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Sample records for fluorescent imaging agent

  1. Breast Cancer Imaging Using the Near-Infrared Fluorescent Agent, CLR1502

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    Melissa L. Korb

    2015-01-01

    Full Text Available Positive margins after breast conservation surgery represent a significant problem in the treatment of breast cancer. The near-infrared fluorescence agent CLR1502 (Cellectar Biosciences, Madison, WI was studied in a preclinical breast cancer model to determine imaging properties and ability to detect small islands of malignancy. Nude mice bearing human breast cancer flank xenografts were given a systemic injection of CLR1502, and imaging was performed using LUNA (Novadaq Technologies Inc., Richmond, BC and Pearl Impulse (LI-COR Biosciences, Lincoln, NE devices. Normal tissues were examined for fluorescence signal, and conventional and fluorescence histology was performed using the Odyssey scanner. Peak tumor to background ratio occurred 2 days after injection with CLR1502. The smallest amount of tumor that was imaged and detected using these devices was 1.9 mg, equivalent to 1.9 × 106 cells. The highest fluorescence signal was seen in tumor and normal lymph node tissue, and the lowest fluorescence signal was seen in muscle and plasma. Human breast cancer tumors can be imaged in vivo with multiple optical imaging platforms using CLR1502. This pilot study supports further investigations of this fluorescent agent for improving surgical resection of malignancies, with the goal of eventual clinical translation.

  2. Quantitative whole body biodistribution of fluorescent-labeled agents by non-invasive tomographic imaging.

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    Kristine O Vasquez

    Full Text Available When small molecules or proteins are injected into live animals, their physical and chemical properties will significantly affect pharmacokinetics, tissue penetration, and the ultimate routes of metabolism and clearance. Fluorescence molecular tomography (FMT offers the ability to non-invasively image and quantify temporal changes in fluorescence throughout the major organ systems of living animals, in a manner analogous to traditional approaches with radiolabeled agents. This approach is best used with biotherapeutics (therapeutic antibodies, or other large proteins or large-scaffold drug-delivery vectors, that are minimally affected by low-level fluorophore conjugation. Application to small molecule drugs should take into account the significant impact of fluorophore labeling on size and physicochemical properties, however, the presents studies show that this technique is readily applied to small molecule agents developed for far-red (FR or near infrared (NIR imaging. Quantification by non-invasive FMT correlated well with both fluorescence from tissue homogenates as well as with planar (2D fluorescence reflectance imaging of excised intact organs (r²  =  0.996 and 0.969, respectively. Dynamic FMT imaging (multiple times from 0 to 24 h performed in live mice after the injection of four different FR/NIR-labeled agents, including immunoglobulin, 20-50 nm nanoparticles, a large vascular imaging agent, and a small molecule integrin antagonist, showed clear differences in the percentage of injected dose per gram of tissue (%ID/g in liver, kidney, and bladder signal. Nanoparticles and IgG1 favored liver over kidney signal, the small molecule integrin-binding agent favored rapid kidney and bladder clearance, and the vascular agent, showed both liver and kidney clearance. Further assessment of the volume of distribution of these agents by fluorescent volume added information regarding their biodistribution and highlighted the relatively poor

  3. Quantitative Imaging of Cell-Permeable Magnetic Resonance Contrast Agents Using X-Ray Fluorescence

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    Paul J. Endres

    2006-10-01

    Full Text Available The inability to transduce cellular membranes is a limitation of current magnetic resonance imaging probes used in biologic and clinical settings. This constraint confines contrast agents to extracellular and vascular regions of the body, drastically reducing their viability for investigating processes and cycles in developmental biology. Conversely, a contrast agent with the ability to permeate cell membranes could be used in visualizing cell patterning, cell fate mapping, gene therapy, and, eventually, noninvasive cancer diagnosis. Therefore, we describe the synthesis and quantitative imaging of four contrast agents with the capability to cross cell membranes in sufficient quantity for detection. Each agent is based on the conjugation of a Gd(III chelator with a cellular transduction moiety. Specifically, we coupled Gd(III–diethylenetriaminepentaacetic acid DTPA and Gd(III–1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid with an 8–amino acid polyarginine oligomer and an amphipathic stilbene molecule, 4-amino-4'-(N,N-dimethylaminostilbene. The imaging modality that provided the best sensitivity and spatial resolution for direct detection of the contrast agents is synchrotron radiation x-ray fluorescence (SR-XRF. Unlike optical microscopy, SR-XRF provides two-dimensional images with resolution 103 better than 153Gd gamma counting, without altering the agent by organic fluorophore conjugation. The transduction efficiency of the intracellular agents was evaluated by T1 analysis and inductively coupled plasma mass spectrometry to determine the efficacy of each chelate-transporter combination.

  4. New generation ICG-based contrast agents for ultrasound-switchable fluorescence imaging

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    Yu, Shuai; Cheng, Bingbing; Yao, Tingfeng; Xu, Cancan; Nguyen, Kytai T.; Hong, Yi; Yuan, Baohong

    2016-10-01

    Recently, we developed a new technology, ultrasound-switchable fluorescence (USF), for high-resolution imaging in centimeter-deep tissues via fluorescence contrast. The success of USF imaging highly relies on excellent contrast agents. ICG-encapsulated poly(N-isopropylacrylamide) nanoparticles (ICG-NPs) are one of the families of the most successful near-infrared (NIR) USF contrast agents. However, the first-generation ICG-NPs have a short shelf life (6 months). In addition, we have conjugated hydroxyl or carboxyl function groups on the ICG-NPs for future molecular targeting. Finally, we have demonstrated the effect of temperature-switching threshold (Tth) and the background temperature (TBG) on the quality of USF images. We estimated that the Tth of the ICG-NPs should be controlled at ~38–40 °C (slightly above the body temperature of 37 °C) for future in vivo USF imaging. Addressing these challenges further reduces the application barriers of USF imaging.

  5. New generation ICG-based contrast agents for ultrasound-switchable fluorescence imaging

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    Yu, Shuai; Cheng, Bingbing; Yao, Tingfeng; Xu, Cancan; Nguyen, Kytai T.; Hong, Yi; Yuan, Baohong

    2016-01-01

    Recently, we developed a new technology, ultrasound-switchable fluorescence (USF), for high-resolution imaging in centimeter-deep tissues via fluorescence contrast. The success of USF imaging highly relies on excellent contrast agents. ICG-encapsulated poly(N-isopropylacrylamide) nanoparticles (ICG-NPs) are one of the families of the most successful near-infrared (NIR) USF contrast agents. However, the first-generation ICG-NPs have a short shelf life (6 months). In addition, we have conjugated hydroxyl or carboxyl function groups on the ICG-NPs for future molecular targeting. Finally, we have demonstrated the effect of temperature-switching threshold (Tth) and the background temperature (TBG) on the quality of USF images. We estimated that the Tth of the ICG-NPs should be controlled at ~38–40 °C (slightly above the body temperature of 37 °C) for future in vivo USF imaging. Addressing these challenges further reduces the application barriers of USF imaging. PMID:27775014

  6. Cell tracking with gadophrin-2: a bifunctional contrast agent for MR imaging, optical imaging, and fluorescence microscopy

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    Daldrup-Link, Heike E. [Department of Radiology, UCSF Medical Center, University of California in San Francisco, 513 Parnassus Ave, CA 94143, San Francisco (United States); Rudelius, Martina; Piontek, Guido; Schlegel, Juergen [Institute of Pathology, Technical University, Munich (Germany); Metz, Stephan; Settles, Marcus; Rummeny, Ernst J. [Department of Radiology, Technical University, Munich (Germany); Pichler, Bernd [Department of Biomedical Engineering, University of California Davis, Davis (United States); Heinzmann, Ulrich [National Research Center for Environment and Health, Technical University, Munich (Germany); Oostendorp, Robert A.J. [3. Clinic of Internal Medicine, Laboratory of Stem Cell Physiology, Technical University, Munich (Germany)

    2004-09-01

    The purpose of this study was to assess the feasibility of use of gadophrin-2 to trace intravenously injected human hematopoietic cells in athymic mice, employing magnetic resonance (MR) imaging, optical imaging (OI), and fluorescence microscopy. Mononuclear peripheral blood cells from GCSF-primed patients were labeled with gadophrin-2 (Schering AG, Berlin, Germany), a paramagnetic and fluorescent metalloporphyrin, using established transfection techniques with cationic liposomes. The labeled cells were evaluated in vitro with electron microscopy and inductively coupled plasma atomic emission spectrometry. Then, 1 x 10{sup 6}-3 x 10{sup 8} labeled cells were injected into 14 nude Balb/c mice and the in vivo cell distribution was evaluated with MR imaging and OI before and 4, 24, and 48 h after intravenous injection (p.i.). Five additional mice served as controls: three mice were untreated controls and two mice were investigated after injection of unlabeled cells. The contrast agent effect was determined quantitatively for MR imaging by calculating signal-to-noise-ratio (SNR) data. After completion of in vivo imaging studies, fluorescence microscopy of excised organs was performed. Intracellular cytoplasmatic uptake of gadophrin-2 was confirmed by electron microscopy. Spectrometry determined an uptake of 31.56 nmol Gd per 10{sup 6} cells. After intravenous injection, the distribution of gadophrin-2 labeled cells in nude mice could be visualized by MR, OI, and fluorescence microscopy. At 4 h p.i., the transplanted cells mainly distributed to lung, liver, and spleen, and 24 h p.i. they also distributed to the bone marrow. Fluorescence microscopy confirmed the distribution of gadophrin-2 labeled cells to these target organs. Gadophrin-2 is suited as a bifunctional contrast agent for MR imaging, OI, and fluorescence microscopy and may be used to combine the advantages of each individual imaging modality for in vivo tracking of intravenously injected hematopoietic cells

  7. Determining Optimal Fluorescent Agent Concentrations in Dental Adhesive Resins for Imaging the Tooth/Restoration Interface.

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    Bim Júnior, Odair; Cebim, Marco A; Atta, Maria T; Machado, Camila M; Francisconi-Dos-Rios, Luciana F; Wang, Linda

    2017-02-01

    Fluorescent dyes like Rhodamine B (RB) have been used to identify the spatial distribution of adhesive restorative materials in the tooth/restoration interface. Potential effects of the addition of RB to dental adhesives were addressed in the past, but no further information is available on how to determine suitable concentrations of RB in these bonding agents for imaging in the confocal laser scanning microscope. This study provides systematical strategies for adding RB to viscous dental adhesive resins, focusing on the determination of the lowest range of dye concentrations necessary to achieve an acceptable image of the dentin/adhesive interface. It was demonstrated that optimized images of the resin distribution in dentin can be produced with 0.1-0.02 mg/mL of RB in the (tested) adhesives. Our approaches took into account aspects related to the dye concentration, photophysical parameters in different host media, specimen composition and morphology to develop a rational use of the fluorescent agent with the resin-based materials. Information gained from this work can help optimize labeling methods using dispersions of low-molecular-weight dyes in different monomer blend systems.

  8. Imaging Primary Mouse Sarcomas After Radiation Therapy Using Cathepsin-Activatable Fluorescent Imaging Agents

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    Cuneo, Kyle C. [Department of Radiation Oncology, Duke University School of Medicine, Durham, North Carolina (United States); Mito, Jeffrey K.; Javid, Melodi P. [Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, North Carolina (United States); Ferrer, Jorge M. [Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Kim, Yongbaek [Department of Clinical Pathology, College of Veterinary Medicine, Seoul National University, Seoul (Korea, Republic of); Lee, W. David [The David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Bawendi, Moungi G. [Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Brigman, Brian E. [Department of Orthopedic Surgery, Duke University School of Medicine, Durham, North Carolina (United States); Kirsch, David G., E-mail: david.kirsch@duke.edu [Department of Radiation Oncology, Duke University School of Medicine, Durham, North Carolina (United States); Department of Pharmacology and Cancer Biology, Duke University School of Medicine, Durham, North Carolina (United States)

    2013-05-01

    Purpose: Cathepsin-activated fluorescent probes can detect tumors in mice and in canine patients. We previously showed that these probes can detect microscopic residual sarcoma in the tumor bed of mice during gross total resection. Many patients with soft tissue sarcoma (STS) and other tumors undergo radiation therapy (RT) before surgery. This study assesses the effect of RT on the ability of cathepsin-activated probes to differentiate between normal and cancerous tissue. Methods and Materials: A genetically engineered mouse model of STS was used to generate primary hind limb sarcomas that were treated with hypofractionated RT. Mice were injected intravenously with cathepsin-activated fluorescent probes, and various tissues, including the tumor, were imaged using a hand-held imaging device. Resected tumor and normal muscle samples were harvested to assess cathepsin expression by Western blot. Uptake of activated probe was analyzed by flow cytometry and confocal microscopy. Parallel in vitro studies using mouse sarcoma cells were performed. Results: RT of primary STS in mice and mouse sarcoma cell lines caused no change in probe activation or cathepsin protease expression. Increasing radiation dose resulted in an upward trend in probe activation. Flow cytometry and immunofluorescence showed that a substantial proportion of probe-labeled cells were CD11b-positive tumor-associated immune cells. Conclusions: In this primary murine model of STS, RT did not affect the ability of cathepsin-activated probes to differentiate between tumor and normal muscle. Cathepsin-activated probes labeled tumor cells and tumor-associated macrophages. Our results suggest that it would be feasible to include patients who have received preoperative RT in clinical studies evaluating cathepsin-activated imaging probes.

  9. Time-domain imaging with quench-based fluorescent contrast agents

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    Akers, Walter J.; Solomon, Metasebya; Sudlow, Gail P.; Berezin, Mikhail; Achilefu, Samuel

    2012-03-01

    Quench-based probes utilize unique characteristics of fluorescence resonance energy transfer (FRET) to enhance contrast upon de-quenching. This mechanism has been used in a variety of molecular probes for imaging of cancer related enzyme activity such as matrix metalloproteinases, cathepsins and caspases. While non-fluorescent upon administration, fluorescence can be restored by separation of donor and acceptor, resulting in higher intensity in the presence of activator. Along with decreased quantum yield, FRET also results in altered fluorescence lifetime. Time-domain imaging can further enhance contrast and information yield from quench-based probes. We present in vivo time-domain imaging for detecting activation of quench-based probes. Quench-based probes utilize unique characteristics of fluorescence resonance energy transfer (FRET) to enhance contrast upon de-quenching. This mechanism has been used in a variety of molecular probes for imaging of cancer related enzyme activity such as matrix metalloproteinases, cathepsins and caspases. While non-fluorescent upon administration, fluorescence can be restored by separation of donor and acceptor, resulting in higher intensity in the presence of activator. Along with decreased quantum yield, FRET also results in altered fluorescence lifetime. Time-domain imaging can further enhance contrast and information yield from quench-based probes. We present in vivo time-domain imaging for detecting activation of quench-based probes. Time-domain diffuse optical imaging was performed to assess the FRET and quenching in living mice with orthotopic breast cancer. Tumor contrast enhancement was accompanied by increased fluorescence lifetime after administration of quenched probes selective for matrix metalloproteinases while no significant change was observed for non-quenched probes for integrin receptors. These results demonstrate the utility of timedomain imaging for detection of cancer-related enzyme activity in vivo.

  10. PLGA nanoparticles from nano-emulsion templating as imaging agents: Versatile technology to obtain nanoparticles loaded with fluorescent dyes.

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    Fornaguera, C; Feiner-Gracia, N; Calderó, G; García-Celma, M J; Solans, C

    2016-11-01

    The interest in polymeric nanoparticles as imaging systems for biomedical applications has increased notably in the last decades. In this work, PLGA nanoparticles, prepared from nano-emulsion templating, have been used to prepare novel fluorescent imaging agents. Two model fluorescent dyes were chosen and dissolved in the oil phase of the nano-emulsions together with PLGA. Nano-emulsions were prepared by the phase inversion composition (PIC) low-energy method. Fluorescent dye-loaded nanoparticles were obtained by solvent evaporation of nano-emulsion templates. PLGA nanoparticles loaded with the fluorescent dyes showed hydrodynamic radii lower than 40nm; markedly lower than those reported in previous studies. The small nanoparticle size was attributed to the nano-emulsification strategy used. PLGA nanoparticles showed negative surface charge and enough stability to be used for biomedical imaging purposes. Encapsulation efficiencies were higher than 99%, which was also attributed to the nano-emulsification approach as well as to the low solubility of the dyes in the aqueous component. Release kinetics of both fluorescent dyes from the nanoparticle dispersions was pH-independent and sustained. These results indicate that the dyes could remain encapsulated enough time to reach any organ and that the decrease of the pH produced during cell internalization by the endocytic route would not affect their release. Therefore, it can be assumed that these nanoparticles are appropriate as systemic imaging agents. In addition, in vitro toxicity tests showed that nanoparticles are non-cytotoxic. Consequently, it can be concluded that the preparation of PLGA nanoparticles from nano-emulsion templating represents a very versatile technology that enables obtaining biocompatible, biodegradable and safe imaging agents suitable for biomedical purposes. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Folate-targeted gadolinium-lipid-based nanoparticles as a bimodal contrast agent for tumor fluorescent and magnetic resonance imaging.

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    Nakamura, Taro; Kawano, Kumi; Shiraishi, Kouichi; Yokoyama, Masayuki; Maitani, Yoshie

    2014-01-01

    To enhance tumor magnetic resonance imaging (MRI) signals via the selective accumulation of contrast agents, we prepared folate-modified gadolinium-lipid-based nanoparticles as MRI contrast agents. Folate-modified nanoparticles were comprised of polyethylene glycol (PEG)-lipid, gadolinium diethylenetriamine pentaacetic acid lipid, cationic cholesterol derivatives, folate-conjugated PEG-lipid, and Cy7-PEG-lipid. Folate receptor-mediated cellular nanoparticle association was examined in KB cells, which overexpress the folate receptor. The biodistribution of nanoparticles after their intravenous injection into KB tumor-bearing mice was measured. Mice were imaged through in vivo fluorescence imaging and MRI 24 h after nanoparticle injection, and the intensity enhancement of the tumor MRI signal was evaluated. Increased cellular association of folate-modified nanoparticles was inhibited by excess free folic acid, indicating that nanoparticle association was folate receptor-mediated. Irrespective of folate modification, the amount of nanoparticles in blood 24 h after injection was ca. 10% of the injected dose. Compared with non-modified nanoparticles, folate-modified nanoparticles exhibited significant accumulation in tumor tissues without altering other biodistribution, as well as enhanced tumor fluorescence and MRI signal intensity. The results support the feasibility of MRI- and in vivo fluorescence imaging-based tumor visualization using folate-modified nanoparticles and provide opportunities to develop folate targeting-based imaging applications.

  12. Enhanced two-photon excited fluorescence from imaging agents using true thermal light

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    Jechow, Andreas; Seefeldt, Michael; Kurzke, Henning; Heuer, Axel; Menzel, Ralf

    2013-12-01

    Two-photon excited fluorescence (TPEF) is a standard technique in modern microscopy, but is still affected by photodamage to the probe. It has been proposed that TPEF can be enhanced using entangled photons, but this has proven challenging. Recently, it was shown that some features of entangled photons can be mimicked with thermal light, which finds application in ghost imaging, subwavelength lithography and metrology. Here, we use true thermal light from a superluminescent diode to demonstrate TPEF that is enhanced compared to coherent light, using two common fluorophores and luminescent quantum dots, which suit applications in imaging and microscopy. We find that the TPEF rate is directly proportional to the measured degree of second-order coherence, as predicted by theory. Our results show that photon bunching in thermal light can be exploited in two-photon microscopy, with the photon statistic providing a new degree of freedom.

  13. Efficient labeling in vitro with non-ionic gadolinium magnetic resonance imaging contrast agent and fluorescent transfection agent in bone marrow stromal cells of neonatal rats.

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    Li, Ying-Qin; Tang, Ying; Fu, Rao; Meng, Qiu-Hua; Zhou, Xue; Ling, Ze-Min; Cheng, Xiao; Tian, Su-Wei; Wang, Guo-Jie; Liu, Xue-Guo; Zhou, Li-Hua

    2015-07-01

    Although studies have been undertaken on gadolinium labeling-based molecular imaging in magnetic resonance imaging (MRI), the use of non-ionic gadolinium in the tracking of stem cells remains uncommon. To investigate the efficiency in tracking of stem cells with non-ionic gadolinium as an MRI contrast agent, a rhodamine-conjugated fluorescent reagent was used to label bone marrow stromal cells (BMSCs) of neonatal rats in vitro, and MRI scanning was undertaken. The fluorescent-conjugated cell uptake reagents were able to deliver gadodiamide into BMSCs, and cell uptake was verified using flow cytometry. In addition, the labeled stem cells with paramagnetic contrast medium remained detectable by an MRI monitor for a minimum of 28 days. The present study suggested that this method can be applied efficiently and safely for the labeling and tracking of bone marrow stromal cells in neonatal rats.

  14. A new hydrothermal refluxing route to strong fluorescent carbon dots and its application as fluorescent imaging agent.

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    Zhang, Ye-Yun; Wu, Ming; Wang, Yan-Qin; He, Xi-Wen; Li, Wen-You; Feng, Xi-Zeng

    2013-12-15

    Due to their unique optical and biochemical properties, the water-soluble fluorescent carbon dots (CDs) have attracted a lot of attention recently. Here, strong fluorescent carbon dots with excellent quality have been synthesized by the hydrothermal refluxing method using lactose as carbon source and tris(hydroxymethyl) aminomethane (i.e. Tris) as surface passivation reagent. This facile approach was simple, efficient, economical, green without pollution, and allows large-scale production of CDs without any post-treatment. TEM measurements showed that the resulting particles exhibited an average diameter of 1.5 nm. The obtained CDs possess small particle sizes, good stability in a wide range of pH values (pH 3.5-9.5), high tolerance of salt concentration, strong resistibility to photobleaching, and a fluorescent quantum yield up to 12.5%. The CDs were applied to optical bioimaging of HeLa cells, showing low cytotoxicity and excellent biocompatibility.

  15. Terbium-doped gadolinium oxide nanoparticles prepared by laser ablation in liquid for use as a fluorescence and magnetic resonance imaging dual-modal contrast agent.

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    Chen, Fei; Chen, Min; Yang, Chuan; Liu, Jun; Luo, Ningqi; Yang, Guowei; Chen, Dihu; Li, Li

    2015-01-14

    Dual-modal lanthanide-doped gadolinium nanoparticles (NPs), which exhibit an excellent magnetic resonance imaging (MRI) spatial resolution and high fluorescence imaging (FI) sensitivity, have attracted tremendous attention in biotechnology and nanomedicine applications. In this paper, terbium (Tb) ion doped gadolinium oxide (Gd2O3:Tb) NPs with varied Tb concentrations were synthesized by a laser ablation in liquid (LAL) method. The characterization of the structure, morphology, and composition shows that these NPs are spherical with excellent crystallinity. The effects of Tb ion concentration on the visible green fluorescence and longitudinal relaxivity were investigated, indicating that the fluorescence properties were significantly influenced by the Tb ion concentration, but all samples were still efficient T1-weighted contrast agents. Furthermore, the optimum Tb doping concentration was determined to be 1%. The cell viability, cellular fluorescence imaging and in vivo MRI of this dual-modal nano-probe were studied, with the results revealing that the Gd2O3:Tb NPs did not have a significant cytotoxic effect, making them good candidates for use as a dual-modal contrast agent for MRI and fluorescence imaging.

  16. Self-assembled dual-modality contrast agents for non-invasive stem cell tracking via near-infrared fluorescence and magnetic resonance imaging.

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    Liu, Hong; Tan, Yan; Xie, Lisi; Yang, Lei; Zhao, Jing; Bai, Jingxuan; Huang, Ping; Zhan, Wugen; Wan, Qian; Zou, Chao; Han, Yali; Wang, Zhiyong

    2016-09-15

    Stem cells hold great promise for treating various diseases. However, one of the main drawbacks of stem cell therapy is the lack of non-invasive image-tracking technologies. Although magnetic resonance imaging (MRI) and near-infrared fluorescence (NIRF) imaging have been employed to analyse cellular and subcellular events via the assistance of contrast agents, the sensitivity and temporal resolution of MRI and the spatial resolution of NIRF are still shortcomings. In this study, superparamagnetic iron oxide nanocrystals and IR-780 dyes were co-encapsulated in stearic acid-modified polyethylenimine to form a dual-modality contrast agent with nano-size and positive charge. These resulting agents efficiently labelled stem cells and did not influence the cellular viability and differentiation. Moreover, the labelled cells showed the advantages of dual-modality imaging in vivo.

  17. Fluorescence and Spectral Imaging

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    Ralph S. DaCosta

    2007-01-01

    Full Text Available Early identification of dysplasia remains a critical goal for diagnostic endoscopy since early discovery directly improves patient survival because it allows endoscopic or surgical intervention with disease localized without lymph node involvement. Clinical studies have successfully used tissue autofluorescence with conventional white light endoscopy and biopsy for detecting adenomatous colonic polyps, differentiating benign hyperplastic from adenomas with acceptable sensitivity and specificity. In Barrett's esophagus, the detection of dysplasia remains problematic because of background inflammation, whereas in the squamous esophagus, autofluorescence imaging appears to be more dependable. Point fluorescence spectroscopy, although playing a crucial role in the pioneering mechanistic development of fluorescence endoscopic imaging, does not seem to have a current function in endoscopy because of its nontargeted sampling and suboptimal sensitivity and specificity. Other point spectroscopic modalities, such as Raman spectroscopy and elastic light scattering, continue to be evaluated in clinical studies, but still suffer the significant disadvantages of being random and nonimaging. A recent addition to the fluorescence endoscopic imaging arsenal is the use of confocal fluorescence endomicroscopy, which provides real-time optical biopsy for the first time. To improve detection of dysplasia in the gastrointestinal tract, a new and exciting development has been the use of exogenous fluorescence contrast probes that specifically target a variety of disease-related cellular biomarkers using conventional fluorescent dyes and novel potent fluorescent nanocrystals (i.e., quantum dots. This is an area of great promise, but still in its infancy, and preclinical studies are currently under way.

  18. Boronic acids for fluorescence imaging of carbohydrates.

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    Sun, Xiaolong; Zhai, Wenlei; Fossey, John S; James, Tony D

    2016-02-28

    "Fluorescence imaging" is a particularly exciting and rapidly developing area of research; the annual number of publications in the area has increased ten-fold over the last decade. The rapid increase of interest in fluorescence imaging will necessitate the development of an increasing number of molecular receptors and binding agents in order to meet the demand in this rapidly expanding area. Carbohydrate biomarkers are particularly important targets for fluorescence imaging given their pivotal role in numerous important biological events, including the development and progression of many diseases. Therefore, the development of new fluorescent receptors and binding agents for carbohydrates is and will be increasing in demand. This review highlights the development of fluorescence imaging agents based on boronic acids a particularly promising class of receptors given their strong and selective binding with carbohydrates in aqueous media.

  19. Stroboscopic fluorescence lifetime imaging.

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    Holton, Mark D; Silvestre, Oscar R; Errington, Rachel J; Smith, Paul J; Matthews, Daniel R; Rees, Paul; Summers, Huw D

    2009-03-30

    We report a fluorescence lifetime imaging technique that uses the time integrated response to a periodic optical excitation, eliminating the need for time resolution in detection. A Dirac pulse train of variable period is used to probe the frequency response of the total fluorescence per pulse leading to a frequency roll-off that is dependent on the relaxation rate of the fluorophores. The technique is validated by demonstrating wide-field, realtime, lifetime imaging of the endocytosis of inorganic quantum dots by a cancer cell line. Surface charging of the dots in the intra-cellular environment produces a switch in the fluorescence lifetime from approximately 40 ns to technique offers lifetime based imaging at video rates with standard CCD cameras and has application in probing millisecond cell dynamics and in high throughput imaging assays.

  20. Cyanine 5.5 conjugated nanobubbles as a tumor selective contrast agent for dual ultrasound-fluorescence imaging in a mouse model.

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    Liyi Mai

    Full Text Available Nanobubbles and microbubbles are non-invasive ultrasound imaging contrast agents that may potentially enhance diagnosis of tumors. However, to date, both nanobubbles and microbubbles display poor in vivo tumor-selectivity over non-targeted organs such as liver. We report here cyanine 5.5 conjugated nanobubbles (cy5.5-nanobubbles of a biocompatible chitosan-vitamin C lipid system as a dual ultrasound-fluorescence contrast agent that achieved tumor-selective imaging in a mouse tumor model. Cy5.5-nanobubble suspension contained single bubble spheres and clusters of bubble spheres with the size ranging between 400-800 nm. In the in vivo mouse study, enhancement of ultrasound signals at tumor site was found to persist over 2 h while tumor-selective fluorescence emission was persistently observed over 24 h with intravenous injection of cy5.5-nanobubbles. In vitro cell study indicated that cy5.5-flurescence dye was able to accumulate in cancer cells due to the unique conjugated nanobubble structure. Further in vivo fluorescence study suggested that cy5.5-nanobubbles were mainly located at tumor site and in the bladder of mice. Subsequent analysis confirmed that accumulation of high fluorescence was present at the intact subcutaneous tumor site and in isolated tumor tissue but not in liver tissue post intravenous injection of cy5.5-nanobubbles. All these results led to the conclusion that cy5.5-nanobubbles with unique crosslinked chitosan-vitamin C lipid system have achieved tumor-selective imaging in vivo.

