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Sample records for time bioluminescence imaging

  1. Optimisation of acquisition time in bioluminescence imaging

    Taylor, Shelley L.; Mason, Suzannah K. G.; Glinton, Sophie; Cobbold, Mark; Styles, Iain B.; Dehghani, Hamid

    2015-03-01

    Decreasing the acquisition time in bioluminescence imaging (BLI) and bioluminescence tomography (BLT) will enable animals to be imaged within the window of stable emission of the bioluminescent source, a higher imaging throughput and minimisation of the time which an animal is anaesthetised. This work investigates, through simulation using a heterogeneous mouse model, two methods of decreasing acquisition time: 1. Imaging at fewer wavelengths (a reduction from five to three); and 2. Increasing the bandwidth of filters used for imaging. The results indicate that both methods are viable ways of decreasing the acquisition time without a loss in quantitative accuracy. Importantly, when choosing imaging wavelengths, the spectral attenuation of tissue and emission spectrum of the source must be considered, in order to choose wavelengths at which a high signal can be achieved. Additionally, when increasing the bandwidth of the filters used for imaging, the bandwidth must be accounted for in the reconstruction algorithm.

  2. Bioluminescence Imaging

    Sadikot, Ruxana T.; Blackwell, Timothy S.

    2005-01-01

    Bioluminescence refers to the process of visible light emission in living organisms. Bioluminescence imaging is a powerful methodology that has been developed over the last decade as a tool for molecular imaging of small laboratory animals, enabling the study of ongoing biological processes in vivo. This form of optical imaging is low cost and noninvasive and facilitates real-time analysis of disease processes at the molecular level in living organisms. In this article, we provide a brief int...

  3. Real-Time Bioluminescence Imaging of Nitroreductase in Mouse Model.

    Feng, Ping; Zhang, Huateng; Deng, Quankun; Liu, Wei; Yang, Linghui; Li, Guobo; Chen, Guo; Du, Lupei; Ke, Bowen; Li, Minyong

    2016-06-01

    Nitroreductase (NTR) is an endogenous reductase overexpressed in hypoxic tumors; however, its precise detection in living cells and animals remains a considerable challenge. Herein, we developed three reaction-based probes and a related bioluminescence assay for the real-time NTR detection. The high sensitivity and selectivity of probe 3, combined with its remarkable potential of bioluminescence imaging, affords a valuable approach for in vivo imaging of NTR in a tumor model mouse. PMID:27197544

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

    Jason Alan Gruenhagen

    2003-12-12

    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. Ca{sup 2+} imaging along with ATP imaging revealed that activation of phospholipase C and induction of intracellular Ca{sup 2+} 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{sup +} 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 functionalized Cd

  5. Bioluminescence microscopy using a short focal-length imaging lens

    Ogoh, K; Akiyoshi, R.; May-Maw-Thet,; Sugiyama, T; Dosaka, S; Hatta-Ohashi, Y; Suzuki, H.

    2014-01-01

    Bioluminescence from cells is so dim that bioluminescence microscopy is performed using an ultra low-light imaging camera. Although the image sensor of such cameras has been greatly improved over time, such improvements have not been made commercially available for microscopes until now. Here, we customized the optical system of a microscope for bioluminescence imaging. As a result, bioluminescence images of cells could be captured with a conventional objective lens and colour imaging camera....

  6. Infection routes of Aeromonas salmonicida in rainbow trout monitored in vivo by real-time bioluminescence imaging

    Bartkova, Simona; Kokotovic, Branko; Dalsgaard, Inger

    2016-01-01

    Recent development of imaging tools has facilitated studies of pathogen infections in vivo in real time. This trend can be exemplified by advances in bioluminescence imaging (BLI), an approach that helps to visualize dissemination of pathogens within the same animal over several time points. Here...

  7. BIOLUMINESCENCE IMAGING: PROGRESS AND APPLICATIONS

    Badr, Christian E.; Tannous, Bakhos A

    2011-01-01

    Application of bioluminescence imaging has grown tremendously in the past decade and has significantly contributed to the core conceptual advances in biomedical research. This technology provides valuable means for monitoring of different biological processes for immunology, oncology, virology and neuroscience. In this review, we will discuss current trends in bioluminescence and its application in different fields with emphasis on cancer research.

  8. Real-time monitoring of cariogenic bacteria via bioluminescent imaging: A biodontic hypothesis

    Jafar Kolahi

    2016-01-01

    Full Text Available Introduction: Dental caries (tooth decay remains one of the most common chronic infectious disease in the world. Disclosure of camouflaged cariogenic bacteria will be a great motivation for better oral hygiene. The Hypothesis: At present, lux transposon cassette, Tn4001 luxABCDE Kmr, is available that could be used for stable bioluminescent transformation of a wide range of gram-positive bacteria, e.g. Streptococcus mutans and Lactobacillus. After this step, sensitive charge-coupled device (CCD camera could be used to detect the low levels of light emitted from bioluminescent cariogenic bacteria. Living imaging software would be used for analysis and three-dimensional (3D reconstruction of images. Evaluation of the Hypothesis: Entrance of transgenic organisms into the oral cavity should be done with great caution. Ethical consideration is necessary and primary animal studies are required. The main limitation of this technique will be oxygen. As mentioned previously, bioluminescent reactions need oxygen. Hence, bioluminescent imaging cannot be used for anaerobic bacteria, e.g., Streptococcus sobrinus.

  9. Bioluminescence imaging in live cells and animals.

    Tung, Jack K; Berglund, Ken; Gutekunst, Claire-Anne; Hochgeschwender, Ute; Gross, Robert E

    2016-04-01

    The use of bioluminescent reporters in neuroscience research continues to grow at a rapid pace as their applications and unique advantages over conventional fluorescent reporters become more appreciated. Here, we describe practical methods and principles for detecting and imaging bioluminescence from live cells and animals. We systematically tested various components of our conventional fluorescence microscope to optimize it for long-term bioluminescence imaging. High-resolution bioluminescence images from live neurons were obtained with our microscope setup, which could be continuously captured for several hours with no signs of phototoxicity. Bioluminescence from the mouse brain was also imaged noninvasively through the intact skull with a conventional luminescence imager. These methods demonstrate how bioluminescence can be routinely detected and measured from live cells and animals in a cost-effective way with common reagents and equipment. PMID:27226972

  10. Bioluminescence imaging characteristics and application

    Bioluminescence imaging (BLI) by luciferase gene marked cells or DNA, in the presence of ATP and oxygen, catalytic oxidation reaction of fluorescein luminescence. So that it can directly monitor in vivo cell activity and gene behavior. In this paper, by comparing the BLI and MRI, PET, radiography of the similarities and differences, as well as about their cancer, stem cells and immune cells transportation, apoptosis and other aspects of the application, in order to better provide the basis for promoting the application of BLI. (authors)

  11. Noninvasive Bioluminescence Imaging in Small Animals

    Zinn, Kurt R.; Chaudhuri, Tandra R.; Szafran, April Adams; O’Quinn, Darrell; Weaver, Casey; Dugger, Kari; Lamar, Dale; Kesterson, Robert A.; Wang, Xiangdong; Frank, Stuart J.

    2008-01-01

    There has been a rapid growth of bioluminescence imaging applications in small animal models in recent years, propelled by the availability of instruments, analysis software, reagents, and creative approaches to apply the technology in molecular imaging. Advantages include the sensitivity of the technique as well as its efficiency, relatively low cost, and versatility. Bioluminescence imaging is accomplished by sensitive detection of light emitted following chemical reaction of the luciferase...

  12. In vivo cell tracking with bioluminescence imaging

    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.

  13. In vivo cell tracking with bioluminescence imaging

    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

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

    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.

  15. Bioluminescence in vivo imaging of autoimmune encephalomyelitis predicts disease

    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.

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

    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.

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

    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.

  18. Dynamic bioluminescence imaging for quantitative tumour burden assessment using IV or IP administration of d-luciferin: effect on intensity, time kinetics and repeatability of photon emission

    In vivo bioluminescence imaging (BLI) is a promising technique for non-invasive tumour imaging. d-luciferin can be administrated intraperitonealy or intravenously. This will influence its availability and, therefore, the bioluminescent signal. The aim of this study is to compare the repeatability of BLI measurement after IV versus IP administration of d-luciferin and assess the correlation between photon emission and histological cell count both in vitro and in vivo. Fluc-positive R1M cells were subcutaneously inoculated in nu/nu mice. Dynamic BLI was performed after IV or IP administration of d-luciferin. Maximal photon emission (PEmax) was calculated. For repeatability assessment, every acquisition was repeated after 4 h and analysed using Bland-Altman method. A second group of animals was serially imaged, alternating IV and IP administration up to 21 days. When mice were killed, PEmax after IV administration was correlated with histological cell number. The coefficients of repeatability were 80.2% (IV) versus 95.0% (IP). Time-to-peak is shorter, and its variance lower for IV (p max was 5.6 times higher for IV. A trend was observed towards lower photon emission per cell in larger tumours. IV administration offers better repeatability and better sensitivity when compared to IP. In larger tumours, multiple factors may contribute to underestimation of tumour burden. It might, therefore, be beneficial to test novel therapeutics on small tumours to enable an accurate evaluation of tumour burden. (orig.)

  19. In Vivo Bioluminescence Imaging of the Murine Pathogen Citrobacter rodentium

    Wiles, Siouxsie; Pickard, Karen M.; Peng, Katian; MacDonald, Thomas T.; Frankel, Gad

    2006-01-01

    Citrobacter rodentium is a natural mouse pathogen related to enteropathogenic and enterohemorrhagic Escherichia coli. We have previously utilized bioluminescence imaging (BLI) to determine the in vivo colonization dynamics of C. rodentium. However, due to the oxygen requirement of the bioluminescence system and the colonic localization of C. rodentium, in vivo localization studies were performed using harvested organs. Here, we report the detection of bioluminescent C. rodentium and commensal...

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

    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

  1. Bioluminescence.

    Jones, M. Gail

    1993-01-01

    Describes bioluminescence and the chemistry of how it occurs. Presents information for conducting the following classroom activities: (1) firefly mimic; (2) modeling deep-sea fish; (3) sea fireflies; and (4) the chemistry of light. (PR)

  2. Bioluminescence: a versatile technique for imaging cellular and molecular features

    Paley, Miranda A.

    2016-01-01

    Bioluminescence is a ubiquitous imaging modality for visualizing biological processes in vivo. This technique employs visible light and interfaces readily with most cell and tissue types, making it a versatile technology for preclinical studies. Here we review basic bioluminescence imaging principles, along with applications of the technology that are relevant to the medicinal chemistry community. These include noninvasive cell tracking experiments, analyses of protein function, and methods to visualize small molecule metabolites. In each section, we also discuss how bioluminescent tools have revealed insights into experimental therapies and aided drug discovery. Last, we highlight the development of new bioluminescent tools that will enable more sensitive and multi-component imaging experiments and, thus, expand our broader understanding of living systems.

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

    Burrell-Saward, Hollie; Ward, Theresa H

    2016-01-01

    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. PMID:27284970

  4. Interactive graphic editing tools in bioluminescent imaging simulation

    Li, Hui; Tian, Jie; Luo, Jie; Wang, Ge; Cong, Wenxiang

    2005-04-01

    It is a challenging task to accurately describe complicated biological tissues and bioluminescent sources in bioluminescent imaging simulation. Several graphic editing tools have been developed to efficiently model each part of the bioluminescent simulation environment and to interactively correct or improve the initial models of anatomical structures or bioluminescent sources. There are two major types of graphic editing tools: non-interactive tools and interactive tools. Geometric building blocks (i.e. regular geometric graphics and superquadrics) are applied as non-interactive tools. To a certain extent, complicated anatomical structures and bioluminescent sources can be approximately modeled by combining a sufficient large number of geometric building blocks with Boolean operators. However, those models are too simple to describe the local features and fine changes in 2D/3D irregular contours. Therefore, interactive graphic editing tools have been developed to facilitate the local modifications of any initial surface model. With initial models composed of geometric building blocks, interactive spline mode is applied to conveniently perform dragging and compressing operations on 2D/3D local surface of biological tissues and bioluminescent sources inside the region/volume of interest. Several applications of the interactive graphic editing tools will be presented in this article.

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

    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.

  6. Filtering and deconvolution for bioluminescence imaging of small animals

    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)

  7. The Expanding Toolbox of In Vivo Bioluminescent Imaging

    Xu, Tingting; Close, Dan; Handagama, Winode; Marr, Enolia; Sayler, Gary; Ripp, Steven

    2016-01-01

    In vivo bioluminescent imaging (BLI) permits the visualization of engineered bioluminescence from living cells and tissues to provide a unique perspective toward the understanding of biological processes as they occur within the framework of an authentic in vivo environment. The toolbox of in vivo BLI includes an inventory of luciferase compounds capable of generating bioluminescent light signals along with sophisticated and powerful instrumentation designed to detect and quantify these light signals non-invasively as they emit from the living subject. The information acquired reveals the dynamics of a wide range of biological functions that play key roles in the physiological and pathological control of disease and its therapeutic management. This mini review provides an overview of the tools and applications central to the evolution of in vivo BLI as a core technology in the preclinical imaging disciplines. PMID:27446798

  8. The Expanding Toolbox of In Vivo Bioluminescent Imaging.

    Xu, Tingting; Close, Dan; Handagama, Winode; Marr, Enolia; Sayler, Gary; Ripp, Steven

    2016-01-01

    In vivo bioluminescent imaging (BLI) permits the visualization of engineered bioluminescence from living cells and tissues to provide a unique perspective toward the understanding of biological processes as they occur within the framework of an authentic in vivo environment. The toolbox of in vivo BLI includes an inventory of luciferase compounds capable of generating bioluminescent light signals along with sophisticated and powerful instrumentation designed to detect and quantify these light signals non-invasively as they emit from the living subject. The information acquired reveals the dynamics of a wide range of biological functions that play key roles in the physiological and pathological control of disease and its therapeutic management. This mini review provides an overview of the tools and applications central to the evolution of in vivo BLI as a core technology in the preclinical imaging disciplines. PMID:27446798

  9. Space application research of EMCCDs for bioluminescence imaging

    Zhang, Tao

    The detection of bioluminescense is widely used on the ground, while the detection of bioluminescence in space is still at the stage of detecting bright bioluminescense. With the rapid development of research in Space Life Sciences, it will be necessary to develop a detection technology to detect weak bioluminescense. Compared to other low-light detection techniques for ground, there are more advantages of EMCCDs for space application. Build a space bioluminescence imaging detection system, analysis the feasibility and capability of its will be significant. Co-Author:Xie Zongbao,Zheng Weibo

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

    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.

  11. Development of Quantification Method for Bioluminescence Imaging

    Optical molecular luminescence imaging is widely used for detection and imaging of bio-photons emitted by luminescent luciferase activation. The measured photons in this method provide the degree of molecular alteration or cell numbers with the advantage of high signal-to-noise ratio. To extract useful information from the measured results, the analysis based on a proper quantification method is necessary. In this research, we propose a quantification method presenting linear response of measured light signal to measurement time. We detected the luminescence signal by using lab-made optical imaging equipment of animal light imaging system (ALIS) and different two kinds of light sources. One is three bacterial light-emitting sources containing different number of bacteria. The other is three different non-bacterial light sources emitting very weak light. By using the concept of the candela and the flux, we could derive simplified linear quantification formula. After experimentally measuring light intensity, the data was processed with the proposed quantification function. We could obtain linear response of photon counts to measurement time by applying the pre-determined quantification function. The ratio of the re-calculated photon counts and measurement time present a constant value although different light source was applied. The quantification function for linear response could be applicable to the standard quantification process. The proposed method could be used for the exact quantitative analysis in various light imaging equipment with presenting linear response behavior of constant light emitting sources to measurement time

  12. Infection with adenovirus-mediated luciferase reporter gene in mesenchymal stem cells and bioluminescence imaging

    Objective: To construct adenovirus vector containing firefly luciferase reporter gene (Ad-Luc) and infect bone marrow mesenchymal stem cells (BMSC), then to take bioluminescence imaging in vitro and in vivo for identification. Methods: The luciferase gene was amplified with PCR from psiCHECK-2 plasmid and cloned into the adenoviral shuttle vector (pShuttle-CMV). It was confirmed by Nhe Ⅰ/Xba Ⅰ digestion and sequencing. PShuttle-CMV-Luc and backbone vector (pAdeno) were homologous recombined. Then the recombinant plasmid was packaged in HEK293 cells and the virus titer was detected. The BMSC were infected by the recombinant adenovirus. The bioluminescence imaging in vitro was performed to determine the best multiplicity of infection (MOI), and the relationship between bioluminescence intensity and MOI was analyzed by curve fitting regression analysis. Viability was evaluated via Trypan blue staining. The transfected BMSC (1 × 106) were implanted into the muscles of forelimb of SD rats,and then tracked by bioluminescence imaging in vivo. Cell viability was compared using two-way repeated measures analysis of variance between groups. Results: Enzyme digestion and sequence analysis indicated that Ad-Luc was successfully constructed. The virus titer was 1 × 1010 plaque forming unit (PFU)/ml. The bioluminescence detection in vitro showed that Ad-Luc could infect BMSC high efficiently to express luciferase and the best MOI was 50. The bioluminescence intensity enhanced with increase of MOI (R2 =0.98). No statistically significant difference was found in cell viability between transfected and untransfected BMSC at 1, 3, 5, 7 d. The cell survival rates were (92.5±2.3)% vs (94.1±1.8)%, (91.4±0.9)% vs (92.7±2.0)%, (92.1±1.6)% vs (93.3± 2.4)%, (91.9 ± 1.5)% vs (93.0 ± 3.1)%, respectively (F=4.38, P>0.05). The bioluminescence imaging in vivo showed that BMSC survived 1, 3, 7 d after implantation. However, bioluminescence signal decreased gradually over time

  13. Hyperspectral and multispectral bioluminescence optical tomography for small animal imaging

    For bioluminescence imaging studies in small animals, it is important to be able to accurately localize the three-dimensional (3D) distribution of the underlying bioluminescent source. The spectrum of light produced by the source that escapes the subject varies with the depth of the emission source because of the wavelength-dependence of the optical properties of tissue. Consequently, multispectral or hyperspectral data acquisition should help in the 3D localization of deep sources. In this paper, we describe a framework for fully 3D bioluminescence tomographic image acquisition and reconstruction that exploits spectral information. We describe regularized tomographic reconstruction techniques that use semi-infinite slab or FEM-based diffusion approximations of photon transport through turbid media. Singular value decomposition analysis was used for data dimensionality reduction and to illustrate the advantage of using hyperspectral rather than achromatic data. Simulation studies in an atlas-mouse geometry indicated that sub-millimeter resolution may be attainable given accurate knowledge of the optical properties of the animal. A fixed arrangement of mirrors and a single CCD camera were used for simultaneous acquisition of multispectral imaging data over most of the surface of the animal. Phantom studies conducted using this system demonstrated our ability to accurately localize deep point-like sources and show that a resolution of 1.5 to 2.2 mm for depths up to 6 mm can be achieved. We also include an in vivo study of a mouse with a brain tumour expressing firefly luciferase. Co-registration of the reconstructed 3D bioluminescent image with magnetic resonance images indicated good anatomical localization of the tumour

  14. In Vivo Mouse Bioluminescence Tomography with Radionuclide-Based Imaging Validation

    Lu, Yujie; Machado, Hidevaldo B.; Bao, Qinan; Stout, David; Herschman, Harvey; Chatziioannou, Arion F.

    2010-01-01

    Introduction Bioluminescence imaging, especially planar bioluminescence imaging, has been extensively applied in in vivo preclinical biological research. Bioluminescence tomography (BLT) has the potential to provide more accurate imaging information due to its 3D reconstruction compared with its planar counterpart. Methods In this work, we introduce a positron emission tomography (PET) radionuclide imaging-based strategy to validate the BLT results. X-ray computed tomography, PET, spectrally ...

  15. In vivo fluorescence imaging of the reticuloendothelial system using quantum dots in combination with bioluminescent tumour monitoring

    We characterised in vivo fluorescence imaging (FLI) of the reticuloendothelial system using quantum dots (QD) and investigated its use in combination with in vivo bioluminescence imaging (BLI). In vivo FLI was performed in five mice repeatedly after the intravenous administration of QD without conjugation to targeting ligands. Ex vivo FLI of the excised organs was performed 24 h after QD injection in three mice. Seven days after intravenous inoculation of luciferase-expressing model cells of a haematological malignancy, mice were injected with the QD or saline (n = 5 each), and combined BLI/FLI was performed repeatedly. Additional five mice inoculated with the tumour cells were examined by in vivo BLI/FLI, and the structures harbouring bioluminescent foci were determined by ex vivo BLI. The utility of combining FLI with bioluminescent tumour monitoring was evaluated. In vivo FLI after QD injection allowed long-term, repeated observation of the reticuloendothelial system in individual mice, although fluorescence intensity and image contrast gradually decreased over time. Ex vivo FLI verified selective accumulation in reticuloendothelial structures. The administration of QD did not affect whole-body bioluminescent signal intensities during longitudinal tumour monitoring. In vivo BLI/FLI, accompanied by fusion of both images, improved the accuracy and confidence level of the localisation of the bioluminescent foci. In vivo FLI using QD provides an overview of the reticuloendothelial system in living mice. In combination with bioluminescent tumour monitoring, fluorescent reticuloendothelial imaging is expected to provide valuable information for lesion localisation. (orig.)

  16. Impact of Anesthesia Protocols on In Vivo Bioluminescent Bacteria Imaging Results

    Chuzel, Thomas; Sanchez, Violette; Vandamme, Marc; Martin, Stéphane; Flety, Odile; Pager, Aurélie; Chabanel, Christophe; Magnier, Luc; Foskolos, Marie; Petit, Océane; Rokbi, Bachra; Chereul, Emmanuel

    2015-01-01

    Infectious murine models greatly benefit from optical imaging using bioluminescent bacteria to non-invasively and repeatedly follow in vivo bacterial infection. In this context, one of the most critical parameters is the bioluminescence sensitivity to reliably detect the smallest number of bacteria. Another critical point is the anesthetic approaches that have been demonstrated to impact the bioluminescence flux emission in studies with luciferase-transfected tumor cells. However, this impact...

  17. Assessing laser-tissue damage with bioluminescent imaging

    Wilmink, Gerald J.; Opalenik, Susan R.; Beckham, Josh T.; Davidson, Jeffrey M.; Jansen, Eric D.

    2006-07-01

    Effective medical laser procedures are achieved by selecting laser parameters that minimize undesirable tissue damage. Traditionally, human subjects, animal models, and monolayer cell cultures have been used to study wound healing, tissue damage, and cellular effects of laser radiation. Each of these models has significant limitations, and consequently, a novel skin model is needed. To this end, a highly reproducible human skin model that enables noninvasive and longitudinal studies of gene expression was sought. In this study, we present an organotypic raft model (engineered skin) used in combination with bioluminescent imaging (BLI) techniques. The efficacy of the raft model was validated and characterized by investigating the role of heat shock protein 70 (hsp70) as a sensitive marker of thermal damage. The raft model consists of human cells incorporated into an extracellular matrix. The raft cultures were transfected with an adenovirus containing a murine hsp70 promoter driving transcription of luciferase. The model enables quantitative analysis of spatiotemporal expression of proteins using BLI. Thermal stress was induced on the raft cultures by means of a constant temperature water bath or with a carbon dioxide (CO2) laser (λ=10.6 µm, 0.679 to 2.262 W/cm2, cw, unfocused Gaussian beam, ωL=4.5 mm, 1 min exposure). The bioluminescence was monitored noninvasively with an IVIS 100 Bioluminescent Imaging System. BLI indicated that peak hsp70 expression occurs 4 to 12 h after exposure to thermal stress. A minimum irradiance of 0.679 W/cm2 activated the hsp70 response, and a higher irradiance of 2.262 W/cm2 was associated with a severe reduction in hsp70 response due to tissue ablation. Reverse transcription polymerase chain reaction demonstrated that hsp70 mRNA levels increased with prolonged heating exposures. Enzyme-linked immunosorbent protein assays confirmed that luciferase was an accurate surrogate for hsp70 intracellular protein levels. Hematoxylin and

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

    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.

  19. Confocal Bioluminescence Imaging for Living Tissues with a Caged Substrate of Luciferin.

    Hattori, Mitsuru; Kawamura, Genki; Kojima, Ryosuke; Kamiya, Mako; Urano, Yasuteru; Ozawa, Takeaki

    2016-06-21

    Fluorescence imaging can elucidate morphological organization and coordinal networks, but its background luminescence degrades the image contrast. Our confocal bioluminescence imaging system uses a luciferase caged substrate, with light passing through multipinhole arrays, causing bioluminescence at a focal plane. After a charge-coupled device camera captures luminescence, the imaging system acquires confocal images of multilayered cells with depth information, supporting quantitative analysis of spatial cellular localization in living tissues. PMID:27216493

  20. In vivo bioluminescence and reflectance imaging of multiple organs in bioluminescence reporter mice by bundled-fiber-coupled microscopy.

    Ando, Yoriko; Sakurai, Takashi; Koida, Kowa; Tei, Hajime; Hida, Akiko; Nakao, Kazuki; Natsume, Mistuo; Numano, Rika

    2016-03-01

    Bioluminescence imaging (BLI) is used in biomedical research to monitor biological processes within living organisms. Recently, fiber bundles with high transmittance and density have been developed to detect low light with high resolution. Therefore, we have developed a bundled-fiber-coupled microscope with a highly sensitive cooled-CCD camera that enables the BLI of organs within the mouse body. This is the first report of in vivo BLI of the brain and multiple organs in luciferase-reporter mice using bundled-fiber optics. With reflectance imaging, the structures of blood vessels and organs can be seen clearly with light illumination, and it allowed identification of the structural details of bioluminescence images. This technique can also be applied to clinical diagnostics in a low invasive manner. PMID:27231601

  1. In vivo bioluminescence and reflectance imaging of multiple organs in bioluminescence reporter mice by bundled-fiber-coupled microscopy

    Ando, Yoriko; Sakurai, Takashi; Koida, Kowa; Tei, Hajime; Hida, Akiko; Nakao, Kazuki; Natsume, Mistuo; Numano, Rika

    2016-01-01

    Bioluminescence imaging (BLI) is used in biomedical research to monitor biological processes within living organisms. Recently, fiber bundles with high transmittance and density have been developed to detect low light with high resolution. Therefore, we have developed a bundled-fiber-coupled microscope with a highly sensitive cooled-CCD camera that enables the BLI of organs within the mouse body. This is the first report of in vivo BLI of the brain and multiple organs in luciferase-reporter mice using bundled-fiber optics. With reflectance imaging, the structures of blood vessels and organs can be seen clearly with light illumination, and it allowed identification of the structural details of bioluminescence images. This technique can also be applied to clinical diagnostics in a low invasive manner. PMID:27231601

  2. ATP binding cassette transporters modulate both coelenterazine- and D-luciferin- based bioluminescence imaging

    Huang, Ruimin; Vider, Jelena; Serganova, Inna; Blasberg, Ronald G.

