Jordan; Markovska, Natasha; Dimitrov, D.; Kocev, K.; Dimitrovski, D.

    2000-01-01

    Recently, the interest in building-integrated photovoltaic installations has started to increase within governmental and municipality authorities, as well as some industrial companies. To serve a national public-awareness program of solar electricity promotion and education, the indigenous solar energy potential, optimization of possible PV installation, and three test cases of building-integrated grid-connected experimental facilities have been studied. The results showed the feasibility and performance of the proposed concepts. (Original)

  14. Agroecology : integration with livestock

    OpenAIRE

    Tichit, Muriel; Lecompte, Philippe; Dumont, Bertrand

    2014-01-01

    Livestock systems are a large global asset contributing to food security and poverty alleviation, but livestock supply chains have major environmental impacts at global scale. The scientific literature on agroecology has not yet integrated livestock systems; only 5 percent of the indexed studies concerning agroecology deal with livestock. Following Dumont et al. (2013), we review five principles for integrating livestock systems within the agroecology debate: (i) adopting management practices...

  15. HETEROGENEOUS INTEGRATION TECHNOLOGY

    Science.gov (United States)

    2017-08-24

    9.1.1 FET- BT -HBT Integration........................................................................................... 22 9.1.2 Early GaAs-Si...Fabricated with Different Device Types on Separate Chips are Combined on the Same Chip in 2D SoC [26] 9.1.1 FET- BT -HBT Integration The semiconductor...electronics started with the use of bipolar transistors ( BTs ) but field effect transistors (FETs) were later introduced for digital applications. As

  16. Heterogeneous Integration Technology

    Science.gov (United States)

    2017-05-19

    This type of packaging is used in mobile products where because of space saving features and can be found in smart phones and tables. Figure 69...picture of 8-strata stacked chips with TSVs on interposer layer [128]. ............................. 59 Figure 72: The DARPA SMART program: (a...mounted die is integrated with another package containing stacked chips with bond wire connections [153]. PoP integration is also used heavily in smart

  17. Functional integration over geometries

    International Nuclear Information System (INIS)

    Mottola, E.

    1995-01-01

    The geometric construction of the functional integral over coset spaces M/G is reviewed. The inner product on the cotangent space of infinitesimal deformations of M defines an invariant distance and volume form, or functional integration measure on the full configuration space. Then, by a simple change of coordinates parameterizing the gauge fiber G, the functional measure on the coset space M/G is deduced. This change of integration variables leads to a Jacobian which is entirely equivalent to the Faddeev--Popov determinant of the more traditional gauge fixed approach in non-abelian gauge theory. If the general construction is applied to the case where G is the group of coordinate reparameterizations of spacetime, the continuum functional integral over geometries, i.e. metrics modulo coordinate reparameterizations may be defined. The invariant functional integration measure is used to derive the trace anomaly and effective action for the conformal part of the metric in two and four dimensional spacetime. In two dimensions this approach generates the Polyakov--Liouville action of closed bosonic non-critical string theory. In four dimensions the corresponding effective action leads to novel conclusions on the importance of quantum effects in gravity in the far infrared, and in particular, a dramatic modification of the classical Einstein theory at cosmological distance scales, signaled first by the quantum instability of classical de Sitter spacetime. Finite volume scaling relations for the functional integral of quantum gravity in two and four dimensions are derived, and comparison with the discretized dynamical triangulation approach to the integration over geometries are discussed. Outstanding unsolved problems in both the continuum definition and the simplicial approach to the functional integral over geometries are highlighted

  18. Integrated smart structures wingbox

    Science.gov (United States)

    Simon, Solomon H.

    1993-09-01

    One objective of smart structures development is to demonstrate the ability of a mechanical component to monitor its own structural integrity and health. Achievement of this objective requires the integration of different technologies, i.e.: (1) structures, (2) sensors, and (3) artificial intelligence. We coordinated a team of experts from these three fields. These experts used reliable knowledge towards the forefront of their technologies and combined the appropriate features into an integrated hardware/software smart structures wingbox (SSW) test article. A 1/4 in. hole was drilled into the SSW test article. Although the smart structure had never seen damage of this type, it correctly recognized and located the damage. Based on a knowledge-based simulation, quantification and assessment were also carried out. We have demonstrated that the SSW integrated hardware & software test article can perform six related functions: (1) identification of a defect; (2) location of the defect; (3) quantification of the amount of damage; (4) assessment of performance degradation; (5) continued monitoring in spite of damage; and (6) continuous recording of integrity data. We present the successful results of the integrated test article in this paper, along with plans for future development and deployment of the technology.

  19. APPROACHES TO PUBLIC INTEGRITY

    Directory of Open Access Journals (Sweden)

    Simona-Roxana ULMAN

    2015-08-01

    Full Text Available Both concepts, integrity and public integrity, are always asked qualities in the social and economic environment, but they are never well defined. In this context, finding precise definitions and offering an ample explanation of the concepts are desired and useful for both economic theoretical and practical levels and become the aim of the present paper. Without a good understanding of a concept, no one is capable to attain it. In this context, it is impossible to ask for a public sector of integrity if both public actors and the citizens do not know what it really means. Therefore, the public actors first have to understand what integrity really supposes, and, then, to be capable to respect its principles. Taking into consideration the effects of public integrity or, on its opposite, of the divergence from it, especially represented by the phenomenon of corruption, on the economy at the macro, but also, on the micro level, public integrity in itself becomes a real economic problem that is aimed to be extensively analyzed through our interdisciplinary approach.

  20. Fostering Integrity in Research

    Science.gov (United States)

    McNutt, M. K.

    2017-12-01

    The responsible conduct of research requires that all involved in the research enterprise - researchers, sponsors, reviewers, publishers, and communicators - adhere to a set of integrity principles to protect the public's investment. Given the increasing complexity and globalization of the research enterprise, the National Academies recently re-examined and updated its integrity recommendations for researchers in the report Fostering Integrity in Research, with sponsorship from the National Science Foundation. Major departures from the previous recommendations 25-years earlier are the recognition that practices that were previously classified as merely questionable are indeed detrimental to the responsible conduct of research. Furthermore, the report concludes that there has been insufficient effort to respond to the threats that lapses in research integrity pose to the quality of research products and the reputation of researchers as deserving of the public trust. It recommends the creation of an independent, non-profit entity dedicated to promoting research integrity by serving as a resource and clearing house for expertise, advice, materials, and best practices on fostering research integrity and responding to allegations of research misconduct.