  1. High-affinity Near-infrared Fluorescent Small-molecule Contrast Agents for In Vivo Imaging of Prostate-specific Membrane Antigen

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    Valerie Humblet

    2005-10-01

    Full Text Available Surgical resection remains a definitive treatment for prostate cancer. Yet, prostate cancer surgery is performed without image guidance for tumor margin, extension beyond the capsule and lymph node positivity, and without verification of other occult metastases in the surgical field. Recently, several imaging systems have been described that exploit near-infrared (NIR fluorescent light for sensitive, real-time detection of disease pathology intraoperatively. In this study, we describe a high-affinity (9 nM, single nucleophile-containing, small molecule specific for the active site of the enzyme PSMA. We demonstrate production of a tetra-sulfonated heptamethine indocyanine NIR fluorescent derivative of this molecule using a high-yield LC/MS purification strategy. Interestingly, NIR fluorophore conjugation improves affinity over 20-fold, and we provide mechanistic insight into this observation. We describe the preparative production of enzymatically active PSMA using a baculovirus expression system and an adenovirus that co-expresses PSMA and GFP. We demonstrate sensitive and specific in vitro imaging of endogenous and ectopically expressed PSMA in human cells and in vivo imaging of xenograft tumors. We also discuss chemical strategies for improving performance even further. Taken together, this study describes nearly complete preclinical development of an optically based small-molecule contrast agent for image-guided surgery.

  2. Developments Toward Diagnostic Breast Cancer Imaging Using Near-Infrared Optical Measurements and Fluorescent Contrast Agents1

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    Daniel J. Hawrysz

    2000-09-01

    Full Text Available The use of near-infrared (NIR light to interrogate deep tissues has enormous potential for molecular-based imaging when coupled with NIR excitable dyes. More than a decade has now passed since the initial proposals for NIR optical tomography for breast cancer screening using time-dependent measurements of light propagation in the breast. Much accomplishment in the development of optical mammography has been demonstrated, most recently in the application of time-domain, frequency-domain, and continuous-wave measurements that depend on endogenous contrast owing to angiogenesis and increased hemoglobin absorbance for contrast. Although exciting and promising, the necessity of angiogenesis-mediated absorption contrast for diagnostic optical mammography minimizes the potential for using NIR techniques to assess sentinel lymph node staging, metastatic spread, and multifocality of breast disease, among other applications. In this review, we summarize the progress made in the development of optical mammography, and focus on the emerging work underway in the use of diagnostic contrast agents for the molecular-based, diagnostic imaging of breast.

  3. Dendrimer-encapsulated naphthalocyanine as a single agent-based theranostic nanoplatform for near-infrared fluorescence imaging and combinatorial anticancer phototherapy

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    Taratula, Olena; Schumann, Canan; Duong, Tony; Taylor, Karmin L.; Taratula, Oleh

    2015-02-01

    Multifunctional theranostic platforms capable of concurrent near-infrared (NIR) fluorescence imaging and phototherapies are strongly desired for cancer diagnosis and treatment. However, the integration of separate imaging and therapeutic components into nanocarriers results in complex theranostic systems with limited translational potential. A single agent-based theranostic nanoplatform, therefore, was developed for concurrent NIR fluorescence imaging and combinatorial phototherapy with dual photodynamic (PDT) and photothermal (PTT) therapeutic mechanisms. The transformation of a substituted silicon naphthalocyanine (SiNc) into a biocompatible nanoplatform (SiNc-NP) was achieved by SiNc encapsulation into the hydrophobic interior of a generation 5 polypropylenimine dendrimer following surface modification with polyethylene glycol. Encapsulation provides aqueous solubility to SiNc and preserves its NIR fluorescence, PDT and PTT properties. Moreover, an impressive photostability in the dendrimer-encapsulated SiNc has been detected. Under NIR irradiation (785 nm, 1.3 W cm-2), SiNc-NP manifested robust heat generation capability (ΔT = 40 °C) and efficiently produced reactive oxygen species essential for PTT and PDT, respectively, without releasing SiNc from the nanopaltform. By varying the laser power density from 0.3 W cm-2 to 1.3 W cm-2 the therapeutic mechanism of SiNc-NP could be switched from PDT to combinatorial PDT-PTT treatment. In vitro and in vivo studies confirmed that phototherapy mediated by SiNc can efficiently destroy chemotherapy resistant ovarian cancer cells. Remarkably, solid tumors treated with a single dose of SiNc-NP combined with NIR irradiation were completely eradicated without cancer recurrence. Finally, the efficiency of SiNc-NP as an NIR imaging agent was confirmed by recording the strong fluorescence signal in the tumor, which was not photobleached during the phototherapeutic procedure.Multifunctional theranostic platforms capable of

  4. Development, preclinical safety, formulation, and stability of clinical grade bevacizumab-800CW, a new near infrared fluorescent imaging agent for first in human use.

    Science.gov (United States)

    Ter Weele, Eva J; Terwisscha van Scheltinga, Anton G T; Linssen, Matthijs D; Nagengast, Wouter B; Lindner, Ingo; Jorritsma-Smit, Annelies; de Vries, Elisabeth G E; Kosterink, Jos G W; Lub-de Hooge, Marjolijn N

    2016-07-01

    There is a dire need for better visualization of cancer and analysis of specific targets in vivo. Molecular imaging with fluorescence is gaining more and more attention, as it allows detection of these targets and has advantages over radioactivity, such as no radiation dose, and lower costs. A key challenge in optical imaging however, is translation of the newly developed tracers from pre-clinical phase to clinical application. We describe the development and safety testing of clinical grade bevacizumab-800CW, an antibody-based targeted agent for non-invasive imaging of vascular endothelial growth factor A (VEGF-A). Development included implementing the manufacturing process and analytical methods according to current Good Manufacturing Practice (cGMP), formulation studies, extended characterization and stability testing. For safety pharmacology an extended single dose toxicity study in mice was performed. Bevacizumab-800CW was formulated in isotonic phosphate buffered sodium chloride solution at pH 7. The production was robust and showed a reproducible labeling efficiency, and no impurities. The binding affinity to VEGF-A remained intact. The optimized product meets all release specifications, is stable up to at least 3months and its characteristics did not significantly differ from the unlabeled bevacizumab. Toxicity testing in mice showed no remarkable findings. In conclusion, sterile bevacizumab-800CW (6mg=6ml) can be produced in stock according to current Good Manufacturing Practice. It is ready for first-in-human use.

  5. X-ray fluorescence microscopy demonstrates preferential accumulation of a vanadium-based magnetic resonance imaging contrast agent in murine colonic tumors.

    Science.gov (United States)

    Mustafi, Devkumar; Ward, Jesse; Dougherty, Urszula; Bissonnette, Marc; Hart, John; Vogt, Stefan; Karczmar, Gregory S

    2015-01-01

    Contrast agents that specifically enhance cancers on magnetic resonance imaging (MRI) will allow earlier detection. Vanadium-based chelates (VCs) selectively enhance rodent cancers on MRI, suggesting selective uptake of VCs by cancers. Here we report x-ray fluorescence microscopy (XFM) of VC uptake by murine colon cancer. Colonic tumors in mice treated with azoxymethane/dextran sulfate sodium were identified by MRI. Then a gadolinium-based contrast agent and a VC were injected intravenously; mice were sacrificed and colons sectioned. VC distribution was sampled at 120 minutes after injection to evaluate the long-term accumulation. Gadolinium distribution was sampled at 10 minutes after injection due to its rapid washout. XFM was performed on 72 regions of normal and cancerous colon from five normal mice and four cancer-bearing mice. XFM showed that all gadolinium was extracellular, with similar concentrations in colon cancers and normal colon. In contrast, the average VC concentration was twofold higher in cancers versus normal tissue (p < .002). Cancers also contained numerous "hot spots" with intracellular VC concentrations sixfold higher than the concentration in normal colon (p < .0001). No hot spots were detected in normal colon. This is the first direct demonstration that VCs selectively accumulate in cancer cells and thus may improve cancer detection.

  6. Assessing Photosynthesis by Fluorescence Imaging

    Science.gov (United States)

    Saura, Pedro; Quiles, Maria Jose

    2011-01-01

    This practical paper describes a novel fluorescence imaging experiment to study the three processes of photochemistry, fluorescence and thermal energy dissipation, which compete during the dissipation of excitation energy in photosynthesis. The technique represents a non-invasive tool for revealing and understanding the spatial heterogeneity in…

  7. Development, preclinical safety, formulation, and stability of clinical grade bevacizumab-800CW, a new near infrared fluorescent imaging agent for first in human use

    NARCIS (Netherlands)

    ter Weele, Eva J.; Terwisscha Van Scheltinga, Anton; Linssen, Matthijs D.; Nagengast, Wouter B.; Lindner, Ingo; Jorritsma-Smit, Annelies; Vries, de Elisabeth G. E.; Kosterink, Jos G. W.; Lub-de Hooge, Marjolijn N.

    There is a dire need for better visualization of cancer and analysis of specific targets in vivo. Molecular imaging with fluorescence is gaining more and more attention, as it allows detection of these targets and has advantages over radioactivity, such as no radiation dose, and lower costs. A key

  8. Deep two-photon microscopic imaging through brain tissue using the second singlet state from fluorescent agent chlorophyll α in spinach leaf.

    Science.gov (United States)

    Shi, Lingyan; Rodríguez-Contreras, Adrián; Budansky, Yury; Pu, Yang; Nguyen, Thien An; Alfano, Robert R

    2014-06-01

    Two-photon (2P) excitation of the second singlet (S₂) state was studied to achieve deep optical microscopic imaging in brain tissue when both the excitation (800 nm) and emission (685 nm) wavelengths lie in the "tissue optical window" (650 to 950 nm). S₂ state technique was used to investigate chlorophyll α (Chl α) fluorescence inside a spinach leaf under a thick layer of freshly sliced rat brain tissue in combination with 2P microscopic imaging. Strong emission at the peak wavelength of 685 nm under the 2P S₂ state of Chl α enabled the imaging depth up to 450 μm through rat brain tissue.

  9. New nontoxic double information magnetic and fluorescent MRI agent

    Energy Technology Data Exchange (ETDEWEB)

    Kublickas, Augustinas; Rastenien, Loreta; Bloznelytė-Plėšnienė, Laima; Karalius, Nerijus [Liquid Crystals Laboratory, Institute of Science and Technology, Lithuanian University of Educational Sciences (Lithuania); Franckevinius, Marius [Institute of Physics, Center for Physical Sciences and Technology (Lithuania); Loudos, George [Technological Educational Institute of Athens (Greece); Fahmi, Amir [Materials Science, Rhein-Waal University of Applied Sciences (Germany); Vaisnoras, Rimas [Liquid Crystals Laboratory, Institute of Science and Technology, Lithuanian University of Educational Sciences (Lithuania)

    2015-05-18

    Today sensitivity of the MRI is not enough compared to the nuclear methods, such as positron emission tomography and single photon emission computed tomography. Challenging its extension to the nanometre scale could provide a powerful new tool for the nanosciences and nanomedicine. To achieve this potential, innovative new detection strategies are required to overcome the severe sensitivity limitations of conventional inductive detection techniques. In this regard, we perform embodiment of nanodiamonds in dendrimer matrix as additional fluorescent optical and magnetic (together with Gd (III)) imaging modalities of the MRI. New hybrid system composed of dendrimer-gadolinium Gd (III) - nanodiamond as a new contrast agent for MRI was studied. Poly(propilene-imine) PPI and poly(amidoamine) PAMAM dendrimers with fixed size of nanocavities will be used as host material to protect organism against the toxicity and also to increase relaxivity of contrast agent (resulting in the increases MRI resolution). Nanodiamond as biocompatible platform to functionalize the contrast agent will be used. This bimodal hybrid system enables to use smaller amount of the contrast agent and could permit the decrease of the lateral toxicity. This bimodal hybrid system as MRI agent is providing double information (magnetic and fluorescent) about the damaged cell.

  10. Detection of rheumatoid arthritis in humans by fluorescence imaging

    Science.gov (United States)

    Ebert, Bernd; Dziekan, Thomas; Weissbach, Carmen; Mahler, Marianne; Schirner, Michael; Berliner, Birgitt; Bauer, Daniel; Voigt, Jan; Berliner, Michael; Bahner, Malte L.; Macdonald, Rainer

    2010-02-01

    The blood pool agent indo-cyanine green (ICG) has been investigated in a prospective clinical study for detection of rheumatoid arthritis using fluorescence imaging. Temporal behavior as well as spatial distribution of fluorescence intensity are suited to differentiate healthy and inflamed finger joints after i.v. injection of an ICG bolus.

  11. Multi Spectral Fluorescence Imager (MSFI)

    Science.gov (United States)

    Caron, Allison

    2016-01-01

    Genetic transformation with in vivo reporter genes for fluorescent proteins can be performed on a variety of organisms to address fundamental biological questions. Model organisms that may utilize an ISS imager include unicellular organisms (Saccharomyces cerevisiae), plants (Arabidopsis thaliana), and invertebrates (Caenorhabditis elegans). The multispectral fluorescence imager (MSFI) will have the capability to accommodate 10 cm x 10 cm Petri plates, various sized multi-well culture plates, and other custom culture containers. Features will include programmable temperature and light cycles, ethylene scrubbing (less than 25 ppb), CO2 control (between 400 ppm and ISS-ambient levels in units of 100 ppm) and sufficient airflow to prevent condensation that would interfere with imaging.

  12. Hyperspectral fluorescence imaging with multi wavelength LED excitation

    Science.gov (United States)

    Luthman, A. Siri; Dumitru, Sebastian; Quirós-Gonzalez, Isabel; Bohndiek, Sarah E.

    2016-04-01

    Hyperspectral imaging (HSI) can combine morphological and molecular information, yielding potential for real-time and high throughput multiplexed fluorescent contrast agent imaging. Multiplexed readout from targets, such as cell surface receptors overexpressed in cancer cells, could improve both sensitivity and specificity of tumor identification. There remains, however, a need for compact and cost effective implementations of the technology. We have implemented a low-cost wide-field multiplexed fluorescence imaging system, which combines LED excitation at 590, 655 and 740 nm with a compact commercial solid state HSI system operating in the range 600 - 1000 nm. A key challenge for using reflectance-based HSI is the separation of contrast agent fluorescence from the reflectance of the excitation light. Here, we illustrate how it is possible to address this challenge in software, using two offline reflectance removal methods, prior to least-squares spectral unmixing. We made a quantitative comparison of the methods using data acquired from dilutions of contrast agents prepared in well-plates. We then established the capability of our HSI system for non-invasive in vivo fluorescence imaging in small animals using the optimal reflectance removal method. The HSI presented here enables quantitative unmixing of at least four fluorescent contrast agents (Alexa Fluor 610, 647, 700 and 750) simultaneously in living mice. A successful unmixing of the four fluorescent contrast agents was possible both using the pure contrast agents and with mixtures. The system could in principle also be applied to imaging of ex vivo tissue or intraoperative imaging in a clinical setting. These data suggest a promising approach for developing clinical applications of HSI based on multiplexed fluorescence contrast agent imaging.

  13. Assessment of tumor angiogenesis using fluorescence contrast agents

    Science.gov (United States)

    Chen, Yu; Liu, Qian; Huang, Ping; Hyman, Shay; Intes, Xavier; Lee, William; Chance, Britton

    2003-12-01

    Angiogenesis is an important factor for further tumor growth and thus could be an attractive therapeutic target. Optical imaging can provide a non-invasive way to measure the permeability of tumor blood vessels and assess the tumor vasculature. We have developed a dual-channel near-infrared fluorescence system for simultaneous measurement of the pharmacokinetics of tumorous and normal tissues with exogenous contrast agents. This frequency-domain system consists of the light source (780 nm laser diode), fiber optics, interference filter (830 nm) and the detector (PMT). The fluorescent contrast agent used in this study is Indocyanine Green (ICG), and the normal dosage is 100 μl at a concentration of 5 μM. In vivo animal study is performed on the K1735 melanoma-bearing mouse. The fluorescence signals both tumorous and normal tissues after the bolus injection of ICG through the tail vein are continuously recorded as a function of time. The data is fitted by a double-exponential model to reveal the wash-in and wash-out parameters of different tissues. We observed an elongated wash-out from the tumor compared with normal tissue (leg). The effect of radiation therapy on the tumor vasculature is also discussed.

  14. Subharmonic imaging of contrast agents.

    Science.gov (United States)

    Forsberg, F; Shi, W T; Goldberg, B B

    2000-03-01

    Ultrasound contrast agents promise to improve the sensitivity and specificity of diagnostic ultrasound imaging. It is of great importance to adapt ultrasound equipment for optimal use with contrast agents e.g., by exploiting the nonlinear properties of the contrast microbubbles. Harmonic imaging is one technique that has been extensively studied and is commercially available. However, harmonic imaging is associated with problems, due to second harmonic generation and accumulation within the tissue itself. Given the lack of subharmonic generation in tissue, one alternative is the creation of subharmonic images by transmitting at the fundamental frequency (fo) and receiving at the subharmonic (fo/2). Subharmonic imaging should have a much better lateral resolution and may be suitable for scanning deep-lying structures owing to the higher transmit frequency and the much smaller attenuation of scattered subharmonic signals. In this paper, we will review different aspects of subharmonic imaging including implementation, in-vitro gray-scale imaging and subharmonic aided pressure estimation.

  15. Fluorescence imaging spectrometer optical design

    Science.gov (United States)

    Taiti, A.; Coppo, P.; Battistelli, E.

    2015-09-01

    The optical design of the FLuORescence Imaging Spectrometer (FLORIS) studied for the Fluorescence Explorer (FLEX) mission is discussed. FLEX is a candidate for the ESA's 8th Earth Explorer opportunity mission. FLORIS is a pushbroom hyperspectral imager foreseen to be embarked on board of a medium size satellite, flying in tandem with Sentinel-3 in a Sun synchronous orbit at a height of about 815 km. FLORIS will observe the vegetation fluorescence and reflectance within a spectral range between 500 and 780 nm. Multi-frames acquisitions on matrix detectors during the satellite movement will allow the production of 2D Earth scene images in two different spectral channels, called HR and LR with spectral resolution of 0.3 and 2 nm respectively. A common fore optics is foreseen to enhance by design the spatial co-registration between the two spectral channels, which have the same ground spatial sampling (300 m) and swath (150 km). An overlapped spectral range between the two channels is also introduced to simplify the spectral coregistration. A compact opto-mechanical solution with all spherical and plane optical elements is proposed, and the most significant design rationales are described. The instrument optical architecture foresees a dual Babinet scrambler, a dioptric telescope and two grating spectrometers (HR and LR), each consisting of a modified Offner configuration. The developed design is robust, stable vs temperature, easy to align, showing very high optical quality along the whole field of view. The system gives also excellent correction for transverse chromatic aberration and distortions (keystone and smile).

  16. Quantitative imaging with fluorescent biosensors.

    Science.gov (United States)

    Okumoto, Sakiko; Jones, Alexander; Frommer, Wolf B

    2012-01-01

    Molecular activities are highly dynamic and can occur locally in subcellular domains or compartments. Neighboring cells in the same tissue can exist in different states. Therefore, quantitative information on the cellular and subcellular dynamics of ions, signaling molecules, and metabolites is critical for functional understanding of organisms. Mass spectrometry is generally used for monitoring ions and metabolites; however, its temporal and spatial resolution are limited. Fluorescent proteins have revolutionized many areas of biology-e.g., fluorescent proteins can report on gene expression or protein localization in real time-yet promoter-based reporters are often slow to report physiologically relevant changes such as calcium oscillations. Therefore, novel tools are required that can be deployed in specific cells and targeted to subcellular compartments in order to quantify target molecule dynamics directly. We require tools that can measure enzyme activities, protein dynamics, and biophysical processes (e.g., membrane potential or molecular tension) with subcellular resolution. Today, we have an extensive suite of tools at our disposal to address these challenges, including translocation sensors, fluorescence-intensity sensors, and Förster resonance energy transfer sensors. This review summarizes sensor design principles, provides a database of sensors for more than 70 different analytes/processes, and gives examples of applications in quantitative live cell imaging.

  17. A novel Tc-99 m and fluorescence labeled peptide as a multimodal imaging agent for targeting angiogenesis in a murine tumor model.

    Science.gov (United States)

    Kim, Myoung Hyoun; Kim, Chang Guhn; Kim, Seul-Gi; Kim, Dae-Weung

    2016-11-01

    The serine-aspartic acid-valine (SDV) peptide binds specifically to integrin αV β3 . In the present study, we successfully developed a TAMRA-GHEG-ECG-SDV peptide labeled with both Tc-99 m and TAMRA to target the integrin αV β3 of tumor cells; furthermore, we evaluated the diagnostic performance of Tc-99 m TAMRA-GHEG-ECG-SDV as a dual-modality imaging agent for tumor of the murine model. TAMRA-GHEG-ECG-SDV was synthesized using Fmoc solid-phase peptide synthesis. Radiolabeling of TAMRA-GHEG-ECG-SDV with Tc-99 m was done using ligand exchange methods. Labeling stability and cytotoxicity studies were performed. Gamma camera imaging, biodistribution and ex vivo imaging studies were performed in murine models with HT-1080 and HT-29 tumors. A tumor tissue slide was prepared and analyzed using confocal microscopy. After radiolabeling procedures with Tc-99 m, the Tc-99 m TAMRA-GHEG-ECG-SDV complexes were prepared in high yield (>99%). In the gamma camera imaging study, a substantial uptake of Tc-99 m TAMRA-GHEG-ECG-SDV into HT-1080 tumor (integrin αV β3 positive) and low uptake of Tc-99 m TAMRA-GHEG-ECG-SDV into HT-29 tumor (integrin αV β3 negative) were demonstrated. A competition study revealed that HT-1080 tumor uptake was effectively blocked by the co-injection of an excess concentration of SDV. Specific uptake of Tc-99 m TAMRA-GHEG-ECG-SDV was confirmed by biodistribution, ex vivo imaging and confocal microscopy studies. Our in vivo and in vitro studies revealed substantial uptake of Tc-99 m TAMRA-GHEG-ECG-SDV in the integrin αV β3 -positive tumor. Tc-99 m TAMRA-GHEG-ECG-SDV could be a good candidate for a dual-modality imaging agent targeting tumor angiogenesis. Copyright © 2016 John Wiley & Sons, Ltd.

  18. Fluorescence confocal endomicroscopy in biological imaging

    Science.gov (United States)

    Delaney, Peter; Thomas, Steven; Allen, John; McLaren, Wendy; Murr, Elise; Harris, Martin

    2007-02-01

    In vivo fluorescence microscopic imaging of biological systems in human disease states and animal models is possible with high optical resolution and mega pixel point-scanning performance using optimised off-the-shelf turn-key devices. There are however various trade-offs between tissue access and instrument performance when miniaturising in vivo microscopy systems. A miniature confocal scanning technology that was developed for clinical human endoscopy has been configured into a portable device for direct hand-held interrogation of living tissue in whole animal models (Optiscan FIVE-1 system). Scanning probes of 6.3mm diameter with a distal tip diameter of 5.0mm were constructed either in a 150mm length for accessible tissue, or a 300mm probe for laparoscopic interrogation of internal tissues in larger animal models. Both devices collect fluorescence confocal images (excitation 488 nm; emission >505 or >550 nm) comprised of 1024 x 1204 sampling points/image frame, with lateral resolution 0.7um; axial resolution 7um; FOV 475 x 475um. The operator can dynamically control imaging depth from the tissue surface to approx 250um in 4um steps via an internally integrated zaxis actuator. Further miniaturisation is achieved using an imaging contact probe based on scanning the proximal end of a high-density optical fibre bundle (~30,000 fibres) of sheep and pigs was fluorescently stained with calcein-AM or fluorescein. Surface and sub-surface cellular and sub-cellular details could be readily visualised in vivo at high resolution. In rodent disease models, in vivo endomicroscopy with appropriate fluorescent agents allowed examination of thrombosis formation, tumour microvasculature and liver metastases, diagnosis and staging of ulcerative colitis, liver necrosis and glomerulonephritis. Miniaturised confocal endomicroscopy allows rapid in vivo molecular and subsurface microscopy of normal and pathologic tissue at high resolution in small and large whole animal models

  19. Site-specific tumor-targeted fluorescent contrast agents

    Science.gov (United States)

    Achilefu, Samuel I.; Bugaj, Joseph E.; Dorshow, Richard B.; Jimenez, Hermo N.; Rajagopalan, Raghavan; Wilhelm, R. Randy; Webb, Elizabeth G.; Erion, Jack L.

    2001-01-01

    Site-specific delivery of drugs and contrast agents to tumors protects normal tissues from the cytotoxic effect of drugs, and enhances the contrast between normal and diseased tissues. In optical medicine, biocompatible dyes can be used as photo therapeutics or as contrast agents. Previous studies have shown that the use of covalent or non-covalent dye conjugates of carries such as antibodies, liposomes, and polysaccharides improves the delivery of such molecules to tumors. However, large biomolecules can elicit adverse immunogenic reactions and also result in prolonged blood circulation times, delaying visualization of target tissues. A viable alternative to this strategy is to use small bioactive molecule-dye conjugates. These molecules have several advantages over large biomolecules, including ease of synthesis of a variety of high purity compounds for combinatorial screening of new targets, enhanced diffusivity to solid tumors, and the ability to affect the pharmocokinetics of the conjugates by minor structural changes. Thus, we conjugated a near IR light absorbing dye to bioactive peptides that specifically target over expressed tumor receptors in established rat tumor lines. High tumor uptake of the conjugates was obtained without loss of either the peptide receptor affinity or the dye fluorescence. These findings demonstrate the efficacy of a small peptide-dye conjugate strategy for in vivo tumor imaging. Site-specific delivery of photodynamic therapy agents may also benefit form this approach.

  20. Detection of Leukotriene Receptor CysLT1R in Inflammatory Diseases by Molecular Imaging with Near-Infrared Fluorescence-Based Contrast Agents

    Directory of Open Access Journals (Sweden)

    Corinna Busch

    2011-03-01

    Full Text Available As leukotriene D4 receptor CysLT1R upregulation is an early event in inflammatory processes, specific detection of CysLT1R via molecular imaging might be a promising diagnostic tool for inflammatory diseases. We coupled a specific anti-CysLT1R IgG antibody to near-infrared (NIR hemicyanine fluorophore DY-734. The fluorophore was also coupled to unspecific rabbit-IgG antibody or corresponding Fab fragments. Expression of CysLT1R in HL-60 human promyelocytic leukemia cells in vitro could be proven by reverse transcriptase—polymerase chain reaction (PCR, real-time PCR, and flow cytometry. Detection of the probes by flow cytometry showed that CysLT1R*DY-734 probe binds distinctly stronger to HL-60 cells than IgG*DY-734. Induction of ear edema in mice was conducted to test signaling of the synthesized probes in vivo. A markedly higher fluorescence intensity was observed in the edematous region than in the healthy region by a whole-body imaging system. Semiquantitative analysis showed that CysLT1R*DY-734 and Fab-CysLT1R*DY-734 probes bind 1.9- and 1.2-fold stronger, respectively, than the unspecific probes. Biodistribution studies revealed an enrichment of full-length IgG probes in liver and spleen, whereas Fab-containing probes are mostly found in liver and kidneys. Taken together, we present an approach that might improve early diagnosis of inflammatory diseases in the long term.