    2011-01-01

    Bioluminescence imaging (BLI) of luciferase reporters provides a cost-effective and sensitive means to image biological processes. However, transport of luciferase substrates across the cell membrane does affect BLI-readout-intensity from intact living cells.

  3. Characterization of sevoflurane effects on Per2 expression using ex vivo bioluminescence imaging of the suprachiasmatic nucleus in transgenic rats.

    Matsuo, Izumi; Iijima, Norio; Takumi, Ken; Higo, Shimpei; Aikawa, Satoko; Anzai, Megumi; Ishii, Hirotaka; Sakamoto, Atsuhiro; Ozawa, Hitoshi

    2016-06-01

    The inhalation anesthetic sevoflurane suppresses Per2 expression in the suprachiasmatic nucleus (SCN) in rodents. Here, we investigated the intra-SCN regional specificity, time-dependency, and pharmacological basis of sevoflurane-effects. Bioluminescence image was taken from the SCN explants of mPer2 promoter-destabilized luciferase transgenic rats, and each small regions of interest (ROI) of the image was analyzed. Sevoflurane suppressed bioluminescence in all ROIs, suggesting that all regions in the SCN are sensitive to sevoflurane. Clear time-dependency in sevoflurane effects were also observed; application during the trough phase of the bioluminescence cycle suppressed the subsequent increase in bioluminescence and resulted in a phase delay of the cycle; sevoflurane applied during the middle of the ascending phase induced a phase advance; sevoflurane on the descending phase showed no effect. These results indicate that the sevoflurane effect may depend on the intrinsic state of circadian machinery. Finally, we examined the involvement of GABAergic signal transduction in the sevoflurane effect. Co-application of both GABAA and GABAB receptor antagonists completely blocked the effect of sevoflurane on the bioluminescence rhythm, suggesting that sevoflurane inhibits Per2 expression via GABAergic signal transduction. Current study elucidated the anesthetic effects on the molecular mechanisms of circadian rhythm. PMID:26696094

  4. U-SPECT-BioFluo: an integrated radionuclide, bioluminescence, and fluorescence imaging platform

    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 fully integrated bioluminescence-fluorescence-SPECT platform. Next to an optimization in logistics and image fusion, this integration can help improve understanding of the optical imaging (OI) resul...

  5. Tomographic bioluminescence imaging by an iteratively re-weighted minimization

    Wu, Ping; Liu, Kai; Xue, Zhenwen; Guo, Wei; Qin, Chenghu; Tian, Jie

    2012-03-01

    Tomographic bioluminescence imaging (TBI), with visible light emission in living organisms, is an effective way of molecular imaging, which allows for the study of ongoing tumor biological processes in vivo and non-invasively. This newly developed technology enables three-dimensional accuracy localization and quantitative analysis of the target tumor cells in small animal via reconstructing the images acquired by the high-resolution imaging system. Due to the difficulty of reconstruction, which is often referred to an ill-posed inverse problem, continuous efforts are still made to find more practical and efficient approaches. In this paper, an iteratively re-weighted minimization (IRM) has been applied to reconstruct the entire source distribution, which is known as sparse signals, inside the target tissue with the limited outgoing photon density on its boundary. By introducing a weight function into the objective function, we convert the lp norm problem into a more simple form of l2 norm to reduce the computational complexity. The weight function is updated in each iterative step to compute the final optimal solution more efficiently. This method is proved to be robust to different parameters, and mouse experiments are conducted to validate the feasibility of IRM approach, which is also reliable at whole-body imaging.

  6. Multimodal imaging of orthotopic hepatocellular carcinoma using small animal PET, bioluminescence and contrast enhanced CT imaging

    Molecular imaging with small-animal PET and bioluminescence imaging has been used as an important tool in cancer research. One of the disadvantages of these imaging modalities is the lack of anatomic information. To obtain fusion images with both molecular and anatomical information, small-animal PET and bioluminescence images fused with contrast enhance CT image in orthotopic hepatocellular carcinoma (HCC) model. We retrovially transfected dual gene (HSV1-tk and firefly luciferase) to morris hepatoma cells. The expression of HSV1-tk and luciferase was checked by optical imager and in vitro radiolabeled FIAU uptake, respectively and also checked by RT-PCR analysis. MCA-TL cells (5X105/ 0.05 ml) mixed with matrigel (1: 10) injected into left lobe of liver in nude mice. 124I-FIAU-PET, bioluminescence and contrast enhanced CT images were obtained in the orthotopic HCC model and digital whole body autoradiography (DWBA) was performed. Small animal PET image was obtained at 2 h post injection of 124I-FIAU and contrast enhanced CT image was obtained at 3 h post injection of Fenestra LC (0.3 ml). MCA-TL cells showed more specific 124I-FIAU uptake and higher luminescent activity than parental cells. The orthotopic HCC was detected by 124I-FIAU PET, contrast enhanced CT, and BLI and confirmed by DWBA. Registered image in orthotopic HCC t models showed a good correlation of images from both PET and CT. Contrast enhanced CT image delineated margin of HCC. Multimodal imaging with 124I-FIAU PET, bioluminescence and contrast enhanced CT allows a precise and improved detection of tumor in orthotopic hepatocellular carcinoma model. Multimodal imaging is potentially useful for monitoring progression of hepatic metastasis and for the evaluation of cancer treatments

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

    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 (R 2=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 mm3 (R 2=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

  8. In Vivo Bioluminescence Imaging of Tumor Cells Using Optimized Firefly Luciferase luc2

    2013-01-01

    The present study was aimed to establish a tumor cell line stably expressing luciferase luc2, and to develop the technique to observe primary tumor nodes and metastases using in vivo bioluminescence imaging. Materials and Methods. In this research we used pLuc2-N plasmid, lentiviral vector pLVT-1, Colo 26 cell line and BALB/c mice to generate new bioluminescent tumor model. Bioluminescence imaging in vitro и in vivo was carried out on IVIS-Spectrum system (Caliper Life Sciences, USA). Pri...

  9. High resolution in vitro bioluminescence imaging using a multimodal optical system

    Altabella, L.; Gigliotti, C. R.; Perani, L.; Crippa, M. P.; Boschi, F.; Spinelli, A. E.

    2016-01-01

    Bioluminescence in vitro studies are usually performed with dedicated microscopes. In this work, we developed a novel image recovery algorithm and a multimodal system prototype to perform bioluminescence microscopy. We performed a feasibility study using GEANT4 Monte Carlo (MC) simulation of bioluminescent cells acquired at low SNR frames and processed using a Super Resolution Regularization Algorithm (SRRA). The method was also tested using in vitro cell acquisition. The results obtained with MC simulations showed an improvement in the spatial resolution from 90 μ m to 10 μ m and from 110 μ m to 13 μ m for in vitro imaging of mesothelioma cells.

  10. High resolution in vitro bioluminescence imaging using a multimodal optical system

    Bioluminescence in vitro studies are usually performed with dedicated microscopes. In this work, we developed a novel image recovery algorithm and a multimodal system prototype to perform bioluminescence microscopy. We performed a feasibility study using GEANT4 Monte Carlo (MC) simulation of bioluminescent cells acquired at low SNR frames and processed using a Super Resolution Regularization Algorithm (SRRA). The method was also tested using in vitro cell acquisition. The results obtained with MC simulations showed an improvement in the spatial resolution from 90 μm to 10 μm and from 110 μm to 13 μm for in vitro imaging of mesothelioma cells

  11. U-SPECT-BioFluo: an integrated radionuclide, bioluminescence, and fluorescence imaging platform

    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

  12. Bioluminescence Imaging of Chlamydia muridarum Ascending Infection in Mice

    Jessica Campbell; Yumeng Huang; Yuanjun Liu; Robert Schenken; Bernard Arulanandam; Guangming Zhong

    2014-01-01

    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 intrav...

  13. A luciferin analogue generating near-infrared bioluminescence achieves highly sensitive deep-tissue imaging.

    Kuchimaru, Takahiro; Iwano, Satoshi; Kiyama, Masahiro; Mitsumata, Shun; Kadonosono, Tetsuya; Niwa, Haruki; Maki, Shojiro; Kizaka-Kondoh, Shinae

    2016-01-01

    In preclinical cancer research, bioluminescence imaging with firefly luciferase and D-luciferin has become a standard to monitor biological processes both in vitro and in vivo. However, the emission maximum (λmax) of bioluminescence produced by D-luciferin is 562 nm where light is not highly penetrable in biological tissues. This emphasizes a need for developing a red-shifted bioluminescence imaging system to improve detection sensitivity of targets in deep tissue. Here we characterize the bioluminescent properties of the newly synthesized luciferin analogue, AkaLumine-HCl. The bioluminescence produced by AkaLumine-HCl in reactions with native firefly luciferase is in the near-infrared wavelength ranges (λmax=677 nm), and yields significantly increased target-detection sensitivity from deep tissues with maximal signals attained at very low concentrations, as compared with D-luciferin and emerging synthetic luciferin CycLuc1. These characteristics offer a more sensitive and accurate method for non-invasive bioluminescence imaging with native firefly luciferase in various animal models. PMID:27297211

  14. A luciferin analogue generating near-infrared bioluminescence achieves highly sensitive deep-tissue imaging

    Kuchimaru, Takahiro; Iwano, Satoshi; Kiyama, Masahiro; Mitsumata, Shun; Kadonosono, Tetsuya; Niwa, Haruki; Maki, Shojiro; Kizaka-Kondoh, Shinae

    2016-01-01

    In preclinical cancer research, bioluminescence imaging with firefly luciferase and D-luciferin has become a standard to monitor biological processes both in vitro and in vivo. However, the emission maximum (λmax) of bioluminescence produced by D-luciferin is 562 nm where light is not highly penetrable in biological tissues. This emphasizes a need for developing a red-shifted bioluminescence imaging system to improve detection sensitivity of targets in deep tissue. Here we characterize the bioluminescent properties of the newly synthesized luciferin analogue, AkaLumine-HCl. The bioluminescence produced by AkaLumine-HCl in reactions with native firefly luciferase is in the near-infrared wavelength ranges (λmax=677 nm), and yields significantly increased target-detection sensitivity from deep tissues with maximal signals attained at very low concentrations, as compared with D-luciferin and emerging synthetic luciferin CycLuc1. These characteristics offer a more sensitive and accurate method for non-invasive bioluminescence imaging with native firefly luciferase in various animal models. PMID:27297211

  15. Compartmentalization of algal bioluminescence: autofluorescence of bioluminescent particles in the dinoflagellate Gonyaulax as studied with image-intensified video microscopy and flow cytometry

    1985-01-01

    Compartmentalization of specialized functions to discrete locales is a fundamental theme of eucaryotic organization in cells. We report here that bioluminescence of the dinoflagellate alga Gonyaulax originates in vivo from discrete subcellular loci that are intrinsically fluorescent. We demonstrate this localization by comparing the loci of fluorescence and bioluminescence as visualized by image-intensified video microscopy. These fluorescent particles appeared to be the same as the previousl...

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

    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.

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

    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.

  18. Bioluminescence imaging of fungal biofilm development in live animals.

    Vande Velde, Greetje; Kucharíková, Soňa; Van Dijck, Patrick; Himmelreich, Uwe

    2014-01-01

    Fungal biofilms formed on various types of medical implants represent a major problem for hospitalized patients. These biofilms and related infections are usually difficult to treat because of their resistance to the classical antifungal drugs. Animal models are indispensable for investigating host-pathogen interactions and for identifying new antifungal targets related to biofilm development. A limited number of animal models is available that can be used for testing novel antifungal drugs in vivo against C. albicans, one of the most common pathogens causing fungal biofilms. Fungal load in biofilms in these models is traditionally analyzed postmortem, requiring host sacrifice and enumeration of microorganisms from individual biofilms in order to evaluate the amount of colony forming units and the efficacy of antifungal treatment. Bioluminescence imaging (BLI) made compatible with small animal models for in vivo biofilm formation is a valuable noninvasive tool to follow-up biofilm development and its treatment longitudinally, reducing the number of animals needed for such studies. Due to the nondestructive and noninvasive nature of BLI, the imaging procedure can be repeated in the same animal, allowing follow-up of the biofilm growth in vivo without removing the implanted device or detaching the biofilm from its substrate. The method described here introduces BLI of C. albicans biofilm formation in vivo on subcutaneously implanted catheters in mice. One of the main challenges to overcome for BLI of fungi is the hampered intracellular substrate delivery through the fungal cell wall, which is managed by using extracellularly located Gaussia luciferase. Although detecting a quantifiable in vivo BLI signal from biofilms formed on the inside of implanted catheters is challenging, BLI proved to be a practical tool in the study of fungal biofilms. This method describing the use of BLI for in vivo follow-up of device-related fungal biofilm formation has the potential for

  19. Bioluminescence Imaging Reveals Dynamics of Beta Cell Loss in the Non-Obese Diabetic (NOD) Mouse Model

    John Virostko; Armandla Radhika; Greg Poffenberger; Dula, Adrienne N.; Moore, Daniel J.; Alvin C Powers

    2013-01-01

    We generated a mouse model (MIP-Luc-VU-NOD) that enables non-invasive bioluminescence imaging (BLI) of beta cell loss during the progression of autoimmune diabetes and determined the relationship between BLI and disease progression. MIP-Luc-VU-NOD mice displayed insulitis and a decline in bioluminescence with age which correlated with beta cell mass, plasma insulin, and pancreatic insulin content. Bioluminescence declined gradually in female MIP-Luc-VU-NOD mice, reaching less than 50% of the ...

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

    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 o

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

    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.

  2. In vitro influence of hypoxia on bioluminescence imaging in brain tumor cells

    Moriyama, Eduardo H.; Jarvi, Mark; Niedre, Mark; Mocanu, Joseph D.; Moriyama, Yumi; Li, Buhong; Lilge, Lothar; Wilson, Brian C.

    2007-02-01

    Bioluminescence Imaging (BLI) has been employed as an imaging modality to identify and characterize fundamental processes related to cancer development and response at cellular and molecular levels. This technique is based on the reaction of luciferin with oxygen in the presence of luciferase and ATP. A major concern in this technique is that tumors are generally hypoxic, either constitutively and/or as a result of treatment, therefore the oxygen available for the bioluminescence reaction could possibly be reduced to limiting levels, and thus leading to underestimation of the actual number of luciferase-labeled cells during in vivo procedures. In this report, we present the initial in vitro results of the oxygen dependence of the bioluminescence signal in rat gliosarcoma 9L cells tagged with the luciferase gene (9L luc cells). Bioluminescence photon emission from cells exposed to different oxygen tensions was detected by a sensitive CCD camera upon exposure to luciferin. The results showed that bioluminescence signal decreased at administered pO II levels below about 5%, falling by approximately 50% at 0.2% pO II. Additional experiments showed that changes in BLI was due to the cell inability to maintain normal levels of ATP during the hypoxic period reducing the ATP concentration to limiting levels for BLI.

  3. Bioluminescence imaging to monitor the prolongation of stem cell survival by pharmaceutical intervention

    Le, Uyenchi N.; Min, Jung Joon; Moon, Sung Min; Ahn, Young Keun; Kim, Yong Sook; Joo, Soo Yeon; Hong, Moon Hwa; Jeong, Myung Ho; Song, Ho Cheon; Bom, Hee Seung [Chonnam National University Medical School, Gwangju (Korea, Republic of)

    2005-07-01

    The rapid donor cell death and rejection owing to humoral and cellular immune reactions are a basic limitation encountered in stem cell therapy for treatment of cardiovascular disease. We investigated the potential for longitudinal bioluminescence imaging to monitor the survival of transplanted stem cells prolonged by immunosuppressive agents. Embryonic rat H9c2 cardio myoblasts were transfected with adenovirus containing luciferase reporter gene (Ad-CMV-Fluc) in different MOI (1,10,100) and various cell doses (1x10{sup 5} - 5x10{sup 6})followed by injection in the thigh muscle of nude mice (n=6 per group), Other mice (n = 18) were undergone transient immunosuppression provided by either Cyclosporine (5mg/kg) or Tacrolimus (1mg/kg) or Dexamethasone (4mg/kg) beginning 3 days prior to and continuing to 2 weeks after transplantation. Optical bioluminescent imaging was then daily carried out using cooled CCD camera (Xenogen) Viral transfection at MOI 100 and the 5x10{sup 6} cell dose implantation resulted in optimal transgene efficiency. Mice received immunosuppressive agents displayed long-term in vivo reporter gene expression for a time course of 14 days. Tacrolimus (Prograf) and Cyclosporine successfully suppressed the transplanted cell loss in animals, that obviously observed until day 8 as compared to Dexamethasone-treated and non-treated mice (day 1: 1.00E+08 (Prograf), 9.47E+07 (Cys), 5.25E+07 (Dex), and 1.25E+07 p/s/cm{sup 2}/sr (control); day 8: 3.27E+05 (Prograf), 1.02E+05 (Cys), 6.17E+04 (Dex) and 2.73E+04 p/s/cm{sup 2}/sr (control)) and continued expressing bioluminescence until day 13 ( 6.42E+05 (Prograf), 4.99E+05 (Cys), and 4.10E+04 p/s/cm{sup 2}/sr. Induction of immune tolerance using pharmaceutical agents during cardio myoblast transplantation improved long-term donor cell survival in murine muscles. Optical imaging technique is capable of being used for tracking implanted stem cells in myocardium of living subjects over time.

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

    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

  5. Bioluminescence imaging to monitor the prolongation of stem cell survival by pharmaceutical intervention

    The rapid donor cell death and rejection owing to humoral and cellular immune reactions are a basic limitation encountered in stem cell therapy for treatment of cardiovascular disease. We investigated the potential for longitudinal bioluminescence imaging to monitor the survival of transplanted stem cells prolonged by immunosuppressive agents. Embryonic rat H9c2 cardio myoblasts were transfected with adenovirus containing luciferase reporter gene (Ad-CMV-Fluc) in different MOI (1,10,100) and various cell doses (1x105 - 5x106)followed by injection in the thigh muscle of nude mice (n=6 per group), Other mice (n = 18) were undergone transient immunosuppression provided by either Cyclosporine (5mg/kg) or Tacrolimus (1mg/kg) or Dexamethasone (4mg/kg) beginning 3 days prior to and continuing to 2 weeks after transplantation. Optical bioluminescent imaging was then daily carried out using cooled CCD camera (Xenogen) Viral transfection at MOI 100 and the 5x106 cell dose implantation resulted in optimal transgene efficiency. Mice received immunosuppressive agents displayed long-term in vivo reporter gene expression for a time course of 14 days. Tacrolimus (Prograf) and Cyclosporine successfully suppressed the transplanted cell loss in animals, that obviously observed until day 8 as compared to Dexamethasone-treated and non-treated mice (day 1: 1.00E+08 (Prograf), 9.47E+07 (Cys), 5.25E+07 (Dex), and 1.25E+07 p/s/cm2/sr (control); day 8: 3.27E+05 (Prograf), 1.02E+05 (Cys), 6.17E+04 (Dex) and 2.73E+04 p/s/cm2/sr (control)) and continued expressing bioluminescence until day 13 ( 6.42E+05 (Prograf), 4.99E+05 (Cys), and 4.10E+04 p/s/cm2/sr. Induction of immune tolerance using pharmaceutical agents during cardio myoblast transplantation improved long-term donor cell survival in murine muscles. Optical imaging technique is capable of being used for tracking implanted stem cells in myocardium of living subjects over time

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

    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

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

    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 [Chonnam National University Hospital, Gwangju (Korea, Republic of); Choi, Eun Seo [Chosun University, Gwangju (Korea, Republic of)

    2009-08-15

    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

  8. Bioluminescence in vivo imaging of autoimmune encephalomyelitis predicts disease

    Steinman Lawrence; Ho Peggy; Luo Jian; Wyss-Coray Tony

    2008-01-01

    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 co...

  9. Bioluminescence imaging of chondrocytes in rabbits by intraarticular injection of D-luciferin

    Luciferase is one of the most commonly used reporter enzymes in the field of in vivo optical imaging. D-luciferin, the substrate for firefly luciferase has very high cost that allows this kind of experiment limited to small animals such as mice and rats. In this current study, we validated local injection of D-luciferin in the articular capsule for bioluminescence imaging in rabbits. Chondrocytes were cultured and infected by replication-defective adenoviral vector encoding firefly luciferase (Fluc). Chondrocytes expressing Fluc were injected or implanted in the left knee joint. The rabbits underwent optical imaging studies after local injection of D-luciferin at 1, 5, 7, 9 days after cellular administration. We sought whether optimal imaging signals was could be by a cooled CCD camera after local injection of D-luciferin. Imaging signal was not observed from the left knee joint after intraperitoneal injection of D-luciferin (15 mg/kg), whereas it was observed after intraarticular injection. Photon intensity from the left knee joint of rabbits was compared between cell injected and implanted groups after intraarticular injection of D-luciferin. During the period of imaging studies, photon intensity of the cell implanted group was 5-10 times higher than that of the cell injected group. We successfully imaged chondrocytes expressing Fluc after intraarticular injection of D-luciferin. This technique may be further applied to develop new drugs for knee joint disease

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

    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.

  11. 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

    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 14C(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 14C-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 14C-d-luciferin in firefly luciferase-expressing tissues could be observed in vivo. The data obtained with 14C-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

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

    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.

  13. Noninvasive bioluminescence imaging of the dynamics of sanguinarine induced apoptosis via activation of reactive oxygen species

    Dai, Yunpeng; Shi, Yaru; Zeng, Qi; Wang, Fu

    2016-01-01

    Most chemotherapeutic drugs exert their anti-tumor effects primarily by triggering a final pathway leading to apoptosis. Noninvasive imaging of apoptotic events in preclinical models would greatly facilitate the development of apoptosis-inducing compounds and evaluation of their therapeutic efficacy. Here we employed a cyclic firefly luciferase (cFluc) reporter to screen potential pro-apoptotic compounds from a number of natural agents. We demonstrated that sanguinarine (SANG) could induce apoptosis in a dose- and time-dependent manner in UM-SCC-22B head and neck cancer cells. Moreover, SANG-induced apoptosis was associated with the generation of reactive oxygen species (ROS) and activation of c-Jun-N-terminal kinase (JNK) and nuclear factor-kappaB (NF-κB) signal pathways. After intravenous administration with SANG in 22B-cFluc xenograft models, a dramatic increase of luminescence signal can be detected as early as 48 h post-treatment, as revealed by longitudinal bioluminescence imaging in vivo. Remarkable apoptotic cells reflected from ex vivo TUNEL staining confirmed the imaging results. Importantly, SANG treatment caused distinct tumor growth retardation in mice compared with the vehicle-treated group. Taken together, our results showed that SANG is a candidate anti-tumor drug and noninvasive imaging of apoptosis using cFluc reporter could provide a valuable tool for drug development and therapeutic efficacy evaluation. PMID:26968950

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

    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.

  15. Efficacy of Lantibiotic Treatment of Staphylococcus aureus-Induced Skin Infections, Monitored by In Vivo Bioluminescent Imaging.

    van Staden, Anton Du Preez; Heunis, Tiaan; Smith, Carine; Deane, Shelly; Dicks, Leon M T

    2016-07-01

    Staphylococcus aureus is a bacterial pathogen responsible for the majority of skin and soft tissue infections. Antibiotics are losing their efficacy as treatment for skin and soft tissue infections as a result of increased resistance in a variety of pathogens, including S. aureus It is thus imperative to explore alternative antimicrobial treatments to ensure future treatment options for skin and soft tissue infections. A select few lantibiotics, a group of natural defense peptides produced by bacteria, inhibit the growth of numerous clinical S. aureus isolates, including methicillin-resistant strains. In this study, the antimicrobial activities of nisin, clausin, and amyloliquecidin, separately administered, were compared to that of a mupirocin-based ointment, which is commonly used as treatment for S. aureus-induced skin infections. Full-thickness excisional wounds, generated on the dorsal surfaces of mice, were infected with a bioluminescent strain of S. aureus (strain Xen 36). The infections were monitored in real time using in vivo bioluminescent imaging. Lantibiotic treatments significantly reduced the bioluminescence of S. aureus Xen 36 to a level similar to that recorded with mupirocin treatment. Wound closure, however, was more pronounced during lantibiotic treatment. Lantibiotics thus have the potential to be used as an alternative treatment option for S. aureus-induced skin infections. PMID:27067340

  16. Non-invasive visualisation of the development of peritoneal carcinomatosis and tumour regression after 213Bi-radioimmunotherapy using bioluminescence imaging

    Non-invasive imaging of tumour development remains a challenge, especially for tumours in the intraperitoneal cavity. Therefore, the aim of this study was the visualisation of both the development of peritoneal carcinomatosis and tumour regression after radioimmunotherapy with tumour-specific 213Bi-Immunoconjugates, via in vivo bioluminescence imaging of firefly luciferase-transfected cells. Human diffuse-type gastric cancer cells expressing mutant d9-E-cadherin were stably transfected with firefly luciferase (HSC45-M2-luc). For bioluminescence imaging, nude mice were inoculated intraperitoneally with 1 x 107 HSC45-M2-luc cells. On days 4 and 8 after tumour cell inoculation, imaging was performed following D-luciferin injection using a cooled CCD camera with an image intensifier unit. For therapy, mice were injected with 2.7 MBq 213Bi-d9MAb targeting d9-E-cadherin on day 8 after tumour cell inoculation. Bioluminescence images were taken every 4 days to monitor tumour development. After i.p. inoculation of HSC45-M2-luc cells into nude mice, development as well as localisation of peritoneal carcinomatosis could be visualised using bioluminescence imaging. Following 213Bi-d9MAb therapy on day 8 after intraperitoneal inoculation of HSC45-M2-luc cells, small tumour nodules were totally eliminated and larger nodules showed a clear reduction in size on day 12 after tumour cell inoculation. Subsequently a recurrence of tumour mass was observed, starting from the remaining tumour spots. By measuring the mean grey level intensity, tumour development over time could be demonstrated. Non-invasive bioluminescence imaging permits visualisation of the development of peritoneal carcinomatosis, localisation of tumour in the intraperitoneal cavity and evaluation of therapeutic success after 213Bi-d9MAb treatment. (orig.)