  1. Fluorescent ampicillin analogues as multifunctional disguising agents against opsonization

    Science.gov (United States)

    Kotagiri, Nalinikanth; Sakon, Joshua; Han, Haewook; Zharov, Vladimir P.; Kim, Jin-Woo

    2016-06-01

    Cancer nanomedicines are opening new paradigms in cancer management and recent research points to how they can vastly improve imaging and therapy through multimodality and multifunctionality. However, challenges to achieving optimal efficacy are manifold starting from processing materials and evaluating their intended effectiveness on biological tissue, to developing new strategies aimed at improving transport of these materials through the biological milieu to the target tissue. Here, we report a fluorescent derivative of a beta-lactam antibiotic, ampicillin (termed iAmp) and its multifunctional physicobiochemical characteristics and potential as a biocompatible shielding agent and an effective dispersant. Carbon nanotubes (CNTs) were chosen to demonstrate the efficacy of iAmp. CNTs are known for their versatility and have been used extensively for cancer theranostics as photothermal and photoacoustic agents, but have limited solubility in water and biocompatibility. Traditional dispersants are associated with imaging artifacts and are not fully biocompatible. The chemical structure of iAmp is consistent with a deamination product of ampicillin. Although the four-membered lactam ring is intact, it does not retain the antibiotic properties. The iAmp is an effective dispersant and simultaneously serves as a fluorescent label for single-walled CNTs (SWNTs) with minimal photobleaching. The iAmp also enables bioconjugation of SWNTs to bio-ligands such as antibodies through functional carboxyl groups. Viability tests show that iAmp-coated SWNTs have minimal toxicity. Bio-stability tests under physiological conditions reveal that iAmp coating not only remains stable in a biologically relevant environment with high protein and salt concentrations, but also renders SWNTs transparent against nonspecific protein adsorption, also known as protein corona. Mammalian tissue culture studies with macrophages and opsonins validate that iAmp coating affords immunological resistance

  2. Detection of brain tumors using fluorescence diffuse optical tomography and nanoparticles as contrast agents

    Science.gov (United States)

    Fortin, Pierre-Yves; Genevois, Coralie; Koenig, Anne; Heinrich, Emilie; Texier, Isabelle; Couillaud, Franck

    2012-12-01

    Near-infrared fluorescence-enhanced diffuse optical tomography (fDOT) is used to localize tumors in mice using fluorescent nanoparticles as a blood pool contrast agent. The infrared dye DiR is loaded in the lipid core of nontargeted nanoparticles (DiR-lipidots) and injected systemically via the tail vein in mice bearing U87 tumors. Distribution and time-course of DiR-lipidots are followed using in vivo fluorescence reflectance imaging and reveal enhanced fluorescent signal within the subcutaneous tumors up to seven days due to the enhanced permeability and retention effect. Tumor growth into the brain is followed using bioluminescent imaging, and tumor localization is further determined by magnetic resonance imaging. The fDOT provides three-dimensional fluorescent maps that allow for consistent localization for both subcutaneous and brain tumors.

  3. Intelligent Design of Nano-Scale Molecular Imaging Agents

    Directory of Open Access Journals (Sweden)

    Takeaki Ozawa

    2012-12-01

    Full Text Available Visual representation and quantification of biological processes at the cellular and subcellular levels within living subjects are gaining great interest in life science to address frontier issues in pathology and physiology. As intact living subjects do not emit any optical signature, visual representation usually exploits nano-scale imaging agents as the source of image contrast. Many imaging agents have been developed for this purpose, some of which exert nonspecific, passive, and physical interaction with a target. Current research interest in molecular imaging has mainly shifted to fabrication of smartly integrated, specific, and versatile agents that emit fluorescence or luminescence as an optical readout. These agents include luminescent quantum dots (QDs, biofunctional antibodies, and multifunctional nanoparticles. Furthermore, genetically encoded nano-imaging agents embedding fluorescent proteins or luciferases are now gaining popularity. These agents are generated by integrative design of the components, such as luciferase, flexible linker, and receptor to exert a specific on–off switching in the complex context of living subjects. In the present review, we provide an overview of the basic concepts, smart design, and practical contribution of recent nano-scale imaging agents, especially with respect to genetically encoded imaging agents.

  4. Microencapsulation of self-healing agents containing a fluorescent dye

    Directory of Open Access Journals (Sweden)

    J. K. Lee

    2013-01-01

    Full Text Available Two different self-healing agent candidates, endo-dicyclopentadiene (endo-DCPD and 5-ethylidene-2-norbornene (ENB, containing a fluorescent dye surrounded by a melamine–urea–formaldehyde (MUF shell were microencapsulated by in-situ polymerization and the resulting microcapsules were characterized in this work. The microcapsules showed a narrow size distribution with a spherical shape and rough outer and smooth inner surfaces for both healing agent systems. Shell thicknesses of the microcapsules were ~880±80 nm for endo-DCPD and ~620±60 nm for ENB. The incorporation of a fluorescent dye as tracer into self-healing agents did not disturb the formation of microcapsules. The release of self-healing liquid into the induced crack from ruptured microcapsules in an epoxy coating layer was observed using a fluorescence microscopy. The use of a fluorescent dye is very effective in the observation of a damage site.

  5. S - and N-alkylating agents diminish the fluorescence of fluorescent dye-stained DNA.

    Science.gov (United States)

    Giesche, Robert; John, Harald; Kehe, Kai; Schmidt, Annette; Popp, Tanja; Balzuweit, Frank; Thiermann, Horst; Gudermann, Thomas; Steinritz, Dirk

    2017-01-25

    Sulfur mustard (SM), a chemical warfare agent, causes DNA alkylation, which is believed to be the main cause of its toxicity. SM DNA adducts are commonly used to verify exposure to this vesicant. However, the required analytical state-of-the-art mass-spectrometry methods are complex, use delicate instruments, are not mobile, and require laboratory infrastructure that is most likely not available in conflict zones. Attempts have thus been made to develop rapid detection methods that can be used in the field. The analysis of SM DNA adducts (HETE-G) by immunodetection is a convenient and suitable method. For a diagnostic assessment, HETE-G levels must be determined in relation to the total DNA in the sample. Total DNA can be easily visualized by the use of fluorescent DNA dyes. This study examines whether SM and related compounds affect total DNA staining, an issue that has not been investigated before. After pure DNA was extracted from human keratinocytes (HaCaT cells), DNA was exposed to different S- and N-alkylating agents. Our experiments revealed a significant, dose-dependent decrease in the fluorescence signal of fluorescent dye-stained DNA after exposure to alkylating agents. After mass spectrometry and additional fluorescence measurements ruled out covalent modifications of ethidium bromide (EthBr) by SM, we assumed that DNA crosslinks caused DNA condensation and thereby impaired access of the fluorescent dyes to the DNA. DNA digestion by restriction enzymes restored fluorescence, a fact that strengthened our hypothesis. However, monofunctional agents, which are unable to crosslink DNA, also decreased the fluorescence signal. In subsequent experiments, we demonstrated that protons produced during DNA alkylation caused a pH decrease that was found responsible for the reduction in fluorescence. The use of an appropriate buffer system eliminated the adverse effect of alkylating agents on DNA staining with fluorescent dyes. An appropriate buffer system is thus

  6. Fluorescence goggle for intraoperative breast cancer imaging

    Science.gov (United States)

    Liu, Yang; Bauer, Adam Q.; Akers, Walter; Sudlow, Gail; Liang, Kexian; Charanya, Tauseef; Mondal, Suman; Culver, Joseph P.; Achilefu, Samuel

    2012-03-01

    We have developed a fluorescence goggle device for intraoperative oncologic imaging. With our system design, the surgeon can directly visualize the fluorescence information from the eyepieces in real time without any additional monitor, which can improve one's coordination and surgical accuracy. In conjunction with targeting fluorescent dyes, the goggle device can successfully detect tumor margins and small nodules that are not obvious to naked eye. This can potentially decrease the incidence of incomplete resection.

  7. High speed multispectral fluorescence lifetime imaging

    NARCIS (Netherlands)

    Fereidouni, F.; Reitsma, K.; Gerritsen, H.C.

    2013-01-01

    We report a spectrally resolved fluorescence lifetime imaging system based on time gated single photon detection with a fixed gate width of 200 ps and 7 spectral channels. Time gated systems can operate at high count rates but usually have large gate widths and sample only part of the fluorescence d

  8. Moxifloxacin: Clinically compatible contrast agent for multiphoton imaging

    Science.gov (United States)

    Wang, Taejun; Jang, Won Hyuk; Lee, Seunghun; Yoon, Calvin J.; Lee, Jun Ho; Kim, Bumju; Hwang, Sekyu; Hong, Chun-Pyo; Yoon, Yeoreum; Lee, Gilgu; Le, Viet-Hoan; Bok, Seoyeon; Ahn, G.-One; Lee, Jaewook; Gho, Yong Song; Chung, Euiheon; Kim, Sungjee; Jang, Myoung Ho; Myung, Seung-Jae; Kim, Myoung Joon; So, Peter T. C.; Kim, Ki Hean

    2016-06-01

    Multiphoton microscopy (MPM) is a nonlinear fluorescence microscopic technique widely used for cellular imaging of thick tissues and live animals in biological studies. However, MPM application to human tissues is limited by weak endogenous fluorescence in tissue and cytotoxicity of exogenous probes. Herein, we describe the applications of moxifloxacin, an FDA-approved antibiotic, as a cell-labeling agent for MPM. Moxifloxacin has bright intrinsic multiphoton fluorescence, good tissue penetration and high intracellular concentration. MPM with moxifloxacin was demonstrated in various cell lines, and animal tissues of cornea, skin, small intestine and bladder. Clinical application is promising since imaging based on moxifloxacin labeling could be 10 times faster than imaging based on endogenous fluorescence.

  9. Imaging an atomic beam using fluorescence

    Institute of Scientific and Technical Information of China (English)

    Ming He(何明); Jin Wang(王谨); Mingsheng Zhan(詹明生)

    2003-01-01

    A fluorescence detection scheme is applied to image an atomic beam. Using two laser diodes as the sources of detection light and pumping light respectively, the fluorescence image of the atomic beam is then observed by a commercial CCD-camera, which is corresponding to the atomic state and velocity distribution. The detection scheme has a great utilization in the experiments of cold atoms and atomic optics.

  10. Cancer detection by quantitative fluorescence image analysis.

    Science.gov (United States)

    Parry, W L; Hemstreet, G P

    1988-02-01

    Quantitative fluorescence image analysis is a rapidly evolving biophysical cytochemical technology with the potential for multiple clinical and basic research applications. We report the application of this technique for bladder cancer detection and discuss its potential usefulness as an adjunct to methods used currently by urologists for the diagnosis and management of bladder cancer. Quantitative fluorescence image analysis is a cytological method that incorporates 2 diagnostic techniques, quantitation of nuclear deoxyribonucleic acid and morphometric analysis, in a single semiautomated system to facilitate the identification of rare events, that is individual cancer cells. When compared to routine cytopathology for detection of bladder cancer in symptomatic patients, quantitative fluorescence image analysis demonstrated greater sensitivity (76 versus 33 per cent) for the detection of low grade transitional cell carcinoma. The specificity of quantitative fluorescence image analysis in a small control group was 94 per cent and with the manual method for quantitation of absolute nuclear fluorescence intensity in the screening of high risk asymptomatic subjects the specificity was 96.7 per cent. The more familiar flow cytometry is another fluorescence technique for measurement of nuclear deoxyribonucleic acid. However, rather than identifying individual cancer cells, flow cytometry identifies cellular pattern distributions, that is the ratio of normal to abnormal cells. Numerous studies by others have shown that flow cytometry is a sensitive method to monitor patients with diagnosed urological disease. Based upon results in separate quantitative fluorescence image analysis and flow cytometry studies, it appears that these 2 fluorescence techniques may be complementary tools for urological screening, diagnosis and management, and that they also may be useful separately or in combination to elucidate the oncogenic process, determine the biological potential of tumors

  11. Cubosomes for in vivo fluorescence lifetime imaging

    Science.gov (United States)

    Biffi, Stefania; Andolfi, Laura; Caltagirone, Claudia; Garrovo, Chiara; Falchi, Angela M.; Lippolis, Vito; Lorenzon, Andrea; Macor, Paolo; Meli, Valeria; Monduzzi, Maura; Obiols-Rabasa, Marc; Petrizza, Luca; Prodi, Luca; Rosa, Antonella; Schmidt, Judith; Talmon, Yeshayahu; Murgia, Sergio

    2017-02-01

    Herein we provided the first proof of principle for in vivo fluorescence optical imaging application using monoolein-based cubosomes in a healthy mouse animal model. This formulation, administered at a non-cytotoxic concentration, was capable of providing both exogenous contrast for NIR fluorescence imaging with very high efficiency and chemospecific information upon lifetime analysis. Time-resolved measurements of fluorescence after the intravenous injection of cubosomes revealed that the dye rapidly accumulated mainly in the liver, while lifetimes profiles obtained in vivo allowed for discriminating between free dye or dye embedded within the cubosome nanostructure after injection.

  12. Cubosomes for in vivo fluorescence lifetime imaging.

    Science.gov (United States)

    Biffi, Stefania; Andolfi, Laura; Caltagirone, Claudia; Garrovo, Chiara; Falchi, Angela M; Lippolis, Vito; Lorenzon, Andrea; Macor, Paolo; Meli, Valeria; Monduzzi, Maura; Obiols-Rabasa, Marc; Petrizza, Luca; Prodi, Luca; Rosa, Antonella; Schmidt, Judith; Talmon, Yeshayahu; Murgia, Sergio

    2017-02-03

    Herein we provided the first proof of principle for in vivo fluorescence optical imaging application using monoolein-based cubosomes in a healthy mouse animal model. This formulation, administered at a non-cytotoxic concentration, was capable of providing both exogenous contrast for NIR fluorescence imaging with very high efficiency and chemospecific information upon lifetime analysis. Time-resolved measurements of fluorescence after the intravenous injection of cubosomes revealed that the dye rapidly accumulated mainly in the liver, while lifetimes profiles obtained in vivo allowed for discriminating between free dye or dye embedded within the cubosome nanostructure after injection.

  13. Multiparameter fluorescence spectroscopic imaging of cell function

    Science.gov (United States)

    Bright, Gary R.

    1994-08-01

    The ability to quantitate physiological parameters in single living cells using fluorescence spectroscopic imaging has expanded our understanding of many cell regulatory processes. Previous studies have focussed on the measurement of single parameters, such as the concentration of calcium, and more recently two parameters, such as calcium and pH using fluorescence ratio imaging. The complexity of the interrelationships among cell biochemical reactions suggests a need to extend the measurement scheme to several parameters. Expansion of the number of parameters involves several complexities associated with fluorescent probe selection and instrumentation design as well as the processing and management of the data. A system has been assembled which provides maximum flexibility in multiparameter fluorescence imaging measurements. The system provides multiple combinations of excitation, dichroic mirror, and emission wavelengths. It has automatic acquisition of any number of parameters. The number of parameters is primarily limited by the selection of fluorescent probes with nonoverlapping spectra. We demonstrate the utility of the system by the coordinated monitoring of stimulated changes in the concentrations of calcium, magnesium, and pH using fluorescence ratio imaging coupled with a conventional transmitted light image of single smooth muscle cells. The results demonstrate coordinated changes in some instances but uncoordinated changes in others.

  14. A portable fluorescence microscopic imaging system for cholecystectomy

    Science.gov (United States)

    Ye, Jian; Yang, Chaoyu; Gan, Qi; Ma, Rong; Zhang, Zeshu; Chang, Shufang; Shao, Pengfei; Zhang, Shiwu; Liu, Chenhai; Xu, Ronald

    2016-03-01

    In this paper we proposed a portable fluorescence microscopic imaging system to prevent iatrogenic biliary injuries from occurring during cholecystectomy due to misidentification of the cystic structures. The system consisted of a light source module, a CMOS camera, a Raspberry Pi computer and a 5 inch HDMI LCD. Specifically, the light source module was composed of 690 nm and 850 nm LEDs, allowing the CMOS camera to simultaneously acquire both fluorescence and background images. The system was controlled by Raspberry Pi using Python programming with the OpenCV library under Linux. We chose Indocyanine green(ICG) as a fluorescent contrast agent and then tested fluorescence intensities of the ICG aqueous solution at different concentration levels by our fluorescence microscopic system compared with the commercial Xenogen IVIS system. The spatial resolution of the proposed fluorescence microscopic imaging system was measured by a 1951 USAF resolution target and the dynamic response was evaluated quantitatively with an automatic displacement platform. Finally, we verified the technical feasibility of the proposed system in mouse models of bile duct, performing both correct and incorrect gallbladder resection. Our experiments showed that the proposed system can provide clear visualization of the confluence between the cystic duct and common bile duct or common hepatic duct, suggesting that this is a potential method for guiding cholecystectomy. The proposed portable system only cost a total of $300, potentially promoting its use in resource-limited settings.

  15. Hyperspectral Fluorescence and Reflectance Imaging Instrument

    Science.gov (United States)

    Ryan, Robert E.; O'Neal, S. Duane; Lanoue, Mark; Russell, Jeffrey

    2008-01-01

    The system is a single hyperspectral imaging instrument that has the unique capability to acquire both fluorescence and reflectance high-spatial-resolution data that is inherently spatially and spectrally registered. Potential uses of this instrument include plant stress monitoring, counterfeit document detection, biomedical imaging, forensic imaging, and general materials identification. Until now, reflectance and fluorescence spectral imaging have been performed by separate instruments. Neither a reflectance spectral image nor a fluorescence spectral image alone yields as much information about a target surface as does a combination of the two modalities. Before this system was developed, to benefit from this combination, analysts needed to perform time-consuming post-processing efforts to co-register the reflective and fluorescence information. With this instrument, the inherent spatial and spectral registration of the reflectance and fluorescence images minimizes the need for this post-processing step. The main challenge for this technology is to detect the fluorescence signal in the presence of a much stronger reflectance signal. To meet this challenge, the instrument modulates artificial light sources from ultraviolet through the visible to the near-infrared part of the spectrum; in this way, both the reflective and fluorescence signals can be measured through differencing processes to optimize fluorescence and reflectance spectra as needed. The main functional components of the instrument are a hyperspectral imager, an illumination system, and an image-plane scanner. The hyperspectral imager is a one-dimensional (line) imaging spectrometer that includes a spectrally dispersive element and a two-dimensional focal plane detector array. The spectral range of the current imaging spectrometer is between 400 to 1,000 nm, and the wavelength resolution is approximately 3 nm. The illumination system consists of narrowband blue, ultraviolet, and other discrete

  16. MMP-2/9-Specific Activatable Lifetime Imaging Agent

    Directory of Open Access Journals (Sweden)

    Marcus T.M. Rood

    2015-05-01

    Full Text Available Optical (molecular imaging can benefit from a combination of the high signal-to-background ratio of activatable fluorescence imaging with the high specificity of luminescence lifetime imaging. To allow for this combination, both imaging techniques were integrated in a single imaging agent, a so-called activatable lifetime imaging agent. Important in the design of this imaging agent is the use of two luminophores that are tethered by a specific peptide with a hairpin-motive that ensured close proximity of the two while also having a specific amino acid sequence available for enzymatic cleavage by tumor-related MMP-2/9. Ir(ppy3 and Cy5 were used because in close proximity the emission intensities of both luminophores were quenched and the influence of Cy5 shortens the Ir(ppy3 luminescence lifetime from 98 ns to 30 ns. Upon cleavage in vitro, both effects are undone, yielding an increase in Ir(ppy3 and Cy5 luminescence and a restoration of Ir(ppy3 luminescence lifetime to 94 ns. As a reference for the luminescence activation, a similar imaging agent with the more common Cy3-Cy5 fluorophore pair was used. Our findings underline that the combination of enzymatic signal activation with lifetime imaging is possible and that it provides a promising method in the design of future disease specific imaging agents.

  17. Fluorescence Imaging of Fast Retrograde Axonal Transport in Living Animals

    Directory of Open Access Journals (Sweden)

    Dawid Schellingerhout

    2009-11-01

    Full Text Available Our purpose was to enable an in vivo imaging technology that can assess the anatomy and function of peripheral nerve tissue (neurography. To do this, we designed and tested a fluorescently labeled molecular probe based on the nontoxic C fragment of tetanus toxin (TTc. TTc was purified, labeled, and subjected to immunoassays and cell uptake assays. The compound was then injected into C57BL/6 mice (N = 60 for in vivo imaging and histologic studies. Image analysis and immunohistochemistry were performed. We found that TTc could be labeled with fluorescent moieties without loss of immunoreactivity or biologic potency in cell uptake assays. In vivo fluorescent imaging experiments demonstrated uptake and retrograde transport of the compound along the course of the sciatic nerve and in the spinal cord. Ex vivo imaging and immunohistochemical studies confirmed the presence of TTc in the sciatic nerve and spinal cord, whereas control animals injected with human serum albumin did not exhibit these features. We have demonstrated neurography with a fluorescently labeled molecular imaging contrast agent based on the TTc.

  18. Fluorescence Lifetime Imaging System for in Vivo Studies

    Directory of Open Access Journals (Sweden)

    Moinuddin Hassan

    2007-07-01

    Full Text Available In this article, a fluorescence lifetime imaging system for small animals is presented. Data were collected by scanning a region of interest with a measurement head, a linear fiber array with fixed separations between a single source fiber and several detection fibers. The goal was to localize tumors and monitor their progression using specific fluorescent markers. We chose a near-infrared contrast agent, Alexa Fluor 750 (Invitrogen Corp., Carlsbad, CA. Preliminary results show that the fluorescence lifetime for this dye was sensitive to the immediate environment of the fluorophore (in particular, pH, making it a promising candidate for reporting physiologic changes around a fluorophore. To quantify the intrinsic lifetime of deeply embedded fluorophores, we performed phantom experiments to investigate the contribution of photon migration effects on observed lifetime by calculating the fluorescence intensity decay time. A previously proposed theoretical model of migration, based on random walk theory, is also substantiated by new experimental data. The developed experimental system has been used for in vivo mouse imaging with Alexa Fluor 750 contrast agent conjugated to tumor-specific antibodies (trastuzumab [Herceptin]. Three-dimensional mapping of the fluorescence lifetime indicates lower lifetime values in superficial breast cancer tumors in mice.

  19. 3-D Image Analysis of Fluorescent Drug Binding

    Directory of Open Access Journals (Sweden)

    M. Raquel Miquel

    2005-01-01

    Full Text Available Fluorescent ligands provide the means of studying receptors in whole tissues using confocal laser scanning microscopy and have advantages over antibody- or non-fluorescence-based method. Confocal microscopy provides large volumes of images to be measured. Histogram analysis of 3-D image volumes is proposed as a method of graphically displaying large amounts of volumetric image data to be quickly analyzed and compared. The fluorescent ligand BODIPY FL-prazosin (QAPB was used in mouse aorta. Histogram analysis reports the amount of ligand-receptor binding under different conditions and the technique is sensitive enough to detect changes in receptor availability after antagonist incubation or genetic manipulations. QAPB binding was concentration dependent, causing concentration-related rightward shifts in the histogram. In the presence of 10 μM phenoxybenzamine (blocking agent, the QAPB (50 nM histogram overlaps the autofluorescence curve. The histogram obtained for the 1D knockout aorta lay to the left of that of control and 1B knockout aorta, indicating a reduction in 1D receptors. We have shown, for the first time, that it is possible to graphically display binding of a fluorescent drug to a biological tissue. Although our application is specific to adrenergic receptors, the general method could be applied to any volumetric, fluorescence-image-based assay.

  20. Multiphoton fluorescence lifetime imaging of human hair.

    Science.gov (United States)

    Ehlers, Alexander; Riemann, Iris; Stark, Martin; König, Karsten

    2007-02-01

    In vivo and in vitro multiphoton imaging was used to perform high resolution optical sectioning of human hair by nonlinear excitation of endogenous as well as exogenous fluorophores. Multiphoton fluorescence lifetime imaging (FLIM) based on time-resolved single photon counting and near-infrared femtosecond laser pulse excitation was employed to analyze the various fluorescent hair components. Time-resolved multiphoton imaging of intratissue pigments has the potential (i) to identify endogenous keratin and melanin, (ii) to obtain information on intrahair dye accumulation, (iii) to study bleaching effects, and (iv) to monitor the intratissue diffusion of pharmaceutical and cosmetical components along hair shafts.

  1. Fluorescence imaging of soybean flavonol isolines

    Science.gov (United States)

    Kim, Moon S.; Lee, Edward H.; Mulchi, Charles L.; McMurtrey, James E., III; Chappelle, Emmett W.; Rowland, Randy A.

    1998-07-01

    Experiments were conducted to characterize the fluorescence emission of leaves from four soybean ('Harosoy') plants containing different concentrations of flavonols (kaempferol glycosides). The investigation utilized genetically mutated soybean flavonol isolines grown in a constant environment, thus limiting factors known to affect fluorescence emission characteristics other than different kaempferol glycosides concentrations. Flavonol isolines included OX922, OX941, OX942, OX944. The first two isolines contain kaempferol (K) glycosides; K3, K6, and K9, and the latter two did not have K3, K6, and K9. A fluorescence imaging system (FIS) was used to characterize steady state florescence images of the sample leaves measured at wavelengths centered at 450, 550, 680, and 740 nm with an excitation at 360 nm. Images taken with FIS greatly complement non-imaging fluorescence measurements by characterizing the spatial variation of fluorescence within leaves. We also acquired fluorescence emission spectra to characterize spectral features of the soybean flavonol isolines. The emission spectral shape of the fluorescence emission characteristics were not significantly different between the soybeans that contain kaempferol glycosides and the ones that do not contain kaempferol glycosides. Typical emission maxima of green vegetation in the blue, green, red, and far-red bands were noticed in all four soybean isolines. However, plants containing kaempferol glycosides, OX922 and OX941 had significantly lower intensities throughout the wavelength regions. These results imply that fluorescence emission intensities in the fluorescence emission bands studied are significantly affected by the presence and absence of kaempferol glycosides concentrations (UV radiation screening compounds). Pure kaempferol glycoside dissolved in solution show minimal fluorescence emission when excited with the absorption maximum radiation at 365 nm. However, a broad band emission can be seen in the green

  2. Laser-induced fluorescence imaging of bacteria

    Science.gov (United States)

    Hilton, Peter J.

    1998-12-01

    This paper outlines a method for optically detecting bacteria on various backgrounds, such as meat, by imaging their laser induced auto-fluorescence response. This method can potentially operate in real-time, which is many times faster than current bacterial detection methods, which require culturing of bacterial samples. This paper describes the imaging technique employed whereby a laser spot is scanned across an object while capturing, filtering, and digitizing the returned light. Preliminary results of the bacterial auto-fluorescence are reported and plans for future research are discussed. The results to date are encouraging with six of the eight bacterial strains investigated exhibiting auto-fluorescence when excited at 488 nm. Discrimination of these bacterial strains against red meat is shown and techniques for reducing background fluorescence discussed.

  3. Non-invasive imaging of skin cancer with fluorescence lifetime imaging using two photon tomography

    Science.gov (United States)

    Patalay, Rakesh; Talbot, Clifford; Alexandrov, Yuriy; Munro, Ian; Breunig, Hans Georg; König, Karsten; Warren, Sean; Neil, Mark A. A.; French, Paul M. W.; Chu, Anthony; Stamp, Gordon W.; Dunsby, Christopher

    2011-07-01

    Multispectral fluorescence lifetime imaging (FLIM) using two photon microscopy as a non-invasive technique for the diagnosis of skin lesions is described. Skin contains fluorophores including elastin, keratin, collagen, FAD and NADH. This endogenous contrast allows tissue to be imaged without the addition of exogenous agents and allows the in vivo state of cells and tissues to be studied. A modified DermaInspect® multiphoton tomography system was used to excite autofluorescence at 760 nm in vivo and on freshly excised ex vivo tissue. This instrument simultaneously acquires fluorescence lifetime images in four spectral channels between 360-655 nm using time-correlated single photon counting and can also provide hyperspectral images. The multispectral fluorescence lifetime images were spatially segmented and binned to determine lifetimes for each cell by fitting to a double exponential lifetime model. A comparative analysis between the cellular lifetimes from different diagnoses demonstrates significant diagnostic potential.

  4. Technique of Hadamard transform microscope fluorescence image analysis

    Institute of Scientific and Technical Information of China (English)

    梅二文; 顾文芳; 曾晓斌; 陈观铨; 曾云鹗

    1995-01-01

    Hadamard transform spatial multiplexed imaging technique is combined with fluorescence microscope and an instrument of Hadamard transform microscope fluorescence image analysis is developed. Images acquired by this instrument can provide a lot of useful information simultaneously, including three-dimensional Hadamard transform microscope cell fluorescence image, the fluorescence intensity and fluorescence distribution of a cell, the background signal intensity and the signal/noise ratio, etc.

  5. Agents described in the Molecular Imaging and Contrast Agent Database for imaging carbonic anhydrase IX expression.

    Science.gov (United States)

    Sneddon, Deborah; Poulsen, Sally-Ann

    2014-10-01

    Carbonic anhydrase IX (CA IX) is selectively expressed in a range of hypoxic tumours and is a validated endogenous hypoxia marker with prognostic significance; hence, CA IX is of great interest as a molecular imaging target in oncology. In this review, we present an overview of the different imaging agents and imaging modalities that have been applied for the in vivo detection of CA IX. The imaging agents reviewed are all entries in the Molecular Imaging and Contrast Agent Database (MICAD) and comprise antibody, antibody fragments and small molecule imaging agents. The effectiveness of these agents for imaging CA IX in vivo gave variable performance; however, a number of agents proved very capable. As molecular imaging has become indispensable in current medical practice we anticipate that the clinical significance of CA IX will see continued development and improvements in imaging agents for targeting this enzyme.

  6. Fluorescent and scattering contrast agents in a mouse model of colorectal cancer

    Science.gov (United States)

    Winkler, Amy M.; Rice, Photini F. S.; Troutman, Timothy S.; Backer, Marina V.; Backer, Joseph M.; Drezek, Rebekah A.; Romanowski, Marek; Barton, Jennifer K.

    2008-02-01

    In previous work we have demonstrated the utility of laser-induced fluorescence (LIF) and optical coherence tomography (OCT) to identify adenoma in mouse models of colorectal cancer with high sensitivity and specificity. However, improved sensitivity to early disease, as well as the ability to distinguish confounders (e.g. fecal contamination, natural variations in mucosal thickness), is desired. In this study, we investigated the signal enhancement of fluorescent and scattering contrast agents in the colons of AOM-treated mice. The fluorescent tracer scVEGF/Cy, targeted to receptors for vascular endothelial growth factor, was visualized on a dual modality OCT/LIF endoscopic system with 1300-nm center wavelength OCT source and 635-nm LIF excitation. Scattering agents were tested with an 890-nm center wavelength endoscopic OCT system. Agents included nanoshells, 120-nm in diameter, and nanorods, 20-nm in diameter by 80-nm in length. Following imaging, colons were excised. Tissue treated with fluorophore was imaged on an epifluorescence microscope. Histological sections were obtained and stained with H&E and silver enhancer to verify disease and identify regions of gold uptake, respectively. Non-specific signal enhancement was observed with the scattering contrast agents. Specificity for adenoma was seen with the scVEGF/Cy dye.

  7. Targeted Agents for Imaging and Therapy

    NARCIS (Netherlands)

    Grüll, H.; Robillard, M.S.

    2005-01-01

    Molecular Imaging allows the visualization of biological processesin vivo, offering new chances for healthcare with respect to early diagnosis and improved therapy. The new field of molecular imaging isboosted by more sensitive imaging systems and the emergence of targeted imaging agents that hom

  8. Fluorescein Derivatives in Intravital Fluorescence Imaging

    Directory of Open Access Journals (Sweden)

    Michael S. Roberts

    2013-08-01

    Full Text Available Intravital fluorescence microscopy enables the direct imaging of fluorophores in vivo and advanced techniques such as fluorescence lifetime imaging (FLIM enable the simultaneous detection of multiple fluorophores. Consequently, it is now possible to record distribution and metabolism of a chemical in vivo and to optimise the delivery of fluorophores in vivo. Recent clinical applications with fluorescein and other intravital fluorescent stains have occurred in neurosurgery, dermatology [including photodynamic therapy (PDT] and endomicroscopy. Potential uses have been identified in periodontal disease, skin graft and cancer surgery. Animal studies have demonstrated that diseased tissue can be specifically stained with fluorophore conjugates. This review focuses on the fluorescein derived fluorophores in common clinical use and provides examples of novel applications from studies in tissue samples.

  9. A review of imaging agent development.

    Science.gov (United States)

    Agdeppa, Eric D; Spilker, Mary E

    2009-06-01

    This educational review highlights the processes, opportunities, and challenges encountered in the discovery and development of imaging agents, mainly positron emission tomography and single-photon emission computed tomography tracers. While the development of imaging agents parallels the drug development process, unique criteria are needed to identify opportunities for new agents. Imaging agent development has the flexibility to pursue functional or nonfunctional targets as long as they play a role in the specific disease or mechanism of interest and meet imageability requirements. However, their innovation is tempered by relatively small markets for diagnostic imaging agents, intellectual property challenges, radiolabeling constraints, and adequate target concentrations for imaging. At the same time, preclinical imaging is becoming a key translational tool for proof of mechanism and concept studies. Pharmaceutical and imaging industries face a common bottleneck in the form of the limited number of trials one company can possibly perform. However, microdosing and theranostics are evidence that partnerships between pharmaceutical and imaging companies can accelerate clinical translation of tracers and therapeutic interventions. This manuscript will comment on these aspects to provide an educational review of the discovery and development processes for imaging agents.

  10. A new fluorescent imaging of renal inflammation with RCP.

    Science.gov (United States)

    Nakamura, Kentaro; Tabata, Yasuhiko

    2010-12-20

    The objective of this study is to design a fluorescent imaging agent with R-Gel, one of the recombinant polymers (RCP), for renal inflammation. The R-Gel based on human type I collagen has multiple Arg-Gly-Asp (RGD) motifs which are ligands for some types of integrin receptors on the cell surface. After intravenous administration of R-Gel labeled by Cy7 of a fluorescent dye to three animal models of nephritis mousse, interstitial nephritis (by using UUO model mice), glomerulonephritis (HIGA mice), and ischemia-reperfusion injured kidney (I/R mice), the extent of fluorescent imaging at the renal inflammation was assessed. The Cy7-labeled R-Gel was accumulated in the inflammation site to a significantly greater extent than in the normal one at 24h after administration. The renal pattern of fluorescent imaging was similar to that of administration anti-Mac1 antibody. Taken together, it is conceivable that the R-Gel was targeted to macrophages infiltrated into the inflammation site of kidney. Copyright © 2010 Elsevier B.V. All rights reserved.

  11. Dendrimer-entrapped metal colloids as imaging agents.

    Science.gov (United States)

    Li, Du; Wen, Shihui; Shi, Xiangyang

    2015-01-01

    This review reports the recent advances in dendrimer-entrapped metal colloids as contrast agents for biomedical imaging applications. The versatile dendrimer scaffolds with 3-dimensional spherical shape, highly branched internal cavity, tunable surface conjugation chemistry, and excellent biocompatibility and nonimmunogenicity afford their uses as templates to create multifunctional dendrimer-entrapped metal colloids for mono- or multi- mode molecular imaging applications. In particular, multifunctional dendrimer-entrapped gold nanoparticles with different surface modifications have been used for fluorescence imaging, targeted tumor computed tomography (CT) imaging, enhanced blood pool CT imaging, dual mode CT/MR imaging, and tumor theranostics (combined CT imaging and chemotherapy) will be introduced and discussed in detail. © 2015 Wiley Periodicals, Inc.

  12. Fluorescence imaging of experimental rheumatoid arthritis in vivo using a fast flying-spot scanner

    Science.gov (United States)

    Berger, J.; Voigt, J.; Seifert, F.; Ebert, B.; Macdonald, R.; Gemeinhardt, I.; Gemeinhardt, O.; Schnorr, J.; Taupitz, M.; Vater, A.; Vollmer, S.; Licha, K.; Schirner, M.

    2007-07-01

    We have developed a flying-spot scanner for fluorescence imaging of rheumatoid arthritis in the near infrared (NIR) spectral range following intravenous administration of contrast agents. The new imaging system has been characterized with respect to linearity, dynamic range and spatial resolution with the help of fluorescent phantoms. In vivo experiments were performed on an animal model of rheumatoid arthritis. Finally, NIR-fluorescence images of early stages of joint inflammation have been compared with findings from contrast enhanced MR imaging and histology.

  13. Multifunctional photosensitizer-based contrast agents for photoacoustic imaging.

    Science.gov (United States)

    Ho, Chris Jun Hui; Balasundaram, Ghayathri; Driessen, Wouter; McLaren, Ross; Wong, Chi Lok; Dinish, U S; Attia, Amalina Binte Ebrahim; Ntziachristos, Vasilis; Olivo, Malini

    2014-06-18

    Photoacoustic imaging is a novel hybrid imaging modality combining the high spatial resolution of optical imaging with the high penetration depth of ultrasound imaging. Here, for the first time, we evaluate the efficacy of various photosensitizers that are widely used as photodynamic therapeutic (PDT) agents as photoacoustic contrast agents. Photoacoustic imaging of photosensitizers exhibits advantages over fluorescence imaging, which is prone to photobleaching and autofluorescence interference. In this work, we examined the photoacoustic activity of 5 photosensitizers: zinc phthalocyanine, protoporphyrin IX, 2,4-bis [4-(N,N-dibenzylamino)-2,6-dihydroxyphenyl] squaraine, chlorin e6 and methylene blue in phantoms, among which zinc phthalocyanine showed the highest photoacoustic activity. Subsequently, we evaluated its tumor localization efficiency and biodistribution at multiple time points in a murine model using photoacoustic imaging. We observed that the probe localized at the tumor within 10 minutes post injection, reaching peak accumulation around 1 hour and was cleared within 24 hours, thus, demonstrating the potential of photosensitizers as photoacoustic imaging contrast agents in vivo. This means that the known advantages of photosensitizers such as preferential tumor uptake and PDT efficacy can be combined with photoacoustic imaging capabilities to achieve longitudinal monitoring of cancer progression and therapy in vivo.

  14. Multifunctional Photosensitizer-Based Contrast Agents for Photoacoustic Imaging

    Science.gov (United States)

    Ho, Chris Jun Hui; Balasundaram, Ghayathri; Driessen, Wouter; McLaren, Ross; Wong, Chi Lok; Dinish, U. S.; Attia, Amalina Binte Ebrahim; Ntziachristos, Vasilis; Olivo, Malini

    2014-06-01

    Photoacoustic imaging is a novel hybrid imaging modality combining the high spatial resolution of optical imaging with the high penetration depth of ultrasound imaging. Here, for the first time, we evaluate the efficacy of various photosensitizers that are widely used as photodynamic therapeutic (PDT) agents as photoacoustic contrast agents. Photoacoustic imaging of photosensitizers exhibits advantages over fluorescence imaging, which is prone to photobleaching and autofluorescence interference. In this work, we examined the photoacoustic activity of 5 photosensitizers: zinc phthalocyanine, protoporphyrin IX, 2,4-bis [4-(N,N-dibenzylamino)-2,6-dihydroxyphenyl] squaraine, chlorin e6 and methylene blue in phantoms, among which zinc phthalocyanine showed the highest photoacoustic activity. Subsequently, we evaluated its tumor localization efficiency and biodistribution at multiple time points in a murine model using photoacoustic imaging. We observed that the probe localized at the tumor within 10 minutes post injection, reaching peak accumulation around 1 hour and was cleared within 24 hours, thus, demonstrating the potential of photosensitizers as photoacoustic imaging contrast agents in vivo. This means that the known advantages of photosensitizers such as preferential tumor uptake and PDT efficacy can be combined with photoacoustic imaging capabilities to achieve longitudinal monitoring of cancer progression and therapy in vivo.

  15. Developing an imaging bi-spectrometer for fluorescent materials

    Science.gov (United States)

    Mohammadi, Mahnaz

    Fluorescent effects have been observed for thousands of years. Stokes, in 1852, began the science of fluorescence culminating in his law of fluorescence, which explained that fluorescence emission occurs at longer wavelengths than the excitation wavelength. This phenomenon is observed extensively in the art world. Daylight fluorescent colors known as Day-GloRTM have become an artistic medium since the 1960s. Modern artists exploit these saturated and brilliant colors to glitter their painting. Multipsectral imaging as a noninvasive technique has been used for archiving by museums and cultural-heritage institutions for about a decade. The complex fluorescence phenomenon has been often ignored in the multispectral projects. The ignored fluorescence results in errors in digital imaging of artwork containing fluorescent colors. The illuminant-dependency of the fluorescence radiance makes the fluorescence colorimetry and consequently spectral imaging more complex. In this dissertation an abridged imaging bi-spectrometer for artwork containing both fluorescent and non-fluorescent colors was developed. The method developed included two stages of reconstruction of the spectral reflected radiance factor and prediction of the fluorescent radiance factor. The estimation of the reflected radiance factor as a light source independent component was achieved by imaging with a series of short-wavelength cutoff filters placed in the illumination path. The fluorescent radiance factor, a light source dependent component, was estimated based on a proposed model, the abridged two-monochromator method. The abridged two-monochromator method was developed for reconstructing the bi-spectral matrix of a fluorescent color based on a calibrated UV-fluorescence imaging. In this way, one could predict the fluorescence radiance factor under any desired illuminant and consequently a better color evaluation and rendering could be obtained. Furthermore, this method easily fitted in a general system

  16. Near-Infrared Squaraine Dye Encapsulated Micelles for in Vivo Fluorescence and Photoacoustic Bimodal Imaging.

    Science.gov (United States)

    Sreejith, Sivaramapanicker; Joseph, James; Lin, Manjing; Menon, Nishanth Venugopal; Borah, Parijat; Ng, Hao Jun; Loong, Yun Xian; Kang, Yuejun; Yu, Sidney Wing-Kwong; Zhao, Yanli

    2015-06-23

    Combined near-infrared (NIR) fluorescence and photoacoustic imaging techniques present promising capabilities for noninvasive visualization of biological structures. Development of bimodal noninvasive optical imaging approaches by combining NIR fluorescence and photoacoustic tomography demands suitable NIR-active exogenous contrast agents. If the aggregation and photobleaching are prevented, squaraine dyes are ideal candidates for fluorescence and photoacoustic imaging. Herein, we report rational selection, preparation, and micelle encapsulation of an NIR-absorbing squaraine dye (D1) for in vivo fluorescence and photoacoustic bimodal imaging. D1 was encapsulated inside micelles constructed from a biocompatible nonionic surfactant (Pluoronic F-127) to obtain D1-encapsulated micelles (D1(micelle)) in aqueous conditions. The micelle encapsulation retains both the photophysical features and chemical stability of D1. D1(micelle) exhibits high photostability and low cytotoxicity in biological conditions. Unique properties of D1(micelle) in the NIR window of 800-900 nm enable the development of a squaraine-based exogenous contrast agent for fluorescence and photoacoustic bimodal imaging above 820 nm. In vivo imaging using D1(micelle), as demonstrated by fluorescence and photoacoustic tomography experiments in live mice, shows contrast-enhanced deep tissue imaging capability. The usage of D1(micelle) proven by preclinical experiments in rodents reveals its excellent applicability for NIR fluorescence and photoacoustic bimodal imaging.

  17. Liposome imaging agents in personalized medicine

    DEFF Research Database (Denmark)

    Petersen, Anncatrine Luisa; Hansen, Anders Elias; Gabizon, Alberto;

    2012-01-01

    that selectively localize in tumor tissue can transport both drugs and imaging agents, which allows for a theranostic approach with great potential in personalized medicine. Radiolabeling of liposomes have for many years been used in preclinical studies for evaluating liposome in vivo performance and has been...... start to consider how to use imaging for patient selection and treatment monitoring in connection to nanocarrier based medicines. Nanocarrier imaging agents could furthermore have interesting properties for disease diagnostics and staging. Here, we review the major advances in the development...... of radiolabeled liposomes for imaging as a tool in personalized medicine....

  18. Peptide-based MRI contrast agent and near-infrared fluorescent probe for intratumoral legumain detection.

    Science.gov (United States)

    Chen, Yu-Jen; Wu, Shou-Cheng; Chen, Chung-Yung; Tzou, Shey-Cherng; Cheng, Tian-Lu; Huang, Ying-Fang; Yuan, Shyng-Shiou; Wang, Yun-Ming

    2014-01-01

    Recent studies suggest that intratumoral legumain promotes tumorigenesis. To monitor legumain activity in tumors, we developed a new MRI contrast agent ([Gd-NBCB-TTDA-Leg(L)]) and a NIR fluorescence probe (CyTE777-Leg(L)-CyTE807). The MRI contrast agent was prepared by introduction of cyclobutyl and benzyl group residues to TTDA (3,6,10-tri(carboxymethyl)-3,6,10-triaza-dodecanedioic acid), followed by the attachment of a legumain-specific substrate peptide (Leg(L)). The NIR fluorescence probe was designed by conjugating two NIR fluorochromes (CyTE777 and CyTE807) with Leg(L). Peptide cleavage of the MRI contrast agent by legumain can increase its hydrophobicity and promote rotational correlation time (τ(R)). Peptide cleavage of the NIR probes by the legumain relieves the self quench of the probe. Peptide cleavage of the MRI contrast agent and the NIR fluorescence probe by legumain were confirmed by T1 relaxometric studies and by fluorescence studies, respectively. In vivo MR images showed that [Gd-NBCB-TTDA-Leg(L)] attained 55.3 fold (254.2% versus 4.6%, at 2.0 h post-injection) higher imaging enhancement, as compared with control contrast agent bearing a noncleaveable peptide ([Gd-NBCB-TTDA-Leg(D)], in the CT-26 (legumain(+)) tumors. Similarly, optical imaging probe CyTE777-Leg(L)-CyTE807 attained 15.2 fold (3.34 × 10(9) photons/min versus 0.22 × 10(9) photons/min, at 24.0 h post-injection) higher imaging enhancement in the CT-26 (legumain(+)) tumors, compared to a NIR control probe (CyTE777-Leg(D)-CyTE807). These data indicate that the [Gd-NBCB-TTDA-Leg(L)] and the CyTE777-Leg(L)-CyTE807 probes may be promising tools to image the legumain-expressing cancers for diagnoses and targeted treatments.

  19. Neurosurgical confocal endomicroscopy: A review of contrast agents, confocal systems, and future imaging modalities

    Directory of Open Access Journals (Sweden)

    Aqib H Zehri

    2014-01-01

    Full Text Available Background: The clinical application of fluorescent contrast agents (fluorescein, indocyanine green, and aminolevulinic acid with intraoperative microscopy has led to advances in intraoperative brain tumor imaging. Their properties, mechanism of action, history of use, and safety are analyzed in this report along with a review of current laser scanning confocal endomicroscopy systems. Additional imaging modalities with potential neurosurgical utility are also analyzed. Methods: A comprehensive literature search was performed utilizing PubMed and key words: In vivo confocal microscopy, confocal endomicroscopy, fluorescence imaging, in vivo diagnostics/neoplasm, in vivo molecular imaging, and optical imaging. Articles were reviewed that discussed clinically available fluorophores in neurosurgery, confocal endomicroscopy instrumentation, confocal microscopy systems, and intraoperative cancer diagnostics. Results: Current clinically available fluorescent contrast agents have specific properties that provide microscopic delineation of tumors when imaged with laser scanning confocal endomicroscopes. Other imaging modalities such as coherent anti-Stokes Raman scattering (CARS microscopy, confocal reflectance microscopy, fluorescent lifetime imaging (FLIM, two-photon microscopy, and second harmonic generation may also have potential in neurosurgical applications. Conclusion: In addition to guiding tumor resection, intraoperative fluorescence and microscopy have the potential to facilitate tumor identification and complement frozen section analysis during surgery by providing real-time histological assessment. Further research, including clinical trials, is necessary to test the efficacy of fluorescent contrast agents and optical imaging instrumentation in order to establish their role in neurosurgery.

  20. Polymer-encapsulated organic nanoparticles for fluorescence and photoacoustic imaging.

    Science.gov (United States)

    Li, Kai; Liu, Bin

    2014-09-21

    Polymer encapsulated organic nanoparticles have recently attracted increasing attention in the biomedical field because of their unique optical properties, easy fabrication and outstanding performance as imaging and therapeutic agents. Of particular importance is the polymer encapsulated nanoparticles containing conjugated polymers (CP) or fluorogens with aggregation induced emission (AIE) characteristics as the core, which have shown significant advantages in terms of tunable brightness, superb photo- and physical stability, good biocompatibility, potential biodegradability and facile surface functionalization. In this review, we summarize the latest advances in the development of polymer encapsulated CP and AIE fluorogen nanoparticles, including preparation methods, material design and matrix selection, nanoparticle fabrication and surface functionalization for fluorescence and photoacoustic imaging. We also discuss their specific applications in cell labeling, targeted in vitro and in vivo imaging, blood vessel imaging, cell tracing, inflammation monitoring and molecular imaging. We specially focus on strategies to fine-tune the nanoparticle property (e.g. size and fluorescence quantum yield) through precise engineering of the organic cores and careful selection of polymer matrices. The review also highlights the merits and limitations of these nanoparticles as well as strategies used to overcome the limitations. The challenges and perspectives for the future development of polymer encapsulated organic nanoparticles are also discussed.

  1. Fluorescence lifetime imaging of oxygen in living cells

    NARCIS (Netherlands)

    Gerritsen, H.C.; Sanders, R.; Draaijer, A.; Ince, C.; Levine, Y.K.

    1997-01-01

    The usefulness of the fluorescent probe ruthenium tris(2,2′-dipyridyl) dichloride hydrate (RTDP) for the quantitative imaging of oxygen in single cells was investigated utilizing fluorescence life-time imaging. The results indicate that the fluorescence behavior of RTDP in the presence of oxygen can

  2. Enhancing contrast and quantitation by spatial frequency domain fluorescence molecular imaging

    Science.gov (United States)

    Sun, Jessica; Hathi, Deep; Zhou, Haiying; Shokeen, Monica; Akers, Walter J.

    2016-03-01

    Optical imaging with fluorescent contrast agents is highly sensitive for molecular imaging but is limited in depth to a few centimeters below the skin. Planar fluorescence imaging with full-field, uniform illumination and scientific camera image capture provides a portable and robust configuration for real-time, sensitive fluorescence detection with scalable resolution, but is inherently surface weighted and therefore limited in depth to a few millimeters. At the NIR region (700-1000 nm), tissue absorption and autofluorescence are relatively reduced, increasing depth penetration and reducing background signal, respectively. Optical imaging resolution scales with depth, limiting microscopic resolution with multiphoton microscopy and optical coherence tomography to skin and peri-tumoral tissues are not uniform, varying in thickness and color, complicating subsurface fluorescence measurements. Diffuse optical imaging methods have been developed that better quantify optical signals relative to faster full-field planar reflectance imaging, but require long scan times, complex instrumentation, and reconstruction algorithms. Here we report a novel strategy for rapid measurement of subsurface fluorescence using structured light illumination to improve quantitation of deep-seated fluorescence molecular probe accumulation. This technique, in combination with highly specific, tumor-avid fluorescent molecular probes, will easily integrate noninvasive diagnostics for superficial cancers and fluorescence guided surgery.

  3. Creating Panoramic Images for Bladder Fluorescence Endoscopy

    Directory of Open Access Journals (Sweden)

    A. Behrens

    2008-01-01

    Full Text Available The medical diagnostic analysis and therapy of urinary bladder cancer based on endoscopes are state of the art in urological medicine. Due to the limited field of view of endoscopes, the physician can examine only a small part of the whole operating field at once. This constraint makes visual control and navigation difficult, especially in hollow organs. A panoramic image, covering a larger field of view, can overcome this difficulty. Directly motivated by a physician we developed an image mosaicing algorithm for endoscopic bladder fluorescence video sequences. In this paper, we present an approach which is capable of stitching single endoscopic video images to a combined panoramic image. Based on SIFT features we estimate a 2-D homography for each image pair, using an affine model and an iterative model-fitting algorithm. We then apply the stitching process and perform a mutual linear interpolation. Our panoramic image results show a correct stitching and lead to a better overview and understanding of the operation field. 

  4. Performance evaluation of spot detection algorithms in fluorescence microscopy images

    CSIR Research Space (South Africa)

    Mabaso, M

    2012-10-01

    Full Text Available Detection of messenger Ribonucleic Acid (mRNA) spots in fluorescence microscopy images is of great importance for biologists seeking better understanding of cell functionality. Fluorescence microscopy and specific staining methods make biological...

  5. Smartphone microendoscopy for high resolution fluorescence imaging

    CERN Document Server

    Hong, Xiangqian; Mugler, Dale H; Yu, Bing

    2016-01-01

    High resolution optical endoscopes are increasingly used in diagnosis of various medical conditions of internal organs, such as the gastrointestinal tracts, but they are too expensive for use in resource-poor settings. On the other hand, smartphones with high resolution cameras and Internet access have become more affordable, enabling them to diffuse into most rural areas and developing countries in the past decade. In this letter we describe a smartphone microendoscope that can take fluorescence images with a spatial resolution of 3.1 {\\mu}m. Images collected from ex vivo, in vitro and in vivo samples using the device are also presented. The compact and cost-effective smartphone microendoscope may be envisaged as a powerful tool for detecting pre-cancerous lesions of internal organs in low and middle income countries.

  6. A novel multiwavelength fluorescence image-guided surgery imaging system

    Science.gov (United States)

    Volpi, D.; Tullis, I. D. C.; Laios, A.; Pathiraja, P. N. J.; Haldar, K.; Ahmed, A. A.; Vojnovic, B.

    2014-02-01

    We describe the development and performance analysis of two clinical near-infrared fluorescence image-guided surgery (FIGS) devices that aim to overcome some of the limitations of current FIGS systems. The devices operate in a widefield-imaging mode and can work (1) in conjunction with a laparoscope, during minimally invasive surgery, and (2) as a hand-held, open surgery imaging system. In both cases, narrow-band excitation light, delivered at multiple wavelengths, is efficiently combined with white reflectance light. Light is delivered to ~100 cm2 surgical field at 1-2 mW/cm2 for white light and 3-7 mW/cm2 (depending on wavelength) of red - near infrared excitation, at a typical working distance of 350 mm for the hand-held device and 100 mm for the laparoscope. A single, sensitive, miniaturized color camera collects both fluorescence and white reflectance light. The use of a single imager eliminates image alignment and software overlay complexity. A novel filtering and illumination arrangement allows simultaneous detection of white reflectance and fluorescence emission from multiple dyes in real-time. We will present both fluorescence detection sensitivity modeling and practical performance data. We have demonstrated the efficiency and the advantages of the devices both pre-clinically and during live surgery on humans. Both the hand-held and the laparoscopic systems have proved to be reliable and beneficial in an ongoing clinical trial involving sentinel lymph node detection in gynecological cancers. We will show preliminary results using two clinically approved dyes, Methylene blue and indocyanine green. We anticipate that this technology can be integrated and routinely used in a larger variety of surgical procedures.

  7. Carbon Quantum Dots for Zebrafish Fluorescence Imaging

    Science.gov (United States)

    Kang, Yan-Fei; Li, Yu-Hao; Fang, Yang-Wu; Xu, Yang; Wei, Xiao-Mi; Yin, Xue-Bo

    2015-07-01

    Carbon quantum dots (C-QDs) are becoming a desirable alternative to metal-based QDs and dye probes owing to their high biocompatibility, low toxicity, ease of preparation, and unique photophysical properties. Herein, we describe fluorescence bioimaging of zebrafish using C-QDs as probe in terms of the preparation of C-QDs, zebrafish husbandry, embryo harvesting, and introduction of C-QDs into embryos and larvae by soaking and microinjection. The multicolor of C-QDs was validated with their imaging for zebrafish embryo. The distribution of C-QDs in zebrafish embryos and larvae were successfully observed from their fluorescence emission. the bio-toxicity of C-QDs was tested with zebrafish as model and C-QDs do not interfere to the development of zebrafish embryo. All of the results confirmed the high biocompatibility and low toxicity of C-QDs as imaging probe. The absorption, distribution, metabolism and excretion route (ADME) of C-QDs in zebrafish was revealed by their distribution. Our work provides the useful information for the researchers interested in studying with zebrafish as a model and the applications of C-QDs. The operations related zebrafish are suitable for the study of the toxicity, adverse effects, transport, and biocompatibility of nanomaterials as well as for drug screening with zebrafish as model.

  8. Imaging carious dental tissues with multiphoton fluorescence lifetime imaging microscopy

    Science.gov (United States)

    Lin, Po-Yen; Lyu, Hong-Chou; Hsu, Chin-Ying Stephen; Chang, Chia-Seng; Kao, Fu-Jen

    2011-01-01

    In this study, multiphoton excitation was utilized to image normal and carious dental tissues noninvasively. Unique structures in dental tissues were identified using the available multimodality (second harmonic, autofluorescence, and fluorescence lifetime analysis) without labeling. The collagen in dentin exhibits a strong second harmonic response. Both dentin and enamel emit strong autofluorescence that reveals in detail morphological features (such as dentinal tubules and enamel rods) and, despite their very similar spectral profiles, can be differentiated by lifetime analysis. Specifically, the carious dental tissue exhibits a greatly reduced autofluorescence lifetime, which result is consistent with the degree of demineralization, determined by micro-computed tomography. Our findings suggest that two-photon excited fluorescence lifetime imaging may be a promising tool for diagnosing and monitoring dental caries. PMID:21326645

  9. Development of a Hybrid Nanoprobe for Triple-Modality MR/SPECT/Optical Fluorescence Imaging

    Science.gov (United States)

    Madru, Renata; Svenmarker, Pontus; Ingvar, Christian; Ståhlberg, Freddy; Engels, Stefan-Andersson; Knutsson, Linda; Strand, Sven-Erik

    2014-01-01

    Hybrid clinical imaging is an emerging technology, which improves disease diagnosis by combining already existing technologies. With the combination of high-resolution morphological imaging, i.e., MRI/CT, and high-sensitive molecular detection offered by SPECT/PET/Optical, physicians can detect disease progression at an early stage and design patient-specific treatments. To fully exploit the possibilities of hybrid imaging a hybrid probe compatible with each imaging technology is required. Here, we present a hybrid nanoprobe for triple modality MR/SPECT/Fluorescence imaging. Our imaging agent is comprised of superparamagnetic iron oxide nanoparticles (SPIONs), labeled with 99mTc and an Alexa fluorophore (AF), together forming 99mTc-AF-SPIONs. The agent was stable in human serum, and, after subcutaneous injection in the hind paw of Wistar rats, showed to be highly specific by accumulating in the sentinel lymph node. All three modalities clearly visualized the imaging agent. Our results show that a single imaging agent can be used for hybrid imaging. The use of a single hybrid contrast agent permits simultaneous hybrid imaging and, more conventionally, allow for single modality imaging at different time points. For example, a hybrid contrast agent enables pre-operative planning, intra-operative guidance, and post-operative evaluation with the same contrast agent. PMID:26852675

  10. In vivo tomographic imaging with fluorescence and MRI using tumor-targeted dual-labeled nanoparticles

    Directory of Open Access Journals (Sweden)

    Zhang Y

    2013-12-01

    Full Text Available Yue Zhang,1 Bin Zhang,1 Fei Liu,1,2 Jianwen Luo,1,3 Jing Bai1 1Department of Biomedical Engineering, School of Medicine, 2Tsinghua-Peking Center for Life Sciences, 3Center for Biomedical Imaging Research, Tsinghua University, Beijing, People's Republic of China Abstract: Dual-modality imaging combines the complementary advantages of different modalities, and offers the prospect of improved preclinical research. The combination of fluorescence imaging and magnetic resonance imaging (MRI provides cross-validated information and direct comparison between these modalities. Here, we report on the application of a novel tumor-targeted, dual-labeled nanoparticle (NP, utilizing iron oxide as the MRI contrast agent and near infrared (NIR dye Cy5.5 as the fluorescent agent. Results of in vitro experiments verified the specificity of the NP to tumor cells. In vivo tumor targeting and uptake of the NPs in a mouse model were visualized by fluorescence and MR imaging collected at different time points. Quantitative analysis was carried out to evaluate the efficacy of MRI contrast enhancement. Furthermore, tomographic images were also acquired using both imaging modalities and cross-validated information of tumor location and size between these two modalities was revealed. The results demonstrate that the use of dual-labeled NPs can facilitate the dual-modal detection of tumors, information cross-validation, and direct comparison by combing fluorescence molecular tomography (FMT and MRI. Keywords: dual-modality, fluorescence molecular tomography (FMT, magnetic resonance imaging (MRI, nanoparticle

  11. Synthesis and evaluation of new salicylaldehyde-2-picolinylhydrazone Schiff base compounds of Ru(II), Rh(III) and Ir(III) as in vitro antitumor, antibacterial and fluorescence imaging agents.