  17. Recovering 3D tumor locations from 2D bioluminescence images and registration with CT images

    Huang, Xiaolei; Metaxas, Dimitris N.; Menon, Lata G.; Mayer-Kuckuk, Philipp; Bertino, Joseph R.; Banerjee, Debabrata

    2006-02-01

    In this paper, we introduce a novel and efficient algorithm for reconstructing the 3D locations of tumor sites from a set of 2D bioluminescence images which are taken by a same camera but after continually rotating the object by a small angle. Our approach requires a much simpler set up than those using multiple cameras, and the algorithmic steps in our framework are efficient and robust enough to facilitate its use in analyzing the repeated imaging of a same animal transplanted with gene marked cells. In order to visualize in 3D the structure of the tumor, we also co-register the BLI-reconstructed crude structure with detailed anatomical structure extracted from high-resolution microCT on a single platform. We present our method using both phantom studies and real studies on small animals.

  18. Live Cell Bioluminescence Imaging in Temporal Reaction of G Protein-Coupled Receptor for High-Throughput Screening and Analysis.

    Hattori, Mitsuru; Ozawa, Takeaki

    2016-01-01

    G protein-coupled receptors (GPCRs) are notable targets of basic therapeutics. Many screening methods have been established to identify novel agents for GPCR signaling in a high-throughput manner. However, information related to the temporal reaction of GPCR with specific ligands remains poor. We recently developed a bioluminescence method for the quantitative detection of the interaction between GPCR and β-arrestin using split luciferase complementation. To monitor time-course variation of the interactions, a new imaging system contributes to the accurate evaluation of drugs for GPCRs in a high-throughput manner. PMID:27424906

  19. Design and development of high bioluminescent resonance energy transfer efficiency hybrid-imaging constructs.

    Kumar, Manoj; Kovalski, Letícia; Broyles, David; Hunt, Eric A; Daftarian, Pirouz; Dikici, Emre; Daunert, Sylvia; Deo, Sapna K

    2016-04-01

    Here we describe the design and construction of an imaging construct with high bioluminescent resonance energy transfer (BRET) efficiency that is composed of multiple quantum dots (QDs; λem = 655 nm) self-assembled onto a bioluminescent protein, Renilla luciferase (Rluc). This is facilitated by the streptavidin-biotin interaction, allowing the facile formation of a hybrid-imaging construct (HIC) comprising up to six QDs (acceptor) grafted onto a light-emitting Rluc (donor) core. The resulting assembly of multiple acceptors surrounding a donor permits this construct to exhibit high resonance energy transfer efficiency (∼64.8%). The HIC was characterized using fluorescence excitation anisotropy measurements and high-resolution transmission electron microscopy. To demonstrate the application of our construct, a generation-5 (G5) polyamidoamine dendrimer (PAMAM) nanocarrier was loaded with our HIC for in vitro and in vivo imaging. We envision that this design of multiple acceptors and bioluminescent donor will lead to the development of new BRET-based systems useful in sensing, imaging, and other bioanalytical applications. PMID:26772160

  20. Quantitative bioluminescence imaging of transgene expression in intact porcine antral follicles in vitro

    Jung, Song-Yi; Willard, Scott T

    2014-01-01

    Background The porcine oocyte maturation in vivo occurs within the ovarian follicle and is regulated by the interactions between oocytes and surrounding follicular components, including theca, granulosa, and cumulus cells, and follicular fluid. Therefore, the antral follicle is an essential microenvironment for efficient oocyte maturation and its developmental competence. Quantitative bioluminescence imaging of firefly luciferase reporter genes in an intact antral follicle would allow investi...

  1. Design and Synthesis of an Alkynyl Luciferin Analogue for Bioluminescence Imaging.

    Steinhardt, Rachel C; O'Neill, Jessica M; Rathbun, Colin M; McCutcheon, David C; Paley, Miranda A; Prescher, Jennifer A

    2016-03-01

    Herein, the synthesis and characterization of an alkyne-modified luciferin is reported. This bioluminescent probe was accessed using C-H activation methodology and was found to be stable in solution and capable of light production with firefly luciferase. The luciferin analogue was also cell permeant and emitted more redshifted light than d-luciferin, the native luciferase substrate. Based on these features, the alkynyl luciferin will be useful for a variety of imaging applications. PMID:26784889

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

    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.

  3. Construction of a bioluminescence reporter plasmid for Francisella tularensis

    Bina, Xiaowen R.; Miller, Mark A.; James E Bina

    2010-01-01

    A Francisella tularensis shuttle vector that constitutively expresses the Photorhabdus luminescens lux operon in type A and type B strains of F. tularensis was constructed. The bioluminescence reporter plasmid was introduced into the live vaccine strain of F. tularensis and used to follow F. tularensis growth in a murine intranasal challenge model in real time by bioluminescence imaging. The results show that the new bioluminescence reporter plasmid represents a useful tool for tularemia rese...

  4. Firefly Luciferase Mutants Allow Substrate-Selective Bioluminescence Imaging in the Mouse Brain.

    Adams, Spencer T; Mofford, David M; Reddy, G S Kiran Kumar; Miller, Stephen C

    2016-04-11

    Bioluminescence imaging is a powerful approach for visualizing specific events occurring inside live mice. Animals can be made to glow in response to the expression of a gene, the activity of an enzyme, or the growth of a tumor. But bioluminescence requires the interaction of a luciferase enzyme with a small-molecule luciferin, and its scope has been limited by the mere handful of natural combinations. Herein, we show that mutants of firefly luciferase can discriminate between natural and synthetic substrates in the brains of live mice. When using adeno-associated viral (AAV) vectors to express luciferases in the brain, we found that mutant luciferases that are inactive or weakly active with d-luciferin can light up brightly when treated with the aminoluciferins CycLuc1 and CycLuc2 or their respective FAAH-sensitive luciferin amides. Further development of selective luciferases promises to expand the power of bioluminescence and allow multiple events to be imaged in the same live animal. PMID:26991209

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

    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.

  6. A bright cyan-excitable orange fluorescent protein facilitates dual-emission microscopy and enhances bioluminescence imaging in vivo.

    Chu, Jun; Oh, Younghee; Sens, Alex; Ataie, Niloufar; Dana, Hod; Macklin, John J; Laviv, Tal; Welf, Erik S; Dean, Kevin M; Zhang, Feijie; Kim, Benjamin B; Tang, Clement Tran; Hu, Michelle; Baird, Michelle A; Davidson, Michael W; Kay, Mark A; Fiolka, Reto; Yasuda, Ryohei; Kim, Douglas S; Ng, Ho-Leung; Lin, Michael Z

    2016-07-01

    Orange-red fluorescent proteins (FPs) are widely used in biomedical research for multiplexed epifluorescence microscopy with GFP-based probes, but their different excitation requirements make multiplexing with new advanced microscopy methods difficult. Separately, orange-red FPs are useful for deep-tissue imaging in mammals owing to the relative tissue transmissibility of orange-red light, but their dependence on illumination limits their sensitivity as reporters in deep tissues. Here we describe CyOFP1, a bright, engineered, orange-red FP that is excitable by cyan light. We show that CyOFP1 enables single-excitation multiplexed imaging with GFP-based probes in single-photon and two-photon microscopy, including time-lapse imaging in light-sheet systems. CyOFP1 also serves as an efficient acceptor for resonance energy transfer from the highly catalytic blue-emitting luciferase NanoLuc. An optimized fusion of CyOFP1 and NanoLuc, called Antares, functions as a highly sensitive bioluminescent reporter in vivo, producing substantially brighter signals from deep tissues than firefly luciferase and other bioluminescent proteins. PMID:27240196

  7. Autonomously Bioluminescent Mammalian Cells For Continuous And Real-Time Monitoring Of Cytotoxicity

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

    2013-01-01

    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 fac...

  8. Bioluminescence imaging in a medium-sized animal by local injection of d-luciferin

    Luciferase is one of the most commonly used reporter enzymes in the field of molecular imaging. D-luciferin is known as the substrate for luciferase enzyme and its cost is very expensive. Therefore, the bioluminescence molecular imaging study has been allowed in small animals such as mice and rats. In this current study, we validated local injection of D-luciferin in articular capsule for bioluminescence imaging in rabbits. Chondrocytes were cultured and infected by replication-defective adenoviral vector encoding firefly luciferase. And then was performed different method of chondrocyte cell injection and transplantation into the knee of rabbits. The rabbits underwent imaging by cooled CCD camera after local injection of D-luciferin (3mg) into experimental knee joint as well as contralateral normal knee joint on days 1, 5, 7, 9. We sought whether optimal imaging signal was acquired by using cooled CCD camera after local injection of D-luciferin. We successfully visualized injected or transplanted cells in knee joint by local injection of D-luciferin. Total photon flux (7.86E+08 p/s/cm2/sr) from the knee joint transplanted with cells approximately increased 10-fold more than (9.43E+07p/s/cm2/sr) that from injected knee joints until 7 day. Imaging signal was observed in transplanted joints until day 9 after surgery while signal from injected knee was observed by day 7 after injection. We successfully carried out bioluminescence imaging study with medium sized animal by local injection of small amount of D-luciferin. Survival of chondrocytes were prolonged when surgically transplanted in joints than when directly injected in joint space

  9. Bioluminescence imaging of leukemia cell lines in vitro and in mouse xenografts: effects of monoclonal and polyclonal cell populations on intensity and kinetics of photon emission

    Christoph Sandra

    2013-01-01

    Full Text Available Abstract Background We investigated the utility of bioluminescence imaging (BLI using firefly luciferase in monoclonal and polyclonal populations of leukemia cells in vitro and in vivo. Methods Monoclonal and polyclonal human lymphoid and myeloid leukemia cell lines transduced with firefly luciferase were used for BLI. Results Kinetics and dynamics of bioluminescence signal were cell line dependent. Luciferase expression decreased significantly over time in polyclonal leukemia cells in vitro. Transplantation of polyclonal luciferase-tagged cells in mice resulted in inconsistent signal intensity. After selection of monoclonal cell populations, luciferase activity was stable, equal kinetic and dynamic of bioluminescence intensity and strong correlation between cell number and light emission in vitro were observed. We obtained an equal development of leukemia burden detected by luciferase activity in NOD-scid-gamma mice after transplantation of monoclonal populations. Conclusion The use of monoclonal leukemia cells selected for stable and equal luciferase activity is recommended for experiments in vitro and xenograft mouse models. The findings are highly significant for bioluminescence imaging focused on pre-clinical drug development.

  10. Detection and quantitation of circulating tumor cell dynamics by bioluminescence imaging in an orthotopic mammary carcinoma model.

    Laura Sarah Sasportas

    Full Text Available Circulating tumor cells (CTCs have been detected in the bloodstream of both early-stage and advanced cancer patients. However, very little is know about the dynamics of CTCs during cancer progression and the clinical relevance of longitudinal CTC enumeration. To address this, we developed a simple bioluminescence imaging assay to detect CTCs in mouse models of metastasis. In a 4T1 orthotopic metastatic mammary carcinoma mouse model, we demonstrated that this quantitative method offers sensitivity down to 2 CTCs in 0.1-1mL blood samples and high specificity for CTCs originating from the primary tumor, independently of their epithelial status. In this model, we simultaneously monitored blood CTC dynamics, primary tumor growth, and lung metastasis progression over the course of 24 days. Early in tumor development, we observed low numbers of CTCs in blood samples (10-15 cells/100 µL and demonstrated that CTC dynamics correlate with viable primary tumor growth. To our knowledge, these data represent the first reported use of bioluminescence imaging to detect CTCs and quantify their dynamics in any cancer mouse model. This new assay is opening the door to the study of CTC dynamics in a variety of animal models. These studies may inform clinical decision on the appropriate timing of blood sampling and value of longitudinal CTC enumeration in cancer patients.

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

    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

  12. Accounting for systematic errors in bioluminescence imaging to improve quantitative accuracy

    Taylor, Shelley L.; Perry, Tracey A.; Styles, Iain B.; Cobbold, Mark; Dehghani, Hamid

    2015-07-01

    Bioluminescence imaging (BLI) is a widely used pre-clinical imaging technique, but there are a number of limitations to its quantitative accuracy. This work uses an animal model to demonstrate some significant limitations of BLI and presents processing methods and algorithms which overcome these limitations, increasing the quantitative accuracy of the technique. The position of the imaging subject and source depth are both shown to affect the measured luminescence intensity. Free Space Modelling is used to eliminate the systematic error due to the camera/subject geometry, removing the dependence of luminescence intensity on animal position. Bioluminescence tomography (BLT) is then used to provide additional information about the depth and intensity of the source. A substantial limitation in the number of sources identified using BLI is also presented. It is shown that when a given source is at a significant depth, it can appear as multiple sources when imaged using BLI, while the use of BLT recovers the true number of sources present.

  13. Quantification of bioluminescence images of point source objects using diffusion theory models

    A simple approach for estimating the location and power of a bioluminescent point source inside tissue is reported. The strategy consists of using a diffuse reflectance image at the emission wavelength to determine the optical properties of the tissue. Following this, bioluminescence images are modelled using a single point source and the optical properties from the reflectance image, and the depth and power are iteratively adjusted to find the best agreement with the experimental image. The forward models for light propagation are based on the diffusion approximation, with appropriate boundary conditions. The method was tested using Monte Carlo simulations, Intralipid tissue-simulating phantoms and ex vivo chicken muscle. Monte Carlo data showed that depth could be recovered within 6% for depth 4-12 mm, and the corresponding relative source power within 12%. In Intralipid, the depth could be estimated within 8% for depth 4-12 mm, and the relative source power, within 20%. For ex vivo tissue samples, source depths of 4.5 and 10 mm and their relative powers were correctly identified

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

    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.

  15. In Vivo Bioluminescent Imaging (BLI: Noninvasive Visualization and Interrogation of Biological Processes in Living Animals

    Steven Ripp

    2010-12-01

    Full Text Available In vivo bioluminescent imaging (BLI is increasingly being utilized as a method for modern biological research. This process, which involves the noninvasive interrogation of living animals using light emitted from luciferase-expressing bioreporter cells, has been applied to study a wide range of biomolecular functions such as gene function, drug discovery and development, cellular trafficking, protein-protein interactions, and especially tumorigenesis, cancer treatment, and disease progression. This article will review the various bioreporter/biosensor integrations of BLI and discuss how BLI is being applied towards a new visual understanding of biological processes within the living organism.

  16. 18F-fluoride PET imaging in a nude rat model of bone metastasis from breast cancer: Comparison with 18F-FDG and bioluminescence imaging

    Introduction: Clinically-relevant animal models and appropriate imaging diagnostic tools are essential to study cancer and develop novel therapeutics. We evaluated a model of bone metastasis in nude rats by micro-PET and bioluminescence imaging. Methods: A bone metastasis model was produced by intracardiac injection of osteotropic MDA-MB-231Bo-Luc human breast cancer cells into nude rats. Bioluminescence imaging and micro-PET scans using 18F-FDG and 18F-fluoride were acquired serially for 5 weeks. We correlated bioluminescence imaging, 18F-FDG and 18F-fluoride PET images, and histological slides. Results: Multiple bone metastases were successfully evaluated by bioluminescence imaging and 18F-FDG and 18F-fluoride PET scans. Bioluminescence photon flux increased exponentially on weekly follow-up. 18F-FDG PET revealed increased FDG uptake at the spine and bilaterally in the hind legs in week 2 images, and showed a progressive pattern up to 4 weeks that correlated with bioluminescence imaging. 18F-fluoride PET showed minimal abnormal findings in week 2 images, but it showed an irregular pattern at the spine from week 3 or 4 images. On quantitative analysis with standardized uptake values, a pattern of gradual increase was observed from week 2 to week 4 in both 18F-FDG PET and fluoride PET. Histopathological examination confirmed the formation of osteolytic metastasis and necrosis of the distal femur, which appeared as a photon defect on PET scans. Conclusion: Developing bone metastasis from breast cancer in a nude rat model was successfully evaluated with an animal PET imaging system and bioluminescence imaging. This nude rat model of bone metastasis, which can be evaluated by PET imaging, may be a valuable tool for evaluating early responses to novel therapeutics

  17. Novel bioluminescent quantitative detection of nucleic acid amplification in real-time.

    Olga A Gandelman

    Full Text Available BACKGROUND: The real-time monitoring of polynucleotide amplification is at the core of most molecular assays. This conventionally relies on fluorescent detection of the amplicon produced, requiring complex and costly hardware, often restricting it to specialised laboratories. PRINCIPAL FINDINGS: Here we report the first real-time, closed-tube luminescent reporter system for nucleic acid amplification technologies (NAATs enabling the progress of amplification to be continuously monitored using simple light measuring equipment. The Bioluminescent Assay in Real-Time (BART continuously reports through bioluminescent output the exponential increase of inorganic pyrophosphate (PPi produced during the isothermal amplification of a specific nucleic acid target. BART relies on the coupled conversion of inorganic pyrophosphate (PPi produced stoichiometrically during nucleic acid synthesis to ATP by the enzyme ATP sulfurylase, and can therefore be coupled to a wide range of isothermal NAATs. During nucleic acid amplification, enzymatic conversion of PPi released during DNA synthesis into ATP is continuously monitored through the bioluminescence generated by thermostable firefly luciferase. The assay shows a unique kinetic signature for nucleic acid amplifications with a readily identifiable light output peak, whose timing is proportional to the concentration of original target nucleic acid. This allows qualitative and quantitative analysis of specific targets, and readily differentiates between negative and positive samples. Since quantitation in BART is based on determination of time-to-peak rather than absolute intensity of light emission, complex or highly sensitive light detectors are not required. CONCLUSIONS: The combined chemistries of the BART reporter and amplification require only a constant temperature maintained by a heating block and are shown to be robust in the analysis of clinical samples. Since monitoring the BART reaction requires only a

  18. Bioluminescence imaging of cord blood derived mesenchymal stem cell transplanatation into myocardium

    Min, Jung Joon; Ahn, Young Keun; Moon, Sung Min; Lim, Sang Yup; Yun, Kyung Ho; Heo, Young Jun; Song, Ho Chun; Jeong, Myung Ho; Bom, Hee Seung [School of Medicine, Chonnam National University, Gwangju (Korea, Republic of)

    2004-07-01

    The conventional method of analyzing myocardial cell transplanation relies on postmortem histology. We sought to demonstrate the feasibility of longitudinal monitoring transplanted cell survival in living animals using optical imaging techniques. Umblical cord blood was collected upon delivery with informed consent. Umblical mononuclear cells were obtained by negative immuno-depletion of CD3, CD14, CD19, CD38, CD66b, and glycophorin- A positive cells, followed by Ficoll- Paque density gradient centrifugation, and plated in non-coated tissue culture flasks in expansion medium. Cells were allowed to adhere overnight, thereafter non-adherent cells were washed out with medium changes. After getting the MSCs, they were transfected [multiplicity of infection (MOl) = 40) with Ad-CMV-Fluc overnight. Rats (n=4) underwent intramyocardial injection of 5 x 10{sup 5} MSCs expressing firefly luciferase (Fluc) reporter gene. Optical bioluminescence imaging was performed using the charged-coupled device camera (Xenogen) from the 1st day of transplantion. Cardiac bioluminescence signals were present from 2nd day of transplantation. Cardiac signals were clearly present at day 2 (9.2x10{sup 3}p/s/cm{sup 2}/sr). The signal reduced from day 3. The locations, magnitude, and survival duration of cord blood derived MSCs were monitored noninvasively. With further development, molecular imaging studies should add critical insights into cardiac cell transplantation.

  19. Bioluminescence imaging of cord blood derived mesenchymal stem cell transplanatation into myocardium

    The conventional method of analyzing myocardial cell transplanation relies on postmortem histology. We sought to demonstrate the feasibility of longitudinal monitoring transplanted cell survival in living animals using optical imaging techniques. Umblical cord blood was collected upon delivery with informed consent. Umblical mononuclear cells were obtained by negative immuno-depletion of CD3, CD14, CD19, CD38, CD66b, and glycophorin- A positive cells, followed by Ficoll- Paque density gradient centrifugation, and plated in non-coated tissue culture flasks in expansion medium. Cells were allowed to adhere overnight, thereafter non-adherent cells were washed out with medium changes. After getting the MSCs, they were transfected [multiplicity of infection (MOl) = 40) with Ad-CMV-Fluc overnight. Rats (n=4) underwent intramyocardial injection of 5 x 105 MSCs expressing firefly luciferase (Fluc) reporter gene. Optical bioluminescence imaging was performed using the charged-coupled device camera (Xenogen) from the 1st day of transplantion. Cardiac bioluminescence signals were present from 2nd day of transplantation. Cardiac signals were clearly present at day 2 (9.2x103p/s/cm2/sr). The signal reduced from day 3. The locations, magnitude, and survival duration of cord blood derived MSCs were monitored noninvasively. With further development, molecular imaging studies should add critical insights into cardiac cell transplantation

  20. Relation between deep bioluminescence and oceanographic variables: A statistical analysis using time-frequency decompositions

    Martini, S.; Nerini, D.; Tamburini, C.

    2014-09-01

    We consider the statistical analysis of a 1.7-year high-frequency sampled time series, between 2009 and 2010, recorded at the ANTARES observatory in the deep NW Mediterranean Sea (2475 m depth). The objective was to estimate relationships between bioluminescence and environmental time series (temperature, salinity and current speed). As this entire dataset is characterized by non-linearity and non-stationarity, two time-frequency decomposition methods (wavelet and Hilbert-Huang) were used. These mathematical methods are dedicated to the analysis of a signal at various time and frequencies scales. This work propose some statistical tools dedicated to the study of relationships between two time series. Our study highlights three events of high bioluminescence activity in March 2009, December 2009 and March 2010. We demonstrate that the two events occurring in March 2009 and 2010 are correlated to the arrival of newly formed deep water masses at frequencies of approximately 4.8×10-7 (period of 24.1 days). In contrast, the event in December 2009 is only correlated with current speed at frequencies of approximately 1.9×10-6 (period of 6.0 days). The use of both wavelet and Hilbert-Huang transformations has proven to be successful for the analysis of multivariate time series. These methods are well-suited in a context of the increasing number of long time series recorded in oceanography.

  1. Bioluminescence Imaging of NADPH Oxidase Activity in Different Animal Models

    Han, Wei; Li, Hui; Segal, Brahm H.; Blackwell, Timothy S.

    2012-01-01

    NADPH oxidase is a critical enzyme that mediates antibacterial and antifungal host defense. In addition to its role in antimicrobial host defense, NADPH oxidase has critical signaling functions that modulate the inflammatory response 1. Thus, the development of a method to measure in "real-time" the kinetics of NADPH oxidase-derived ROS generation is expected to be a valuable research tool to understand mechanisms relevant to host defense, inflammation, and injury.

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

    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 presente

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

    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

  4. Assessing the effect of EPO on tumor oxygenation and radioresponsiveness via in vivo bioluminescence imaging

    Evaluating tumor kill by volume measurement lacks sensitivity while in vivo-in vitro and histological assays are unsuitable for serial measurements. In vivo bioluminescence imaging (BI) nondestructively measures the number of metabolically active cells containing luciferase (LUC) over time. In this paper, the effect of erythropoietin (EPO) on tumor oxygenation and radioresponsivenessis is studied using BI and conventional methods. Murine adenocarcinoma cells, transfected with the LUC gene, were placed in the flank of BALB/C mice. EPO 1 u/g or saline was injected sc tiw for two weeks, starting the day of transplant. Mice then underwent irradiation (XRT) or pO2 measurement with an optical probe. In BI, mice were injected with luciferin and total photon flux (TPF) measured with a CCD camera. In vitro, cells were plated, irradiated and incubated at 37 deg C. Initial hematocrit was 47% (n=119) vs. 61% in EPO-treated mice (n=23, p2 (6.4 vs. 4.7 mm Hg, p=0.04) than controls. For 1-3x7 Gy, TPF was stable for 2 days after the start of XRT, then fell precipitously. Two weeks post XRT, TPF was 10-5 the initial value and a nidus of LUC activity persisted for months in most tumors. Tumor volume decreased only 1-2 orders of magnitude. For 3x7 Gy, tumor regrew in 1/11 EPO-TM and controls (p=NS.) For 1x7 Gy, tumors regrew in 4/6 EPO-TM and 2/4 controls (p=NS). TPF did not increase with tumor regrowth. Recurrent tumors exhibited lower median pO2 (2.1 mm Hg, p=.003) and higher hypoxic fraction than controls. A clonogenic assay yielded D10 = 3.7 Gy with all colonies expressing LUC. The TPF of 0-Gy treated wells rose significantly over incubation, while that of wells treated to 10 Gy was unchanged. Though EPO improved tumor oxygenation, no effect on XRT-mediated cell kill was seen. BI measured tumor killing in vivo over a broad dynamic range. The results suggest that cell killing in vivo is a multistep process, amplified by humoral factors

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

    Alexandrakis, George; Rannou, Fernando R; Chatziioannou, Arion F

    2005-01-01

    The feasibility and limits in performing tomographic bioluminescence imaging with a combined optical-PET (OPET) system was 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 °C. The TOAST finite-element code was employed to simulate the diffuse transport of photons emitted from biolumi...