    Science.gov (United States)

    Palepu, Narasinga Rao; Nongbri, S L; Premkumar, J Richard; Verma, Akalesh Kumar; Bhattacharjee, Kaushik; Joshi, S R; Forbes, Scott; Mozharivskyj, Yurij; Thounaojam, Romita; Aguan, K; Kollipara, Mohan Rao

    2015-06-01

    Reaction of salicylaldehyde-2-picolinylhydrazone (HL) Schiff base ligand with precursor compounds [{(p-cymene)RuCl2}2] 1, [{(C6H6)RuCl2}2] 2, [{Cp*RhCl2}2] 3 and [{Cp*IrCl2}2] 4 yielded the corresponding neutral mononuclear compounds 5-8, respectively. The in vitro antitumor evaluation of the compounds 1-8 against Dalton's ascites lymphoma (DL) cells by fluorescence-based apoptosis study and by their half-maximal inhibitory concentration (IC50) values revealed the high antitumor activity of compounds 3, 4, 5 and 6. Compounds 1-8 render comparatively lower apoptotic effect than that of cisplatin on model non-tumor cells, i.e., peripheral blood mononuclear cells (PBMC). The antibacterial evaluation of compounds 5-8 by agar well-diffusion method revealed that compound 6 is significantly effective against all the eight bacterial species considered with zone of inhibition up to 35 mm. Fluorescence imaging study of compounds 5-8 with plasmid circular DNA (pcDNA) and HeLa RNA demonstrated their fluorescence imaging property upon binding with nucleic acids. The docking study with some key enzymes associated with the propagation of cancer such as ribonucleotide reductase, thymidylate synthase, thymidylate phosphorylase and topoisomerase II revealed strong interactions between proteins and compounds 5-8. Conformational analysis by density functional theory (DFT) study has corroborated our experimental observation of the N, N binding mode of ligand. Compounds 5-8 exhibited a HOMO (highest occupied molecular orbital)-LUMO (lowest unoccupied molecular orbital) energy gap 2.99-3.04 eV. Half-sandwich ruthenium, rhodium and iridium compounds were obtained by treatment of metal precursors with salicylaldehyde-2-picolinylhydrazone (HL) by in situ metal-mediated deprotonation of the ligand. Compounds under investigation have shown potential antitumor, antibacterial and fluorescence imaging properties. Arene ruthenium compounds exhibited higher activity compared to that of Cp

  12. Metal complex-based templates and nanostructures for magnetic resonance/optical multimodal imaging agents

    NARCIS (Netherlands)

    Galindo Millan, J.J.

    2012-01-01

    In this thesis, new approaches directed towards simple and functional imaging agents (IAs) for magnetic resonance (MR) and fluorescence multimodal imaging are proposed. In Chapter 3, hybrid silver nanostructures (hAgNSs), grown using a polyamino carboxylic acid scaffold, namely 1,4,7,10-tetraaza-1-(

  13. Cellular image segmentation using n-agent cooperative game theory

    Science.gov (United States)

    Dimock, Ian B.; Wan, Justin W. L.

    2016-03-01

    Image segmentation is an important problem in computer vision and has significant applications in the segmentation of cellular images. Many different imaging techniques exist and produce a variety of image properties which pose difficulties to image segmentation routines. Bright-field images are particularly challenging because of the non-uniform shape of the cells, the low contrast between cells and background, and imaging artifacts such as halos and broken edges. Classical segmentation techniques often produce poor results on these challenging images. Previous attempts at bright-field imaging are often limited in scope to the images that they segment. In this paper, we introduce a new algorithm for automatically segmenting cellular images. The algorithm incorporates two game theoretic models which allow each pixel to act as an independent agent with the goal of selecting their best labelling strategy. In the non-cooperative model, the pixels choose strategies greedily based only on local information. In the cooperative model, the pixels can form coalitions, which select labelling strategies that benefit the entire group. Combining these two models produces a method which allows the pixels to balance both local and global information when selecting their label. With the addition of k-means and active contour techniques for initialization and post-processing purposes, we achieve a robust segmentation routine. The algorithm is applied to several cell image datasets including bright-field images, fluorescent images and simulated images. Experiments show that the algorithm produces good segmentation results across the variety of datasets which differ in cell density, cell shape, contrast, and noise levels.

  14. Augmented microscopy: real-time overlay of bright-field and near-infrared fluorescence images.

    Science.gov (United States)

    Watson, Jeffrey R; Gainer, Christian F; Martirosyan, Nikolay; Skoch, Jesse; Lemole, G Michael; Anton, Rein; Romanowski, Marek

    2015-10-01

    Intraoperative applications of near-infrared (NIR) fluorescent contrast agents can be aided by instrumentation capable of merging the view of surgical field with that of NIR fluorescence. We demonstrate augmented microscopy, an intraoperative imaging technique in which bright-field (real) and electronically processed NIR fluorescence (synthetic) images are merged within the optical path of a stereomicroscope. Under luminance of 100,000 lx, representing typical illumination of the surgical field, the augmented microscope detects 189 nM concentration of indocyanine green and produces a composite of the real and synthetic images within the eyepiece of the microscope at 20 fps. Augmentation described here can be implemented as an add-on module to visualize NIR contrast agents, laser beams, or various types of electronic data within the surgical microscopes commonly used in neurosurgical, cerebrovascular, otolaryngological, and ophthalmic procedures.

  15. Nanoprobes for super-resolution fluorescence imaging at the nanoscale

    Institute of Scientific and Technical Information of China (English)

    HOU ShangGuo; LIANG Le; DENG SuHui; CHEN JianFang; HUANG Qing; CHENG Ya; FAN ChunHai

    2014-01-01

    Compared with other imaging techniques,fluorescence microscopy has become an essential tool to study cell biology due to its high compatibility with living cells.Owing to the resolution limit set by the diffraction of light,fluorescence microscopy could not resolve the nanostructures in the range of〈200 nm.Recently,many techniques have been emerged to overcome the diffraction barrier,providing nanometer spatial resolution.In the course of development,the progress in fluorescent probes has helped to promote the development of the high-resolution fluorescence nanoscopy.Here,we describe the contributions of the fluorescent probes to far-field super resolution imaging,focusing on concepts of the existing super-resolution nanoscopy based on the photophysics of fluorescent nanoprobes,like photoswitching,bleaching and blinking.Fluorescent probe technology is crucial in the design and implementation of super-resolution imaging methods.

  16. IRDye78 Conjugates for Near-Infrared Fluorescence Imaging

    Directory of Open Access Journals (Sweden)

    Atif Zaheer

    2002-10-01

    Full Text Available The detection of human malignancies by near-infrared (NIR fluorescence will require the conjugation of cancer-specific ligands to NIR fluorophores that have optimal photoproperties and pharmacokinetics. IRDye78, a tetra-sulfonated heptamethine indocyanine NIR fluorophore, meets most of the criteria for an in vivo imaging agent, and is available as an N-hydroxysuccinimide ester for conjugation to low-molecular-weight ligands. However, IRDye78 has a high charge-to-mass ratio, complicating purification of conjugates. It also has a potentially labile linkage between fluorophore and ligand. We have developed an ion-pairing purification strategy for IRDye78 that can be performed with a standard C18 column under neutral conditions, thus preserving the stability of fluorophore, ligand, and conjugate. By employing parallel evaporative light scatter and absorbance detectors, all reactants and products are identified, and conjugate purity is maximized. We describe reversible and irreversible conversions of IRDye78 that can occur during sample purification, and describe methods for preserving conjugate stability. Using seven ligands, spanning several classes of small molecules and peptides (neutral, charged, and/or hydrophobic, we illustrate the robustness of these methods, and confirm that IRDye78 conjugates so purified retain bioactivity and permit NIR fluorescence imaging of specific targets.

  17. Fluorescence Imaging Study of Impinging Underexpanded Jets

    Science.gov (United States)

    Inman, Jennifer A.; Danehy, Paul M.; Nowak, Robert J.; Alderfer, David W.

    2008-01-01

    An experiment was designed to create a simplified simulation of the flow through a hole in the surface of a hypersonic aerospace vehicle and the subsequent impingement of the flow on internal structures. In addition to planar laser-induced fluorescence (PLIF) flow visualization, pressure measurements were recorded on the surface of an impingement target. The PLIF images themselves provide quantitative spatial information about structure of the impinging jets. The images also help in the interpretation of impingement surface pressure profiles by highlighting the flow structures corresponding to distinctive features of these pressure profiles. The shape of the pressure distribution along the impingement surface was found to be double-peaked in cases with a sufficiently high jet-exit-to-ambient pressure ratio so as to have a Mach disk, as well as in cases where a flow feature called a recirculation bubble formed at the impingement surface. The formation of a recirculation bubble was in turn found to depend very sensitively upon the jet-exit-to-ambient pressure ratio. The pressure measured at the surface was typically less than half the nozzle plenum pressure at low jet pressure ratios and decreased with increasing jet pressure ratios. Angled impingement cases showed that impingement at a 60deg angle resulted in up to a factor of three increase in maximum pressure at the plate compared to normal incidence.

  18. Ultrasmall near-infrared gold nanoclusters for tumor fluorescence imaging in vivo

    Science.gov (United States)

    Wu, Xu; He, Xiaoxiao; Wang, Kemin; Xie, Can; Zhou, Bing; Qing, Zhihe

    2010-10-01

    In this paper, we explore the possibility of using ultrasmall near-infrared (NIR) gold nanoclusters (AuNCs) as novel contrast imaging agents for tumor fluorescence imaging in vivo. The fluorescence imaging signal of the tail vein administrated AuNCs in living organisms can spectrally be well distinguished from the background with maximum emission wavelength at about 710 nm, and the high photostability of AuNCs promises continuous imaging in vivo. The uptake of AuNCs by the reticuloendothelial system is relatively low in comparison with other nanoparticle-based contrast imaging agents due to their ultrasmall hydrodynamic size (~2.7 nm). Through the body weight change analysis, the results show that the body weight of the mice administrated with AuNCs has not been changed obviously in comparison with that of the control mice injected with PBS. Furthermore, using MDA-MB-45 and Hela tumor xenograft models, in vivo and ex vivo imaging studies show that the ultrasmall NIR AuNCs are able to be highly accumulated in the tumor areas, thanks to the enhanced permeability and retention (EPR) effects. And the tumor-to-background ratio is about 15 for 6 h postinjection. The results indicate that the ultrasmall NIR AuNCs appear as very promising contrast imaging agents for in vivo fluorescence tumor imaging.

  19. Clinical application of several tumor imaging agents

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Neoplasms is one of the main diseases for harming health.It is difficult to prevent the neoplasms because the factors of bringing out them are complex.To raise survival rate the early diagnosis of tumors is very important.Radionuclide imaging is useful to detect recurrent or residual diseaseand to identificate benign or malignant tumor.Several tumorimaging agents as following have clinical significance indiagnosing tumors.

  20. Phase-sensitive fluorescent imaging with coherent reconstruction

    CERN Document Server

    Field, Jeffrey J; Bartels, Randy A

    2015-01-01

    Optical imaging plays a critical role in advancing our understanding of three dimensional dynamics of biological systems. Coherent imaging (CI) methods exploit spatial phase information, encoded through propagation of coherent signal light emerging from a specimen, to extract a three-dimensional representation of the object from a single high-speed measurement. Until now, CI methods could not be applied to incoherent light, severely limiting their ability to image the most powerful biological probes available - fluorescent molecules - with sufficient speed and volume to observe important processes, such as neural processing in live specimens. We introduce a new imaging technique that transfers the spatial propagation phase of coherent illumination light to incoherent fluorescent light emission. The transfer of propagation phase allows CI techniques to be applied to fluorescent light imaging, and leads to large increases in imaging speed and depth of field. With this advance, biological imaging of fluorescent ...

  1. Ultrasound contrast agents for ultrasound molecular imaging.

    Science.gov (United States)

    Tranquart, F; Arditi, M; Bettinger, T; Frinking, P; Hyvelin, J M; Nunn, A; Pochon, S; Tardy, I

    2014-11-01

    Ultrasound is a real-time imaging technique which is widely used in many clinical applications for its capacity to provide anatomic information with high spatial and temporal resolution. The advent of ultrasound contrast agents in combination with contrast-specific imaging modes has given access to perfusion assessments at an organ level, leading to an improved diagnostic accuracy. More recently, the development of biologically-targeted ultrasound contrast agents has expanded the role of ultrasound even further into molecular imaging applications. Ultrasound molecular imaging can be used to visualize the expression of intravascular markers, and to assess their local presence over time and/or during therapeutic treatment. Major applications are in the field of inflammation and neoangiogenesis due to the strictly intravascular presence of microbubbles. Various technologies have been investigated for attaching the targeting moiety to the shell from simple biotin-avidin constructs to more elaborated insertion within the shell through attachment to PEG residues. This important improvement has allowed a clinical translation of initial pre-clinical investigations, opening the way for an early detection and an accurate characterization of lesions in patients. The combination of anatomic, functional and molecular information/data provided by contrast ultrasound is a powerful tool which is still in its infancy due to the lack of agents suitable for clinical use. The advantages of ultrasound techniques combined with the molecular signature of lesions will represent a significant advance in imaging in the field of personalized medicine. © Georg Thieme Verlag KG Stuttgart · New York.

  2. Preparation and Characterization of Highly Fluorescent, Glutathione-coated Near Infrared Quantum Dots for in Vivo Fluorescence Imaging

    Directory of Open Access Journals (Sweden)

    Yoshichika Yoshioka

    2008-10-01

    Full Text Available Fluorescent probes that emit in the near-infrared (NIR, 700-1,300 nm region are suitable as optical contrast agents for in vivo fluorescence imaging because of low scattering and absorption of the NIR light in tissues. Recently, NIR quantum dots (QDs have become a new class of fluorescent materials that can be used for in vivo imaging. Compared with traditional organic fluorescent dyes, QDs have several unique advantages such as size- and composition-tunable emission, high brightness, narrow emission bands, large Stokes shifts, and high resistance to photobleaching. In this paper, we report a facile method for the preparation of highly fluorescent, water-soluble glutathione (GSH-coated NIR QDs for in vivo imaging. GSH-coated NIR QDs (GSH-QDs were prepared by surface modification of hydrophobic CdSeTe/CdS (core/shell QDs. The hydrophobic surface of the CdSeTe/CdS QDs was exchanged with GSH in tetrahydrofuran-water. The resulting GSH-QDs were monodisperse particles and stable in PBS (phosphate buffered saline, pH = 7.4. The GSH-QDs (800 nm emission were highly fluorescent in aqueous solutions (quantum yield = 22% in PBS buffer, and their hydrodynamic diameter was less than 10 nm, which is comparable to the size of proteins. The cellular uptake and viability for the GSH-QDs were examined using HeLa and HEK 293 cells. When the cells were incubated with aqueous solutions of the GSH-QDs (10 nM, the QDs were taken into the cells and distributed in the perinuclear region of both cells. After 12 hrs incubation of 4 nM of GSH-QDs, the viabilities of HeLa and HEK 293 cells were ca. 80 and 50%, respectively. As a biomedical utility of the GSH-QDs, in vivo NIRfluorescence imaging of a lymph node in a mouse is presented.

  3. Multispectral fluorescence imaging techniques for nondestructive food safety inspection

    Science.gov (United States)

    Kim, Moon S.; Lefcourt, Alan M.; Chen, Yud-Ren

    2004-03-01

    The use of spectral sensing has gained acceptance as a rapid means for nondestructive inspection of postharvest food produce. Current technologies generally use color or a single wavelength camera technology. The applicability and sensitivity of these techniques can be expanded through the use of multiple wavelengths. Reflectance in the Vis/NIR is the prevalent spectral technique. Fluorescence, compared to reflectance, is regarded as a more sensitive technique due to its dynamic responses to subtle changes in biological entities. Our laboratory has been exploring fluorescence as a potential means for detection of quality and wholesomeness of food products. Applications of fluorescence sensing require an understanding of the spectral characteristics emanating from constituents and potential contaminants. A number of factors affecting fluorescence emission characteristics are discussed. Because of relatively low fluorescence quantum yield from biological samples, a system with a powerful pulse light source such as a laser coupled with a gated detection device is used to harvest fluorescence, in the presence of ambient light. Several fluorescence sensor platforms developed in our laboratory, including hyperspectral imaging, and laser-induced fluorescence (LIF) and steady-state fluorescence imaging systems with multispectral capabilities are presented. We demonstrate the potential uses of recently developed fluorescence imaging platforms in food safety inspection of apples contaminated with animal feces.

  4. Advances in Magnetic Resonance Imaging Contrast Agents for Biomarker Detection

    Science.gov (United States)

    Sinharay, Sanhita; Pagel, Mark D.

    2016-06-01

    Recent advances in magnetic resonance imaging (MRI) contrast agents have provided new capabilities for biomarker detection through molecular imaging. MRI contrast agents based on the T2 exchange mechanism have more recently expanded the armamentarium of agents for molecular imaging. Compared with T1 and T2* agents, T2 exchange agents have a slower chemical exchange rate, which improves the ability to design these MRI contrast agents with greater specificity for detecting the intended biomarker. MRI contrast agents that are detected through chemical exchange saturation transfer (CEST) have even slower chemical exchange rates. Another emerging class of MRI contrast agents uses hyperpolarized 13C to detect the agent with outstanding sensitivity. These hyperpolarized 13C agents can be used to track metabolism and monitor characteristics of the tissue microenvironment. Together, these various MRI contrast agents provide excellent opportunities to develop molecular imaging for biomarker detection.

  5. Fluorescence microscopy studies of a peripheral-benzodiazepine-receptor-targeted molecular probe for brain tumor imaging

    Science.gov (United States)

    Marcu, Laura; Vernier, P. Thomas; Manning, H. Charles; Salemi, Sarah; Li, Aimin; Craft, Cheryl M.; Gundersen, Martin A.; Bornhop, Darryl J.

    2003-10-01

    This study investigates the potential of a new multi-modal lanthanide chelate complex for specifically targeting brain tumor cells. We report here results from ongoing studies of up-take, sub-cellular localization and binding specificity of this new molecular imaging probe. Fluorescence microscopy investigations in living rat C6 glioma tumor cells demonstrate that the new imaging agent has affinity for glioma cells and binds to mitochondria.

  6. Functional imaging in bulk tissue specimens using optical emission tomography: fluorescence preservation during optical clearing

    Science.gov (United States)

    Sakhalkar, H. S.; Dewhirst, M.; Oliver, T.; Cao, Y.; Oldham, M.

    2007-04-01

    Optical emission computed tomography (optical-ECT) is a technique for imaging the three-dimensional (3D) distribution of fluorescent probes in biological tissue specimens with high contrast and spatial resolution. In optical-ECT, functional information can be imaged by (i) systemic application of functional labels (e.g. fluorophore labelled proteins) and/or (ii) endogenous expression of fluorescent reporter proteins (e.g. red fluorescent protein (RFP), green fluorescent protein (GFP)) in vivo. An essential prerequisite for optical-ECT is optical clearing, a procedure where tissue specimens are made transparent to light by sequential perfusion with fixing, dehydrating and clearing agents. In this study, we investigate clearing protocols involving a selection of common fixing (4% buffered paraformaldehyde (PFA), methanol and ethanol), dehydrating (methanol and ethanol) and clearing agents (methyl salicylate and benzyl-alcohol-benzyl-benzoate (BABB)) in order to determine a 'fluorescence friendly' clearing procedure. Cell culture experiments were employed to optimize the sequence of chemical treatments that best preserve fluorescence. Texas red (TxRed), fluorescein isothiocyanate (FITC), RFP and GFP were tested as fluorophores and fluorescent reporter proteins of interest. Fluorescent and control cells were imaged on a microscope using a DSred2 and FITC filter set. The most promising clearing protocols of cell culture experiments were applied to whole xenograft tumour specimens, to test their effectiveness in large unsectioned samples. Fluorescence of TxRed/FITC fluorophores was not found to be significantly affected by any of the test clearing protocols. RFP and GFP fluorescence, however, was found to be significantly greater when cell fixation was in ethanol. Fixation in either PFA or methanol resulted in diminished fluorescence. After ethanol fixation, the RFP and GFP fluorescence proved remarkably robust to subsequent exposure to either methyl salicylate or BABB

  7. Detection of coalescing agents in water-borne latex emulsions using an environment sensitive fluorescent probe

    NARCIS (Netherlands)

    Raja, T.N.; Brouwer, A.M.; Biemans, K.; Nabuurs, T.; Tennebroek, R.

    2010-01-01

    In this paper we report the determination of partitioning of coalescing agents (organic co-solvents) in water-borne latex emulsions by means of a fluorescence method. An environment-sensitive fluorescent probe 1 was copolymerized via emulsion polymerization. The presence of organic co-solvents insid

  8. Synthesis of fluorescent and low cytotoxicity phenol formaldehyde resin (PFR)@Ag composites for cell imaging and antibacterial activity.

    Science.gov (United States)

    Yang, Ping; Dong, Hao; Xia, Jun; Xu, Andong; Shi, Jianjun; He, Jie; Ding, Jianzhong; Li, Dewei

    2015-12-01

    Ag nanoparticles (NPs) were loaded onto the surface of phenol formaldehyde resin (PFR) NPs without any reducing agent. The as-synthesized PFR@Ag composites have low cytotoxicity, which makes them promising antibacterial agents. Furthermore, the good fluorescence of PFR could be used for cell imaging.

  9. Silica micro/nanospheres for theranostics: from bimodal MRI and fluorescent imaging probes to cancer therapy

    Directory of Open Access Journals (Sweden)

    Shanka Walia

    2015-02-01

    Full Text Available Nano-theranostics offer remarkable potential for future biomedical technology with simultaneous applications for diagnosis and therapy of disease sites. Through smart and careful chemical modifications of the nanoparticle surface, these can be converted to multifunctional tiny objects which in turn can be used as vehicle for delivering multimodal imaging agents and therapeutic material to specific target sites in vivo. In this sense, bimodal imaging probes that simultaneously enable magnetic resonance imaging and fluorescence imaging have gained tremendous attention because disease sites can be characterized quick and precisely through synergistic multimodal imaging. But such hybrid nanocomposite materials have limitations such as low chemical stability (magnetic component and harsh cytotoxic effects (fluorescent component and, hence, require a biocompatible protecting agent. Silica micro/nanospheres have shown promise as protecting agent due to the high stability and low toxicity. This review will cover a full description of MRI-active and fluorescent multifunctional silica micro/nanospheres including the design of the probe, different characterization methods and their application in imaging and treatment in cancer.

  10. Refractive Index Sensing of Green Fluorescent Proteins in Living Cells Using Fluorescence Lifetime Imaging Microscopy

    NARCIS (Netherlands)

    Manen, van Henk-Jan; Verkuijlen, Paul; Wittendorp, Paul; Subramaniam, Vinod; Berg, van den Timo K.; Roos, Dirk; Otto, Cees

    2008-01-01

    We show that fluorescence lifetime imaging microscopy (FLIM) of green fluorescent protein (GFP) molecules in cells can be used to report on the local refractive index of intracellular GFP. We expressed GFP fusion constructs of Rac2 and gp91phox, which are both subunits of the phagocyte NADPH oxidase

  11. Multimodality Imaging Probe for Positron Emission Tomography and Fluorescence Imaging Studies

    Directory of Open Access Journals (Sweden)

    Suresh K. Pandey

    2014-05-01

    Full Text Available Our goal is to develop multimodality imaging agents for use in cell tracking studies by positron emission tomography (PET and optical imaging (OI. For this purpose, bovine serum albumin (BSA was complexed with biotin (histologic studies, 5(6- carboxyfluorescein, succinimidyl ester (FAM SE (OI studies, and diethylenetriamine pentaacetic acid (DTPA for chelating gallium 68 (PET studies. For synthesis of BSA-biotin-FAM-DTPA, BSA was coupled to (+-biotin N-hydroxysuccinimide ester (biotin-NHSI. BSA- biotin was treated with DTPA-anhydride and biotin-BSA-DTPA was reacted with FAM. The biotin-BSA-DTPA-FAM was reacted with gallium chloride 3 to 5 mCi eluted from the generator using 0.1 N HCl and was passed through basic resin (AG 11 A8 and 150 mCi (100 μL, pH 7–8 was incubated with 0.1 mg of FAM conjugate (100 μL at room temperature for 15 minutes to give 66Ga-BSA-biotin-DTPA-FAM. A shaved C57 black mouse was injected with FAM conjugate (50 μL at one flank and FAM-68Ga (50 μL, 30 mCi at the other. Immediately after injection, the mouse was placed in a fluorescence imaging system (Kodak In-Vivo F, Bruker Biospin Co., Woodbridge, CT and imaged (Λex: 465 nm, Λem: 535 nm, time: 8 seconds, Xenon Light Source, Kodak. The same mouse was then placed under an Inveon microPET scanner (Siemens Medical Solutions, Knoxville, TN injected (intravenously with 25 μCi of 18F and after a half-hour (to allow sufficient bone uptake was imaged for 30 minutes. Molecular weight determined using matrix-associated laser desorption ionization (MALDI for the BSA sample was 66,485 Da and for biotin-BSA was 67,116 Da, indicating two biotin moieties per BSA molecule; for biotin-BSA-DTPA was 81,584 Da, indicating an average of 30 DTPA moieties per BSA molecule; and for FAM conjugate was 82,383 Da, indicating an average of 1.7 fluorescent moieties per BSA molecule. Fluorescence imaging clearly showed localization of FAM conjugate and FAM-68Ga at respective flanks of the mouse

  12. Intraoperative fluorescence imaging for personalized brain tumor resection: Current state and future directions

    Directory of Open Access Journals (Sweden)

    Evgenii Belykh

    2016-10-01

    Full Text Available Introduction: Fluorescence-guided surgery is one of the rapidly emerging methods of surgical theranostics. In this review, we summarize current fluorescence techniques used in neurosurgical practice for brain tumor patients, as well as future applications of recent laboratory and translational studies.Methods: Review of the literature.Results: A wide spectrum of fluorophores that have been tested for brain surgery is reviewed. Beginning with a fluorescein sodium application in 1948 by Moore, fluorescence guided brain tumor surgery is either routinely applied in some centers or is under active study in clinical trials. Besides the trinity of commonly used drugs (fluorescein sodium, 5-ALA and ICG, less studied fluorescent stains, such as tetracyclines, cancer-selective alkylphosphocholine analogs, cresyl violet, acridine orange, and acriflavine can be used for rapid tumor detection and pathological tissue examination. Other emerging agents such as activity-based probes and targeted molecular probes that can provide biomolecular specificity for surgical visualization and treatment are reviewed. Furthermore, we review available engineering and optical solutions for fluorescent surgical visualization. Instruments for fluorescent-guided surgery are divided into wide-field imaging systems and hand-held probes. Recent advancements in quantitative fluorescence-guided surgery are discussed.Conclusion: We are standing on the doorstep of the era of marker-assisted tumor management. Innovations in the fields of surgical optics, computer image analysis, and molecular bioengineering are advancing fluorescence-guided tumor resection paradigms, leading to cell-level approaches to visualization and resection of brain tumors.

  13. Fluorescent metal nanoshell and CK19 detection on single cell image

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Jian, E-mail: jian@cfs.biomet.umaryland.edu [Center for Fluorescence Spectroscopy, University of Maryland School of Medicine, Department of Biochemistry and Molecular Biology, 725 West Lombard Street, Baltimore, MD 21201 (United States); Fu, Yi [Center for Fluorescence Spectroscopy, University of Maryland School of Medicine, Department of Biochemistry and Molecular Biology, 725 West Lombard Street, Baltimore, MD 21201 (United States); Li, Ge [Division of Molecular Pathology, Department of Pathology, University of Maryland School of Medicine, 10 South Pine Street, Baltimore, MD 21201 (United States); Lakowicz, Joseph R. [Center for Fluorescence Spectroscopy, University of Maryland School of Medicine, Department of Biochemistry and Molecular Biology, 725 West Lombard Street, Baltimore, MD 21201 (United States); Zhao, Richard Y., E-mail: rzhao@som.umaryland.edu [Division of Molecular Pathology, Department of Pathology, University of Maryland School of Medicine, 10 South Pine Street, Baltimore, MD 21201 (United States); Department of Microbiology-Immunology, University of Maryland School of Medicine, 10 South Pine Street, Baltimore, MD 21201 (United States); Institute of Human Virology, University of Maryland School of Medicine, 10 South Pine Street, Baltimore, MD 21201 (United States)

    2011-09-16

    Highlights: {yields} Novel metal nanoshell as fluorescence imaging agent. {yields} Fluorescent mAb-metal complex with enhanced intensity and shortened lifetime. {yields} Immuno-interactions of mAb-metal complexes with CK19 molecules on CNCAP and HeLa cell surfaces. {yields} Isolation of conjugated mAb-metal complexes from cellular autofluorescence on cell image. -- Abstract: In this article, we report the synthesis strategy and optical properties of a novel type of fluorescence metal nanoshell when it was used as imaging agent for fluorescence cell imaging. The metal nanoshells were made with 40 nm silica cores and 10 nm silver shells. Unlike typical fluorescence metal nanoshells which contain the organic dyes in the cores, novel metal nanoshells were composed of Cy5-labelled monoclonal anti-CK19 antibodies (mAbs) on the external surfaces of shells. Optical measurements to the single nanoparticles showed that in comparison with the metal free labelled mAbs, the mAb-Ag complexes displayed significantly enhanced emission intensity and dramatically shortened lifetime due to near-field interactions of fluorophores with metal. These metal nanoshells were found to be able to immunoreact with target cytokeratin 19 (CK19) molecules on the surfaces of LNCAP and HeLa cells. Fluorescence cell images were recorded on a time-resolved confocal microscope. The emissions from the metal nanoprobes could be clearly isolated from the cellular autofluorescence backgrounds on the cell images as either individuals or small clusters due to their stronger emission intensities and shorter lifetimes. These emission signals could also be precisely counted on single cell images. The count number may provide an approach for quantifying the target molecules in the cells.

  14. Zinc phthalocyanine labelled polyethylene glycol: preparation, characterization, interaction with bovine serum albumin and near infrared fluorescence imaging in vivo.

    Science.gov (United States)

    Lv, Feng; Cao, Bo; Cui, Yanli; Liu, Tianjun

    2012-05-25

    Zinc phthalocyanine labelled polyethylene glycol was prepared to track and monitor the in vivo fate of polyethylene glycol. The chemical structures were characterized by nuclear magnetic resonance and infrared spectroscopy. Their light stability and fluorescence quantum yield were evaluated by UV-Visible and fluorescence spectroscopy methods. The interaction of zinc phthalocyanine labelled polyethylene glycol with bovine serum albumin was evaluated by fluorescence titration and isothermal titration calorimetry methods. Optical imaging in vivo, organ aggregation as well as distribution of fluorescence experiments for tracking polyethylene glycol were performed with zinc phthalocyanine labelled polyethylene glycol as fluorescent agent. Results show that zinc phthalocyanine labelled polyethylene glycol has good optical stability and high emission ability in the near infrared region. Imaging results demonstrate that zinc phthalocyanine labelled polyethylene glycol can track and monitor the in vivo process by near infrared fluorescence imaging, which implies its potential in biomaterials evaluation in vivo by a real-time noninvasive method.

  15. Clinical multi-colour fluorescence imaging of malignant tumours - initial experience

    Energy Technology Data Exchange (ETDEWEB)

    Svanberg, K.; Wang, I. [Lund Univ. Hospital (Sweden). Lund Medical Laser Centre]|[Lund Univ. Hospital (Sweden). Dept. of Oncology; Colleen, S. [Lund Univ. Hospital (Sweden). Lund Medical Laser Centre]|[Lund Univ. Hospital (Sweden). Dept. of Urology; Idvall, I. [Lund Univ. Hospital (Sweden). Lund Medical Laser Centre]|[Lund Univ. Hospital (Sweden). Dept. of Pathology; Ingvar, C. [Lund Univ. Hospital (Sweden). Lund Medical Laser Centre]|[Lund Univ. Hospital (Sweden). Dept. of Surgery; Rydell, R. [Lund Univ. Hospital (Sweden). Lund Medical Laser Centre]|[Lund Univ. Hospital (Sweden). Dept. of Oto-Rhino-Laryngology; Jocham, D. [Luebeck Univ. (Germany). Abt. fuer Urologie; Diddens, H. [Luebeck Univ. (Germany). Medizinisches Laser Zentrum; Bown, S.; Gregory, G. [National Medical Laser Centre, Dept. of Surgery, Rayne Inst., London (United Kingdom); Montan, S. [Spectraphos AB, Ideon, Lund (Sweden); Andersson-Engels, S.; Svanberg, S. [Lund Univ. Hospital (Sweden). Lund Medical Laser Centre]|[Lund Inst. of Technology (Sweden). Dept. of Physics

    1998-01-01

    The purpose of this study was to present a new technique for non-invasive tumour detection based on tissue fluorescence imaging. A clinically adapted multi-colour fluorescence system was employed in the real-time imaging of malignant tumours of the skin, breast, head and neck region, and urinary bladder. Tumour detection was based on the contrast displayed in fluorescence between normal and malignant tissue, related to the selective uptake of tumour-marking agents and natural chromophore differences between various tissues. In order to demarcate basal cell carcinomas of the skin, ALA was applied topically 4-6 h before the fluorescence investigation. For urinary bladder tumour visualisation, ALA was instilled into the bladder 1-2 h prior to the study. Malignant and premalignant lesions in the head and neck region were imaged after i.v. injection of HPD (Photofrin). The tumour imaging system was coupled to an endoscope. Fluorescence light emission from the tissue surface was induced with 100-ns-long optical pulses at 390 nm, generated from a frequency-doubled alexandrite laser. With the use of special image-splitting optics, the tumour fluorescence, intensified in a micro-channel plate, was imaged in 3 selected wavelength bands. These 3 images were processed together to form a new optimised-contrast image of the tumour. This image, updated at a rate of about 3 frames/s was mixed with a normal colour video image of the tissue. A clear demarcation from normal surrounding tissue was found during in vivo measurements of superficial bladder carcinoma, basal cell carcinoma of the skin, and leukoplakia with dysplasia of the lip, and in vitro investigations of resected breast cancer. (orig./MG).

  16. Hyperspectral imaging fluorescence excitation scanning for colon cancer detection

    Science.gov (United States)

    Leavesley, Silas J.; Walters, Mikayla; Lopez, Carmen; Baker, Thomas; Favreau, Peter F.; Rich, Thomas C.; Rider, Paul F.; Boudreaux, Carole W.

    2016-10-01

    Optical spectroscopy and hyperspectral imaging have shown the potential to discriminate between cancerous and noncancerous tissue with high sensitivity and specificity. However, to date, these techniques have not been effectively translated to real-time endoscope platforms. Hyperspectral imaging of the fluorescence excitation spectrum represents new technology that may be well suited for endoscopic implementation. However, the feasibility of detecting differences between normal and cancerous mucosa using fluorescence excitation-scanning hyperspectral imaging has not been evaluated. The goal of this study was to evaluate the initial feasibility of using fluorescence excitation-scanning hyperspectral imaging for measuring changes in fluorescence excitation spectrum concurrent with colonic adenocarcinoma using a small pre-pilot-scale sample size. Ex vivo analysis was performed using resected pairs of colorectal adenocarcinoma and normal mucosa. Adenocarcinoma was confirmed by histologic evaluation of hematoxylin and eosin (H&E) permanent sections. Specimens were imaged using a custom hyperspectral imaging fluorescence excitation-scanning microscope system. Results demonstrated consistent spectral differences between normal and cancerous tissues over the fluorescence excitation range of 390 to 450 nm that could be the basis for wavelength-dependent detection of colorectal cancers. Hence, excitation-scanning hyperspectral imaging may offer an alternative approach for discriminating adenocarcinoma from surrounding normal colonic mucosa, but further studies will be required to evaluate the accuracy of this approach using a larger patient cohort.

  17. Research progress of magnetic resonance imaging contrast agents

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Magnetic resonance imaging (MRI) is a clinical diagnostic modality, which has become popular in hospitals around the world. Approximately 30% of MRI exams include the use of contrast agents. The research progress of the paramagnetic resonance imaging contrast agents was described briefly. Three important approaches in the soluble paramagnetic resonance imaging contrast agents design including nonionic, tissue-specific and macromolecular contrast agents were investigated. In addition, the problems in the research and development in future were discussed.

  18. A framework for creating realistic synthetic fluorescence microscopy image sequences

    CSIR Research Space (South Africa)

    Mabaso, M

    2016-02-01

    Full Text Available of the 9th International Joint Conference on Biomedical Engineering Systems and Technologies, Rome, Italy. 21-23 February, 2016 A Framework for Creating Realistic Synthetic Fluorescence Microscopy Image Sequences Matsilele Mabaso1, Daniel Withey1...

  19. PARPi-FL - a Fluorescent PARP1 Inhibitor for Glioblastoma Imaging

    Directory of Open Access Journals (Sweden)

    Christopher P. Irwin

    2014-05-01

    Full Text Available New intravital optical imaging technologies have revolutionized our understanding of mammalian biology and continue to evolve rapidly. However, there are only a limited number of imaging probes available to date. In this study, we investigated in mouse models of glioblastoma whether a fluorescent small molecule inhibitor of the DNA repair enzyme PARP1, PARPi-FL, can be used as an imaging agent to detect glioblastomas in vivo. We demonstrated that PARPi-FL has appropriate biophysical properties, low toxicity at concentrations used for imaging, high stability in vivo, and accumulates selectively in glioblastomas due to high PARP1 expression. Importantly, subcutaneous and orthotopic glioblastoma xenografts were imaged with high contrast clearly defining tumor tissue from normal surrounding tissue. This research represents a step toward exploring and developing PARPi-FL as an optical intraoperative imaging agent for PARP1 in the clinic.

  20. Timing and Operating Mode Design for Time-Gated Fluorescence Lifetime Imaging Microscopy

    OpenAIRE

    Chao Liu; Xinwei Wang; Yan Zhou; Yuliang Liu

    2013-01-01

    Steady-state fluorence imaging and time-resolved fluorescence imaging are two important areas in fluorescence imaging research. Fluorescence lifetime imaging is an absolute measurement method which is independent of excitation laser intensity, fluorophore concentration, and photobleaching compared to fluorescence intensity imaging techniques. Time-gated fluorescence lifetime imaging microscopy (FLIM) can provide high resolution and high imaging frame during mature FLIM methods. An abstract ti...

  1. Image processing for drift compensation in fluorescence microscopy

    DEFF Research Database (Denmark)

    Petersen, Steffen; Thiagarajan, Viruthachalam; Coutinho, Isabel

    2013-01-01

    Fluorescence microscopy is characterized by low background noise, thus a fluorescent object appears as an area of high signal/noise. Thermal gradients may result in apparent motion of the object, leading to a blurred image. Here, we have developed an image processing methodology that may remove....../reduce blur significantly for any type of microscopy. A total of ~100 images were acquired with a pixel size of 30 nm. The acquisition time for each image was approximately 1second. We can quantity the drift in X and Y using the sub pixel accuracy computed centroid location of an image object in each frame....... We can measure drifts down to approximately 10 nm in size and a drift-compensated image can therefore be reconstructed on a grid of the same size using the “Shift and Add” approach leading to an image of identical size asthe individual image. We have also reconstructed the image using a 3 fold larger...

  2. Fluorogen-based reporters for fluorescence imaging: a review

    Science.gov (United States)

    Jullien, Ludovic; Gautier, Arnaud

    2015-12-01

    Fluorescence bioimaging has recently jumped into a new area of spatiotemporal resolution and sensitivity thanks to synergistic advances in both optical physics and probe/biosensor design. This review focuses on the recent development of genetically encodable fluorescent reporters that bind endogenously present or exogenously applied fluorogenic chromophores (so-called fluorogens) and activate their fluorescence. We highlight the innovative engineering and design that gave rise to these new natural and synthetic fluorescent reporters, and describe some of the emerging applications in imaging and biosensing.

  3. Fluorescence Image Analyzer - FLIMA: software for quantitative analysis of fluorescence in situ hybridization.

    Science.gov (United States)

    Silva, H C M; Martins-Júnior, M M C; Ribeiro, L B; Matoso, D A

    2017-03-30

    The Fluorescence Image Analyzer (FLIMA) software was developed for the quantitative analysis of images generated by fluorescence in situ hybridization (FISH). Currently, the images of FISH are examined without a coefficient that enables a comparison between them. Through GD Graphics Library, the FLIMA software calculates the amount of pixels on image and recognizes each present color. The coefficient generated by the algorithm shows the percentage of marks (probes) hybridized on the chromosomes. This software can be used for any type of image generated by a fluorescence microscope and is able to quantify digoxigenin probes exhibiting a red color, biotin probes exhibiting a green color, and double-FISH probes (digoxigenin and biotin used together), where the white color is displayed.

  4. Optical imaging as an expansion of nuclear medicine: Cerenkov-based luminescence vs fluorescence-based luminescence

    Energy Technology Data Exchange (ETDEWEB)

    Chin, Patrick T.K.; Welling, Mick M.; Leeuwen, Fijs W.B. van [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology, P.O. Box 9600, Leiden (Netherlands); Meskers, Stefan C.J. [Eindhoven University of Technology, Molecular Materials and Nanosystems, P.O. Box 513, Eindhoven (Netherlands); Valdes Olmos, Renato A. [Leiden University Medical Center, Interventional Molecular Imaging Laboratory, Department of Radiology, P.O. Box 9600, Leiden (Netherlands); Netherlands Cancer Institute-Antoni van Leeuwenhoek Hospital, Department of Nuclear Medicine, Amsterdam (Netherlands); Tanke, Hans [Leiden University Medical Center, Department of Molecular Cell Biology, P.O. Box 9600, Leiden (Netherlands)

    2013-08-15

    Integration of optical imaging technologies can further strengthen the field of radioguided surgery. Rather than using two separate chemical entities to achieve this extension, hybrid imaging agents can be used that contain both radionuclear and optical properties. Two types of such hybrid imaging agents are available: (1) hybrid imaging agents generated by Cerenkov luminescence imaging (CLI) of {beta}-emitters and (2) hybrid imaging agents that contain both a radioactive moiety and a fluorescent dye. One major challenge clinicians are now facing is to determine the potential value of these approaches. With this tutorial review we intend to clarify the differences between the two approaches and highlight the clinical potential of hybrid imaging during image-guided surgery applications. (orig.)

  5. Use of fluorescence lifetime imaging (FLIM) for latent fingerprints detection

    Science.gov (United States)

    Wang, Peng; Chao, Zhi Xia; Seah, Leong K.; Murukeshan, Vadakke M.

    2005-04-01

    Fluorescence lifetime imaging (FLIM) in frequency domain enables the mapping of the spatial distribution of fluorescence lifetimes of a specimen. FLIM can provide unique information about fluorophores and hence is widely used in biology and for medical diagnostics. In this paper, a theoretical analysis for the fluorescence lifetime determination of latent fingerprint samples is described, which is followed by the feasibility study of using FLIM in frequency domain for latent fingerprints detection. Experiments are carried out with fingerprint on green paper substrate and postcard substrate treated with certain fluorescent powder. The total phase lag and demodulation factor are calculated to determine the lifetimes pixel by pixel. The resulting fluorescence lifetime image of fingerprint revealed an improvement in the contrast, and was able to detect the latent fingerprint clearly.

  6. Fluorescence polarization imaging for delineating nonmelanoma skin cancers

    Science.gov (United States)

    Yaroslavsky, A. N.; Neel, V.; Anderson, R. R.

    2004-09-01

    We present a method for detecting nonmelanoma skin cancers using exogenous fluorescence polarization. We built an automated system that permits exogenous fluorescence polarization imaging. It includes a tunable linearly polarized monochromatic light source and a CCD camera equipped with a rotating linear polarizer and a filter to reject excitation light. Two fluorophores that are retained in tumors, toluidine blue and methylene blue, are employed. We demonstrate that fluorescence polarization imaging can be used for accurate delineation of nonmelanoma cancers. The results suggest that this optical technique may be suitable for real-time noninvasive demarcation of epithelial cancers.

  7. Noninvasive measurement of pharmacokinetics by near-infrared fluorescence imaging in the eye of mice

    Science.gov (United States)

    Dobosz, Michael; Strobel, Steffen; Stubenrauch, Kay-Gunnar; Osl, Franz; Scheuer, Werner

    2014-01-01

    Purpose: For generating preclinical pharmacokinetics (PKs) of compounds, blood is drawn at different time points and levels are quantified by different analytical methods. In order to receive statistically meaningful data, 3 to 5 animals are used for each time point to get serum peak-level and half-life of the compound. Both characteristics are determined by data interpolation, which may influence the accuracy of these values. We provide a method that allows continuous monitoring of blood levels noninvasively by measuring the fluorescence intensity of labeled compounds in the eye and other body regions of anesthetized mice. Procedures: The method evaluation was performed with four different fluorescent compounds: (i) indocyanine green, a nontargeting dye; (ii) OsteoSense750, a bone targeting agent; (iii) tumor targeting Trastuzumab-Alexa750; and (iv) its F(-alxea750 fragment. The latter was used for a direct comparison between fluorescence imaging and classical blood analysis using enzyme-linked immunosorbent assay (ELISA). Results: We found an excellent correlation between blood levels measured by noninvasive eye imaging with the results generated by classical methods. A strong correlation between eye imaging and ELISA was demonstrated for the F( fragment. Whole body imaging revealed a compound accumulation in the expected regions (e.g., liver, bone). Conclusions: The combination of eye and whole body fluorescence imaging enables the simultaneous measurement of blood PKs and biodistribution of fluorescent-labeled compounds.

  8. Chlorophyll Fluorescence Imaging Uncovers Photosynthetic Fingerprint of Citrus Huanglongbing

    Directory of Open Access Journals (Sweden)

    Haiyan Cen

    2017-08-01

    Full Text Available Huanglongbing (HLB is one of the most destructive diseases of citrus, which has posed a serious threat to the global citrus production. This research was aimed to explore the use of chlorophyll fluorescence imaging combined with feature selection to characterize and detect the HLB disease. Chlorophyll fluorescence images of citrus leaf samples were measured by an in-house chlorophyll fluorescence imaging system. The commonly used chlorophyll fluorescence parameters provided the first screening of HLB disease. To further explore the photosynthetic fingerprint of HLB infected leaves, three feature selection methods combined with the supervised classifiers were employed to identify the unique fluorescence signature of HLB and perform the three-class classification (i.e., healthy, HLB infected, and nutrient deficient leaves. Unlike the commonly used fluorescence parameters, this novel data-driven approach by using the combination of the mean fluorescence parameters and image features gave the best classification performance with the accuracy of 97%, and presented a better interpretation for the spatial heterogeneity of photochemical and non-photochemical components in HLB infected citrus leaves. These results imply the potential of the proposed approach for the citrus HLB disease diagnosis, and also provide a valuable insight for the photosynthetic response to the HLB disease.

  9. Chlorophyll Fluorescence Imaging Uncovers Photosynthetic Fingerprint of Citrus Huanglongbing.

    Science.gov (United States)

    Cen, Haiyan; Weng, Haiyong; Yao, Jieni; He, Mubin; Lv, Jingwen; Hua, Shijia; Li, Hongye; He, Yong

    2017-01-01

    Huanglongbing (HLB) is one of the most destructive diseases of citrus, which has posed a serious threat to the global citrus production. This research was aimed to explore the use of chlorophyll fluorescence imaging combined with feature selection to characterize and detect the HLB disease. Chlorophyll fluorescence images of citrus leaf samples were measured by an in-house chlorophyll fluorescence imaging system. The commonly used chlorophyll fluorescence parameters provided the first screening of HLB disease. To further explore the photosynthetic fingerprint of HLB infected leaves, three feature selection methods combined with the supervised classifiers were employed to identify the unique fluorescence signature of HLB and perform the three-class classification (i.e., healthy, HLB infected, and nutrient deficient leaves). Unlike the commonly used fluorescence parameters, this novel data-driven approach by using the combination of the mean fluorescence parameters and image features gave the best classification performance with the accuracy of 97%, and presented a better interpretation for the spatial heterogeneity of photochemical and non-photochemical components in HLB infected citrus leaves. These results imply the potential of the proposed approach for the citrus HLB disease diagnosis, and also provide a valuable insight for the photosynthetic response to the HLB disease.

  10. Near-infrared fluorescent probes in cancer imaging and therapy: an emerging field

    Directory of Open Access Journals (Sweden)

    Yi XM

    2014-03-01

    Full Text Available Xiaomin Yi, Fuli Wang, Weijun Qin, Xiaojian Yang, Jianlin Yuan Department of Urology, Xijing Hospital, Fourth Military Medical University, Xi'an, Shaanxi, People's Republic of China Abstract: Near-infrared fluorescence (NIRF imaging is an attractive modality for early cancer detection with high sensitivity and multi-detection capability. Due to convenient modification by conjugating with moieties of interests, NIRF probes are ideal candidates for cancer targeted imaging. Additionally, the combinatory application of NIRF imaging and other imaging modalities that can delineate anatomical structures extends fluorometric determination of biomedical information. Moreover, nanoparticles loaded with NIRF dyes and anticancer agents contribute to the synergistic management of cancer, which integrates the advantage of imaging and therapeutic functions to achieve the ultimate goal of simultaneous diagnosis and treatment. Appropriate probe design with targeting moieties can retain the original properties of NIRF and pharmacokinetics. In recent years, great efforts have been made to develop new NIRF probes with better photostability and strong fluorescence emission, leading to the discovery of numerous novel NIRF probes with fine photophysical properties. Some of these probes exhibit tumoricidal activities upon light radiation, which holds great promise in photothermal therapy, photodynamic therapy, and photoimmunotherapy. This review aims to provide a timely and concise update on emerging NIRF dyes and multifunctional agents. Their potential uses as agents for cancer specific imaging, lymph node mapping, and therapeutics are included. Recent advances of NIRF dyes in clinical use are also summarized. Keywords: near infrared dyes, nanoparticles, imaging, cancer targeting, cancer therapy

  11. Multiplexed Spectral Imaging of 120 Different Fluorescent Labels.

    Directory of Open Access Journals (Sweden)

    Alex M Valm

    Full Text Available The number of fluorescent labels that can unambiguously be distinguished in a single image when acquired through band pass filters is severely limited by the spectral overlap of available fluorophores. The recent development of spectral microscopy and the application of linear unmixing algorithms to spectrally recorded image data have allowed simultaneous imaging of fluorophores with highly overlapping spectra. However, the number of distinguishable fluorophores is still limited by the unavoidable decrease in signal to noise ratio when fluorescence signals are fractionated over multiple wavelength bins. Here we present a spectral image analysis algorithm to greatly expand the number of distinguishable objects labeled with binary combinations of fluorophores. Our algorithm utilizes a priori knowledge about labeled specimens and imposes a binary label constraint on the unmixing solution. We have applied our labeling and analysis strategy to identify microbes labeled by fluorescence in situ hybridization and here demonstrate the ability to distinguish 120 differently labeled microbes in a single image.

  12. Photobleaching correction in fluorescence microscopy images

    Energy Technology Data Exchange (ETDEWEB)

    Vicente, Nathalie B; Diaz Zamboni, Javier E; Adur, Javier F; Paravani, Enrique V; Casco, Victor H [Microscopy Laboratory, School of Engineering - Bioengineering, National University of Entre Rios (UNER), Ruta 11, Km 10 (3101), Oro Verde, Entre Rios (Argentina)

    2007-11-15

    Fluorophores are used to detect molecular expression by highly specific antigen-antibody reactions in fluorescence microscopy techniques. A portion of the fluorophore emits fluorescence when irradiated with electromagnetic waves of particular wavelengths, enabling its detection. Photobleaching irreversibly destroys fluorophores stimulated by radiation within the excitation spectrum, thus eliminating potentially useful information. Since this process may not be completely prevented, techniques have been developed to slow it down or to correct resulting alterations (mainly, the decrease in fluorescent signal). In the present work, the correction by photobleaching curve was studied using E-cadherin (a cell-cell adhesion molecule) expression in Bufo arenarum embryos. Significant improvements were observed when applying this simple, inexpensive and fast technique.

  13. DNA as sensors and imaging agents for metal ions.

    Science.gov (United States)

    Xiang, Yu; Lu, Yi

    2014-02-17

    Increasing interest in detecting metal ions in many chemical and biomedical fields has created demands for developing sensors and imaging agents for metal ions with high sensitivity and selectivity. This review covers recent progress in DNA-based sensors and imaging agents for metal ions. Through both combinatorial selection and rational design, a number of metal-ion-dependent DNAzymes and metal-ion-binding DNA structures that can selectively recognize specific metal ions have been obtained. By attachment of these DNA molecules with signal reporters such as fluorophores, chromophores, electrochemical tags, and Raman tags, a number of DNA-based sensors for both diamagnetic and paramagnetic metal ions have been developed for fluorescent, colorimetric, electrochemical, and surface Raman detection. These sensors are highly sensitive (with a detection limit down to 11 ppt) and selective (with selectivity up to millions-fold) toward specific metal ions. In addition, through further development to simplify the operation, such as the use of "dipstick tests", portable fluorometers, computer-readable disks, and widely available glucose meters, these sensors have been applied for on-site and real-time environmental monitoring and point-of-care medical diagnostics. The use of these sensors for in situ cellular imaging has also been reported. The generality of the combinatorial selection to obtain DNAzymes for almost any metal ion in any oxidation state and the ease of modification of the DNA with different signal reporters make DNA an emerging and promising class of molecules for metal-ion sensing and imaging in many fields of applications.

  14. Wide field-of-view fluorescence imaging of coral reefs.

    Science.gov (United States)

    Treibitz, Tali; Neal, Benjamin P; Kline, David I; Beijbom, Oscar; Roberts, Paul L D; Mitchell, B Greg; Kriegman, David

    2015-01-13

    Coral reefs globally are declining rapidly because of both local and global stressors. Improved monitoring tools are urgently needed to understand the changes that are occurring at appropriate temporal and spatial scales. Coral fluorescence imaging tools have the potential to improve both ecological and physiological assessments. Although fluorescence imaging is regularly used for laboratory studies of corals, it has not yet been used for large-scale in situ assessments. Current obstacles to effective underwater fluorescence surveying include limited field-of-view due to low camera sensitivity, the need for nighttime deployment because of ambient light contamination, and the need for custom multispectral narrow band imaging systems to separate the signal into meaningful fluorescence bands. Here we describe the Fluorescence Imaging System (FluorIS), based on a consumer camera modified for greatly increased sensitivity to chlorophyll-a fluorescence, and we show high spectral correlation between acquired images and in situ spectrometer measurements. This system greatly facilitates underwater wide field-of-view fluorophore surveying during both night and day, and potentially enables improvements in semi-automated segmentation of live corals in coral reef photographs and juvenile coral surveys.

  15. Optical imaging of breast tumors and of gastrointestinal cancer by laser-induced fluorescence.

    Science.gov (United States)

    Ebert, Bernd; Grosenick, Dirk

    2013-01-01

    Optical imaging offers a high potential for noninvasive detection of cancer in humans. Recent advances in instrumentation for diffuse optical imaging have led to new capabilities for the detection of cancer in highly scattering tissue such as the female breast. We review recent developments in the detection of breast cancer in humans by fluorescent contrast agents. So far, the unspecific contrast agents indocyanine green (ICG) and omocyanine have been applied, whereas molecular probes for direct targeted imaging of this disease are still in preclinical research. We discuss recent improvements in the differentiation of malignant and benign lesions with ICG based on its enhanced extravasation in breast cancer. Whereas fluorescence imaging in thick tissue layers is hampered by strong light scattering, tissue surfaces can be investigated with high spatial resolution. As an example for superficial tumors, lesions of the gastrointestinal tract (GI) are discussed. In these investigations, protoporphyrin IX is used as a tumor-specific (due to its strong enhancement in tumor cells) target for spectroscopic identification and imaging. We present a time-gated method for fluorescence imaging and spectroscopy with strong suppression of tissue autofluorescence and show results on patients with Barrett's esophagus and with colitis ulcerosa.