  6. In Vivo Bioluminescent Imaging of ATP-Binding Cassette Transporter-Mediated Efflux at the Blood-Brain Barrier.

    Bakhsheshian, Joshua; Wei, Bih-Rong; Hall, Matthew D; Simpson, R Mark; Gottesman, Michael M

    2016-01-01

    We provide a detailed protocol for imaging ATP-binding cassette subfamily G member 2 (ABCG2) function at the blood-brain barrier (BBB) of transgenic mice. D-Luciferin is specifically transported by ABCG2 found on the apical side of endothelial cells at the BBB. The luciferase-luciferin enzymatic reaction produces bioluminescence, which allows a direct measurement of ABCG2 function at the BBB. Therefore bioluminescence imaging (BLI) correlates with ABCG2 function at the BBB and this can be measured by administering luciferin in a mouse model that expresses luciferase in the brain parenchyma. BLI allows for a relatively low-cost alternative for studying transporter function in vivo compared to other strategies such as positron emission tomography. This method for imaging ABCG2 function at the BBB can be used to investigate pharmacokinetic inhibition of the transporter. PMID:27424909

  7. Fast iterative image reconstruction methods for fully 3D multispectral bioluminescence tomography

    We investigate fast iterative image reconstruction methods for fully 3D multispectral bioluminescence tomography for applications in small animal imaging. Our forward model uses a diffusion approximation for optically inhomogeneous tissue, which we solve using a finite element method (FEM). We examine two approaches to incorporating the forward model into the solution of the inverse problem. In a conventional direct calculation approach one computes the full forward model by repeated solution of the FEM problem, once for each potential source location. We describe an alternative on-the-fly approach where one does not explicitly solve for the full forward model. Instead, the solution to the forward problem is included implicitly in the formulation of the inverse problem, and the FEM problem is solved at each iteration for the current image estimate. We evaluate the convergence speeds of several representative iterative algorithms. We compare the computation cost of those two approaches, concluding that the on-the-fly approach can lead to substantial reductions in total cost when combined with a rapidly converging iterative algorithm

  8. Metabolic imaging in microregions of tumors and normal tissues with bioluminescence and photon counting

    A method has been developed for metabolic imaging on a microscopic level in tumors, tumor spheroids, and normal tissues. The technique makes it possible to determine the spatial distribution of glucose, lactate, and ATP in absolute terms at similar locations within tissues or cell aggregates. The substrate distributions are registered in serial cryostat sections from tissue cryobiopsies or from frozen spheroids with the use of bioluminescence reactions. The light emission is measured directly by a special imaging photon counting system enabling on-line image analysis. The technique has been applied to human breast cancer xenografts, to spheroids originating from a human colon adenocarcinoma, and to skeletal rat muscle. Preliminary data obtained indicate that heterogeneities in the substrate distributions measured are much more pronounced in tumors than in normal tissue. There was no obvious correlation among the three quantities measured at similar locations within the tissues. The distribution of ATP corresponded well with the histological structure of larger spheroids; values were low in the necrotic center and high in the viable rim of these cell aggregates

  9. In Vivo Tracking of Systemically Administered Allogeneic Bone Marrow Mesenchymal Stem Cells in Normal Rats through Bioluminescence Imaging

    Cao, Juan; Hou, Shike; Ding, Hui; Liu, Ziquan; Song, Meijuan; Qin, Xiaojing; Wang, Xue; Yu, Mengyang; Sun, Zhiguang; Liu, Jinyang; Sun, Shuli; Xiao, Peixin

    2016-01-01

    Recently, mesenchymal stem cells (MSCs) are increasingly used as a panacea for multiple types of disease short of effective treatment. Dozens of clinical trials published demonstrated strikingly positive therapeutic effects of MSCs. However, as a specific agent, little research has focused on the dynamic distribution of MSCs after in vivo administration. In this study, we track systemically transplanted allogeneic bone marrow mesenchymal stem cells (BMSCs) in normal rats through bioluminescence imaging (BLI) in real time. Ex vivo organ imaging, immunohistochemistry (IHC), and RT-PCR were conducted to verify the histological distribution of BMSCs. Our results showed that BMSCs home to the dorsal skin apart from the lungs and kidneys after tail vein injection and could not be detected 14 days later. Allogeneic BMSCs mainly appeared not at the parenchymatous organs but at the subepidermal connective tissue and adipose tissue in healthy rats. There were no significant MSCs-related adverse effects except for transient decrease in neutrophils. These findings will provide experimental evidences for a better understanding of the biocharacteristics of BMSCs.

  10. Physicochemical characterization and in vivo bioluminescence imaging of nanostructured lipid carriers for targeting the brain: apomorphine as a model drug

    Hsu, Shu-Hui; Wen, Chih-Jen; Al-Suwayeh, S. A.; Chang, Hui-Wen; Yen, Tzu-Chen; Fang, Jia-You

    2010-10-01

    Nanostructured lipid carriers (NLCs) were prepared to investigate whether the duration of brain targeting and accumulation of drugs in the brain can be improved by intravenous delivery. NLCs were developed using cetyl palmitate as the lipid matrix, squalene as the cationic surfactant, and Pluronic F68, polysorbate 80 and polyethylene glycol as the interfacial additives. Solid lipid nanoparticles (SLNs) and lipid emulsions (LEs) were also prepared for comparison. An anti-Parkinson's drug, apomorphine, was used as the model drug. Nuclear magnetic resonance and differential scanning calorimetry showed possible interactions between the solid and liquid lipids in the inner core. The lipid nanoparticles with different compositions were characterized by mean size, zeta potential, apomorphine encapsulation and in vitro drug release. NLCs were 370-430 nm in size, which was between the sizes of the SLNs and LEs. A cationic surfactant was used to produce a positive surface charge of 42-50 mV. The base form of apomorphine was successfully entrapped by NLCs with an entrapment percentage of > 60%. The loading of apomorphine in nanoparticles resulted in a slower release behavior compared to the aqueous solution, with LEs showing the lowest release. In vivo real-time bioluminescence imaging of the rat brain revealed that NLCs could be targeted, through certain vessels, to selected brain regions. This effect was further confirmed by imaging the entire brain and brain slices. The results indicated that NLCs with moderate additives are a promising controlled-release and drug-targeting system.

  11. In Vivo Tracking of Systemically Administered Allogeneic Bone Marrow Mesenchymal Stem Cells in Normal Rats through Bioluminescence Imaging.

    Cao, Juan; Hou, Shike; Ding, Hui; Liu, Ziquan; Song, Meijuan; Qin, Xiaojing; Wang, Xue; Yu, Mengyang; Sun, Zhiguang; Liu, Jinyang; Sun, Shuli; Xiao, Peixin; Lv, Qi; Fan, Haojun

    2016-01-01

    Recently, mesenchymal stem cells (MSCs) are increasingly used as a panacea for multiple types of disease short of effective treatment. Dozens of clinical trials published demonstrated strikingly positive therapeutic effects of MSCs. However, as a specific agent, little research has focused on the dynamic distribution of MSCs after in vivo administration. In this study, we track systemically transplanted allogeneic bone marrow mesenchymal stem cells (BMSCs) in normal rats through bioluminescence imaging (BLI) in real time. Ex vivo organ imaging, immunohistochemistry (IHC), and RT-PCR were conducted to verify the histological distribution of BMSCs. Our results showed that BMSCs home to the dorsal skin apart from the lungs and kidneys after tail vein injection and could not be detected 14 days later. Allogeneic BMSCs mainly appeared not at the parenchymatous organs but at the subepidermal connective tissue and adipose tissue in healthy rats. There were no significant MSCs-related adverse effects except for transient decrease in neutrophils. These findings will provide experimental evidences for a better understanding of the biocharacteristics of BMSCs. PMID:27610137

  12. Physicochemical characterization and in vivo bioluminescence imaging of nanostructured lipid carriers for targeting the brain: apomorphine as a model drug

    Hsu, Shu-Hui [Department of Pharmacy, Chia Nan University of Pharmacy and Science, Tainan 717, Taiwan (China); Wen, Chih-Jen; Yen, Tzu-Chen [Animal Molecular Imaging Center, Chang Gung Memorial Hospital, Kweishan, Taoyuan 333, Taiwan (China); Al-Suwayeh, S A; Fang, Jia-You [Department of Pharmaceutics, College of Pharmacy, King Saud University, Riyadh (Saudi Arabia); Chang, Hui-Wen, E-mail: fajy@mail.cgu.edu.tw [Pharmaceutics Laboratory, Graduate Institute of Natural Products, Chang Gung University, Kweishan, Taoyuan 333, Taiwan (China)

    2010-10-08

    Nanostructured lipid carriers (NLCs) were prepared to investigate whether the duration of brain targeting and accumulation of drugs in the brain can be improved by intravenous delivery. NLCs were developed using cetyl palmitate as the lipid matrix, squalene as the cationic surfactant, and Pluronic F68, polysorbate 80 and polyethylene glycol as the interfacial additives. Solid lipid nanoparticles (SLNs) and lipid emulsions (LEs) were also prepared for comparison. An anti-Parkinson's drug, apomorphine, was used as the model drug. Nuclear magnetic resonance and differential scanning calorimetry showed possible interactions between the solid and liquid lipids in the inner core. The lipid nanoparticles with different compositions were characterized by mean size, zeta potential, apomorphine encapsulation and in vitro drug release. NLCs were 370-430 nm in size, which was between the sizes of the SLNs and LEs. A cationic surfactant was used to produce a positive surface charge of 42-50 mV. The base form of apomorphine was successfully entrapped by NLCs with an entrapment percentage of > 60%. The loading of apomorphine in nanoparticles resulted in a slower release behavior compared to the aqueous solution, with LEs showing the lowest release. In vivo real-time bioluminescence imaging of the rat brain revealed that NLCs could be targeted, through certain vessels, to selected brain regions. This effect was further confirmed by imaging the entire brain and brain slices. The results indicated that NLCs with moderate additives are a promising controlled-release and drug-targeting system.

  13. Physicochemical characterization and in vivo bioluminescence imaging of nanostructured lipid carriers for targeting the brain: apomorphine as a model drug

    Nanostructured lipid carriers (NLCs) were prepared to investigate whether the duration of brain targeting and accumulation of drugs in the brain can be improved by intravenous delivery. NLCs were developed using cetyl palmitate as the lipid matrix, squalene as the cationic surfactant, and Pluronic F68, polysorbate 80 and polyethylene glycol as the interfacial additives. Solid lipid nanoparticles (SLNs) and lipid emulsions (LEs) were also prepared for comparison. An anti-Parkinson's drug, apomorphine, was used as the model drug. Nuclear magnetic resonance and differential scanning calorimetry showed possible interactions between the solid and liquid lipids in the inner core. The lipid nanoparticles with different compositions were characterized by mean size, zeta potential, apomorphine encapsulation and in vitro drug release. NLCs were 370-430 nm in size, which was between the sizes of the SLNs and LEs. A cationic surfactant was used to produce a positive surface charge of 42-50 mV. The base form of apomorphine was successfully entrapped by NLCs with an entrapment percentage of > 60%. The loading of apomorphine in nanoparticles resulted in a slower release behavior compared to the aqueous solution, with LEs showing the lowest release. In vivo real-time bioluminescence imaging of the rat brain revealed that NLCs could be targeted, through certain vessels, to selected brain regions. This effect was further confirmed by imaging the entire brain and brain slices. The results indicated that NLCs with moderate additives are a promising controlled-release and drug-targeting system.

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

    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.

  15. Development of bioluminescent Salmonella strains for use in food safety

    Bailey R Hartford

    2008-01-01

    Full Text Available Abstract Background Salmonella can reside in healthy animals without the manifestation of any adverse effects on the carrier. If raw products of animal origin are not handled properly during processing or cooked to a proper temperature during preparation, salmonellosis can occur. In this research, we developed bioluminescent Salmonella strains that can be used for real-time monitoring of the pathogen's growth on food products. To accomplish this, twelve Salmonella strains from the broiler production continuum were transformed with the broad host range plasmid pAKlux1, and a chicken skin attachment model was developed. Results Salmonella strains carrying pAKlux1 constitutively expressed the luxCDABE operon and were therefore detectable using bioluminescence. Strains were characterized in terms of bioluminescence properties and plasmid stability. To assess the usefulness of bioluminescent Salmonella strains in food safety studies, we developed an attachment model using chicken skin. The effect of washing on attachment of Salmonella strains to chicken skin was tested using bioluminescent strains, which revealed the attachment properties of each strain. Conclusion This study demonstrated that bioluminescence is a sensitive and effective tool to detect Salmonella on food products in real-time. Bioluminescence imaging is a promising technology that can be utilized to evaluate new food safety measures for reducing Salmonella contamination on food products.

  16. Bioluminescence imaging of transplanted human endothelial colony-forming cells in an ischemic mouse model.

    Ding, Jie; Zhao, Zhen; Wang, Chao; Wang, Cong-Xiao; Li, Pei-Cheng; Qian, Cheng; Teng, Gao-Jun

    2016-07-01

    Ischemic strokes are devastating events responsible for high mortality and morbidity worldwide each year. Endothelial colony-forming cell (ECFC) therapy holds promise for stroke treatment; however, grafted ECFCs need to be monitored better understand their biological behavior in vivo, so as to evaluate their safety and successful delivery. The objectives of this study are to visualize the fate of infused human cord blood derived ECFCs via bioluminescence imaging (BLI) in an ischemic stroke mouse model and to determine the therapeutic effects of ECFC transplantation. ECFCs derived from human umbilical cord blood were infected with lentivirus carrying enhanced green fluorescent protein (eGFP) and firefly luciferase (Luc2) double fusion reporter gene. Labeled ECFCs were grafted into a photothrombotic ischemic stroke mouse model via intra-arterial injection though the left cardiac ventricle. The homing of infused cells and functional recovery of stroke mice were evaluated using BLI, neurological scoring, and immunohistochemistry. Significantly, BLI signals were highest in the brain on day 1 and decreased steadily until day 14. GFP-positive cells were also found surrounding infarct border zones in brain sections using immunohistochemical staining, suggesting that ECFCs properly homed to the ischemic brain tissue. Using a modified neurological severity score assay and histological analysis of brain slices with CD31 immunostaining in brain tissue, double cortin analysis, and the TdT-mediated dUTP nick end labeling (TUNEL) assay, we demonstrated functional restoration, improved angiogenesis, neurogenesis, and decreased apoptosis in ischemic mice after ECFC infusion. Collectively, our data support that ECFCs may be a promising therapeutic agent for stroke. PMID:27038754

  17. A new multicolor bioluminescence imaging platform to investigate NF-κB activity and apoptosis in human breast cancer cells.

    Laura Mezzanotte

    Full Text Available BACKGROUND: Evaluation of novel drugs for clinical development depends on screening technologies and informative preclinical models. Here we developed a multicolor bioluminescent imaging platform to simultaneously investigate transcription factor NF-κB signaling and apoptosis. METHODS: The human breast cancer cell line (MDA-MB-231 was genetically modified to express green, red and blue light emitting luciferases to monitor cell number and viability, NF-κB promoter activity and to perform specific cell sorting and detection, respectively. The pro-luciferin substrate Z-DEVD-animoluciferin was employed to determine apoptotic caspase 3/7 activity. We used the cell line for the in vitro evaluation of natural compounds and in vivo optical imaging of tumor necrosis factor TNFα-induced NF-κB activation. RESULTS: Celastrol, resveratrol, sulphoraphane and curcumin inhibited the NF-κB promoter activity significantly and in a dose dependent manner. All compounds except resveratrol induced caspase 3/7 dependent apoptosis. Multicolor bioluminescence in vivo imaging allowed the investigation of tumor growth and NF-κB induction in a mouse model of breast cancer. CONCLUSION: Our new method provides an imaging platform for the identification, validation, screening and optimization of compounds acting on NF-κB signaling and apoptosis both in vitro and in vivo.

  18. A New Multicolor Bioluminescence Imaging Platform to Investigate NF-κB Activity and Apoptosis in Human Breast Cancer Cells

    Mezzanotte, Laura; An, Na; Mol, Isabel M.; Löwik, Clemens W. G. M.; Kaijzel, Eric L.

    2014-01-01

    Background Evaluation of novel drugs for clinical development depends on screening technologies and informative preclinical models. Here we developed a multicolor bioluminescent imaging platform to simultaneously investigate transcription factor NF-κB signaling and apoptosis. Methods The human breast cancer cell line (MDA-MB-231) was genetically modified to express green, red and blue light emitting luciferases to monitor cell number and viability, NF-κB promoter activity and to perform specific cell sorting and detection, respectively. The pro-luciferin substrate Z-DEVD-animoluciferin was employed to determine apoptotic caspase 3/7 activity. We used the cell line for the in vitro evaluation of natural compounds and in vivo optical imaging of tumor necrosis factor TNFα-induced NF-κB activation. Results Celastrol, resveratrol, sulphoraphane and curcumin inhibited the NF-κB promoter activity significantly and in a dose dependent manner. All compounds except resveratrol induced caspase 3/7 dependent apoptosis. Multicolor bioluminescence in vivo imaging allowed the investigation of tumor growth and NF-κB induction in a mouse model of breast cancer. Conclusion Our new method provides an imaging platform for the identification, validation, screening and optimization of compounds acting on NF-κB signaling and apoptosis both in vitro and in vivo. PMID:24465597

  19. Three-dimensional localization of in vivo bioluminescent source based on multispectral imaging

    Feng, Jinchao; Jia, Kebin; Tian, Jie; Yan, Guorui; Zhu, Shouping

    2009-02-01

    Bioluminescence tomography (BLT) is a novel in vivo technique in small animal studies, which can reveal the molecular and cellular information at the whole-body small animal level. At present, there is an increasing interest in multispectral bioluminescence tomography, since multispectral data acquisition could improve the BLT performance significantly. In view to the ill-posedness of BLT problem, we develop an optimal permissible source region strategy to constrain the possible solution of the source by utilizing spectrum character of bioluminescent source. Then a linear system to link the measured data with the unknown light source variables is established by utilizing the optimal permissible region strategy based on adaptive finite element analysis. Furthermore, singular value decomposition analysis is used for data dimensionality reduction and improving computational efficiency in multispectral case. The reconstructed speed and stability benefit from adaptive finite element, the permissible region strategy and singular value decomposition. In the numerical simulation, the heterogeneous phantom experiment has been used to evaluate the performance of the proposed algorithm with the Monte Carlo based synthetic data. The reconstruction results demonstrate the merits and potential of our methodology for localizing bioluminescent source.

  20. Establishment of cell strains stably-transfected with luciferase gene mediated by retrovirus and their detection with bioluminescence imaging system

    Hai-juan WANG

    2012-05-01

    Full Text Available Objective  To establish cell strains stably transfected with Luciferase gene (Luc2, which was mediated by retrovirus, and explore the relationship between the number of Luc2-positive cells and light flux from bioluminescence imaging system by experiments in vitro and in vivo. Methods  We co-transfected pMX-Luc2 plasmid and pMD.G plasmid into 293T gag-pol cells to get retrovirus expressing Luc2 gene. Stable Luc2 positive cell lines were generated and screened by transduction of Retro-Luc2 in mouse colon cancer cell line CT26, human non-small cell lung cancer cell line NCI-H446, human colon cancer cell line HT-29, human ovarian carcinoma cell line SKOV3 and human hepatocellular carcinoma cell line SMMC-7721, all of them were identified by bioluminescence imaging system. Different numbers of SKOV3-Luc2 cells ranging from 10 to 10000 were plated onto culture dishes. Two xenograft models of ovarian cancer were reproduced by subcutaneous injection of 200μl SKOV3-Luc2 cell suspension with different concentrations (1×107, 5×106, 2.5×106, 1×106, 5×105, 2.5×105, 1×105 and 5×104/ml into 16 sites on the back of 4 nude mice, or intravenous injection of 1×106 or 3 ×106 SKOV3-Luc2 cells into the tail vein. Light flux value of SKOV3-Luc2 cells in dishes and in mice was measured by bioluminescence imaging system. Results  Retro-Luc2 was constructed successfully and expressed Luc2 stably in transduced CT26, NCI-H446, HT-29, SKOV3 and SMMC-7721 cell lines. Light flux was correlated in a linear manner with the number of Luc2-positive cells in dishes and in mice (R2=0.944, β=0.972; R2=0.991, β=0.996; R2=0.351, β=0.628; P < 0.01. Conclusion  Luc2-positive cell lines could be established rapidly and accurately by infecting tumor cells with retrovirus expressing Luc2. The number of Luc2 positive cells is significantly related in a linear manner to light flux from bioluminescence imaging system in vitro and in vivo.

  1. Comprehensive assessment of host responses to ionizing radiation by nuclear factor-κB bioluminescence imaging-guided transcriptomic analysis.

    Chung-Ta Chang

    Full Text Available The aim of this study was to analyze the host responses to ionizing radiation by nuclear factor-κB (NF-κB bioluminescence imaging-guided transcriptomic tool. Transgenic mice carrying the NF-κB-driven luciferase gene were exposed to a single dose of 8.5 Gy total-body irradiation. In vivo imaging showed that a maximal NF-κB-dependent bioluminescent intensity was observed at 3 h after irradiation and ex vivo imaging showed that liver, intestine, and brain displayed strong NF-κB activations. Microarray analysis of these organs showed that irradiation altered gene expression signatures in an organ-specific manner and several pathways associated with metabolism and immune system were significantly altered. Additionally, the upregulation of fatty acid binding protein 4, serum amyloid A2, and serum amyloid A3 genes, which participate in both inflammation and lipid metabolism, suggested that irradiation might affect the cross pathways of metabolism and inflammation. Moreover, the alteration of chemokine (CC-motif ligand 5, chemokine (CC-motif ligand 20, and Jagged 1 genes, which are involved in the inflammation and enterocyte proliferation, suggested that these genes might be involved in the radiation enteropathy. In conclusion, this report describes the comprehensive evaluation of host responses to ionizing radiation. Our findings provide the fundamental information about the in vivo NF-κB activity and transcriptomic pattern after irradiation. Moreover, novel targets involved in radiation injury are also suggested.

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

    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

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

    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

  4. Development of a Multicolor Bioluminescence Imaging Platform to Simultaneously Investigate Transcription Factor NF-κB Signaling and Apoptosis.

    Knol-Blankevoort, Vicky T; Mezzanotte, Laura; Rabelink, Martijn J W E; Löwik, Clemens W G M; Kaijzel, Eric L

    2016-01-01

    Here we describe a novel multicolor bioluminescent imaging platform that enables us to simultaneously investigate transcription factor nuclear factor-κB (NF-κB) signalling and apoptosis. We genetically modified the human breast cancer cell line MDA-MB-231 to express green, red, and blue light-emitting luciferases to monitor cell number and viability, NF-κB promoter activity, and to enable specific cell sorting and detection, respectively. Z-DEVD-animoluciferin, the pro-luciferin substrate, was used to determine apoptotic caspase 3/7 activity. We used this multicolored cell line for the in vitro evaluation of natural compounds and in vivo optical imaging of tumor necrosis factor (TNFα)-induced NF-κB activation (Mezzanotte et al., PLoS One 9:e85550, 2014). PMID:27424911

  5. Destabilized bioluminescent proteins

    Allen, Michael S.; Rakesh, Gupta; Gary, Sayler S.

    2007-07-31

    Purified nucleic acids, vectors and cells containing a gene cassette encoding at least one modified bioluminescent protein, wherein the modification includes the addition of a peptide sequence. The duration of bioluminescence emitted by the modified bioluminescent protein is shorter than the duration of bioluminescence emitted by an unmodified form of the bioluminescent protein.

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

    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

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

    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

  8. In vivo bioluminescence imaging of hyperglycemia exacerbating stem cells on choroidal neovascularization in mice

    Gao, Xiang; Wang, Yu; Hou, Hui-Yuan; Lyu, Yang; Wang, Hai-Yan; Yao, Li-Bo; Zhang, Jian; Cao, Feng; Wang, Yu-Sheng

    2016-01-01

    AIM To investigate the influence of hyperglycemia on the severity of choroidal neovascularization (CNV), especially the involvement of bone marrow-derived cells (BMCs) and underlying mechanisms. METHODS BMCs from firefly luciferase (Fluc)/green fluorescent protein (GFP) double transgenic mice were transplanted into C57BL/6J wide-type mice. The recipient mice were injected intraperitoneally with streptozotocin (STZ) daily for 5 consecutive days to induce diabetes mellitus (DM), followed by CNV laser photocoagulation. The BMCs recruitment in CNV exposed to hyperglycemia was firstly examined in Fluc/GFP chimeric mice by in vivo optical bioluminescence imaging (BLI) and in vitro Fluc assays. The CNV severity was evaluated by H&E staining and choroidal flatmount. The expression of vascular endothelial growth factor (VEGF) and stromal cell derived factor-1 (SDF-1) was detected by Western Blot. RESULTS BLI showed that the BMCs exerted dynamic effects in CNV model in Fluc/GFP chimeric mice exposed to hyperglycemia. The signal intensity of transplanted Fluc+GFP+ BMCs in the DM chimeric mice was significantly higher than that in the control chimeric mice with CNV induction at days 5, 7, 14 and 21 (121861.67±9948.81 vs 144998.33±13787.13 photons/second/cm2/sr for control and DM mice, P5d<0.05; 178791.67±30350.8 vs 240166.67±22605.3, P7d<0.05; 124176.67±16253.52 vs 196376.67±18556.79, P14d<0.05; 97951.60±10343.09 vs 119510.00±14383.76, P21d<0.05), which was consistent with in vitro Fluc assay at day 7 [relative light units of Fluc (RLU1)], 215.00±52.05 vs 707.33±88.65, P<0.05; RLU1/ relative light units of renilla luciferase (RLU2), 0.90±0.17 vs 1.83±0.17, P<0.05]. The CNVs in the DM mice were wider than those in the control group at days 5, 7, 14 and 21 (147.83±17.36 vs 220.33±20.17 µm, P5d<0.05; 212.17±24.63 vs 326.83±19.49, P7d<0.05; 163.17±18.24 vs 265.17±20.55, P14d<0.05; 132.00±10.88 vs 205.33±12.98, P21d<0.05). The average area of CNV in the DM

  9. A Bioluminescence Assay System for Imaging Metal Cationic Activities in Urban Aerosols.

    Kim, Sung-Bae; Naganawa, Ryuichi; Murata, Shingo; Nakayama, Takayoshi; Miller, Simon; Senda, Toshiya

    2016-01-01

    A bioluminescence-based assay system was fabricated for an efficient determination of the activities of air pollutants. The following four components were integrated into this assay system: (1) an 8-channel assay platform uniquely designed for simultaneously sensing multiple optical samples, (2) single-chain probes illuminating toxic chemicals or heavy metal cations from air pollutants, (3) a microfluidic system for circulating medium mimicking the human body, and (4) the software manimulating the above system. In the protocol, we briefly introduce how to integrate the components into the system and the application to the illumination of the metal cationic activities in air pollutants. PMID:27424913

  10. Improved Reconstruction Quality of Bioluminescent Images by Combining SP3 Equations and Bregman Iteration Method

    Qiang Wu

    2013-01-01

    Full Text Available Bioluminescence tomography (BLT has a great potential to provide a powerful tool for tumor detection, monitoring tumor therapy progress, and drug development; developing new reconstruction algorithms will advance the technique to practical applications. In the paper, we propose a BLT reconstruction algorithm by combining SP3 equations and Bregman iteration method to improve the quality of reconstructed sources. The numerical results for homogeneous and heterogeneous phantoms are very encouraging and give significant improvement over the algorithms without the use of SP3 equations and Bregman iteration method.

  11. Inflammatory modulating effects of low level laser therapy on iNOS expression by means of bioluminescence imaging

    Moriyama, Yumi; Moriyama, Eduardo H.; Blackmore, Kristina; Akens, Margarete K.; Lilge, Lothar

    2005-09-01

    This study investigates the efficacy of low level laser therapy (LLLT) in modulating inducible nitric oxide synthase (iNOS) expression as molecular marker of the inflammation signaling pathway. LLLT was mediated by different therapeutic wavelengths using transgenic animals with the luciferase gene under control of the iNOS gene expression. Inflammation in 30 transgenic mice (iNOS-luc mice, from FVB strain) was induced by intra-articular injection of Zymosan-A in both knee joints. Four experimental groups were treated with one of four different wavelengths (λ=635, 785, 808 and 905nm) and one not laser-irradiated control group. Laser treatment (25 mW cm-2, 5 J cm-2) was applied to the knees 15 minutes after inflammation induction. Measurements of iNOS expression were performed at multiple times (0, 3, 5, 7, 9 and 24h) post-LLLT by measuring the bioluminescence signal using a highly sensitive charge-coupled device (CCD) camera. The responsivity of BLI was sufficient to demonstrate a significant increase in bioluminescence signals after laser irradiation of 635nm when compared to non-irradiated animals and the other LLLT treated groups, showing the wavelength-dependence of LLLT on iNOS expression during the acute inflammatory process.

  12. Chemiluminescence and bioluminescence microbe detection

    Taylor, R. E.; Chappelle, E.; Picciolo, G. L.; Jeffers, E. L.; Thomas, R. R.

    1978-01-01

    Automated biosensors for online use with NASA Water Monitoring System employs bioluminescence and chemiluminescence techniques to rapidly measure microbe contamination of water samples. System eliminates standard laboratory procedures requiring time duration of 24 hours or longer.

  13. A novel triple-modality reporter gene for whole-body fluorescent, bioluminescent, and nuclear noninvasive imaging

    Ponomarev, Vladimir; Vider, Jelena; Shavrin, Aleksander; Ageyeva, Ludmila; Tourkova, Vilia [Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, NY 10021, New York (United States); Doubrovin, Michael; Serganova, Inna; Beresten, Tatiana; Ivanova, Anna; Blasberg, Ronald [Department of Neurology, Memorial Sloan-Kettering Cancer Center, New York (United States); Balatoni, Julius [Radiochemistry/Cyclotron Core Facility, Memorial Sloan-Kettering Cancer Center, New York (United States); Bornmann, William [Organic Chemistry Synthesis Core Facility, Memorial Sloan-Kettering Cancer Center, New York (United States); Gelovani Tjuvajev, Juri [Department of Radiology, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, NY 10021, New York (United States); MD Anderson Cancer Center, 1515 Holcombe Road, Box 0057, TX 77030, Houston (United States)

    2004-05-01

    Two genetic reporter systems were developed for multimodality reporter gene imaging of different molecular-genetic processes using fluorescence, bioluminescence (BLI), and nuclear imaging techniques. The eGFP cDNA was fused at the N-terminus with HSV1-tk cDNA bearing a nuclear export signal from MAPKK (NES-HSV1-tk) or with truncation at the N-terminus of the first 45 amino acids ({delta}45HSV1-tk) and with firefly luciferase at the C-terminus. A single fusion protein with three functional subunits is formed following transcription and translation from a single open reading frame. The NES-TGL (NES-TGL) or {delta}45HSV1-tk/GFP/luciferase ({delta}45-TGL) triple-fusion gene cDNAs were cloned into a MoMLV-based retrovirus, which was used for transduction of U87 human glioma cells. The integrity, fluorescence, bioluminescence, and enzymatic activity of the TGL reporter proteins were assessed in vitro. The predicted molecular weight of the fusion proteins (130 kDa) was confirmed by western blot. The U87-NES-TGL and U87-{delta}45-TGL cells had cytoplasmic green fluorescence. The in vitro BLI was 7- and 13-fold higher in U87-NES-TGL and U87-{delta}45-TGL cells compared to nontransduced control cells. The Ki of {sup 14}C-FIAU was 0.49{+-}0.02, 0.51{+-}0.03, and 0.003{+-}0.001 ml/min/g in U87-NES-TGL, U87-{delta}45-TGL, and wild-type U87 cells, respectively. Multimodality in vivo imaging studies were performed in nu/nu mice bearing multiple s.c. xenografts established from U87-NES-TGL, U87-{delta}45-TGL, and wild-type U87 cells. BLI was performed after administration of d-luciferin (150 mg/kg i.v.). Gamma camera or PET imaging was conducted at 2 h after i.v. administration of [{sup 131}I]FIAU (7.4 MBq/animal) or [{sup 124}I]FIAU (7.4 MBq/animal), respectively. Whole-body fluorescence imaging was performed in parallel with the BLI and radiotracer imaging studies. In vivo BLI and gamma camera imaging showed specific localization of luminescence and radioactivity to the TGL

  14. A novel triple-modality reporter gene for whole-body fluorescent, bioluminescent, and nuclear noninvasive imaging

    Two genetic reporter systems were developed for multimodality reporter gene imaging of different molecular-genetic processes using fluorescence, bioluminescence (BLI), and nuclear imaging techniques. The eGFP cDNA was fused at the N-terminus with HSV1-tk cDNA bearing a nuclear export signal from MAPKK (NES-HSV1-tk) or with truncation at the N-terminus of the first 45 amino acids (Δ45HSV1-tk) and with firefly luciferase at the C-terminus. A single fusion protein with three functional subunits is formed following transcription and translation from a single open reading frame. The NES-TGL (NES-TGL) or Δ45HSV1-tk/GFP/luciferase (Δ45-TGL) triple-fusion gene cDNAs were cloned into a MoMLV-based retrovirus, which was used for transduction of U87 human glioma cells. The integrity, fluorescence, bioluminescence, and enzymatic activity of the TGL reporter proteins were assessed in vitro. The predicted molecular weight of the fusion proteins (130 kDa) was confirmed by western blot. The U87-NES-TGL and U87-Δ45-TGL cells had cytoplasmic green fluorescence. The in vitro BLI was 7- and 13-fold higher in U87-NES-TGL and U87-Δ45-TGL cells compared to nontransduced control cells. The Ki of 14C-FIAU was 0.49±0.02, 0.51±0.03, and 0.003±0.001 ml/min/g in U87-NES-TGL, U87-Δ45-TGL, and wild-type U87 cells, respectively. Multimodality in vivo imaging studies were performed in nu/nu mice bearing multiple s.c. xenografts established from U87-NES-TGL, U87-Δ45-TGL, and wild-type U87 cells. BLI was performed after administration of d-luciferin (150 mg/kg i.v.). Gamma camera or PET imaging was conducted at 2 h after i.v. administration of [131I]FIAU (7.4 MBq/animal) or [124I]FIAU (7.4 MBq/animal), respectively. Whole-body fluorescence imaging was performed in parallel with the BLI and radiotracer imaging studies. In vivo BLI and gamma camera imaging showed specific localization of luminescence and radioactivity to the TGL transduced xenografts with background levels of activity in the

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

    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.

  16. 一种生物在体荧光成像的自适应分割算法%Adaptive Segmentation Algorithm of Bioluminescent Image

    常志军; 杨鑫

    2011-01-01

    Bioluminescence imaging is regarded as an imaging modality with a high performance, low cost and good prospect in molecular imaging technique. This paper proposes a new adaptive segmentation algorithm, which is based on the characteristics and the application requirements of the bioluminescent images. The adaptive segmentation is realized by performing the normalized processing, connectivity operation and the interested regions distinguishing on bioluminescent images. Experimental results show that this algorithm can get better segmentation results in the ease of weak signal, low signal-to-noise ratio and multiple light sources, so it is a kind of effective segmentation algorithm of bioluminescent images.%生物在体荧光成像是新兴分子影像技术中性能高、费用低、前景好的一种成像模态.针对生物在体荧光图像的特点和应用需求,提出一种全新的自适应图像分割算法.通过对荧光图像的归一化处理、连通性操作、感兴趣区域区分实现自适应分割.实验结果表明,该算法能够在弱信号、低信噪比、多光源的情况下得到较理想的分割结果,是一种有效的荧光图像分割算法.

  17. Repeated and Widespread Evolution of Bioluminescence in Marine Fishes.

    Davis, Matthew P; Sparks, John S; Smith, W Leo

    2016-01-01

    Bioluminescence is primarily a marine phenomenon with 80% of metazoan bioluminescent genera occurring in the world's oceans. Here we show that bioluminescence has evolved repeatedly and is phylogenetically widespread across ray-finned fishes. We recover 27 independent evolutionary events of bioluminescence, all among marine fish lineages. This finding indicates that bioluminescence has evolved many more times than previously hypothesized across fishes and the tree of life. Our exploration of the macroevolutionary patterns of bioluminescent lineages indicates that the present day diversity of some inshore and deep-sea bioluminescent fish lineages that use bioluminescence for communication, feeding, and reproduction exhibit exceptional species richness given clade age. We show that exceptional species richness occurs particularly in deep-sea fishes with intrinsic bioluminescent systems and both shallow water and deep-sea lineages with luminescent systems used for communication. PMID:27276229

  18. Repeated and Widespread Evolution of Bioluminescence in Marine Fishes

    Davis, Matthew P.; Sparks, John S.; Smith, W. Leo

    2016-01-01

    Bioluminescence is primarily a marine phenomenon with 80% of metazoan bioluminescent genera occurring in the world’s oceans. Here we show that bioluminescence has evolved repeatedly and is phylogenetically widespread across ray-finned fishes. We recover 27 independent evolutionary events of bioluminescence, all among marine fish lineages. This finding indicates that bioluminescence has evolved many more times than previously hypothesized across fishes and the tree of life. Our exploration of the macroevolutionary patterns of bioluminescent lineages indicates that the present day diversity of some inshore and deep-sea bioluminescent fish lineages that use bioluminescence for communication, feeding, and reproduction exhibit exceptional species richness given clade age. We show that exceptional species richness occurs particularly in deep-sea fishes with intrinsic bioluminescent systems and both shallow water and deep-sea lineages with luminescent systems used for communication. PMID:27276229

  19. Observations of in situ deep-sea marine bioluminescence with a high-speed, high-resolution sCMOS camera

    Phillips, Brennan T.; Gruber, David F.; Vasan, Ganesh; Roman, Christopher N.; Pieribone, Vincent A.; Sparks, John S.

    2016-05-01

    Observing and measuring marine bioluminescence in situ presents unique challenges, characterized by the difficult task of approaching and imaging weakly illuminated bodies in a three-dimensional environment. To address this problem, a scientific complementary-metal-oxide-semiconductor (sCMOS) microscopy camera was outfitted for deep-sea imaging of marine bioluminescence. This system was deployed on multiple 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 2500 m. 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 of light production in several zooplankton groups was observed, and luminescent responses at different depths were quantified as intensity vs. time. These initial results signify a clear advancement in the bioluminescent imaging methods available for observation and experimentation in the deep-sea.

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

    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.

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

    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

  2. A Dual-Color Far-Red to Near-Infrared Firefly Luciferin Analogue Designed for Multiparametric Bioluminescence Imaging**

    Jathoul, A. P.; Grounds, H.; Anderson, J.; Pule, M. A.

    2014-01-01

    Red-shifted bioluminescent emitters allow improved in vivo tissue penetration and signal quantification, and have led to the development of beetle luciferin analogues that elicit red-shifted bioluminescence with firefly luciferase. However, unlike natural luciferin, none have been shown to emit different colors with different luciferases. We have synthesized and tested the first dual color, far-red to near infrared (nIR) emitting analogue of beetle luciferin, which akin to natural luciferin e...

  3. 生物发光成像的特点及应用%Bioluminescence imaging characteristics and application

    杨丽平; 赵敬湘; 裴雪涛

    2009-01-01

    生物发光成像(BLI)是通过荧光素酶基因标记细胞或DNA,在ATP及氧气存在条件下,催化荧光素的氧化反应而发光,从而能够直接监控活体内的细胞活动和基因行为.该文通过比较BLI与MRI,PET、放射成像的异同,以及BLI在肿瘤、干细胞和免疫细胞运输、细胞凋亡等方面的应用,为更好地推广BLI的应用提供依据.%Bioluminescence imaging (BLI) by luciferase gene marked cells or DNA, in the presence of ATP and oxygen, catalytic oxidation reaction of fluorescein luminescence. So that it can directly monitor in vivo cell activity and gene behavior. In this paper, by comparing the BLI and MRI, PET, radiography of the similarities and differences, as well as about their cancer, stem cells and immune cells transportation, apoptosis and other aspects of the application, in order to better provide the basis for promoting the application of BLI.

  4. Intracellular Self-Assembly of Cyclic d-Luciferin Nanoparticles for Persistent Bioluminescence Imaging of Fatty Acid Amide Hydrolase.

    Yuan, Yue; Wang, Fuqiang; Tang, Wei; Ding, Zhanling; Wang, Lin; Liang, Lili; Zheng, Zhen; Zhang, Huafeng; Liang, Gaolin

    2016-07-26

    Fatty acid amide hydrolase (FAAH) overexpression induces several disorder symptoms in nerve systems, and therefore long-term tracing of FAAH activity in vivo is of high importance but remains challenging. Current bioluminescence (BL) methods are limited in detecting FAAH activity within 5 h. Herein, by rational design of a latent BL probe (d-Cys-Lys-CBT)2 (1), we developed a "smart" method of intracellular reduction-controlled self-assembly and FAAH-directed disassembly of its cyclic d-luciferin-based nanoparticles (i.e., 1-NPs) for persistent BL imaging of FAAH activity in vitro, in cells, and in vivo. Using aminoluciferin methyl amide (AMA), Lys-amino-d-luciferin (Lys-Luc), and amino-d-luciferin (NH2-Luc) as control BL probes, we validated that the persistent BL of 1 from luciferase-expressing cells or tumors was controlled by the activity of intracellular FAAH. With the property of long-term tracing of FAAH activity in vivo of 1, we envision that our BL precursor 1 could probably be applied for in vivo screening of FAAH inhibitors and the diagnosis of their related diseases (or disorders) in the future. PMID:27348334

  5. Real-Time Monitoring of Bacterial Infection In Vivo: Development of Bioluminescent Staphylococcal Foreign-Body and Deep-Thigh-Wound Mouse Infection Models

    Kuklin, Nelly A.; Pancari, Gregory D.; Tobery, Timothy W.; Cope, Leslie; Jackson, Jesse; Gill, Charles; Overbye, Karen; Francis, Kevin P.; Yu, Jun; Montgomery, Donna; Anderson, Annaliesa S.; McClements, William; Jansen, Kathrin U.

    2003-01-01

    Staphylococcal infections associated with catheter and prosthetic implants are difficult to eradicate and often lead to chronic infections. Development of novel antibacterial therapies requires simple, reliable, and relevant models for infection. Using bioluminescent Staphylococcus aureus, we have adapted the existing foreign-body and deep-wound mouse models of staphylococcal infection to allow real-time monitoring of the bacterial colonization of catheters or tissues. This approach also enab...

  6. Imaging tumor angiogenesis in breast cancer experimental lung metastasis with positron emission tomography, near-infrared fluorescence, and bioluminescence.

    Zhang, Yin; Hong, Hao; Nayak, Tapas R; Valdovinos, Hector F; Myklejord, Duane V; Theuer, Charles P; Barnhart, Todd E; Cai, Weibo

    2013-07-01

    The goal of this study was to develop a molecular imaging agent that can allow for both positron emission tomography (PET) and near-infrared fluorescence (NIRF) imaging of CD105 expression in metastatic breast cancer. TRC105, a chimeric anti-CD105 monoclonal antibody, was labeled with both a NIRF dye (i.e., IRDye 800CW) and (64)Cu to yield (64)Cu-NOTA-TRC105-800CW. Flow cytometry analysis revealed no difference in CD105 binding affinity/specificity between TRC105 and NOTA-TRC105-800CW. Serial bioluminescence imaging (BLI) was carried out to non-invasively monitor the lung tumor burden in BALB/c mice, after intravenous injection of firefly luciferase-transfected 4T1 (i.e., fLuc-4T1) murine breast cancer cells to establish the experimental lung metastasis model. Serial PET imaging revealed that fLuc-4T1 lung tumor uptake of (64)Cu-NOTA-TRC105-800CW was 11.9 ± 1.2, 13.9 ± 3.9, and 13.4 ± 2.1 %ID/g at 4, 24, and 48 h post-injection respectively (n = 3). Biodistribution studies, blocking fLuc-4T1 lung tumor uptake with excess TRC105, control experiments with (64)Cu-NOTA-cetuximab-800CW (which served as an isotype-matched control), ex vivo BLI/PET/NIRF imaging, autoradiography, and histology all confirmed CD105 specificity of (64)Cu-NOTA-TRC105-800CW. Successful PET/NIRF imaging of tumor angiogenesis (i.e., CD105 expression) in the breast cancer experimental lung metastasis model warrants further investigation and clinical translation of dual-labeled TRC105-based agents, which can potentially enable early detection of small metastases and image-guided surgery for tumor removal. PMID:23471463

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

    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

  8. Rational development of an attenuated recombinant cyprinid herpesvirus 3 vaccine using prokaryotic mutagenesis and in vivo bioluminescent imaging.

    Boutier, Maxime; Ronsmans, Maygane; Ouyang, Ping; Fournier, Guillaume; Reschner, Anca; Rakus, Krzysztof; Wilkie, Gavin S; Farnir, Frédéric; Bayrou, Calixte; Lieffrig, François; Li, Hong; Desmecht, Daniel; Davison, Andrew J; Vanderplasschen, Alain

    2015-02-01

    Cyprinid herpesvirus 3 (CyHV 3) is causing severe economic losses worldwide in common and koi carp industries, and a safe and efficacious attenuated vaccine compatible with mass vaccination is needed. We produced single deleted recombinants using prokaryotic mutagenesis. When producing a recombinant lacking open reading frame 134 (ORF134), we unexpectedly obtained a clone with additional deletion of ORF56 and ORF57. This triple deleted recombinant replicated efficiently in vitro and expressed an in vivo safety/efficacy profile compatible with use as an attenuated vaccine. To determine the role of the double ORF56-57 deletion in the phenotype and to improve further the quality of the vaccine candidate, a series of deleted recombinants was produced and tested in vivo. These experiments led to the selection of a double deleted recombinant lacking ORF56 and ORF57 as a vaccine candidate. The safety and efficacy of this strain were studied using an in vivo bioluminescent imaging system (IVIS), qPCR, and histopathological examination, which demonstrated that it enters fish via skin infection similar to the wild type strain. However, compared to the parental wild type strain, the vaccine candidate replicated at lower levels and spread less efficiently to secondary sites of infection. Transmission experiments allowing water contamination with or without additional physical contact between fish demonstrated that the vaccine candidate has a reduced ability to spread from vaccinated fish to naïve sentinel cohabitants. Finally, IVIS analyses demonstrated that the vaccine candidate induces a protective mucosal immune response at the portal of entry. Thus, the present study is the first to report the rational development of a recombinant attenuated vaccine against CyHV 3 for mass vaccination of carp. We also demonstrated the relevance of the CyHV 3 carp model for studying alloherpesvirus transmission and mucosal immunity in teleost skin. PMID:25700279

  9. Development of a Chinchilla Model To Allow Direct, Continuous, Biophotonic Imaging of Bioluminescent Nontypeable Haemophilus influenzae during Experimental Otitis Media

    Novotny, Laura A.; Mason, Kevin M.; Bakaletz, Lauren O.

    2005-01-01

    We transformed a nontypeable Haemophilus influenzae clinical isolate with a plasmid containing the luxCDABE operon driven by the H. influenzae outer membrane protein P2 promoter. Herein, we demonstrate the ability to detect bioluminescence and to monitor infection within the nasopharynges, eustachian tubes, and middle ears of chinchillas after intranasal and transbullar challenges.

  10. Molecular Imaging Using Fluorescence and Bioluminescence to Reveal Tissue Response to Laser-Mediated Thermal Injury

    Mackanos, Mark A.; Jansen, E. Duco; Contag, Christopher H.

    For decades biological investigation has focused on a reductionist approach, which has greatly advanced our understanding of the biological process, but has also served to move the analysis further and further away from the living body. This was necessary as we sought to identify the cells, genes, mutations and/or etiological agents that were associated with a given process. The information generated through these approaches can now be used to advance more integrative strategies in which specific cellular and molecular events can be studied in context of the functional circulation and intact organ systems of living animals, and humans. Essential tools for integrative analyses of biology include imaging modalities that enable visualization of structure and function in the living body. The relatively recent development of molecular probes as exogenous contrast agents and reporter genes that encode proteins with unique properties that can be distinguished from tissues and cells has ushered in a new set of approaches that are being called molecular imaging.

  11. zebraflash transgenic lines for in vivo bioluminescence imaging of stem cells and regeneration in adult zebrafish

    Chen, Chen-Hui; Durand, Ellen; Wang, Jinhu; Zon, Leonard I.; Poss, Kenneth D.

    2013-01-01

    The zebrafish has become a standard model system for stem cell and tissue regeneration research, based on powerful genetics, high tissue regenerative capacity and low maintenance costs. Yet, these studies can be challenged by current limitations of tissue visualization techniques in adult animals. Here we describe new imaging methodology and present several ubiquitous and tissue-specific luciferase-based transgenic lines, which we have termed zebraflash, that facilitate the assessment of rege...

  12. Bioluminescent imaging of ABCG2 efflux activity at the blood-placenta barrier

    Kumar, Jeyan S.; Bih-Rong Wei; Madigan, James P.; R Mark Simpson; Hall, Matthew D; Gottesman, Michael M.

    2016-01-01

    Physiologic barriers such as the blood placenta barrier (BPB) and the blood brain barrier protect the underlying parenchyma from pathogens and toxins. ATP-binding cassette (ABC) transporters are transmembrane proteins found at these barriers, and function to efflux xenobiotics and maintain chemical homeostasis. Despite the plethora of ex vivo and in vitro data showing the function and expression of ABC transporters, no imaging modality exists to study ABC transporter activity in vivo at the B...

  13. Monitoring the Growth of an Orthotopic Tumour Xenograft Model: Multi-Modal Imaging Assessment with Benchtop MRI (1T), High-Field MRI (9.4T), Ultrasound and Bioluminescence

    Stuckey, Daniel J.; David, Anna L.; Pedley, R. Barbara; Lythgoe, Mark F.; Siow, Bernard; Walker-Samuel, Simon

    2016-01-01

    Background Research using orthotopic and transgenic models of cancer requires imaging methods to non-invasively quantify tumour burden. As the choice of appropriate imaging modality is wide-ranging, this study aimed to compare low-field (1T) magnetic resonance imaging (MRI), a novel and relatively low-cost system, against established preclinical techniques: bioluminescence imaging (BLI), ultrasound imaging (US), and high-field (9.4T) MRI. Methods A model of colorectal metastasis to the liver was established in eight mice, which were imaged with each modality over four weeks post-implantation. Tumour burden was assessed from manually segmented regions. Results All four imaging systems provided sufficient contrast to detect tumours in all of the mice after two weeks. No significant difference was detected between tumour doubling times estimated by low-field MRI, ultrasound imaging or high-field MRI. A strong correlation was measured between high-field MRI estimates of tumour burden and all the other modalities (p < 0.001, Pearson). Conclusion These results suggest that both low-field MRI and ultrasound imaging are accurate modalities for characterising the growth of preclinical tumour models. PMID:27223614

  14. MEASUREMENTS OF BIOLUMINESCENCE IN DEEP SEA

    Chikawa, M.; Kitamura, T; Nakagawa, Nakagawa; Yamamoto, I.; Wada, T.; Okei, K; Yamashita, Y.

    1996-01-01

    [Abstract] We have designed and built a photon counting system which measures low intensities of bioluminescence in deep sea. The system comprises a CCD-TV camera, two-dimensional image intensifier and video cassette recorder. Using this system we measured the vertical profile of bioluminescence in situ at the Suruga Trough and Nankai Trough to a depth of 3600 m and analyzed cultivated them.

  15. 腺病毒介导荧光素酶报告基因感染间充质干细胞的研究%Infection with adenovirus-mediated luciferase reporter gene in mesenchymal stem cells and bioluminescence imaging

    王一帆; 夏睿; 郭玉林; 郜发宝

    2013-01-01

    .PShuttle-CMV-Luc and backbone vector (pAdeno) were homologous recombined.Then the recombinant plasmid was packaged in HEK293 cells and the virus titer was detected.The BMSC were infected by the recombinant adenovirus.The bioluminescence imaging in vitro was performed to determine the best multiplicity of infection (MOI),and the relationship between bioluminescence intensity and MOI was analyzed by curve fitting regression analysis.Viability was evaluated via Trypan blue staining.The transfected BMSC (l× 106) were implanted into the muscles of forelimb of SD rats,and then tracked by bioluminescence imaging in vivo.Cell viability was compared using two-way repeated measures analysis of variance between groups.Results Enzyme digestion and sequence analysis indicated that Ad-Luc was successfully constructed.The virus titer was 1 × 1010 plaque forming unit (PFU)/ml.The bioluminescence detection in vitro showed that Ad-Luc could infect BMSC high efficiently to express luciferase and the best MOI was 50.The bioluminescence intensity enhanced with increase of MOI (R2 =0.98).No statistically significant difference was found in cell viability between transfected and untransfected BMSC at 1,3,5,7 d.The cell survival rates were (92.5±2.3)% vs (94.1±1.8)%,(91.4±0.9)% vs (92.7±2.0)%,(92.1±1.6)% vs (93.3± 2.4) %,(91.9 ± 1.5) % vs (93.0 ± 3.1) %,respectively (F =4.38,P > 0.05).The bioluminescence imaging in vivo showed that BMSC survived 1,3,7 d after implantation.However,bioluminescence signal decreased gradually over time.Conclusion It is feasible to apply the optical reporter gene imaging for tracing transplanted stem cells in vitro and in vivo due to the effective transformation of luciferase reporter gene into BMSC by adenovirus vector.

  16. Self-illuminating quantum dots for non-invasive bioluminescence imaging of mammalian

    Background: The fertility performance of animals is still a mystery and the full comprehension of mammalian gametes maturation and early embryonic development remains to be elucidated. The recent development in nanotechnology offers a new opportunity for real-time study of reproductive cells in thei...

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

    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

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

    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.

  19. Bioluminescent Probe for Detecting Mercury(II) in Living Mice.

    Jiang, Tianyu; Ke, Bowen; Chen, Hui; Wang, Weishan; Du, Lupei; Yang, Keqian; Li, Minyong

    2016-08-01

    A novel bioluminescence probe for mercury(II) was obtained on the basis of the distinct deprotection reaction of dithioacetal to decanal, so as to display suitable sensitivity and selectivity toward mercury(II) over other ions with bacterial bioluminescence signal. These experimental results indicated such a probe was a novel promising method for mercury(II) bioluminescence imaging in environmental and life sciences ex vivo and in vivo. PMID:27412583

  20. Spectrally resolved bioluminescence tomography using the reciprocity approach

    Dehghani, Hamid; Davis, Scott C.; Pogue, Brian W.