  16. Intraoperative Near-infrared Imaging for Parathyroid Gland Identification by Auto-fluorescence: A Feasibility Study.

    Science.gov (United States)

    De Leeuw, Frederic; Breuskin, Ingrid; Abbaci, Muriel; Casiraghi, Odile; Mirghani, Haïtham; Ben Lakhdar, Aïcha; Laplace-Builhé, Corinne; Hartl, Dana

    2016-09-01

    Parathyroid glands (PGs) can be particularly hard to distinguish from surrounding tissue and thus can be damaged or removed during thyroidectomy. Postoperative hypoparathyroidism is the most common complication after thyroidectomy. Very recently, it has been found that the parathyroid tissue shows near-infrared (NIR) auto-fluorescence which could be used for intraoperative detection, without any use of contrast agents. The work described here presents a histological validation ex vivo of the NIR imaging procedure and evaluates intraoperative PG detection by NIR auto-fluorescence using for the first time to our knowledge a commercially available clinical NIR imaging device. Ex vivo study on resected operative specimens combined with a prospective in vivo study of consecutive patients who underwent total or partial thyroid, or parathyroid surgery at a comprehensive cancer center. During surgery, any tissue suspected to be a potential PG by the surgeon was imaged with the Fluobeam 800 (®) system. NIR imaging was compared to conventional histology (ex vivo) and/or visual identification by the surgeon (in vivo). We have validated NIR auto-fluorescence with an ex vivo study including 28 specimens. Sensitivity and specificity were 94.1 and 80 %, respectively. Intraoperative NIR imaging was performed in 35 patients and 81 parathyroids were identified. In 80/81 cases, the fluorescence signal was subjectively obvious on real-time visualization. We determined that PG fluorescence is 2.93 ± 1.59 times greater than thyroid fluorescence in vivo. Real-time NIR imaging based on parathyroid auto-fluorescence is fast, safe, and non-invasive and shows very encouraging results, for intraoperative parathyroid identification.

  17. Dual-Modal Nanoprobes for Imaging of Mesenchymal Stem Cell Transplant by MRI and Fluorescence Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Sung, Chang Kyu; Hong, Kyung Ah; Lin, Shun Mei [Seoul Metropolitan Boramae Medical Center, Seoul (Korea, Republic of)] (and others)

    2009-12-15

    To determine the feasibility of labeling human mesenchymal stem cells (hMSCs) with bifunctional nanoparticles and assessing their potential as imaging probes in the monitoring of hMSC transplantation. The T1 and T2 relaxivities of the nanoparticles (MNP SiO{sub 2}[RITC]-PEG) were measured at 1.5T and 3T magnetic resonance scanner. Using hMSCs and the nanoparticles, labeling efficiency, toxicity, and proliferation were assessed. Confocal laser scanning microscopy and transmission electron microscopy were used to specify the intracellular localization of the endocytosed iron nanoparticles. We also observed in vitro and in vivo visualization of the labeled hMSCs with a 3T MR scanner and optical imaging. MNP SiO{sub 2}(RITC)-PEG showed both superparamagnetic and fluorescent properties. The r{sub 1} and r{sub 2} relaxivity values of the MNP SiO{sub 2}(RITC)-PEG were 0.33 and 398 mM{sup -1} s{sup -1} at 1.5T, respectively, and 0.29 and 453 mM{sup -1} s{sup -1} at 3T, respectively. The effective internalization of MNP SiO{sub 2}(RITC)-PEG into hMSCs was observed by confocal laser scanning fluorescence microscopy. The transmission electron microscopy images showed that MNP SiO{sub 2}(RITC)-PEG was internalized into the cells and mainly resided in the cytoplasm. The viability and proliferation of MNP SiO{sub 2}(RITC)-PEG-labeled hMSCs were not significantly different from the control cells. MNP SiO{sub 2}(RITC)-PEG-labeled hMSCs were observed in vitro and in vivo with optical and MR imaging. MNP SiO{sub 2}(RITC)-PEG can be a useful contrast agent for stem cell imaging, which is suitable for a bimodal detection by MRI and optical imaging.

  18. Advance of Molecular Imaging Technology and Targeted Imaging Agent in Imaging and Therapy

    Directory of Open Access Journals (Sweden)

    Zhi-Yi Chen

    2014-01-01

    Full Text Available Molecular imaging is an emerging field that integrates advanced imaging technology with cellular and molecular biology. It can realize noninvasive and real time visualization, measurement of physiological or pathological process in the living organism at the cellular and molecular level, providing an effective method of information acquiring for diagnosis, therapy, and drug development and evaluating treatment of efficacy. Molecular imaging requires high resolution and high sensitive instruments and specific imaging agents that link the imaging signal with molecular event. Recently, the application of new emerging chemical technology and nanotechnology has stimulated the development of imaging agents. Nanoparticles modified with small molecule, peptide, antibody, and aptamer have been extensively applied for preclinical studies. Therapeutic drug or gene is incorporated into nanoparticles to construct multifunctional imaging agents which allow for theranostic applications. In this review, we will discuss the characteristics of molecular imaging, the novel imaging agent including targeted imaging agent and multifunctional imaging agent, as well as cite some examples of their application in molecular imaging and therapy.

  19. Double agents and secret agents: the emerging fields of exogenous chemical exchange saturation transfer and T2-exchange magnetic resonance imaging contrast agents for molecular imaging.

    Science.gov (United States)

    Daryaei, Iman; Pagel, Mark D

    2015-01-01

    Two relatively new types of exogenous magnetic resonance imaging contrast agents may provide greater impact for molecular imaging by providing greater specificity for detecting molecular imaging biomarkers. Exogenous chemical exchange saturation transfer (CEST) agents rely on the selective saturation of the magnetization of a proton on an agent, followed by chemical exchange of a proton from the agent to water. The selective detection of a biomarker-responsive CEST signal and an unresponsive CEST signal, followed by the ratiometric comparison of these signals, can improve biomarker specificity. We refer to this improvement as a "double-agent" approach to molecular imaging. Exogenous T2-exchange agents also rely on chemical exchange of protons between the agent and water, especially with an intermediate rate that lies between the slow exchange rates of CEST agents and the fast exchange rates of traditional T1 and T2 agents. Because of this intermediate exchange rate, these agents have been relatively unknown and have acted as "secret agents" in the contrast agent research field. This review exposes these secret agents and describes the merits of double agents through examples of exogenous agents that detect enzyme activity, nucleic acids and gene expression, metabolites, ions, redox state, temperature, and pH. Future directions are also provided for improving both types of contrast agents for improved molecular imaging and clinical translation. Therefore, this review provides an overview of two new types of exogenous contrast agents that are becoming useful tools within the armamentarium of molecular imaging.

  20. Photoacoustic tomography of human hepatic malignancies using intraoperative indocyanine green fluorescence imaging.

    Directory of Open Access Journals (Sweden)

    Akinori Miyata

    Full Text Available Recently, fluorescence imaging following the preoperative intravenous injection of indocyanine green has been used in clinical settings to identify hepatic malignancies during surgery. The aim of this study was to evaluate the ability of photoacoustic tomography using indocyanine green as a contrast agent to produce representative fluorescence images of hepatic tumors by visualizing the spatial distribution of indocyanine green on ultrasonographic images. Indocyanine green (0.5 mg/kg, intravenous was preoperatively administered to 9 patients undergoing hepatectomy. Intraoperatively, photoacoustic tomography was performed on the surface of the resected hepatic specimens (n = 10 under excitation with an 800 nm pulse laser. In 4 hepatocellular carcinoma nodules, photoacoustic imaging identified indocyanine green accumulation in the cancerous tissue. In contrast, in one hepatocellular carcinoma nodule and five adenocarcinoma foci (one intrahepatic cholangiocarcinoma and 4 colorectal liver metastases, photoacoustic imaging delineated indocyanine green accumulation not in the cancerous tissue but rather in the peri-cancerous hepatic parenchyma. Although photoacoustic tomography enabled to visualize spatial distribution of ICG on ultrasonographic images, which was consistent with fluorescence images on cut surfaces of the resected specimens, photoacoustic signals of ICG-containing tissues decreased approximately by 40% even at 4 mm depth from liver surfaces. Photoacoustic tomography using indocyanine green also failed to identify any hepatocellular carcinoma nodules from the body surface of model mice with non-alcoholic steatohepatitis. In conclusion, photoacoustic tomography has a potential to enhance cancer detectability and differential diagnosis by ultrasonographic examinations and intraoperative fluorescence imaging through visualization of stasis of bile-excreting imaging agents in and/or around hepatic tumors. However, further technical

  1. Photoacoustic tomography of human hepatic malignancies using intraoperative indocyanine green fluorescence imaging.

    Science.gov (United States)

    Miyata, Akinori; Ishizawa, Takeaki; Kamiya, Mako; Shimizu, Atsushi; Kaneko, Junichi; Ijichi, Hideaki; Shibahara, Junji; Fukayama, Masashi; Midorikawa, Yutaka; Urano, Yasuteru; Kokudo, Norihiro

    2014-01-01

    Recently, fluorescence imaging following the preoperative intravenous injection of indocyanine green has been used in clinical settings to identify hepatic malignancies during surgery. The aim of this study was to evaluate the ability of photoacoustic tomography using indocyanine green as a contrast agent to produce representative fluorescence images of hepatic tumors by visualizing the spatial distribution of indocyanine green on ultrasonographic images. Indocyanine green (0.5 mg/kg, intravenous) was preoperatively administered to 9 patients undergoing hepatectomy. Intraoperatively, photoacoustic tomography was performed on the surface of the resected hepatic specimens (n = 10) under excitation with an 800 nm pulse laser. In 4 hepatocellular carcinoma nodules, photoacoustic imaging identified indocyanine green accumulation in the cancerous tissue. In contrast, in one hepatocellular carcinoma nodule and five adenocarcinoma foci (one intrahepatic cholangiocarcinoma and 4 colorectal liver metastases), photoacoustic imaging delineated indocyanine green accumulation not in the cancerous tissue but rather in the peri-cancerous hepatic parenchyma. Although photoacoustic tomography enabled to visualize spatial distribution of ICG on ultrasonographic images, which was consistent with fluorescence images on cut surfaces of the resected specimens, photoacoustic signals of ICG-containing tissues decreased approximately by 40% even at 4 mm depth from liver surfaces. Photoacoustic tomography using indocyanine green also failed to identify any hepatocellular carcinoma nodules from the body surface of model mice with non-alcoholic steatohepatitis. In conclusion, photoacoustic tomography has a potential to enhance cancer detectability and differential diagnosis by ultrasonographic examinations and intraoperative fluorescence imaging through visualization of stasis of bile-excreting imaging agents in and/or around hepatic tumors. However, further technical advances are needed

  2. Tumor Targeting and Pharmacokinetics of a Near-Infrared Fluorescent-Labeled δ-Opioid Receptor Antagonist Agent, Dmt-Tic-Cy5.

    Science.gov (United States)

    Huynh, Amanda Shanks; Estrella, Veronica; Stark, Valerie E; Cohen, Allison S; Chen, Tingan; Casagni, Todd J; Josan, Jatinder S; Lloyd, Mark C; Johnson, Joseph; Kim, Jongphil; Hruby, Victor J; Vagner, Josef; Morse, David L

    2016-02-01

    Fluorescence molecular imaging can be employed for the development of novel cancer targeting agents. Herein, we investigated the pharmacokinetics (PK) and cellular uptake of Dmt-Tic-Cy5, a delta-opioid receptor (δOR) antagonist-fluorescent dye conjugate, as a tumor-targeting molecular imaging agent. δOR expression is observed normally in the CNS, and pathologically in some tumors, including lung liver and breast cancers. In vitro, in vivo, and ex vivo experiments were conducted to image and quantify the fluorescence signal associated with Dmt-Tic-Cy5 over time using in vitro and intravital fluorescence microscopy and small animal fluorescence imaging of tumor-bearing mice. We observed specific retention of Dmt-Tic-Cy5 in tumors with maximum uptake in δOR-expressing positive tumors at 3 h and observable persistence for >96 h; clearance from δOR nonexpressing negative tumors by 6 h; and systemic clearance from normal organs by 24 h. Live-cell and intravital fluorescence microscopy demonstrated that Dmt-Tic-Cy5 had sustained cell-surface binding lasting at least 24 h with gradual internalization over the initial 6 h following administration. Dmt-Tic-Cy5 is a δOR-targeted agent that exhibits long-lasting and specific signal in δOR-expressing tumors, is rapidly cleared from systemic circulation, and is not retained in non-δOR-expressing tissues. Hence, Dmt-Tic-Cy5 has potential as a fluorescent tumor imaging agent.

  3. Fluorescence and Bioluminescence Imaging of Orthotopic Brain Tumors in Mice.

    Science.gov (United States)

    McKinnon, Emilie; Moore, Alfred; Dixit, Suraj; Zhu, Yun; Broome, Ann-Marie

    2017-01-01

    Optical imaging strategies, such as fluorescence and bioluminescence imaging, are non-invasive, in vivo whole body imaging techniques utilized to study cancer. Optical imaging is widely used in preclinical work because of its ease of use and cost-friendliness. It also provides the opportunity to study animals and biological responses longitudinally over time. Important considerations include depth of tissue penetration, photon scattering, absorption and the choice of light emitting probe, all of which affect the resolution (image quality and data information) and the signal to noise ratio of the image. We describe how to use bioluminescence and fluorescence imaging to track a chemotherapeutic delivery nanocarrier conjugated with a fluorophore to determine its localization in vivo.

  4. Near-infrared fluorescent probes in cancer imaging and therapy: an emerging field.

    Science.gov (United States)

    Yi, Xiaomin; Wang, Fuli; Qin, Weijun; Yang, Xiaojian; Yuan, Jianlin

    2014-01-01

    Near-infrared fluorescence (NIRF) imaging is an attractive modality for early cancer detection with high sensitivity and multi-detection capability. Due to convenient modification by conjugating with moieties of interests, NIRF probes are ideal candidates for cancer targeted imaging. Additionally, the combinatory application of NIRF imaging and other imaging modalities that can delineate anatomical structures extends fluorometric determination of biomedical information. Moreover, nanoparticles loaded with NIRF dyes and anticancer agents contribute to the synergistic management of cancer, which integrates the advantage of imaging and therapeutic functions to achieve the ultimate goal of simultaneous diagnosis and treatment. Appropriate probe design with targeting moieties can retain the original properties of NIRF and pharmacokinetics. In recent years, great efforts have been made to develop new NIRF probes with better photostability and strong fluorescence emission, leading to the discovery of numerous novel NIRF probes with fine photophysical properties. Some of these probes exhibit tumoricidal activities upon light radiation, which holds great promise in photothermal therapy, photodynamic therapy, and photoimmunotherapy. This review aims to provide a timely and concise update on emerging NIRF dyes and multifunctional agents. Their potential uses as agents for cancer specific imaging, lymph node mapping, and therapeutics are included. Recent advances of NIRF dyes in clinical use are also summarized.

  5. A dual oxygenation and fluorescence imaging platform for reconstructive surgery

    Science.gov (United States)

    Ashitate, Yoshitomo; Nguyen, John N.; Venugopal, Vivek; Stockdale, Alan; Neacsu, Florin; Kettenring, Frank; Lee, Bernard T.; Frangioni, John V.; Gioux, Sylvain

    2013-03-01

    There is a pressing clinical need to provide image guidance during surgery. Currently, assessment of tissue that needs to be resected or avoided is performed subjectively, leading to a large number of failures, patient morbidity, and increased healthcare costs. Because near-infrared (NIR) optical imaging is safe, noncontact, inexpensive, and can provide relatively deep information (several mm), it offers unparalleled capabilities for providing image guidance during surgery. These capabilities are well illustrated through the clinical translation of fluorescence imaging during oncologic surgery. In this work, we introduce a novel imaging platform that combines two complementary NIR optical modalities: oxygenation imaging and fluorescence imaging. We validated this platform during facial reconstructive surgery on large animals approaching the size of humans. We demonstrate that NIR fluorescence imaging provides identification of perforator arteries, assesses arterial perfusion, and can detect thrombosis, while oxygenation imaging permits the passive monitoring of tissue vital status, as well as the detection and origin of vascular compromise simultaneously. Together, the two methods provide a comprehensive approach to identifying problems and intervening in real time during surgery before irreparable damage occurs. Taken together, this novel platform provides fully integrated and clinically friendly endogenous and exogenous NIR optical imaging for improved image-guided intervention during surgery.

  6. Carbon dots of different composition and surface functionalization: cytotoxicity issues relevant to fluorescence cell imaging.

    Science.gov (United States)

    Wang, Yanli; Anilkumar, Parambath; Cao, Li; Liu, Jia-Hui; Luo, Pengju G; Tackett, Kenneth N; Sahu, Sushant; Wang, Ping; Wang, Xin; Sun, Ya-Ping

    2011-11-01

    Nanoscale carbon particles have emerged as versatile precursors for a new class of highly fluorescent nanomaterials that resemble semiconductor quantum dots. The surface-passivated fluorescent carbon nanoparticles, dubbed 'carbon dots', were already demonstrated for their potential optical bioimaging applications in vitro and in vivo. In this study, we conducted a systematic cytotoxicity evaluation on the carbon dots prepared by various combinations of precursor carbon nanoparticles and molecules for the particle surface functionalization. The results suggested that the cytotoxicity of carbon dots was dependent on the selection of surface passivation molecules. Those dots showing more significant cytotoxicity at higher concentrations were also evaluated for their effects on the fluorescence imaging of live cells. The implications of the results on the eventual use of carbon dots as cell imaging agents are discussed.

  7. Facile Synthesis of Biocompatible Fluorescent Nanoparticles for Cellular Imaging and Targeted Detection of Cancer Cells.

    Science.gov (United States)

    Tang, Fu; Wang, Chun; Wang, Xiaoyu; Li, Lidong

    2015-11-18

    In this work, we report the facile synthesis of functional core-shell structured nanoparticles with fluorescence enhancement, which show specific targeting of cancer cells. Biopolymer poly-l-lysine was used to coat the silver core with various shell thicknesses. Then, the nanoparticles were functionalized with folic acid as a targeting agent for folic acid receptor. The metal-enhanced fluorescence effect was observed when the fluorophore (5-(and-6)-carboxyfluorescein-succinimidyl ester) was conjugated to the modified nanoparticle surface. Cellular imaging assay of the nanoparticles in folic acid receptor-positive cancer cells showed their excellent biocompatibility and selectivity. The as-prepared functional nanoparticles demonstrate the efficiency of the metal-enhanced fluorescence effect and provide an alternative approach for the cellular imaging and targeting of cancer cells.

  8. Fluorescence lifetime imaging of oxygen in dental biofilm

    Science.gov (United States)

    Gerritsen, Hans C.; de Grauw, Cees J.

    2000-12-01

    Dental biofilm consists of micro-colonies of bacteria embedded in a matrix of polysaccharides and salivary proteins. pH and oxygen concentration are of great importance in dental biofilm. Both can be measured using fluorescence techniques. The imaging of dental biofilm is complicated by the thickness of the biofilms that can be up to several hundred micrometers thick. Here, we employed a combination of two-photon excitation microscopy with fluorescence lifetime imaging to quantify the oxygen concentration in dental biofilm. Collisional quenching of fluorescent probes by molecular oxygen leads to a reduction of the fluorescence lifetime of the probe. We employed this mechanism to measure the oxygen concentration distribution in dental biofilm by means of fluorescence lifetime imaging. Here, TRIS Ruthenium chloride hydrate was used as an oxygen probe. A calibration procedure on buffers was use to measure the lifetime response of this Ruthenium probe. The results are in agreement with the Stern-Volmer equation. A linear relation was found between the ratio of the unquenched and the quenched lifetime and the oxygen concentration. The biofilm fluorescence lifetime imaging results show a strong oxygen gradient at the buffer - biofilm interface and the average oxygen concentration in the biofilm amounted to 50 μM.

  9. The Three-region Method of Color Matching in the Presence of Fluorescent Whitening Agents

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    An expression of reflectance of fluorescent dyes in emissionregion was used in this article. Through the analysis of thesppectral radiance factor of fluorescent whitening agents (FWA), the color matching of FWA was studied and the three-region method of color matching method was put forward. The method was proved to be reliable for the dyes used and was easy to apply for the color matching of FWAs.

  10. Improving Surgical Resection of Metastatic Liver Tumors With Near-Infrared Optical-Guided Fluorescence Imaging.

    Science.gov (United States)

    Barabino, Gabriele; Porcheron, Jack; Cottier, Michèle; Cuilleron, Muriel; Coutard, Jean-Guillaume; Berger, Michel; Molliex, Serge; Beauchesne, Brigitte; Phelip, Jean Marc; Grichine, Alexei; Coll, Jean-Luc

    2016-08-01

    Objective The aim of this study was to investigate the feasibility and future clinical applications of near-infrared (NIR) fluorescence imaging to guide liver resection surgery for metastatic cancer to improve resection margins. Summary Background Data A subset of patients with metastatic hepatic tumors can be cured by surgery. The degree of long-term and disease-free survival is related to the quality of surgery, with the best resection defined as "R0" (complete removal of all tumor cells, as evidenced by microscopic examination of the margins). Although intraoperative ultrasonography can evaluate the surgical margins, surgeons need a new tool to perfect the surgical outcome. Methods A preliminary study was performed on 3 patients. We used NIR imaging postoperatively "ex vivo" on the resected liver tissue. The liver tumors were preoperatively labelled by intravenously injecting the patient with indocyanine green (ICG), a NIR fluorescent agent (24 hours before surgery, 0.25 mg/kg). Fluorescent images were obtained using a miniaturized fluorescence imaging system (FluoStic, Fluoptics, Grenoble, France). Results After liver resection, the surgical specimens from each patient were sliced into 10-mm sections in the operating room and analyzed with the FluoStic. All metastatic tumors presented rim-type fluorescence. Two specimens had incomplete rim fluorescence. The pathologist confirmed the presence of R1 margins (microscopic residual resection), even though the ultrasonographic analysis indicated that the result was R0. Conclusions Surgical liver resection guided by NIR fluorescence can help detect potentially uncertain anatomical areas that may be missed by preoperative imaging and by ultrasonography during surgery. These preliminary results will need to be confirmed in a larger prospective patient series.

  11. Review of fluorescence guided surgery systems: identification of key performance capabilities beyond indocyanine green imaging

    Science.gov (United States)

    DSouza, Alisha V.; Lin, Huiyun; Henderson, Eric R.; Samkoe, Kimberley S.; Pogue, Brian W.

    2016-08-01

    There is growing interest in using fluorescence imaging instruments to guide surgery, and the leading options for open-field imaging are reviewed here. While the clinical fluorescence-guided surgery (FGS) field has been focused predominantly on indocyanine green (ICG) imaging, there is accelerated development of more specific molecular tracers. These agents should help advance new indications for which FGS presents a paradigm shift in how molecular information is provided for resection decisions. There has been a steady growth in commercially marketed FGS systems, each with their own differentiated performance characteristics and specifications. A set of desirable criteria is presented to guide the evaluation of instruments, including: (i) real-time overlay of white-light and fluorescence images, (ii) operation within ambient room lighting, (iii) nanomolar-level sensitivity, (iv) quantitative capabilities, (v) simultaneous multiple fluorophore imaging, and (vi) ergonomic utility for open surgery. In this review, United States Food and Drug Administration 510(k) cleared commercial systems and some leading premarket FGS research systems were evaluated to illustrate the continual increase in this performance feature base. Generally, the systems designed for ICG-only imaging have sufficient sensitivity to ICG, but a fraction of the other desired features listed above, with both lower sensitivity and dynamic range. In comparison, the emerging research systems targeted for use with molecular agents have unique capabilities that will be essential for successful clinical imaging studies with low-concentration agents or where superior rejection of ambient light is needed. There is no perfect imaging system, but the feature differences among them are important differentiators in their utility, as outlined in the data and tables here.

  12. In vivo cellular imaging using fluorescent proteins - Methods and Protocols

    Directory of Open Access Journals (Sweden)

    M. Monti

    2012-12-01

    Full Text Available The discovery and genetic engineering of fluorescent proteins has revolutionized cell biology. What was previously invisible to the cell often can be made visible with the use of fluorescent proteins. With this words, Robert M. Hoffman introduces In vivo Cellular Imaging Using Fluorescent proteins, the eighteen chapters book dedicated to the description of how fluorescence proteins have changed the way to analyze cellular processes in vivo. Modern researches aim to study new and less invasive methods able to follow the behavior of different cell types in different biological contexts: for example, how cancer cells migrate or how they respond to different therapies. Also, in vivo systems can help researchers to better understand animal embryonic development so as how fluorescence proteins may be used to monitor different processes in living organisms at the molecular and cellular level.

  13. Fluorescent Dendrimer Nanoconjugates as Advanced Probes for Biological Imaging

    Science.gov (United States)

    Reilly, Daniel; Kim, Sung Hoon; Katzenellenbogen, John A.; Schroeder, Charles M.

    2014-03-01

    Recent advances in fluorescence microscopy have enabled improvements in spatial resolution for biological imaging. However, there is a strong need for development of advanced fluorescent probes to enable a molecular-scale understanding of biological events. In this work, we report the development of a new class of probes for fluorescence imaging based on dye-conjugated dendrimer nanoconjugates. We utilize molecular-scale dendritic scaffolds as fluorescent probes, thereby enabling conjugation of multiple dyes and linkers to the scaffold periphery. In particular, we use polyamidoamine dendrimers as molecular scaffolds, wherein dye conjugation can be varied over a wide range. Single molecule fluorescence imaging shows that dendrimer nanoconjugates are far brighter than single fluorophores, resulting in increased localization precision. In addition, we further developed a new set of remarkably photostable probes by conjugating photoprotective triplet state quenchers directly onto the dendritic scaffold. We observe large increases in the photobleaching times compared to single dyes and reduced transient dark states (blinking). Overall, we believe that these new probes will allow for single molecule imaging over long time scales, enabling new vistas in biological imaging.

  14. Chitosan oligosaccharide based Gd-DTPA complex as a potential bimodal magnetic resonance imaging contrast agent.

    Science.gov (United States)

    Huang, Yan; Cao, Juan; Zhang, Qi; Lu, Zheng-rong; Hua, Ming-qing; Zhang, Xiao-yan; Gao, Hu

    2016-01-01

    A new gadolinium diethylenetriamine pentaacetic acid (DTPA) complex (Gd-DTPA-DMABA-CS11) as a potential bimodal magnetic resonance imaging (MRI) contrast agent with fluorescence was synthesized. It was synthesized by the incorporation of 4-dimethylaminobenzaldehyde (DMABA) and chitosan oligosaccharide (CSn; n=11) with low polydispersity index to DTPA anhydride and then chelated with gadolinium chloride. The structure was characterized by Fourier transform infrared (FTIR), (1)H NMR, elemental analysis and size exclusion chromatography (SEC). MRI measurements in vitro were evaluated. The results indicated that Gd-DTPA-DMABA-CS11 provided higher molar longitudinal relaxivity (r1) (12.95mM(-1)·s(-1)) than that of commercial Gd-DTPA (3.63mM(-1)·s(-1)) at 0.5T. Gd-DTPA-DMABA-CS11 also emitted fluorescence, and the intensity was much stronger than that of Gd-DTPA. Therefore, it can be meanwhile used in fluorescent imaging for improving the sensitivity in clinic diagnosis. Gd-DTPA-DMABA-CS11 as a potential contrast agent is preliminarily stable in vitro. The results of thermodynamic action between Gd-DTPA-DMABA-CS11 and bovine serum albumin (BSA) illustrated that the binding process was exothermic and spontaneous, and the main force was van der Waals' interaction and hydrogen bond. The preliminary study suggested that Gd-DTPA-DMABA-CS11 could be used in both magnetic resonance and fluorescent imaging as a promising bimodal contrast agent.

  15. Modulated electron-multiplied fluorescence lifetime imaging microscope: all-solid-state camera for fluorescence lifetime imaging

    NARCIS (Netherlands)

    Zhao, Q.; Schelen, B.; Schouten, R., et al.

    2012-01-01

    We have built an all-solid-state camera that is directly modulated at the pixel level for frequency-domain fluorescence lifetime imaging microscopy (FLIM) measurements. This novel camera eliminates the need for an image intensifier through the use of an application-specific charge coupled device des

  16. Enhanced speed in fluorescence imaging using beat frequency multiplexing

    Science.gov (United States)

    Mikami, Hideharu; Kobayashi, Hirofumi; Wang, Yisen; Hamad, Syed; Ozeki, Yasuyuki; Goda, Keisuke

    2016-03-01

    Fluorescence imaging using radiofrequency-tagged emission (FIRE) is an emerging technique that enables higher imaging speed (namely, temporal resolution) in fluorescence microscopy compared to conventional fluorescence imaging techniques such as confocal microscopy and wide-field microscopy. It works based on the principle that it uses multiple intensity-modulated fields in an interferometric setup as excitation fields and applies frequency-division multiplexing to fluorescence signals. Unfortunately, despite its high potential, FIRE has limited imaging speed due to two practical limitations: signal bandwidth and signal detection efficiency. The signal bandwidth is limited by that of an acousto-optic deflector (AOD) employed in the setup, which is typically 100-200 MHz for the spectral range of fluorescence excitation (400-600 nm). The signal detection efficiency is limited by poor spatial mode-matching between two interfering fields to produce a modulated excitation field. Here we present a method to overcome these limitations and thus to achieve higher imaging speed than the prior version of FIRE. Our method achieves an increase in signal bandwidth by a factor of two and nearly optimal mode matching, which enables the imaging speed limited by the lifetime of the target fluorophore rather than the imaging system itself. The higher bandwidth and better signal detection efficiency work synergistically because higher bandwidth requires higher signal levels to avoid the contribution of shot noise and amplifier noise to the fluorescence signal. Due to its unprecedentedly high-speed performance, our method has a wide variety of applications in cancer detection, drug discovery, and regenerative medicine.