    2008-01-01

    Spectrally resolved bioluminescence optical tomography is an approach to recover images of, for example, Luciferase activity within a volume using multiwavelength emission data from internal bioluminescence sources. The underlying problem of uniqueness associated with nonspectrally resolved intensity-based bioluminescence tomography is demonstrated and it is shown that using a non-negative constraint inverse algorithm, an accurate solution for the source distribution can be calculated from th...

  1. Quantum dot-NanoLuc bioluminescence resonance energy transfer enables tumor imaging and lymph node mapping in vivo.

    Kamkaew, Anyanee; Sun, Haiyan; England, Christopher G; Cheng, Liang; Liu, Zhuang; Cai, Weibo

    2016-05-19

    A small luciferase protein (Nluc) was conjugated to QDs as a bioluminescence resonance energy transfer (BRET) pair. The conjugate showed 76% BRET efficiency and lymph node mapping was successfully performed. The cRGD peptide was conjugated to QD-Nluc for tumor targeting. The self-illuminating QD-Nluc showed excellent energy transfer in a living system and offered an optimal tumor-to-background ratio (>85). PMID:27157466

  2. Dynamic Observation on In vivo Bioluminescence Imaging of Experimental Metastatic Animal Models in Nude Mice%实验性肿瘤细胞转移动物模型的活体成像观察

    闫明霞; 朱淼鑫; 刘蕾; 李静; 林河春; 赵方瑜; 姚明

    2012-01-01

    Objective To observe the tumor metastasis in deep organisms of the nude mice by in vivo bioluminescence imaging system. Methods The SMMC-7721-GFP/Luc cells with different concentrations were intravenously inoculated into the tail vein and spleen of the BALB/c-nu/nu mice, the distribution and expression of luciferase in nude mice were monitored by in vivo bioluminescence imaging system. Results The experimental metastatic animal models had been successfully established. The distribution and expression of luciferase ascended with cell concentration increased and decreased with the passage of time. Conclusion The in vivo bioluminescence imaging system may monitor the in vivo growth and metastasis of tumors and provide for studying the mechanisms of tumor metastasis and development of anticancer drug.%目的 利用小动物活体成像系统观察肿瘤细胞在动物体内的转移情况.方法 分别将不同浓度的绿色荧光蛋白(GPF)和荧光素酶(luciferase,Luc)双标的SMMC-7721细胞接种入裸小鼠尾静脉和脾,建立实验性转移动物模型,采用活体成像技术监测不同浓度的细胞在小鼠体内的转移情况,动态观察同一细胞于不同时间点在小鼠体内的转移情况.结果 成功建立了尾静脉接种肺转移及脾内接种肝转移的实验性转移动物模型,经小动物活体成像系统检测发现,随着接种细胞浓度的增加,荧光素的表达面积和强度逐渐增加,二者呈正比关系;随着接种时间的延长,荧光素的表达面积和强度逐渐减弱,二者呈成反比关系.结论 活体荧光成像系统可较好地观测肿瘤在动物体内深部脏器的转移情况,它将为肿瘤转移机制、抗转移治疗等研究提供有益的帮助.

  3. Real-time monitoring of bacterial infection in vivo: development of bioluminescent staphylococcal foreign-body and deep-thigh-wound mouse infection models.

    Kuklin, Nelly A; Pancari, Gregory D; Tobery, Timothy W; Cope, Leslie; Jackson, Jesse; Gill, Charles; Overbye, Karen; Francis, Kevin P; Yu, Jun; Montgomery, Donna; Anderson, Annaliesa S; McClements, William; Jansen, Kathrin U

    2003-09-01

    Staphylococcal infections associated with catheter and prosthetic implants are difficult to eradicate and often lead to chronic infections. Development of novel antibacterial therapies requires simple, reliable, and relevant models for infection. Using bioluminescent Staphylococcus aureus, we have adapted the existing foreign-body and deep-wound mouse models of staphylococcal infection to allow real-time monitoring of the bacterial colonization of catheters or tissues. This approach also enables kinetic measurements of bacterial growth and clearance in each infected animal. Persistence of infection was observed throughout the course of the study until termination of the experiment at day 16 in a deep-wound model and day 21 in the foreign-body model, providing sufficient time to test the effects of antibacterial compounds. The usefulness of both animal models was assessed by using linezolid as a test compound and comparing bioluminescent measurements to bacterial counts. In the foreign-body model, a three-dose antibiotic regimen (2, 5, and 24 h after infection) resulted in a decrease in both luminescence and bacterial counts recovered from the implant compared to those of the mock-treated infected mice. In addition, linezolid treatment prevented the formation of subcutaneous abscesses, although it did not completely resolve the infection. In the thigh model, the same treatment regimen resulted in complete resolution of the luminescent signal, which correlated with clearance of the bacteria from the thighs. PMID:12936968

  4. Effect of electromagnetic fields on the bacteria bioluminescent activity

    The effect of electromagnetic field with frequency from 36.2 to 55.9 GHz on bioluminescence activity of bacterium were investigated. Electromagnetic field results in decrease of bioluminescence, which depends from frequency. The electromagnetic field adaptation time is higher of intrinsic time parameters of bioluminescence system. The effect has nonthermal nature. It is suggested that electromagnetic field influence connects with structure rearrangements near cell emitter. 8 refs.; 3 figs

  5. aequorine bioluminescence response to calcium in vitro and in cerebral cortex

    Tricoire, Ludovic

    2006-01-01

    During my PhD, I investigated in vitro the calcium-dependent bioluminescence of thephotoprotein aequorin and then used its bioluminescence to image neuronal activities in theneocortical network. This genetically encoded calcium sensor can be expressed in specific cell types and its bioluminescence is not toxic and exhibit a high signal/noise ratio.I first search for mutations modifying aequorin bioluminescence, using a randommutagenesis and in vitro evolution approach. I isolated mutants show...

  6. Luciferase expression and bioluminescence does not affect tumor cell growth in vitro or in vivo

    Rasko John EJ

    2010-11-01

    Full Text Available Abstract Live animal imaging is becoming an increasingly common technique for accurate and quantitative assessment of tumor burden over time. Bioluminescence imaging systems rely on a bioluminescent signal from tumor cells, typically generated from expression of the firefly luciferase gene. However, previous reports have suggested that either a high level of luciferase or the resultant light reaction produced upon addition of D-luciferin substrate can have a negative influence on tumor cell growth. To address this issue, we designed an expression vector that allows simultaneous fluorescence and luminescence imaging. Using fluorescence activated cell sorting (FACS, we generated clonal cell populations from a human breast cancer (MCF-7 and a mouse melanoma (B16-F10 cell line that stably expressed different levels of luciferase. We then compared the growth capabilities of these clones in vitro by MTT proliferation assay and in vivo by bioluminescence imaging of tumor growth in live mice. Surprisingly, we found that neither the amount of luciferase nor biophotonic activity was sufficient to inhibit tumor cell growth, in vitro or in vivo. These results suggest that luciferase toxicity is not a necessary consideration when designing bioluminescence experiments, and therefore our approach can be used to rapidly generate high levels of luciferase expression for sensitive imaging experiments.

  7. Bioluminescence in the high Arctic during the polar night

    Berge, Jørgen; Båtnes, Anna Solvang; Johnsen, Geir; Blackwell, Susan; Mark A. Moline

    2012-01-01

    This study examines the composition and activity of the planktonic community during the polar night in the high Arctic Kongsfjord, Svalbard. Our results are the first published evidence of bioluminescence among zooplankton during the Arctic polar night. The observations were collected by a bathyphotometer detecting bioluminescence, integrated into an autonomous underwater vehicle, to determine the concentration and intensity of bioluminescent flashes as a function of time of day and depth. To...

  8. Bioluminescence imaging of point sources implanted in small animals post mortem: evaluation of a method for estimating source strength and depth

    The performance of a simple approach for the in vivo reconstruction of bioluminescent point sources in small animals was evaluated. The method uses the diffusion approximation as a forward model of light propagation from a point source in a homogeneous tissue to find the source depth and power. The optical properties of the tissue are estimated from reflectance images obtained at the same location on the animal. It was possible to localize point sources implanted in mice, 2-8 mm deep, to within 1 mm. The same performance was achieved for sources implanted in rat abdomens when the effects of tissue surface curvature were eliminated. The source power was reconstructed within a factor of 2 of the true power for the given range of depths, even though the apparent brightness of the source varied by several orders of magnitude. The study also showed that reconstructions using optical properties measured in situ were superior to those based on data in the literature

  9. In vivo functional calcium imaging of induced or spontaneous activity in the fly brain using a GFP-apoaequorin-based bioluminescent approach.

    Minocci, Daiana; Carbognin, Elena; Murmu, Meena Sriti; Martin, Jean-René

    2013-07-01

    Different optical imaging techniques have been developed to study neuronal activity with the goal of deciphering the neural code underlying neurophysiological functions. Because of several constraints inherent in these techniques as well as difficulties interpreting the results, the majority of these studies have been dedicated more to sensory modalities than to the spontaneous activity of the central brain. Recently, a novel bioluminescence approach based on GFP-aequorin (GA) (GFP: Green fluorescent Protein), has been developed, allowing us to functionally record in-vivo neuronal activity. Taking advantage of the particular characteristics of GA, which does not require light excitation, we report that we can record induced and/or the spontaneous Ca(2+)-activity continuously over long periods. Targeting GA to the mushrooms-bodies (MBs), a structure implicated in learning/memory and sleep, we have shown that GA is sensitive enough to detect odor-induced Ca(2+)-activity in Kenyon cells (KCs). It has been possible to reveal two particular peaks of spontaneous activity during overnight recording in the MBs. Other peaks of spontaneous activity have been recorded in flies expressing GA pan-neurally. Similarly, expression in the glial cells has revealed that these cells exhibit a cell-autonomous Ca(2+)-activity. These results demonstrate that bioluminescence imaging is a useful tool for studying Ca(2+)-activity in neuronal and/or glial cells and for functional mapping of the neurophysiological processes in the fly brain. These findings provide a framework for investigating the biological meaning of spontaneous neuronal activity. This article is part of a Special Issue entitled: 12th European Symposium on Calcium. PMID:23287020

  10. High-throughput and quantitative approaches for measuring circadian rhythms in cyanobacteria using bioluminescence.

    Shultzaberger, Ryan K; Paddock, Mark L; Katsuki, Takeo; Greenspan, Ralph J; Golden, Susan S

    2015-01-01

    The temporal measurement of a bioluminescent reporter has proven to be one of the most powerful tools for characterizing circadian rhythms in the cyanobacterium Synechococcus elongatus. Primarily, two approaches have been used to automate this process: (1) detection of cell culture bioluminescence in 96-well plates by a photomultiplier tube-based plate-cycling luminometer (TopCount Microplate Scintillation and Luminescence Counter, Perkin Elmer) and (2) detection of individual colony bioluminescence by iteratively rotating a Petri dish under a cooled CCD camera using a computer-controlled turntable. Each approach has distinct advantages. The TopCount provides a more quantitative measurement of bioluminescence, enabling the direct comparison of clock output levels among strains. The computer-controlled turntable approach has a shorter set-up time and greater throughput, making it a more powerful phenotypic screening tool. While the latter approach is extremely useful, only a few labs have been able to build such an apparatus because of technical hurdles involved in coordinating and controlling both the camera and the turntable, and in processing the resulting images. This protocol provides instructions on how to construct, use, and process data from a computer-controlled turntable to measure the temporal changes in bioluminescence of individual cyanobacterial colonies. Furthermore, we describe how to prepare samples for use with the TopCount to minimize experimental noise and generate meaningful quantitative measurements of clock output levels for advanced analysis. PMID:25662451

  11. In vivo bioluminescence tomography with a blocking-off finite-difference SP3 method and MRI∕CT coregistration

    Klose, Alexander D.; Beattie, Bradley J.; Dehghani, Hamid; Vider, Lena; Le, Carl; Ponomarev, Vladimir; Blasberg, Ronald

    2009-01-01

    Purpose: Bioluminescence imaging is a research tool for studying gene expression levels in small animal models of human disease. Bioluminescence light, however, is strongly scattered in biological tissue and no direct image of the light-emitting reporter probe’s location can be obtained. Therefore, the authors have developed a linear image reconstruction method for bioluminescence tomography (BLT) that recovers the three-dimensional spatial bioluminescent source distribution in small animals.

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

    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.

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

    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/cm2. 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.

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

    Vick, Binje; Rothenberg, Maja; Sandhöfer, Nadine; Carlet, Michela; Finkenzeller, Cornelia; Krupka, Christina; Grunert, Michaela; Trumpp, Andreas; Corbacioglu, Selim; Ebinger, Martin; André, Maya C; Hiddemann, Wolfgang; Schneider, Stephanie; Subklewe, Marion; Metzeler, Klaus H; Spiekermann, Karsten; Jeremias, Irmela

    2015-01-01

    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. PMID:25793878

  15. Lighting up bioluminescence with coelenterazine: strategies and applications.

    Jiang, Tianyu; Du, Lupei; Li, Minyong

    2016-04-13

    Bioluminescence-based techniques, such as bioluminescence imaging, BRET and dual-luciferase reporter assay systems, have been widely used to examine a myriad of biological processes. Coelenterazine (CTZ), a luciferin or light-producing compound found in bioluminescent organisms, has sparked great curiosity and interest in searching for analogues with improved photochemical properties. This review summarizes the current development of coelenterazine analogues, their bioluminescence properties, and the rational design of caged coelenterazine towards biotargets, as well as their applications in bioassays. It should be emphasized that the design of caged luciferins can provide valuable insight into detailed molecular processes in organisms and will be a trend in the development of bioluminescent molecules. PMID:27009907

  16. Multispectral Bioluminescence Tomography: Methodology and Simulation

    Ge Wang

    2006-02-01

    Full Text Available Bioluminescent imaging has proven to be a valuable tool for monitoring physiological and pathological activities at cellular and molecular levels in living small animals. Using biological techniques, target cells can be tagged with reporters encoding several kinds of luciferase enzymes, which generate characteristic photons in a wide spectrum covering the infrared range. Part of the diffused light can reach the body surface of the small animal, be separated into several spectral bands using appropriate filters, and collected by a sensitive CCD camera. Here we present a bioluminescence tomography (BLT method for a bioluminescent source reconstruction from multispectral data measured on the external surface, and demonstrate the advantages of multispectral BLT in a numerical study using a heterogeneous mouse chest phantom. The results show that the multispectral approach significantly improves the accuracy and stability of the BLT reconstruction even if the data are highly noisy.

  17. 活体生物发光成像技术及其在病毒感染研究中的应用%Bioluminescence in-vivo imaging technology and its application in the study of viral infection-A review

    柴凡; 周耘裔; 肖庚富

    2011-01-01

    生物发光是动物活体光学成像技术之一,因其反应灵敏、操作简单、数据精确,而被广泛地应用于生命科学研究多个领域,观测活体动物体内病毒复制、肿瘤生长等生命过程.生物发光技术采用荧光素酶基因标记细胞或病毒,与外源注射的底物荧光素发生反应,在冷CCD成像系统下显像并进行数据记录、分析.本文简要介绍活体生物发光成像这一新技术的原理,综述了其在发现病毒新的复制位点、研究干扰素及药物对病毒的抑制作用、展示病毒潜伏感染与再激活的历程等方面的应用,并结合自己工作对该技术及其发展前景进行评述.%Bioluminescence imaging is one of the bio-optical imaging techniques which report definite biological event in living animals with genetic modification.With high sensitivity, simple operation and high precision, it is particularly applied in observing vital processes like viral infection and tumor growth in vivo.We summarize the principle of bioluminescence imaging, introduce its application in finding virus replication site, study of interferon( IFN) inhibiting-virus effect and real-time visualization of viral latent infection and reactivation, and preview the trend of bioluminescence imaging technological development.

  18. Bioluminescent imaging of vaccinia virus infection in immunocompetent and immunodeficient rats as a model for human smallpox.

    Liu, Qiang; Fan, Changfa; Zhou, Shuya; Guo, Yanan; Zuo, Qin; Ma, Jian; Liu, Susu; Wu, Xi; Peng, Zexu; Fan, Tao; Guo, Chaoshe; Shen, Yuelei; Huang, Weijin; Li, Baowen; He, Zhengming; Wang, Youchun

    2015-01-01

    Due to the increasing concern of using smallpox virus as biological weapons for terrorist attack, there is renewed interest in studying the pathogenesis of human smallpox and development of new therapies. Animal models are highly demanded for efficacy and safety examination of new vaccines and therapeutic drugs. Here, we demonstrated that both wild type and immunodeficient rats infected with an engineered vaccinia virus carrying Firefly luciferase reporter gene (rTV-Fluc) could recapitulate infectious and clinical features of human smallpox. Vaccinia viral infection in wild type Sprague-Dawley (SD) rats displayed a diffusible pattern in various organs, including liver, head and limbs. The intensity of bioluminescence generated from rTV-Fluc correlated well with viral loads in tissues. Moreover, neutralizing antibodies had a protective effect against virus reinfection. The recombination activating gene 2 (Rag2) knockout rats generated by transcription activator-like effector nucleases (TALENs) technology were further used to examine the infectivity of the rTV-Fluc in immunodeficient populations. Here we demonstrated that Rag2-/- rats were more susceptible to rTV-Fluc than SD rats with a slower virus clearance rate. Therefore, the rTV-Fluc/SD rats and rTV-Fluc/Rag2-/- rats are suitable visualization models, which recapitulate wild type or immunodeficient populations respectively, for testing human smallpox vaccine and antiviral drugs. PMID:26235050

  19. Space-Time Quantum Imaging

    Ronald E. Meyers

    2015-03-01

    Full Text Available We report on an experimental and theoretical investigation of quantum imaging where the images are stored in both space and time. Ghost images of remote objects are produced with either one or two beams of chaotic laser light generated by a rotating ground glass and two sensors measuring the reference field and bucket field at different space-time points. We further observe that the ghost images translate depending on the time delay between the sensor measurements. The ghost imaging experiments are performed both with and without turbulence. A discussion of the physics of the space-time imaging is presented in terms of quantum nonlocal two-photon analysis to support the experimental results. The theoretical model includes certain phase factors of the rotating ground glass. These experiments demonstrated a means to investigate the time and space aspects of ghost imaging and showed that ghost imaging contains more information per measured photon than was previously recognized where multiple ghost images are stored within the same ghost imaging data sets. This suggests new pathways to explore quantum information stored not only in multi-photon coincidence information but also in time delayed multi-photon interference. The research is applicable to making enhanced space-time quantum images and videos of moving objects where the images are stored in both space and time.

  20. 6-hydroxydopamine-induced Parkinson's disease-like degeneration generates acute microgliosis and astrogliosis in the nigrostriatal system but no bioluminescence imaging-detectable alteration in adult neurogenesis.

    Fricke, Inga B; Viel, Thomas; Worlitzer, Maik M; Collmann, Franziska M; Vrachimis, Alexis; Faust, Andreas; Wachsmuth, Lydia; Faber, Cornelius; Dollé, Frédéric; Kuhlmann, Michael T; Schäfers, Klaus; Hermann, Sven; Schwamborn, Jens C; Jacobs, Andreas H

    2016-05-01

    Parkinson's disease is a slowly progressing neurodegenerative disorder caused by loss of dopaminergic neurons in the substantia nigra (SN), leading to severe impairment in motor and non-motor functions. Endogenous subventricular zone (SVZ) neural stem cells constantly give birth to new cells that might serve as a possible source for regeneration in the adult brain. However, neurodegeneration is accompanied by neuroinflammation and dopamine depletion, potentially compromising regeneration. We therefore employed in vivo imaging methods to study striatal deafferentation (N-ω-fluoropropyl-2β-carbomethoxy-3β-(4-[(123) I]iodophenyl)nortropane single photon emission computed tomography, DaTscan(™) ) and neuroinflammation in the SN and striatum (N,N-diethyl-2-(2-(4-(2-[(18) F]fluoroethoxy)phenyl)-5,7-dimethylpyrazolo[1,5-a]pyrimidin-3-yl)acetamide positron emission tomography, [(18) F]DPA-714 PET) in the intranigral 6-hydroxydopamine Parkinson's disease mouse model. Additionally, we transduced cells in the SVZ with a lentivirus encoding firefly luciferase and followed migration of progenitor cells in the SVZ-olfactory bulb axis via bioluminescence imaging under disease and control conditions. We found that activation of microglia in the SN is an acute process accompanying the degeneration of dopaminergic cell bodies in the SN. Dopaminergic deafferentation of the striatum does not influence the generation of doublecortin-positive neuroblasts in the SVZ, but generates chronic astrogliosis in the nigrostriatal system. PMID:26950181

  1. Nanostructured bioluminescent sensor for rapidly detecting thrombin.

    Chen, Longyan; Bao, Yige; Denstedt, John; Zhang, Jin

    2016-03-15

    Thrombin plays a key role in thrombosis and hemostasis. The abnormal level of thrombin in body fluids may lead to different diseases, such as rheumatoid arthritis, glomerulonephritis, etc. Detection of thrombin level in blood and/or urine is one of important methods for medical diagnosis. Here, a bioluminescent sensor is developed for non-invasively and rapidly detecting thrombin in urine. The sensor is assembled through conjugating gold nanoparticles (Au NPs) and a recombinant protein containing Renilla luciferase (pRluc) by a peptide, which is thrombin specific substrate. The luciferase-catalyzed bioluminescence can be quenched by peptide-conjugating Au NPs. In the presence of thrombin, the short peptide conjugating luciferase and Au NPs is digested and cut off, which results in the recovery of bioluminescence due to the release of luciferase from Au NPs. The bioluminescence intensity at 470 nm is observed, and increases with increasing concentration of thrombin. The bioluminescence intensity of this designed sensor is significantly recovered when the thrombin digestion time lasts for 10 min. In addition, a similar linear relationship between luminescence intensity and the concentration of thrombin is found in the range of 8 nM to 8 μM in both buffer and human urine spiked samples. The limit of detection is as low as 80 pM. It is anticipated that our nanosensor could be a promising tool for clinical diagnosis of thrombin in human urine. PMID:26397418

  2. 艾灸抑制小鼠皮下移植瘤生长的活体成像观察%Bioluminescence imaging evaluation of the inhibitory effect of moxibustion in mice bear-ing subcutaneous cancer cell lines

    周俊梅; 韦芳; 刘素君; 张必萌

    2014-01-01

    目的:观察艾灸对小鼠皮下移植瘤的抑制作用。方法:通过慢病毒转染,构建能稳定表达荧光素酶基因的乳腺癌细胞株,建立小鼠皮下移植瘤模型后通过随机区组方法设为对照组和治疗组,治疗组又分为关元组、大椎组。治疗组艾灸隔天一次,每次10min,共7次,对照组不做处理。通过活体成像系统以及体外测量,监测肿瘤生长情况。治疗终点,处死小鼠,HE染色观察肿瘤的病理形态学变化。结果:建立能稳定表达荧光素酶基因的乳腺癌细胞株及小鼠皮下移植瘤模型,细胞数与荧光强度呈线性关系( r=0.899)。活体成像显示:治疗组小鼠荧光强度较对照组明显减弱。体外测量,治疗组肿瘤体积及重量明显小于对照组(P%Objective:To investigate the inhibitory effect of moxibustion in mice bearing subcutaneous cancer cell lines. Methods:Breast cancer cell line 4T1 with stable expression of luciferase gene(4T1/luc)were established by transfection via lentiviral vector. 4T1/luc cells were inoculated into the right leg of mice to prepare subcutaneous tumor model. A total of 15 tumor-bearing mouse were randomly divided into the following groups:the model control group and treatment group,the later includes guanyuan group and dazhui group. Mouse in dazhui and guanyuan group were treated with moxibustion,once another day for 10min,for a total of 7 times. The control group did nothing. The dynamic growth of subcutaneous tumor was observed using bioluminescence imaging system in vivo. At the end of the treatment,morphology of transplanted tumor tissue was observed by HE staining. Results:The stable 4T1/luc cells and subcutaneous tumor model were obtained. Bioluminescence imaging showed that fluorescent intensity of treated group was apparently lower than that of the control group. Compare with control group,the tomor volume and weight of treated group were significantly decreased(P 0

  3. Bioluminescence imaging evaluation of the inhibitory effect of moxibustion in mice bear-ing subcutaneous cancer cell lines%艾灸抑制小鼠皮下移植瘤生长的活体成像观察

    周俊梅; 韦芳; 刘素君; 张必萌

    2014-01-01

    目的:观察艾灸对小鼠皮下移植瘤的抑制作用。方法:通过慢病毒转染,构建能稳定表达荧光素酶基因的乳腺癌细胞株,建立小鼠皮下移植瘤模型后通过随机区组方法设为对照组和治疗组,治疗组又分为关元组、大椎组。治疗组艾灸隔天一次,每次10min,共7次,对照组不做处理。通过活体成像系统以及体外测量,监测肿瘤生长情况。治疗终点,处死小鼠,HE染色观察肿瘤的病理形态学变化。结果:建立能稳定表达荧光素酶基因的乳腺癌细胞株及小鼠皮下移植瘤模型,细胞数与荧光强度呈线性关系( r=0.899)。活体成像显示:治疗组小鼠荧光强度较对照组明显减弱。体外测量,治疗组肿瘤体积及重量明显小于对照组(P%Objective:To investigate the inhibitory effect of moxibustion in mice bearing subcutaneous cancer cell lines. Methods:Breast cancer cell line 4T1 with stable expression of luciferase gene(4T1/luc)were established by transfection via lentiviral vector. 4T1/luc cells were inoculated into the right leg of mice to prepare subcutaneous tumor model. A total of 15 tumor-bearing mouse were randomly divided into the following groups:the model control group and treatment group,the later includes guanyuan group and dazhui group. Mouse in dazhui and guanyuan group were treated with moxibustion,once another day for 10min,for a total of 7 times. The control group did nothing. The dynamic growth of subcutaneous tumor was observed using bioluminescence imaging system in vivo. At the end of the treatment,morphology of transplanted tumor tissue was observed by HE staining. Results:The stable 4T1/luc cells and subcutaneous tumor model were obtained. Bioluminescence imaging showed that fluorescent intensity of treated group was apparently lower than that of the control group. Compare with control group,the tomor volume and weight of treated group were significantly decreased(P 0

  4. la bioluminescence de l'aequorine en réponse au calcium In vitro et dans le Cortex cerebral

    Tricoire, Ludovic

    2006-01-01

    During my PhD, I investigated in vitro the calcium-dependent bioluminescence of thephotoprotein aequorin and then used its bioluminescence to image neuronal activities in theneocortical network. This genetically encoded calcium sensor can be expressed in specific cell types and its bioluminescence is not toxic and exhibit a high signal/noise ratio.I first search for mutations modifying aequorin bioluminescence, using a randommutagenesis and in vitro evolution approach. I isolated mutants show...

  5. A Multichannel Bioluminescence Determination Platform for Bioassays.

    Kim, Sung-Bae; Naganawa, Ryuichi

    2016-01-01

    The present protocol introduces a multichannel bioluminescence determination platform allowing a high sample throughput determination of weak bioluminescence with reduced standard deviations. The platform is designed to carry a multichannel conveyer, an optical filter, and a mirror cap. The platform enables us to near-simultaneously determine ligands in multiple samples without the replacement of the sample tubes. Furthermore, the optical filters beneath the multichannel conveyer are designed to easily discriminate colors during assays. This optical system provides excellent time- and labor-efficiency to users during bioassays. PMID:27424912

  6. Evaluation of the inflammatory potential of implant materials in a mouse model by bioluminescent imaging of intravenously injected bone marrow cells.

    Rais, Bushra; Köster, Mario; Rahim, Muhammad Imran; Pils, Marina; Seitz, Jan-Marten; Hauser, Hansjörg; Wirth, Dagmar; Mueller, Peter P

    2016-09-01

    To evaluate the inflammatory potential of implants a bioluminescent imaging assay was developed using luciferase-expressing bone marrow cells that were injected into the blood circulation of wild-type mice. After subcutaneous implantation of titanium discs as an example for a clinically established biocompatible material, the luminosity was modest. Similarly, low luminosity signals were generated by pure magnesium implants that were used to represent metallic alloys that are presently under investigation as novel degradable implant materials. Increased luminosity was observed in response to degradable polymeric PLGA implants. Surgical wounds induced a basic luminescent response even in the absence of an implant. However, the material-independent response to injury could be minimized using injectable microparticle suspensions. In parallel with the resorption of biodegradable microparticles, the signal induced by PLGA declined faster when compared to non-degradable polystyrene suspensions. By using an interferon type I inducible Mx2 promoter construct to drive luciferase gene expression, the highest luminosity was observed in response to bacteria, indicating that the system could also be employed to monitor implant infections. Overall, labeled bone marrow cells yielded specific, well-defined localized signals that correlated with the inflammatory responses to implants. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2149-2158, 2016. PMID:27102724

  7. Optimization and technology extension of bioluminescence imaging in vivo in small animals%小动物生物发光活体成像的条件优化与技术扩展研究

    王豫; 关华; 宋曼; 王晓迪; 高毅; 周平坤

    2013-01-01

    目的 为发掘生物发光活体成像技术的灵敏性和应用潜力,进行在体检测条件的优化和技术扩展.方法 将稳定高表达荧光素酶的乳腺癌细胞(4T-1-Luc+)和肝癌细胞(Huh-7-Luc+)接种于BALB/c 小鼠体内,通过活体成像仪进行生物发光检测灵敏度和毛发的影响分析,利用生物发光结合X线扩展活体成像技术.结果 毛发对小鼠皮下接种的肿瘤细胞的生物发光成像有干扰作用,在局部剃除毛发小鼠内体可以检测到50个细胞的光信号值.通过建立X结合发光的检测模型,实现了既能检测发光信号值强度大小,又能比较清晰地观测到部分脏器和发光体(病变组织)的部位.结论 实现了生物发光活体成像技术的检测条件优化,建立了X线结合发光新的检测模式,为扩展小动物活体成像的应用范围奠定了基础.%Objective To explore the sensitivity and potential applicability of bioluminescence in vivo imaging technique and to achieve optimization of detection conditions and technique extension . Methods Breast cancer cells 4T-L-Luc or liver cancer cells Huh -7 -Luc+ with stable expression of luciferase were implanted subcutaneously into BALB /c mice. The sensitivity and effect on the hair were investigated using the small animal in vivo imager of bioluminescent signals. Technique extension was performed by combining bioluminescence with X -ray. Results The hair as showed are obvious interference with the detection of bioluminescent signals emitted from the cancer cells implanted in BALB /c mice. The optical signal could be detected in the cellular mass of 50 cells when local hair was removed. The imaging model established by combining X-ray with bioluminescence could not only detect the strength of optical signals emitted , but also clearly locate some organs and the origin of optical signals or pathological tissue . Conclusion Optimization of bioluminescence imaging condition has been achieved . A imaging

  8. Bioluminescence lights the way to food safety

    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.

  9. Time-resolved molecular imaging

    Xu, Junliang; Blaga, Cosmin I.; Agostini, Pierre; DiMauro, Louis F.

    2016-06-01

    Time-resolved molecular imaging is a frontier of ultrafast optical science and physical chemistry. In this article, we review present and future key spectroscopic and microscopic techniques for ultrafast imaging of molecular dynamics and show their differences and connections. The advent of femtosecond lasers and free electron x-ray lasers bring us closer to this goal, which eventually will extend our knowledge about molecular dynamics to the attosecond time domain.

  10. Circadian regulation of bioluminescence in Gonyaulax involves translational control.

    Morse, D.; Milos, P M; Roux, E.; Hastings, J. W.

    1989-01-01

    A 10-fold circadian variation in the amount of luciferin binding protein (LBP) in the marine dinoflagellate Gonyaulax polyedra is reported. This protein binds and stabilizes luciferin, the bioluminescence substrate. In early night phase, when bioluminescence is increasing and LBP levels are rising in the cell, pulse labeling experiments show that LBP is being rapidly synthesized in vivo. At other times, the rate of LBP synthesis is at least 50 times lower, while the rate of synthesis of most ...

  11. Imaging tumor angiogenesis in breast cancer experimental lung metastasis with positron emission tomography, near-infrared fluorescence, and bioluminescence

    Zhang, Yin; Hong, Hao; Nayak, Tapas R.; Valdovinos, Hector F.; Myklejord, Duane V.; Theuer, Charles P.; Barnhart, Todd E.; Cai, Weibo

    2013-01-01

    The goal of this study was to develop a molecular imaging agent that can allow for both positron emission tomography (PET) and near-infrared fluorescence (NIRF) imaging of CD105 expression in metastatic breast cancer. TRC105, a chimeric anti-CD105 monoclonal antibody, was labeled with both a NIRF dye (i.e., IRDye 800CW) and 64Cu to yield 64Cu-NOTA-TRC105-800CW. Flow cytometry analysis revealed no difference in CD105 binding affinity/specificity between TRC105 and NOTA-TRC105-800CW. Serial bio...

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

    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

  13. Action of γ-radiation on bioluminescence of Noctiluca miliaris

    Results of the study in the action of various doses of irradiation on the bioluminescence of Noctiluca miliaris are presented. The doses are found that stimulate the bioluminescence and the dose - effect curves are obtained. It has been shown that stimulation of Noctiluca luminescence by γ-radiation is not of a constant character and extinguishes after a period of time determined by a dose rate

  14. Bioluminescent bacteria: lux genes as environmental biosensors

    Nunes-Halldorson Vânia da Silva; Duran Norma Letícia

    2003-01-01

    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 env...

  15. Targeted gene therapy and in vivo bioluminescent imaging for monitoring postsurgical recurrence and metastasis in mouse models of liver cancer.

    He, Q; Yao, C L; Li, L; Xin, Z; Jing, Z K; Li, L X

    2016-01-01

    We investigated the effects of combined targeted gene therapy on recurrence and metastasis after liver cancer resection in nude mice. Twenty BALB/C mice were randomly divided into control and treatment groups with 10 mice in each group and a male/female ratio of 1:1. Luciferase gene-labeled human primary hepatic carcinoma cell line MHCC97-H was then used to prepare a carcinoma model. An optical in vivo imaging technique (OIIT) was used 10 days later to detect the distribution of tumor cells, followed by partial liver resection and gene therapy. In the treatment group, 100 mL phosphate-buffered saline (PBS) containing 1 x 1012 rAAV/AFP/IL-24 gene viral vectors was injected into liver sections and peritumoral posterior peritoneal tissues; in the control group, the same amount of PBS containing 1 x 1012 empty viral vectors was injected at the same sites. OIIT was then used to detect the in vivo tumor metastasis 21 days later. Luciferase gene-labeled human primary hepatic carcinoma cell line MHCC97-H successfully infected 20 nude mice, and OIIT showed that the two groups exhibited metastasis after local tumor resection, but there were more tumor cells in the control group (P AFP/IL-24 gene therapy can inhibit recurrence after liver cancer resection. PMID:27525931

  16. Measurement of Bacterial Bioluminescence Intensity and Spectrum: Current Physical Techniques and Principles.

    Jia, Kun; Ionescu, Rodica Elena

    2016-01-01

    : Bioluminescence is light production by living organisms, which can be observed in numerous marine creatures and some terrestrial invertebrates. More specifically, bacterial bioluminescence is the "cold light" produced and emitted by bacterial cells, including both wild-type luminescent and genetically engineered bacteria. Because of the lively interplay of synthetic biology, microbiology, toxicology, and biophysics, different configurations of whole-cell biosensors based on bacterial bioluminescence have been designed and are widely used in different fields, such as ecotoxicology, food toxicity, and environmental pollution. This chapter first discusses the background of the bioluminescence phenomenon in terms of optical spectrum. Platforms for bacterial bioluminescence detection using various techniques are then introduced, such as a photomultiplier tube, charge-coupled device (CCD) camera, micro-electro-mechanical systems (MEMS), and complementary metal-oxide-semiconductor (CMOS) based integrated circuit. Furthermore, some typical biochemical methods to optimize the analytical performances of bacterial bioluminescent biosensors/assays are reviewed, followed by a presentation of author's recent work concerning the improved sensitivity of a bioluminescent assay for pesticides. Finally, bacterial bioluminescence as implemented in eukaryotic cells, bioluminescent imaging, and cancer cell therapies is discussed. PMID:25981856

  17. Real-Time Imaging of Gene Delivery and Expression with DNA Nanoparticle Technologies

    Sun, Wenchao; Ziady, Assem G.

    The construction of safe, efficient, and modifiable synthetic DNA nanoparticles is an emerging technology that has achieved important milestones of success in the past 5 years. Advances in chemical conjugation, purification, and controlled synthesis have allowed researchers to produce uniform and stable particles, whose physical characteristics can be well characterized and monitored. As a result of these improvements, DNA nanoparticles have now been cleared for clinical testing, and show good potential for human gene therapy. A very important recent development in the study of DNA nanoparticles is the use of small-animal imaging. Real-time imaging has become a valuable technique for tracking particle biodistribution and gene transfer efficacy. In this chapter, we discuss how bioluminescent, positron emission tomography, and magnetic resonance imaging can be used separately or in concert to study particle delivery, localization, and magnitude of gene expression in vivo.

  18. Bioluminescence tomography with Gaussian prior

    Gao, Hao; Zhao, Hongkai; Cong, Wenxiang; Wang, Ge

    2010-01-01

    Parameterizing the bioluminescent source globally in Gaussians provides several advantages over voxel representation in bioluminescence tomography. It is mathematically unique to recover Gaussians [Med. Phys. 31(8), 2289 (2004)] and practically sufficient to approximate various shapes by Gaussians in diffusive medium. The computational burden is significantly reduced since much fewer unknowns are required. Besides, there are physiological evidences that the source can be modeled by Gaussians....

  19. Microtiter plate tests for segregation of bioluminescent bacteria.

    Šimkus, Remigijus; Meškienė, Rita; Ledas, Žilvinas; Baronas, Romas; Meškys, Rolandas

    2016-02-01

    It has been recently shown that bioluminescence imaging can be usefully applied to provide new insights into bacterial self-organization. In this work we employ bioluminescence imaging to record images of nutrient rich liquid cultures of the lux-gene reporter Escherichia coli in microtiter plate wells. The images show that patterns of inhomogenous bioluminescence form along the three-phase contact lines. The paper analyzes the dependencies of the average number of luminous aggregates (clouds) on various environmental factors. In particular, our results show that optimal (neutral) pH and high aeration rates determine the highest mean number of clouds, and that spatiotemporal patterns do not form in the pH buffered suspensions. In addition, a sigmoidal (switch-like) dependence of the number of aggregates on the rate of aeration was observed. The obtained bioluminescence imaging data was interpreted by employing the Keller-Segel-Fisher (KSF) model of chemotaxis and logistic growth, adapted to systems of metabolically flexible (two-state) bacteria. The modified KSF model successfully simulated the observed switch-like responses. The results of the microtiter plate tests and their simulations indicate that the segregation of bacteria with different activities proceeds in the three-phase contact line region. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26039821

  20. 活体荧光成像对裸鼠肝癌细胞系MHCC97-H原位移植模型的动态量化分析%Dynamic and Quantitative Analysis of Orthotopically Transplanted Nude Mouse Model with MHCC97-H Cells using Bioluminescent Imaging Technology

    曹阳; 韩炜; 刘洋; 张勇; 郭欣; 陈勇

    2013-01-01

    from liver to abdominal cavity, and the total photon flux emitting from tumor cells increased expotetially over time. Then the tumor was confirmed by pathological examination. Conclusion: Dynamic and quantitative analysis used bioluminescent imaging technology can accurately reflect the volume, growth and metastasis of tumors in orthotopically transplanted nude mouse model for HCC, and provide sensitive access to developing the research of mechanism of oncogenesis, growth and metastasis of tumor, and anti-cancer study.

  1. The rapid bioluminescence assay method for content of bacteria in dehydrated vegetable and condiment before radiation

    The microbial colony-forming unit (cfu) in dehydrated vegetable and condiment was determined by using ATP bioluminescence method. The result showed that bioluminescence of ATP was correlative to the microbial cfu obviously. The detecting time was within 1-2 h. This method could be applied to determine micro load of products before irradiation sterilization. (authors)

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

    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.)

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

    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.)

  4. Effect of irradiation on bioluminescence spectrum of microbial ATP

    The effect of irradiation on bioluminescence spectrum of dehydrated cabbage microbial ATP was studied. The results showed that the spectral bandwidth of ATP standard was from 490 to 640 nm and the peak wavelength was at 563 nm. The spectral bandwidths of irradiated dehydrated cabbage microbial ATP and CK did not change. Peak wavelengths of dehydrated cabbage irradiated at different dosages were not significantly different from that of CK. The peaks of bioluminescence spectrum of irradiated samples were higher than that of CK, which may be because of the increasing concentration of ATP, and this effect would be kept for quite a long time after irradiation. (authors)

  5. A novel reconstruction algorithm for bioluminescent tomography based on Bayesian compressive sensing

    Wang, Yaqi; Feng, Jinchao; Jia, Kebin; Sun, Zhonghua; Wei, Huijun

    2016-03-01

    Bioluminescence tomography (BLT) is becoming a promising tool because it can resolve the biodistribution of bioluminescent reporters associated with cellular and subcellular function through several millimeters with to centimeters of tissues in vivo. However, BLT reconstruction is an ill-posed problem. By incorporating sparse a priori information about bioluminescent source, enhanced image quality is obtained for sparsity based reconstruction algorithm. Therefore, sparsity based BLT reconstruction algorithm has a great potential. Here, we proposed a novel reconstruction method based on Bayesian compressive sensing and investigated its feasibility and effectiveness with a heterogeneous phantom. The results demonstrate the potential and merits of the proposed algorithm.

  6. Accounting for filter bandwidth improves the quantitative accuracy of bioluminescence tomography

    Taylor, Shelley L.; Mason, Suzannah K. G.; Glinton, Sophie L.; Cobbold, Mark; Dehghani, Hamid

    2015-09-01

    Bioluminescence imaging is a noninvasive technique whereby surface weighted images of luminescent probes within animals are used to characterize cell count and function. Traditionally, data are collected over the entire emission spectrum of the source using no filters and are used to evaluate cell count/function over the entire spectrum. Alternatively, multispectral data over several wavelengths can be incorporated to perform tomographic reconstruction of source location and intensity. However, bandpass filters used for multispectral data acquisition have a specific bandwidth, which is ignored in the reconstruction. In this work, ignoring the bandwidth is shown to introduce a dependence of the recovered source intensity on the bandwidth of the filters. A method of accounting for the bandwidth of filters used during multispectral data acquisition is presented and its efficacy in increasing the quantitative accuracy of bioluminescence tomography is demonstrated through simulation and experiment. It is demonstrated that while using filters with a large bandwidth can dramatically decrease the data acquisition time, if not accounted for, errors of up to 200% in quantitative accuracy are introduced in two-dimensional planar imaging, even after normalization. For tomographic imaging, the use of this method to account for filter bandwidth dramatically improves the quantitative accuracy.

  7. Circular polarization observed in bioluminescence

    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

  8. Novel Bioluminescent Activatable Reporter for Src Tyrosine Kinase Activity in Living Mice.

    Leng, Weibing; Li, Dezhi; Chen, Liang; Xia, Hongwei; Tang, Qiulin; Chen, Baoqin; Gong, Qiyong; Gao, Fabao; Bi, Feng

    2016-01-01

    Aberrant activation of the Src kinase is implicated in the development of a variety of human malignancies. However, it is almost impossible to monitor Src activity in an in vivo setting with current biochemical techniques. To facilitate the noninvasive investigation of the activity of Src kinase both in vitro and in vivo, we developed a genetically engineered, activatable bioluminescent reporter using split-luciferase complementation. The bioluminescence of this reporter can be used as a surrogate for Src activity in real time. This hybrid luciferase reporter was constructed by sandwiching a Src-dependent conformationally responsive unit (SH2 domain-Srcpep) between the split luciferase fragments. The complementation bioluminescence of this reporter was dependent on the Src activity status. In our study, Src kinase activity in cultured cells and tumor xenografts was monitored quantitatively and dynamically in response to clinical small-molecular kinase inhibitors, dasatinib and saracatinib. This system was also applied for high-throughput screening of Src inhibitors against a kinase inhibitor library in living cells. These results provide unique insights into drug development and pharmacokinetics/phoarmocodynamics of therapeutic drugs targeting Src signaling pathway enabling the optimization of drug administration schedules for maximum benefit. Using both Firefly and Renilla luciferase imaging, we have successfully monitored Src tyrosine kinase activity and Akt serine/threonine kinase activity concurrently in one tumor xenograft. This dual luciferase reporter imaging system will be helpful in exploring the complex signaling networks in vivo. The strategies reported here can also be extended to study and image other important kinases and the cross-talks among them. PMID:26941850

  9. 活体生物发光成像追踪大鼠跟腱内移植干细胞**☆○%Monitoring transplanted stem cells in rat Achilles tendon by in vivo bioluminescent imaging

    黄德清; Gary Balian

    2013-01-01

      BACKGROUND: The mechanisms for the homing, migration, proliferation and differentiation of transplanted adipose tissue derived stem cel s remain unclear. The in vivo bioluminescent imaging system is a newly developed technique for directly detecting the biological behaviors of transplanted cel s in vivo. OBJECTIVE: To demonstrate the feasibility of using in vivo bioluminescent imaging system to monitor the genetical y modified adipose tissue derived stem cel s transplanted in Achil es tendon of rats. METHODS: Adipose tissue derived stem cel s isolated from the abdominal cavity of Sprague-Dawley rat were transduced with an adenovirus containing the luciferase reporter gene (3×1010/L), to observe the influence of transfection on the adipose tissue derived stem cel s. Subsequently, the transfected cel s were implanted into Achil es tendon defects in rats. The in vivo bioluminescent imaging system was used at days 1, 4, 7 and 14 fol owing transplantation to assess the luciferase expression. The cryosections of repaired Achil es tendon of rats were observed under fluorescence microscope at day 28 postoperatively. RESULTS AND CONCLUSION: No influence on the morphology and proliferation of adipose tissue derived stem cel s was observed after transducing in vitro (P > 0.05). On the repaired Achil es tendon, the luciferase gene expression detected with in vivo bioluminescent imaging system at days 1, 4, 7 and 14 was respectively (1.22±0.43)×106, (1.81±0.76)×106, (1.88±0.69)×106 and (0.89±0.26)×105 counts/s (n=6). Abundant adipose tissue derived stem cel s with luciferase expression were also seen in tendon cryosections of this side under fluorescence microscope at day 28. The luciferase gene expression was not detected in the control side. Experimental findings demonstrate that the in vivo bioluminescent imaging system can successful y monitor the fluorogene modified adipose tissue derived stem cel s that are implanted into the rat Achil es tendon, and

  10. Monitoring transplanted stem cells in rat Achilles tendon by in vivo bioluminescent imaging%活体生物发光成像追踪大鼠跟腱内移植干细胞**☆○

    黄德清; Gary Balian

    2013-01-01

      背景:移植脂肪源干细胞在活体内的归巢、迁移、增殖和分化的机制仍未得到充分阐明。活体生物发光活体成像技术是近来发展起来的一种可以直接检测活细胞在动物体内生物学行为的新的技术方法。目的:探讨用活体生物发光成像技术检测大鼠跟腱内移植的经荧光基因修饰的脂肪源干细胞可行性。方法:分离培养大鼠腹腔来源的脂肪源干细胞,用浓度为3×1010 L-1携带虫荧光素酶的腺病毒载体对其进行转染,观察转染对脂肪源干细胞的影响;将转染的脂肪源干细胞移植到大鼠跟腱缺损处,移植后1,4,7,14 d用活体生物发光成像技术检测移植脂肪源干细胞荧光素酶的表达,移植后28 d跟腱标本冰冻切片在荧光显微镜下观察。结果与结论:在体外,腺病毒转染对脂肪源干细胞的生长和增殖无明显影响(P >0.05)。细胞移植后1,4,7,14 d,活体生物发光成像技术在实验侧修复段跟腱检测到的荧光表达强度分别为(1.22±0.43)×106、(1.81±0.76)×106、(1.88±0.69)×106和(0.89±0.26)×105光子/s(n=6)。而对照侧跟腱修复段未检测到荧光表达;移植后28 d,实验侧跟腱冰冻切片在荧光显微镜下见到大量表达荧光的细胞。表明活体生物发光成像技术可成功追踪大鼠跟腱内移植的经荧光基因修饰的脂肪源干细胞。脂肪源干细胞有望成为肌腱组织工程的种子细胞。%  BACKGROUND: The mechanisms for the homing, migration, proliferation and differentiation of transplanted adipose tissue derived stem cel s remain unclear. The in vivo bioluminescent imaging system is a newly developed technique for directly detecting the biological behaviors of transplanted cel s in vivo. OBJECTIVE: To demonstrate the feasibility of using in vivo bioluminescent imaging system to monitor the genetical y modified adipose tissue derived stem cel s transplanted in

  11. Auto-luminescent genetically-encoded ratiometric indicator for real-time Ca2+ imaging at the single cell level.

    Kenta Saito

    Full Text Available BACKGROUND: Efficient bioluminescence resonance energy transfer (BRET from a bioluminescent protein to a fluorescent protein with high fluorescent quantum yield has been utilized to enhance luminescence intensity, allowing single-cell imaging in near real time without external light illumination. METHODOLOGY/PRINCIPAL FINDINGS: We applied BRET to develop an autoluminescent Ca(2+ indicator, BRAC, which is composed of Ca(2+-binding protein, calmodulin, and its target peptide, M13, sandwiched between a yellow fluorescent protein variant, Venus, and an enhanced Renilla luciferase, RLuc8. Adjusting the relative dipole orientation of the luminescent protein's chromophores improved the dynamic range of BRET signal change in BRAC up to 60%, which is the largest dynamic range among BRET-based indicators reported so far. Using BRAC, we demonstrated successful visualization of Ca(2+ dynamics at the single-cell level with temporal resolution at 1 Hz. Moreover, BRAC signals were acquired by ratiometric imaging capable of canceling out Ca(2+-independent signal drifts due to change in cell shape, focus shift, etc. CONCLUSIONS/SIGNIFICANCE: The brightness and large dynamic range of BRAC should facilitate high-sensitive Ca(2+ imaging not only in single live cells but also in small living subjects.

  12. Persistence of a bioluminescent Staphylococcus aureus strain on and around degradable and non-degradable surgical meshes in a murine model.

    Daghighi, Seyedmojtaba; Sjollema, Jelmer; Jaspers, Valery; de Boer, Leonie; Zaat, Sebastian A J; Dijkstra, Rene J B; van Dam, Gooitzen M; van der Mei, Henny C; Busscher, Henk J

    2012-11-01

    Biomaterials are increasingly used for the restoration of human function, but can become infected as a result of peri- or early post-operative bacterial contamination, although biomaterial-associated infections (BAIs) can also initiate at any time from hematogenous spreading of bacteria from an infection elsewhere in the body. Infecting bacteria in BAIs not only seek shelter in their own protective biofilm matrix, but also hide in surrounding tissue. This study compares staphylococcal persistence on and around a degradable and non-degradable surgical mesh through the use of longitudinal bioluminescence imaging in a murine model, including histological evaluation of surrounding tissue after sacrifice. Surgical meshes were first contaminated with bioluminescent Staphylococcus aureus Xen29 and subsequently subcutaneously implanted in mice. Bioluminescent staphylococci persisted on and around non-degradable meshes during the 28-day course of the study, whereas bioluminescence returned to control levels and bacteria disappeared from surrounding tissues once a degradable mesh had fully dissolved. Thus the application of degradable biomaterials yields major advantages with respect to the prevention of BAIs, as dissolution of the implant not only is associated with elimination of the protective biofilm mode of growth of the infecting organisms, but also allows the immune system to clear the surrounding tissue from infecting organisms. PMID:22824527

  13. Expanded palette of Nano-lanterns for real-time multicolor luminescence imaging.

    Takai, Akira; Nakano, Masahiro; Saito, Kenta; Haruno, Remi; Watanabe, Tomonobu M; Ohyanagi, Tatsuya; Jin, Takashi; Okada, Yasushi; Nagai, Takeharu

    2015-04-01

    Fluorescence live imaging has become an essential methodology in modern cell biology. However, fluorescence requires excitation light, which can sometimes cause potential problems, such as autofluorescence, phototoxicity, and photobleaching. Furthermore, combined with recent optogenetic tools, the light illumination can trigger their unintended activation. Because luminescence imaging does not require excitation light, it is a good candidate as an alternative imaging modality to circumvent these problems. The application of luminescence imaging, however, has been limited by the two drawbacks of existing luminescent protein probes, such as luciferases: namely, low brightness and poor color variants. Here, we report the development of bright cyan and orange luminescent proteins by extending our previous development of the bright yellowish-green luminescent protein Nano-lantern. The color change and the enhancement of brightness were both achieved by bioluminescence resonance energy transfer (BRET) from enhanced Renilla luciferase to a fluorescent protein. The brightness of these cyan and orange Nano-lanterns was ∼20 times brighter than wild-type Renilla luciferase, which allowed us to perform multicolor live imaging of intracellular submicron structures. The rapid dynamics of endosomes and peroxisomes were visualized at around 1-s temporal resolution, and the slow dynamics of focal adhesions were continuously imaged for longer than a few hours without photobleaching or photodamage. In addition, we extended the application of these multicolor Nano-lanterns to simultaneous monitoring of multiple gene expression or Ca(2+) dynamics in different cellular compartments in a single cell. PMID:25831507

  14. Bioluminescence Imaging Cells Labeled with Membrane-Anchored Form of Gaussia Luciferase%膜锚定Gaussia萤光素酶的细胞标记及生物荧光成像

    樊炜; 贾帅争; 王怡; 阎少多; 高博; 彭剑淳; 詹林盛

    2011-01-01

    目的:制备表达膜锚定Gaussia萤光素酶(extGluc)报告基因的慢病毒,用于标记细胞.方法:将报告基因extGluc克隆至慢病毒载体pCCsin.PPT.SFFV.IRES.eGFP.Wpre (VeGFP)中,以聚乙烯亚胺(PEI)介导,将慢病毒包装所需4种质粒(pVeGFP-extGLuc、pMDL、pRey、pVSVG),转染293FT细胞,72 h后收集病毒上清进行浓缩,感染293FT细胞,并用流式细胞仪检测病毒滴度,生物荧光成像和化学发光分析extGluc的表达;之后,用收集的慢病毒感染人单核细胞白血病细胞株U937.结果:对经PCR筛选出的阳性克隆所含质粒进行酶切鉴定,表明extGlu报告基因插入载体中;重组慢病毒包装成功且病毒滴度为5×106 TU/mL;用包装的病毒颗粒感染293FT、细胞,生物荧光成像和化学发光证实extGluc的膜定位,且酶活性与细胞数目呈线性相关;病毒颗粒能够感染悬浮细胞U937.结论:包装了extGluc标记的重组慢病毒,可用于标记细胞,为体内监测细胞迁移、聚集和变化提供了一种方法.%Objective: To produce lentivirus expressing reporter gene membrane-anchored form of Gaussia lu-ciferase (extGluc) used for bioluminescence imaging cells. Methods: Reporter gene extGluc was cloned into pCCsin. PPT.SFFV.IRES.eGFP.Wpre(VeGFP). pVeGFP-extGLuc, pMDL, pRev and pVSVG, which were required for packaging, were cotransfected into 293FT cells mediated by polyethylenimine branched. Lentivirus was collected at 72 h post-transfection, concentrated and then was used to infect 293FT cells. Viral titer was determined by flow cytome-try. Luciferase activity was detected by bioluminescence imaging and chemiluminescence. At last, human monocytic leukemia cell line U937 were infected by viral supernatant. Results: PCR and enzyme digestion results indicated that reporter gene extGLuc was successfully cloned into VeGFP. The lentivirus was packaged successfully. The lentivirus titer was 5xlO6 TU/mL. After infecting 293FT with virus particles, ext

  15. A multithread based new sparse matrix method in bioluminescence tomography

    Zhang, Bo; Tian, Jie; Liu, Dan; Sun, Li; Yang, Xin; Han, Dong

    2010-03-01

    Among many molecular imaging modalities, bioluminescence tomography (BLT) stands out as an effective approach for in vivo imaging because of its noninvasive molecular and cellular level detection ability, high sensitivity and low cost in comparison with other imaging technologies. However, there exists the case that large scale problem with large number of points and elements in the structure of mesh standing for the small animal or phantom. And the large scale problem's system matrix generated by the diffuse approximation (DA) model using finite element method (FEM) is large. So there wouldn't be enough random access memory (RAM) for the program and the related inverse problem couldn't be solved. Considering the sparse property of the BLT system matrix, we've developed a new sparse matrix (ZSM) to overcome the problem. And the related algorithms have all been speeded up by multi-thread technologies. Then the inverse problem is solved by Tikhonov regularization method in adaptive finite element (AFE) framework. Finally, the performance of this method is tested on a heterogeneous phantom and the boundary data is obtained through Monte Carlo simulation. During the process of solving the forward model, the ZSM can save more processing time and memory space than the usual way, such as those not using sparse matrix and those using Triples or Cross Linked sparse matrix. Numerical experiments have shown when more CPU cores are used, the processing speed is increased. By incorporating ZSM, BLT can be applied to large scale problems with large system matrix.

  16. Multi-atlas registration and adaptive hexahedral voxel discretization for fast bioluminescence tomography.

    Ren, Shenghan; Hu, Haihong; Li, Gen; Cao, Xu; Zhu, Shouping; Chen, Xueli; Liang, Jimin

    2016-04-01

    Bioluminescence tomography (BLT) has been a valuable optical molecular imaging technique to non-invasively depict the cellular and molecular processes in living animals with high sensitivity and specificity. Due to the inherent ill-posedness of BLT, a priori information of anatomical structure is usually incorporated into the reconstruction. The structural information is usually provided by computed tomography (CT) or magnetic resonance imaging (MRI). In order to obtain better quantitative results, BLT reconstruction with heterogeneous tissues needs to segment the internal organs and discretize them into meshes with the finite element method (FEM). It is time-consuming and difficult to handle the segmentation and discretization problems. In this paper, we present a fast reconstruction method for BLT based on multi-atlas registration and adaptive voxel discretization to relieve the complicated data processing procedure involved in the hybrid BLT/CT system. A multi-atlas registration method is first adopted to estimate the internal organ distribution of the imaged animal. Then, the animal volume is adaptively discretized into hexahedral voxels, which are fed into FEM for the following BLT reconstruction. The proposed method is validated in both numerical simulation and an in vivo study. The results demonstrate that the proposed method can reconstruct the bioluminescence source efficiently with satisfactory accuracy. PMID:27446674

  17. Multi-atlas registration and adaptive hexahedral voxel discretization for fast bioluminescence tomography

    Ren, Shenghan; Hu, Haihong; Li, Gen; Cao, Xu; Zhu, Shouping; Chen, Xueli; Liang, Jimin

    2016-01-01

    Bioluminescence tomography (BLT) has been a valuable optical molecular imaging technique to non-invasively depict the cellular and molecular processes in living animals with high sensitivity and specificity. Due to the inherent ill-posedness of BLT, a priori information of anatomical structure is usually incorporated into the reconstruction. The structural information is usually provided by computed tomography (CT) or magnetic resonance imaging (MRI). In order to obtain better quantitative results, BLT reconstruction with heterogeneous tissues needs to segment the internal organs and discretize them into meshes with the finite element method (FEM). It is time-consuming and difficult to handle the segmentation and discretization problems. In this paper, we present a fast reconstruction method for BLT based on multi-atlas registration and adaptive voxel discretization to relieve the complicated data processing procedure involved in the hybrid BLT/CT system. A multi-atlas registration method is first adopted to estimate the internal organ distribution of the imaged animal. Then, the animal volume is adaptively discretized into hexahedral voxels, which are fed into FEM for the following BLT reconstruction. The proposed method is validated in both numerical simulation and an in vivo study. The results demonstrate that the proposed method can reconstruct the bioluminescence source efficiently with satisfactory accuracy.

  18. Images of time mind, science, reality

    Jaroszkiewicz, George

    2016-01-01

    Have you ever wondered about Time: what it is or how to discuss it? If you have, then you may have been bewildered by the many different views and opinions in many diverse fields to be found, such as physics, mathematics, philosophy, religion, history, and science fiction novels and films. This book will help you unravel fact from fiction. It provides a broad survey of many of these views, these images of time, covering historical, cultural, philosophical, biological, mathematical and physical images of time, including classical and quantum mechanics, special and general relativity and cosmology. This book gives you more than just a review of such images. It provides the reader a basis for judging the scientific soundness of these various images. It develops the reader's critical ability to distinguish Images of Time in terms of its contextual completeness. Differentiating between metaphysical images (which cannot be scientifically validated) and those that could, in principle, be put to empirical test. Showi...

  19. A Nisin Bioassay Based on Bioluminescence

    Wahlström, G.; Saris, P. E. J.

    1999-01-01

    A Lactococcus lactis subsp. lactis strain that can sense the bacteriocin nisin and transduce the signal into bioluminescence was constructed. By using this strain, a bioassay based on bioluminescence was developed for quantification of nisin, for detection of nisin in milk, and for identification of nisin-producing strains. As little as 0.0125 ng of nisin per ml was detected within 3 h by this bioluminescence assay. This detection limit was lower than in previously described methods.

  20. Protein-protein complexation in bioluminescence

    Titushin, Maxim S.; Feng, Yingang; Lee, John; Vysotski, Eugene S.; Liu, Zhi-jie

    2011-01-01

    In this review we summarize the progress made towards understanding the role of protein-protein interactions in the function of various bioluminescence systems of marine organisms, including bacteria, jellyfish and soft corals, with particular focus on methodology used to detect and characterize these interactions. In some bioluminescence systems, protein-protein interactions involve an “accessory protein” whereby a stored substrate is efficiently delivered to the bioluminescent enzyme lucife...

  1. Non-invasive imaging of GFP-luciferase labeled orthotopic prostate cancer model in nude mice using bioluminescence system%可发光可连续检测原位前列腺癌模型的建立

    宋超; 廖文彪; 杨嗣星; 王玲珑

    2012-01-01

    .43 ± 4.56 ) g accordingly.There was a positive linear correlation between in vivo bioluminescent values and ex vivo tumor weight with coefficient being 0.973,P <0.05.Conclusion Our findings demonstrate the ability of the luciferase labeled tumor cells combinated within vivo bioluminesence imaging system is an excellent experimental animal model in tracking the location,magnitude and persistence of luciferase expression cells in human prostate cancer mouse models,which may be usful in non-invasively monitoring on progression of prostate cancer in vivo,even after the metastasis of the tumor.%目的 建立一种可连续定量监测的原位前列腺癌动物模型.方法 利用分子克隆技术构建表达荧光素酶的慢病毒载体,用慢病毒将GFP-Luciferase融合蛋白载体转染到人前列腺癌PC3细胞株;利用稻瘟素Blasticidin筛选获得稳定转染的GFP-Luc-PC3细胞株.将6只雄性裸鼠随机分2组,应用5×106 GFP-Luc-PC3或普通PC3细胞雄性裸鼠皮下注射获得皮下瘤.12周时无菌条件下取荧光素酶标记和未标记的皮下肿瘤组织,将其切成2 mm3大小的组织块,并将组织块随机种植于24只雄性裸鼠前列腺包膜下,实验组、对照组各12只.用IVIS 200光子发射定量分析动态监测肿瘤生长.并于种植后2、4、6、8周分别处死3只裸鼠,取前列腺组织称重.分析肿瘤在体光子值与其重量的相关性.结果 成功建立GFP-Luc-PC3细胞株,该系生长曲线与未标记细胞一致,在荧光素作用下,发光能力与细胞数量呈正相关(r=0.997),其裸鼠皮下成瘤率为100%.原位前列腺癌模型中,所有24只动物均形成前列腺肿瘤,其中GFP-Luc-PC3组可通过IVIS 200系统观察到肿瘤组织的发光情况,PC3组信号不明显.原位肿瘤模型建立后2、4、6、8周,肿瘤在体光子值(光子值/秒,photons/second)依次为(69.13298±2.07900)E+05、(82.66208±1.23100)E+05、(91.942 57±2.32100)E +05、(130.643 40±3.247 00)E+05.

  2. Positive self-image over time

    Santos-Pinto, Luís

    2005-01-01

    This paper incorporates egocentric comparisons into a human capital accumulation model and studies the evolution of positive self image over time. The paper shows that the process of human capital accumulation together with egocentric comparisons imply that positive self image of a cohort is first increasing and then decreasing over time. Additionally, the paper finds that positive self image: (1) peaks earlier in activities where skill depreciation is higher, (2) is smaller in activities whe...

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

    Vacquié-Garcia, Jade; Royer, François; Dragon, Anne-Cécile; Viviant, Morgane; Bailleul, Frédéric; Guinet, Christophe

    2012-01-01

    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. PMID:22952706

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

    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.

  5. BIOLUMINESCENCE TOMOGRAPHY: BIOMEDICAL BACKGROUND, MATHEMATICAL THEORY, AND NUMERICAL APPROXIMATION

    Weimin Han; Ce Wang

    2008-01-01

    Over the last couple of years molecular imaging has been rapidly developed to study physiological and pathological processes in vivo at the cellular and molecular levels. Among molecular imaging modalities, optical imaging stands out for its unique advantages, especially performance and cost-effectiveness. Bioluminescence tomography (BLT) is an emerging optical imaging mode with promising biomedical advantages. In this survey paper, we explain the biomedical significance of BLT, summarize theoretical results on the analysis and numerical solution of a diffusion based BLT model, and comment on a few extensions for the study of BLT.

  6. Mathematical Study and Numerical Simulation of Multispectral Bioluminescence Tomography

    Weimin Han; Wenxiang Cong; Ge Wang

    2006-01-01

    Multispectral bioluminescence tomography (BLT) attracts increasingly more attention in the area of optical molecular imaging. In this paper, we analyze the properties of the solutions to the regularized and discretized multispectral BLT problems. First, we show the solution existence, uniqueness, and its continuous dependence on the data. Then, we introduce stable numerical schemes and derive error estimates for numerical solutions. We report some numerical results to illust...

  7. Experimental Study on Bioluminescence Tomography with Multimodality Fusion

    Lv, Yujie; Tian, Jie; Cong, Wenxiang; Wang, Ge

    2007-01-01

    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 s...

  8. Multi-projection bioluminescence tomography guided system for small animal radiation research platform (SARRP)

    Zhang, Bin; Iordachita, Iulian; Wong, John W.; Wang, Ken Kang-Hsin

    2016-03-01

    Cone beam computed tomography (CBCT) is limited in guiding irradiation for soft tissue targets. As a complementary imaging modality, bioluminescence tomography (BLT) provides strong soft tissue contrast. We developed a dual-use BLT system which consists of an optical assembly, a mobile cart and an independent mouse bed. The system is motorized which can easily dock onto an independent mouse bed operating as a standalone system for longitudinal bioluminescence imaging (BLI)/BLT studies and also dock onto the SARRP for on-line radiation guidance. Our initial tests for the system demonstrate that (i) the imaging depth is 28 mm, (ii) the optical background is sufficiently low and uniform, (iii) the non-uniform response of the optical imaging can be corrected by the flat field correction, and (iv) the imaging acquisition speed was improved by an average of 3.7 times faster than our previous systems. We also presented a geometry calibration procedure to map the planar BLIs acquired at multi-projections onto the surface of the CBCT image. The CBCT is required to generate the mesh for BLT reconstruction and used for treatment planning and radiation delivery. Feasibility study of the geometry calibration was performed on a manual-docking prototype. The mean and maximum mapping accuracy is 0.3 and 0.6 mm. The performance of the proposed motorized dual-use system is expected to be superior to that of the manual-docking prototype because of the mechanism stability. We anticipate the dual-use system as a highly efficient and cost-effective platform to facilitate optical imaging for preclinical radiation research.

  9. Preclinical evaluation of destruxin B as a novel Wnt signaling target suppressing proliferation and metastasis of colorectal cancer using non-invasive bioluminescence imaging

    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.

  10. Preclinical evaluation of destruxin B as a novel Wnt signaling target suppressing proliferation and metastasis of colorectal cancer using non-invasive bioluminescence imaging

    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.

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

    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.

  12. REVIEW OF ENVIRONMENTAL APPLICATIONS OF BIOLUMINESCENCE MEASUREMENTS

    This review of the recent literature on environmental applications of bioluminescence systems will focus on in vivo and in vitro bioluminescence methods that have been utilized to elucidate properties of chemicals, toxic and mutagenic effects, and to estimate biomass. he unifying...

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

    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.

  14. Understanding bioluminescence in dinoflagellates — how far have we come?

    Martha Valiadi; Debora Iglesias-Rodriguez

    2013-01-01

    Some dinoflagellates possess the remarkable genetic, biochemical, and cellular machinery to produce bioluminescence. Bioluminescent species appear to be ubiquitous in surface waters globally and include numerous cosmopolitan and harmful taxa. Nevertheless, bioluminescence remains an enigmatic topic in biology, particularly with regard to the organisms’ lifestyle. In this paper, we review the literature on the cellular mechanisms, molecular evolution, diversity, and ecology of bioluminescence ...

  15. Bioluminescence tomography based on the phase approximation model

    Cong, W.; Wang, G

    2010-01-01

    A reconstruction method of bioluminescence sources is proposed based on a phase approximation model. Compared with the diffuse approximation, this phase approximation model more correctly predicts bioluminescence photon propagation in biological tissues, so that bioluminescence tomography can accurately locate and quantify the distribution of bioluminescence sources. The compressive sensing (CS) technique is applied to regularize the inverse source reconstruction to enhance numerical stabilit...

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

    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

  17. Toward Real Time Uavs' Image Mosaicking

    Mehrdad, S.; Satari, M.; Safdary, M.; Moallem, P.

    2016-06-01

    Anyone knows that sudden catastrophes can instantly do great damage. Fast and accurate acquisition of catastrophe information is an essential task for minimize life and property damage. Compared with other ways of catastrophe data acquisition, UAV based platforms can optimize time, cost and accuracy of the data acquisition, as a result UAVs' data has become the first choice in such condition. In this paper, a novel and fast strategy is proposed for registering and mosaicking of UAVs' image data. Firstly, imprecise image positions are used to find adjoining frames. Then matching process is done by a novel matching method. With keeping Sift in mind, this fast matching method is introduced, which uses images exposure time geometry, SIFT point detector and rBRIEF descriptor vector in order to match points efficiency, and by efficiency we mean not only time efficiency but also elimination of mismatch points. This method uses each image sequence imprecise attitude in order to use Epipolar geometry to both restricting search space of matching and eliminating mismatch points. In consideration of reaching to images imprecise attitude and positions we calibrated the UAV's sensors. After matching process, RANSAC is used to eliminate mismatched tie points. In order to obtain final mosaic, image histograms are equalized and a weighted average method is used to image composition in overlapping areas. The total RMSE over all matching points is 1.72 m.

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

    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.

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

    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. Interactive Real-time Magnetic Resonance Imaging

    Brix, Lau

    Real-time acquisition, reconstruction and interactively changing the slice position using magnetic resonance imaging (MRI) have been possible for years. However, the current clinical use of interactive real-time MRI is limited due to an inherent low spatial and temporal resolution. This PhD project...... regard to optimal sampling strategy for detecting motion in four different anatomies on two different MRI scanner brands. A fully implemented interactive real-time MRI system was exploited in a group of healthy fetuses and proved its eligibility as an alternative diagnostic tool for fetal imaging...

  1. NanoLuc: A Small Luciferase Is Brightening Up the Field of Bioluminescence.

    England, Christopher G; Ehlerding, Emily B; Cai, Weibo

    2016-05-18

    The biomedical field has greatly benefited from the discovery of bioluminescent proteins. Currently, scientists employ bioluminescent systems for numerous biomedical applications, ranging from highly sensitive cellular assays to bioluminescence-based molecular imaging. Traditionally, these systems are based on Firefly and Renilla luciferases; however, the applicability of these enzymes is limited by their size, stability, and luminescence efficiency. NanoLuc (NLuc), a novel bioluminescence platform, offers several advantages over established systems, including enhanced stability, smaller size, and >150-fold increase in luminescence. In addition, the substrate for NLuc displays enhanced stability and lower background activity, opening up new possibilities in the field of bioluminescence imaging. The NLuc system is incredibly versatile and may be utilized for a wide array of applications. The increased sensitivity, high stability, and small size of the NLuc system have the potential to drastically change the field of reporter assays in the future. However, as with all such technology, NLuc has limitations (including a nonideal emission for in vivo applications and its unique substrate) which may cause it to find restricted use in certain areas of molecular biology. As this unique technology continues to broaden, NLuc may have a significant impact in both preclinical and clinical fields, with potential roles in disease detection, molecular imaging, and therapeutic monitoring. This review will present the NLuc technology to the scientific community in a nonbiased manner, allowing the audience to adopt their own views of this novel system. PMID:27045664

  2. Experimental Study on Bioluminescence Tomography with Multimodality Fusion

    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.

  3. Actual imaging time in fetal MRI

    Objective: Safety issues in magnetic resonance imaging (MRI) are important, especially in fetal MRI. However, since basic data with respect of the effective exposure time in fetal MRI are not available, this study aimed to determine the actual imaging time during a fetal MRI study. Methods: 100 fetal MRI studies of singleton pregnancies performed on a 1.5 T system were analysed with respect to study duration (from starting the survey scan until the end of study), the number of sequences acquired, and the actual imaging time, which was calculated by adding up scan time of each sequence. Furthermore, each sequence type was analysed regarding the number of acquisitions, specific absorption rates (SAR), and duration. Results: Mean study duration was 34.6 min (range: 14–58 min; standard deviation (SD): 9.7 min), the average number of sequences acquired was 26.6 (range: 11–44, SD: 6.6). Actual scan time averaged 11.4 min (range: 4–19 min, SD: 4.0 min). Ultrafast T2-weighted and steady-state free-precession sequences accounted for 62.3% of actual scan time, and were distributed over the whole duration of the study. Conclusion: Actual imaging time only accounts for 33% of total study time and is not continuous. The remaining time is consumed by the preparation phases of the scanner, and is spent with planning sequences and the eventual repositioning of the coil and/or pregnant woman. These data may help to more accurately estimate the exposure to radiofrequency deposition and noise during fetal MRI studies.

  4. Gemcitabine upregulates ABCG2/BCRP and modulates the intracellular pharmacokinetic profiles of bioluminescence in pancreatic cancer cells.

    Sun, Yue; Gu, Mancang; Zhu, Lixin; Liu, Junying; Xiong, Yang; Wei, Yinghui; Li, Fanzhu

    2016-03-01

    A lack of methods capable of exploring real-time intracellular drug deposition has since limited the investigation of gemcitabine-induced multidrug resistance in vitro and in vivo. Specifically, resistance induced by D-luciferin, a substrate of the breast cancer resistance protein (ABCG2/BCRP), has recently attracted clinical attention, but further investigation has been limited. Herein, the intracellular pharmacokinetic behavior of D-luciferin was investigated in pancreatic cancer cell lines in real time by using bioluminescence imaging. To achieve this feat, BxPC3 and Panc1 pancreatic cancer cells overexpressing firefly luciferase were treated with gemcitabine in a dose and time gradient manner in vitro. The intracellular pharmacokinetic profiles of each group were then determined through the acquisition of bioluminescent signal intensity of D-luciferin in cells. Simultaneously, key pharmacokinetic parameters including area under the curve, elimination rate constant (K), and mean resident time were calculated according to the noncompartment model. ABCG2 protein levels following gemcitabine treatment were detected through western blot, and gemcitabine showed no significant effect on luciferase activity over dimethyl sulfoxide (DMSO) as a control (P>0.05). However, gemcitabine significantly increased K values while suppressing area under the curve and mean resident time compared with DMSO (Pbioluminescent model and its capability to observe the onset of chemoresistance in real time. PMID:26556627

  5. Analytical Applications of Bioluminescence and Chemiluminescence

    Chappelle, E. W. (Editor); Picciolo, G. L. (Editor)

    1975-01-01

    Bioluminescence and chemiluminescence studies were used to measure the amount of adenosine triphosphate and therefore the amount of energy available. Firefly luciferase - luciferin enzyme system was emphasized. Photometer designs are also considered.

  6. Circadian control sheds light on fungal bioluminescence.

    Oliveira, Anderson G; Stevani, Cassius V; Waldenmaier, Hans E; Viviani, Vadim; Emerson, Jillian M; Loros, Jennifer J; Dunlap, Jay C

    2015-03-30

    Bioluminescence, the creation and emission of light by organisms, affords insight into the lives of organisms doing it. Luminous living things are widespread and access diverse mechanisms to generate and control luminescence [1-5]. Among the least studied bioluminescent organisms are phylogenetically rare fungi-only 71 species, all within the ∼ 9,000 fungi of the temperate and tropical Agaricales order-are reported from among ∼ 100,000 described fungal species [6, 7]. All require oxygen [8] and energy (NADH or NADPH) for bioluminescence and are reported to emit green light (λmax 530 nm) continuously, implying a metabolic function for bioluminescence, perhaps as a byproduct of oxidative metabolism in lignin degradation. Here, however, we report that bioluminescence from the mycelium of Neonothopanus gardneri is controlled by a temperature-compensated circadian clock, the result of cycles in content/activity of the luciferase, reductase, and luciferin that comprise the luminescent system. Because regulation implies an adaptive function for bioluminescence, a controversial question for more than two millennia [8-15], we examined interactions between luminescent fungi and insects [16]. Prosthetic acrylic resin "mushrooms," internally illuminated by a green LED emitting light similar to the bioluminescence, attract staphilinid rove beetles (coleopterans), as well as hemipterans (true bugs), dipterans (flies), and hymenopterans (wasps and ants), at numbers far greater than dark control traps. Thus, circadian control may optimize energy use for when bioluminescence is most visible, attracting insects that can in turn help in spore dispersal, thereby benefitting fungi growing under the forest canopy, where wind flow is greatly reduced. PMID:25802150

  7. Circadian Control Sheds Light on Fungal Bioluminescence

    Oliveira, Anderson G.; Cassius V. Stevani; Waldenmaier, Hans E.; Viviani, Vadim; Emerson, Jillian M.; Loros, Jennifer J.; Jay C Dunlap

    2015-01-01

    Bioluminescence, the creation and emission of light by organisms, affords insight into the lives of organisms doing it. Luminous living things are widespread and access diverse mechanisms to generate and control luminescence [1-5]. Among the least studied bioluminescent organisms are phylogenetically rare fungi – only 71 species, all within the ~9000 fungi of the temperate and tropical Agaricales Order - are reported from among ~100,000 described fungal species [6,7]. All require oxygen [8] a...

  8. Real-time image and video processing

    Kehtarnavaz, Nasser

    2006-01-01

    This book presents an overview of the guidelines and strategies for transitioning an image or video processing algorithm from a research environment into a real-time constrained environment. Such guidelines and strategies are scattered in the literature of various disciplines including image processing, computer engineering, and software engineering, and thus have not previously appeared in one place. By bringing these strategies into one place, the book is intended to serve the greater community of researchers, practicing engineers, industrial professionals, who are interested in taking an im

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

    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

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

    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 l2 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 rather