  17. Fluorescent metal nanoshell and CK19 detection on single cell image.

    Science.gov (United States)

    Zhang, Jian; Fu, Yi; Li, Ge; Lakowicz, Joseph R; Zhao, Richard Y

    2011-09-16

    In this article, we report the synthesis strategy and optical properties of a novel type of fluorescence metal nanoshell when it was used as imaging agent for fluorescence cell imaging. The metal nanoshells were made with 40 nm silica cores and 10nm silver shells. Unlike typical fluorescence metal nanoshells which contain the organic dyes in the cores, novel metal nanoshells were composed of Cy5-labelled monoclonal anti-CK19 antibodies (mAbs) on the external surfaces of shells. Optical measurements to the single nanoparticles showed that in comparison with the metal free labelled mAbs, the mAb-Ag complexes displayed significantly enhanced emission intensity and dramatically shortened lifetime due to near-field interactions of fluorophores with metal. These metal nanoshells were found to be able to immunoreact with target cytokeratin 19 (CK19) molecules on the surfaces of LNCAP and HeLa cells. Fluorescence cell images were recorded on a time-resolved confocal microscope. The emissions from the metal nanoprobes could be clearly isolated from the cellular autofluorescence backgrounds on the cell images as either individuals or small clusters due to their stronger emission intensities and shorter lifetimes. These emission signals could also be precisely counted on single cell images. The count number may provide an approach for quantifying the target molecules in the cells.

  18. Ultra-high-speed fluorescence imaging

    NARCIS (Netherlands)

    Gelderblom, Erik Carl

    2012-01-01

    For several years, ultrasound contrast agents are being investigated for their therapeutic capacity. These gas-filled coated microbubbles have the ability to enhance local intravenous drug delivery. Microbubbles can be loaded with drugs by adding them to the bubble surface of by incorporating them i

  19. Imaging of a targeted PDT drug with fluorescence tomography

    Science.gov (United States)

    Muffoletto, Dan; Gupta, Anurag; Xu, Zhiqiang; Mahrer, Chris; Bauer, Gretchen; Galas, Scott; Pandey, Ravindra K.; Sunar, Ulas

    2009-02-01

    We constructed a whole-body fluorescence tomography instrument to monitor novel bifunctional phototherapeutic drugs (e.g., HPPH-Cyanine dye conjugate) in small animals. The instrument allows dense source and detector sampling with a fast galvo scanner and a CCD detector for improved resolution and sensitivity (Patwardhan et al., 2005). Here we report tissue phantom measurements to evaluate the imaging performance with a newly constructed tomography instrument. Phantom measurements showed that strong fluorescence generated by HPPH-Cyanine dye (HPPH-CD), having high fluorescence quantum yield and long wavelength fluorescence emission, allowed deep tissue imaging. We also report in vivo fluorescence measurements of the conjugate in Nude mice bearing A549 human non-small cell lung carcinoma (NSCLC) tumors at 24 hr post injection to evaluate tumor detection ability of the conjugate. Our results indicate that the HPPH-CD shows preferential uptake in tumors compared to surrounding normal tissue at 24 hr post injection. This study demonstrates a potential use of HPPH-CD in detection (fluorescence imaging) and treatment (PDT) of deeply seated tumors.

  20. Rapid global fitting of large fluorescence lifetime imaging microscopy datasets.

    Directory of Open Access Journals (Sweden)

    Sean C Warren

    Full Text Available Fluorescence lifetime imaging (FLIM is widely applied to obtain quantitative information from fluorescence signals, particularly using Förster Resonant Energy Transfer (FRET measurements to map, for example, protein-protein interactions. Extracting FRET efficiencies or population fractions typically entails fitting data to complex fluorescence decay models but such experiments are frequently photon constrained, particularly for live cell or in vivo imaging, and this leads to unacceptable errors when analysing data on a pixel-wise basis. Lifetimes and population fractions may, however, be more robustly extracted using global analysis to simultaneously fit the fluorescence decay data of all pixels in an image or dataset to a multi-exponential model under the assumption that the lifetime components are invariant across the image (dataset. This approach is often considered to be prohibitively slow and/or computationally expensive but we present here a computationally efficient global analysis algorithm for the analysis of time-correlated single photon counting (TCSPC or time-gated FLIM data based on variable projection. It makes efficient use of both computer processor and memory resources, requiring less than a minute to analyse time series and multiwell plate datasets with hundreds of FLIM images on standard personal computers. This lifetime analysis takes account of repetitive excitation, including fluorescence photons excited by earlier pulses contributing to the fit, and is able to accommodate time-varying backgrounds and instrument response functions. We demonstrate that this global approach allows us to readily fit time-resolved fluorescence data to complex models including a four-exponential model of a FRET system, for which the FRET efficiencies of the two species of a bi-exponential donor are linked, and polarisation-resolved lifetime data, where a fluorescence intensity and bi-exponential anisotropy decay model is applied to the analysis

  1. Compressive Fluorescence Microscopy for Biological and Hyperspectral Imaging

    CERN Document Server

    Studer, Vincent; Chahid, Makhlad; Moussavi, Hamed; Candes, Emmanuel; Dahan, Maxime

    2012-01-01

    The mathematical theory of compressed sensing (CS) asserts that one can acquire signals from measurements whose rate is much lower than the total bandwidth. Whereas the CS theory is now well developed, challenges concerning hardware implementations of CS-based acquisition devices---especially in optics---have only started being addressed. This paper presents an implementation of compressive sensing in fluorescence microscopy and its applications to biomedical imaging. Our CS microscope combines a dynamic structured wide-field illumination and a fast and sensitive single-point fluorescence detection to enable reconstructions of images of fluorescent beads, cells and tissues with undersampling ratios (between the number of pixels and number of measurements) up to 32. We further demonstrate a hyperspectral mode and record images with 128 spectral channels and undersampling ratios up to 64, illustrating the potential benefits of CS acquisition for higher dimensional signals which typically exhibits extreme redund...

  2. Snapshot imaging Fraunhofer line discriminator for detection of plant fluorescence

    Science.gov (United States)

    Gupta Roy, S.; Kudenov, M. W.

    2015-05-01

    Non-invasive quantification of plant health is traditionally accomplished using reflectance based metrics, such as the normalized difference vegetative index (NDVI). However, measuring plant fluorescence (both active and passive) to determine photochemistry of plants has gained importance. Due to better cost efficiency, lower power requirements, and simpler scanning synchronization, detecting passive fluorescence is preferred over active fluorescence. In this paper, we propose a high speed imaging approach for measuring passive plant fluorescence, within the hydrogen alpha Fraunhofer line at ~656 nm, using a Snapshot Imaging Fraunhofer Line Discriminator (SIFOLD). For the first time, the advantage of snapshot imaging for high throughput Fraunhofer Line Discrimination (FLD) is cultivated by our system, which is based on a multiple-image Fourier transform spectrometer and a spatial heterodyne interferometer (SHI). The SHI is a Sagnac interferometer, which is dispersion compensated using blazed diffraction gratings. We present data and techniques for calibrating the SIFOLD to any particular wavelength. This technique can be applied to quantify plant fluorescence at low cost and reduced complexity of data collection.

  3. Tumor-stem cells interactions by fluorescence imaging

    Science.gov (United States)

    Meleshina, Aleksandra V.; Cherkasova, Elena I.; Sergeeva, Ekaterina; Turchin, Ilya V.; Kiseleva, Ekaterina V.; Dashinimaev, Erdem B.; Shirmanova, Marina V.; Zagaynova, Elena V.

    2013-02-01

    Recently, great deal of interest is investigation the function of the stem cells (SC) in tumors. In this study, we studied «recipient-tumor- fluorescent stem cells » system using the methods of in vivo imaging and laser scanning microscopy (LSM). We used adipose-derived adult stem (ADAS) cells of human lentiviral transfected with the gene of fluorescent protein Turbo FP635. ADAS cells were administrated into nude mice with transplanted tumor HeLa Kyoto (human cervical carcinoma) at different stages of tumor growth (0-8 days) intravenously or into tumor. In vivo imaging was performed on the experimental setup for epi - luminescence bioimaging (IAP RAS, Nizhny Novgorod). The results of the imaging showed localization of fluorophore tagged stem cells in the spleen on day 5-9 after injection. The sensitivity of the technique may be improved by spectral separation autofluorescence and fluorescence of stem cells. We compared the results of in vivo imaging and confocal laser scanning microscopy (LSM 510 META, Carl Zeiss, Germany). Internal organs of the animals and tumor tissue were investigated. It was shown that with i.v. injection of ADAS, bright fluorescent structures with spectral characteristics corresponding to TurboFP635 protein are locally accumulated in the marrow, lungs and tumors of animals. These findings indicate that ADAS cells integrate in the animal body with transplanted tumor and can be identified by fluorescence bioimaging techniques in vivo and ex vivo.

  4. Coded Aperture Imaging for Fluorescent X-rays-Biomedical Applications

    Energy Technology Data Exchange (ETDEWEB)

    Haboub, Abdel; MacDowell, Alastair; Marchesini, Stefano; Parkinson, Dilworth

    2013-06-01

    Employing a coded aperture pattern in front of a charge couple device pixilated detector (CCD) allows for imaging of fluorescent x-rays (6-25KeV) being emitted from samples irradiated with x-rays. Coded apertures encode the angular direction of x-rays and allow for a large Numerical Aperture x- ray imaging system. The algorithm to develop the self-supported coded aperture pattern of the Non Two Holes Touching (NTHT) pattern was developed. The algorithms to reconstruct the x-ray image from the encoded pattern recorded were developed by means of modeling and confirmed by experiments. Samples were irradiated by monochromatic synchrotron x-ray radiation, and fluorescent x-rays from several different test metal samples were imaged through the newly developed coded aperture imaging system. By choice of the exciting energy the different metals were speciated.

  5. Whole mount nuclear fluorescent imaging: convenient documentation of embryo morphology.

    Science.gov (United States)

    Sandell, Lisa L; Kurosaka, Hiroshi; Trainor, Paul A

    2012-11-01

    Here, we describe a relatively inexpensive and easy method to produce high quality images that reveal fine topological details of vertebrate embryonic structures. The method relies on nuclear staining of whole mount embryos in combination with confocal microscopy or conventional wide field fluorescent microscopy. In cases where confocal microscopy is used in combination with whole mount nuclear staining, the resulting embryo images can rival the clarity and resolution of images produced by scanning electron microscopy (SEM). The fluorescent nuclear staining may be performed with a variety of cell permeable nuclear dyes, enabling the technique to be performed with multiple standard microscope/illumination or confocal/laser systems. The method may be used to document morphology of embryos of a variety of organisms, as well as individual organs and tissues. Nuclear stain imaging imposes minimal impact on embryonic specimens, enabling imaged specimens to be utilized for additional assays.

  6. Biodegradable polymer based theranostic agents for photoacoustic imaging and cancer therapy

    Science.gov (United States)

    Wang, Yan J.; Strohm, Eric M.; Kolios, Michael C.

    2016-03-01

    In this study, multifunctional theranostic agents for photoacoustic (PA), ultrasound (US), fluorescent imaging, and for therapeutic drug delivery were developed and tested. These agents consisted of a shell made from a biodegradable Poly(lactide-co-glycolic acid) (PLGA) polymer, loaded with perfluorohexane (PFH) liquid and gold nanoparticles (GNPs) in the core, and lipophilic carbocyanines fluorescent dye DiD and therapeutic drug Paclitaxel (PAC) in the shell. Their multifunctional capacity was investigated in an in vitro study. The PLGA/PFH/DiD-GNPs particles were synthesized by a double emulsion technique. The average PLGA particle diameter was 560 nm, with 50 nm diameter silica-coated gold nano-spheres in the shell. MCF7 human breast cancer cells were incubated with PLGA/PFH/DiDGNPs for 24 hours. Fluorescent and PA images were recorded using a fluorescent/PA microscope using a 1000 MHz transducer and a 532 nm pulsed laser. For the particle vaporization and drug delivery test, MCF7 cells were incubated with the PLGA/PFH-GNPs-PAC or PLGA/PFH-GNPs particles for 6, 12 and 24 hours. The effects of particle vaporization and drug delivery inside the cells were examined by irradiating the cells with a laser fluence of 100 mJ/cm2, and cell viability quantified using the MTT assay. The PA images of MCF7 cells containing PLGA/PFH/DiD-GNPs were spatially coincident with the fluorescent images, and confirmed particle uptake. After exposure to the PLGA/PFHGNP- PAC for 6, 12 and 24 hours, the cell survival rate was 43%, 38%, and 36% respectively compared with the control group, confirming drug delivery and release inside the cells. Upon vaporization, cell viability decreased to 20%. The particles show potential as imaging agents and drug delivery vehicles.

  7. APPLICATION OF MODULATED CHLOROPHYLL FLUORESCENCE AND MODULATED CHLOROPHYLL FLUORESCENCE IMAGING IN STUDYING ENVIRONMENTAL STRESSES EFFECT

    Directory of Open Access Journals (Sweden)

    L. Guidi

    2016-03-01

    Full Text Available Chlorophyll (Chl a fluorescence is a widely used tool to monitor the photosynthetic process in plants subjected to environmental stresses.this review reports the theoretical bases of Chl fluorescence, and the significance of the most important Chl fluorescence parameters. it also reportshow these parameters can be utilised to estimate changes in photosystem ii (PSII photochemistry, linear electron flux and energy dissipationmechanisms. the relation between actual PSII photochemistry and CO2 assimilation is discussed, as is the role of photochemical andnon-photochemical quenching in inducing changes in PSII activity. the application of Chl fluorescence imaging to study heterogeneity on leaflamina is also considered. this review summarises only some of the results obtained by this methodology to study the effects of differentenvironmental stresses, namely water and nutrients availability, pollutants, temperature and salinity.

  8. Color-matched esophagus phantom for fluorescent imaging

    Science.gov (United States)

    Yang, Chenying; Hou, Vivian; Nelson, Leonard Y.; Seibel, Eric J.

    2013-02-01

    We developed a stable, reproducible three-dimensional optical phantom for the evaluation of a wide-field endoscopic molecular imaging system. This phantom mimicked a human esophagus structure with flexibility to demonstrate body movements. At the same time, realistic visual appearance and diffuse spectral reflectance properties of the tissue were simulated by a color matching methodology. A photostable dye-in-polymer technology was applied to represent biomarker probed "hot-spot" locations. Furthermore, fluorescent target quantification of the phantom was demonstrated using a 1.2mm ultrathin scanning fiber endoscope with concurrent fluorescence-reflectance imaging.

  9. Fluorescence Lifetime Imaging of Quantum Dot Labeled DNA Microarrays

    Directory of Open Access Journals (Sweden)

    Jonathan G. Terry

    2009-04-01

    Full Text Available Quantum dot (QD labeling combined with fluorescence lifetime imaging microscopy is proposed as a powerful transduction technique for the detection of DNA hybridization events. Fluorescence lifetime analysis of DNA microarray spots of hybridized QD labeled target indicated a characteristic lifetime value of 18.8 ns, compared to 13.3 ns obtained for spots of free QD solution, revealing that QD labels are sensitive to the spot microenvironment. Additionally, time gated detection was shown to improve the microarray image contrast ratio by 1.8, achieving femtomolar target sensitivity. Finally, lifetime multiplexing based on Qdot525 and Alexa430 was demonstrated using a single excitation-detection readout channel.

  10. Magnetic Resonance Imaging Contrast Agents: A Review of Literature

    Directory of Open Access Journals (Sweden)

    Zahra Sahraei

    2015-10-01

    Full Text Available  Magnetic Resonance Imaging (MRI contrast agents most commonly agents used in diagnosing different diseases. Several agents have been ever introduced with different peculiar characteristics. They vary in potency, adverse reaction and other specification, so it is important to select the proper agent in different situations. We conducted a systematic literature search in MEDLINE/PUBMED, Web of Science (ISI, Scopus,Google Scholar by using keywords "gadolinium" and "MRI contrast Medias", "Gadofosvest", "Gadobenate" and "Gadoxetate". The most frequent contrast media agents made based on gadolinium (Gd. These are divided into two categories based on the structure of their chelating parts, linear agents and macrocyclic agents. All characteristics of contrast media factors, including efficiency, kinetic properties, stability, side effects and the rate of resolution are directly related to the structure of chelating part of that formulation.In vitro data has shown that the macrocyclic compounds are the most stable Gd-CA as they do not bind to serum proteins, they all possess similar and relatively low relaxivity and the prevalence of Nephrogenic Systemic Fibrosis (NSF has decreased by increasing the use of macrocyclic agents in recent years. No cases of NSF have been recorded after the administration of any of the high-relaxivity protein interacting agents, the vascular imaging agent gadofosveset trisodium (Ablavar, the hepatic imaging agent gadoxetate meglumine (Eovist, and the multipurpose agent gadobenate dimeglumine (MultiHance. In pregnancy and lactating women, stable macrocyclic agent is recommended.

  11. 3D Beam Reconstruction by Fluorescence Imaging

    CERN Document Server

    Radwell, Neal; Franke-Arnold, Sonja

    2013-01-01

    We present a technique for mapping the complete 3D spatial intensity profile of a laser beam from its fluorescence in an atomic vapour. We propagate shaped light through a rubidium vapour cell and record the resonant scattering from the side. From a single measurement we obtain a camera limited resolution of 200 x 200 transverse points and 659 longitudinal points. In constrast to invasive methods in which the camera is placed in the beam path, our method is capable of measuring patterns formed by counterpropagating laser beams. It has high resolution in all 3 dimensions, is fast and can be completely automated. The technique has applications in areas which require complex beam shapes, such as optical tweezers, atom trapping and pattern formation.

  12. Small animal optoacoustic tomography system for molecular imaging of contrast agents

    Science.gov (United States)

    Su, Richard; Liopo, Anton; Ermilov, Sergey A.; Oraevsky, Alexander A.

    2016-03-01

    We developed a new and improved Laser Optoacoustic Imaging System, LOIS-3D for preclinical research applications in small animal models. The advancements include (i) a new stabilized imaging module with a more homogeneous illumination of the mouse yielding a better spatial resolution (research applications, such as imaging vascularization and measuring hemoglobin / oxyhemoglobin distribution in the organs as well as imaging exogenous or endogenous optoacoustic contrast agents. As examples, we present in vivo experiments using phantoms and mice with and without tumor injected with contrast agents with indocyanine green (ICG). LOIS-3D was capable of detecting ~1-2 pmole of the ICG, in tissues with relatively low blood content. With its high sensitivity and excellent spatial resolution LOIS-3D is an advanced alternative to fluorescence and bioluminescence based modalities for molecular imaging in live mice.

  13. Quasi-real-time fluorescence imaging with lifetime dependent contrast

    Science.gov (United States)

    Jiang, Pei-Chi; Grundfest, Warren S.; Stafsudd, Oscar M.

    2011-08-01

    Conventional fluorescence lifetime imaging requires complicated algorithms to extract lifetimes of fluorophores and acquisition of multiple data points at progressively longer delay times to characterize tissues. To address diminishing signal-to-noise ratios at these progressively longer time delays, we report a time-resolved fluorescence imaging method, normalized fluorescence yield imaging that does not require the extraction of lifetimes. The concept is to extract the ``contrast'' instead of the lifetime value of the fluorophores by using simple mathematical algorithms. This process converts differences in decay times directly to different intensities. The technique was verified experimentally using a gated iCCD camera and an ultraviolet light-emitting diode light source. It was shown that this method can distinguish between chemical dyes (Fluorescein and Rhodamine-B) and biomedical samples, such as powders of elastin and collagen. Good contrast was obtained between fluorophores that varied by less than 6% in lifetime. Additionally, it was shown that long gate times up to 16 ns achieve good contrast depending upon the samples to be studied. These results support the feasibility of time-resolved fluorescence imaging without lifetime extraction, which has a potential clinical role in noninvasive real-time imaging.

  14. Inherently fluorescent polystyrene microspheres for coating, sensing and cellular imaging.

    Science.gov (United States)

    Qu, Jian-Bo; Xu, Yu-Liang; Liu, Yu; Wang, Yanan; Sui, Yuanhong; Liu, Jian-Guo; Wang, Xiaojuan

    2017-04-01

    Commercially available polystyrene (PS) fluorescent microspheres are widely used in biological field for tracing, in vivo imaging and calibration of flow cytometry, among other applications. However, these particles do suffer from some drawbacks such as the leakage and photobleaching of organic dyes within them. In the present study, inherently fluorescent properties of PS based microspheres have been explored for the first time. Here we find that a simple chloromethylation reaction endows the polystyrene particles with inherent fluorescence without any subsequent conjugation of an external fluorophore. A possible mechanism for fluorescence is elucidated by synthesizing and investigating p-ethylbenzyl chloride, a compound with similar structure. Significantly, no photobleaching or leaking issues were observed owing to the stable structure of the microspheres. Chloromethylated PS (CMPS) microspheres can keep their perpetual blue fluorescence even in dry powder state making them attractive as a potential coating material. Furthermore, the chloromethyl groups on CMPS microspheres make them very convenient for further functionalization. Poly(ethylene glycol) (PEG) grafted microspheres showed good biocompatibility and negligible cytotoxicity, and could be used to image intracellular Fe(3+) due to the selective fluorescence quenching effect of aqueous Fe(3+) in cytoplasm.

  15. Enhanced 3D fluorescence live cell imaging on nanoplasmonic substrate

    Energy Technology Data Exchange (ETDEWEB)

    Gartia, Manas Ranjan [Department of Nuclear, Plasma and Radiological Engineering, University of Illinois, Urbana, IL 61801 (United States); Hsiao, Austin; Logan Liu, G [Department of Bioengineering, University of Illinois, Urbana, IL 61801 (United States); Sivaguru, Mayandi [Institute for Genomic Biology, University of Illinois, Urbana, IL 61801 (United States); Chen Yi, E-mail: loganliu@illinois.edu [Department of Electrical and Computer Engineering, University of Illinois, Urbana, IL 61801 (United States)

    2011-09-07

    We have created a randomly distributed nanocone substrate on silicon coated with silver for surface-plasmon-enhanced fluorescence detection and 3D cell imaging. Optical characterization of the nanocone substrate showed it can support several plasmonic modes (in the 300-800 nm wavelength range) that can be coupled to a fluorophore on the surface of the substrate, which gives rise to the enhanced fluorescence. Spectral analysis suggests that a nanocone substrate can create more excitons and shorter lifetime in the model fluorophore Rhodamine 6G (R6G) due to plasmon resonance energy transfer from the nanocone substrate to the nearby fluorophore. We observed three-dimensional fluorescence enhancement on our substrate shown from the confocal fluorescence imaging of chinese hamster ovary (CHO) cells grown on the substrate. The fluorescence intensity from the fluorophores bound on the cell membrane was amplified more than 100-fold as compared to that on a glass substrate. We believe that strong scattering within the nanostructured area coupled with random scattering inside the cell resulted in the observed three-dimensional enhancement in fluorescence with higher photostability on the substrate surface.

  16. Detection of coalescing agents in water-borne latex emulsions using an environment sensitive fluorescent probe.

    Science.gov (United States)

    Raja, Tanzeela Nazir; Brouwer, Albert M; Biemans, Koen; Nabuurs, Tijs; Tennebroek, Ronald

    2010-07-30

    In this paper we report the determination of partitioning of coalescing agents (organic co-solvents) in water-borne latex emulsions by means of a fluorescence method. An environment-sensitive fluorescent probe was copolymerized via emulsion polymerization. The presence of organic co-solvents inside the polymer particles is revealed by the photophysical properties of the probe. In particular, the position of the fluorescence emission maximum of co-polymerized can be used to measure the amount of coalescing agent present in the polymer particles. The spectral shifts are shown to be due to the softening of the matrix, rather than to solvation of the probe by the added co-solvent.

  17. Doped semiconductor nanocrystal based fluorescent cellular imaging probes.

    Science.gov (United States)

    Maity, Amit Ranjan; Palmal, Sharbari; Basiruddin, S K; Karan, Niladri Sekhar; Sarkar, Suresh; Pradhan, Narayan; Jana, Nikhil R

    2013-06-21

    Doped semiconductor nanocrystals such as Mn doped ZnS, Mn doped ZnSe and Cu doped InZnS, are considered as new classes of fluorescent biological probes with low toxicity. Although the synthesis in high quality of such nanomaterials is now well established, transforming them into functional fluorescent probes remains a challenge. Here we report a fluorescent cellular imaging probe made of high quality doped semiconductor nanocrystals. We have identified two different coating approaches suitable for transforming the as synthesized hydrophobic doped semiconductor nanocrystals into water-soluble functional nanoparticles. Following these approaches we have synthesized TAT-peptide- and folate-functionalized nanoparticles of 10-80 nm hydrodynamic diameter and used them as a fluorescent cell label. The results shows that doped semiconductor nanocrystals can be an attractive alternative for conventional cadmium based quantum dots with low toxicity.

  18. Optimization of microsatellite DNA Gelred fluorescence imaging ...

    African Journals Online (AJOL)

    user1

    2012-10-11

    Oct 11, 2012 ... In order to explore the best microsatellite DNA Gelred imaging technology, this study .... The cycling parameters were: 4 min at 94°C, followed by 35 cycles of ..... double-strand breaks in mammalian cells. Nucleic Acids Res.

  19. Hybrid material as contrast agent in magnetic resonance images

    OpenAIRE

    Botella Asunción, Pablo; Cabrera García, Alejandro

    2015-01-01

    [EN] The invention relates to a contrast agent of magnetic resonance based on a hybrid material formed by an organo-metallic core derived from Prussian blue and a silica cover, and optionally, molecules of a poly(ethylene glycol), a fluorescent agent, a radio nucleus and/or a substance that directs to specific receptors, cells or tissues, joined by covalent bonding to the surface of the inorganic cover.

  20. Hybrid material as contrast agent in magnetic resonance images

    OpenAIRE

    Botella Asunción, Pablo; Cabrera García, Alejandro

    2015-01-01

    [EN] The invention relates to a contrast agent of magnetic resonance based on a hybrid material formed by an organo-metallic core derived from Prussian blue and a silica cover, and optionally, molecules of a poly(ethylene glycol), a fluorescent agent, a radio nucleus and/or a substance that directs to specific receptors, cells or tissues, joined by covalent bonding to the surface of the inorganic cover.

  1. Multispectral fluorescence imaging of human ovarian and fallopian tube tissue for early-stage cancer detection

    Science.gov (United States)

    Tate, Tyler H.; Baggett, Brenda; Rice, Photini F. S.; Koevary, Jennifer Watson; Orsinger, Gabriel V.; Nymeyer, Ariel C.; Welge, Weston A.; Saboda, Kathylynn; Roe, Denise J.; Hatch, Kenneth D.; Chambers, Setsuko K.; Utzinger, Urs; Barton, Jennifer Kehlet

    2016-05-01

    With early detection, 5-year survival rates for ovarian cancer exceed 90%, yet no effective early screening method exists. Emerging consensus suggests over 50% of the most lethal form of the disease originates in the fallopian tube. Twenty-eight women undergoing oophorectomy or debulking surgery provided informed consent for the use of surgical discard tissue samples for multispectral fluorescence imaging. Using multiple ultraviolet and visible excitation wavelengths and emissions bands, 12 fluorescence and 6 reflectance images of 47 ovarian and 31 fallopian tube tissue samples were recorded. After imaging, each sample was fixed, sectioned, and stained for pathological evaluation. Univariate logistic regression showed cancerous tissue samples had significantly lower intensity than noncancerous tissue for 17 image types. The predictive power of multiple image types was evaluated using multivariate logistic regression (MLR) and quadratic discriminant analysis (QDA). Two MLR models each using two image types had receiver operating characteristic curves with area under the curve exceeding 0.9. QDA determined 56 image type combinations with perfect resubstituting using as few as five image types. Adaption of the system for future in vivo fallopian tube and ovary endoscopic imaging is possible, which may enable sensitive detection of ovarian cancer with no exogenous contrast agents.

  2. A support vector machine approach to the automatic identification of fluorescence spectra emitted by biological agents

    Science.gov (United States)

    Gelfusa, M.; Murari, A.; Lungaroni, M.; Malizia, A.; Parracino, S.; Peluso, E.; Cenciarelli, O.; Carestia, M.; Pizzoferrato, R.; Vega, J.; Gaudio, P.

    2016-10-01

    Two of the major new concerns of modern societies are biosecurity and biosafety. Several biological agents (BAs) such as toxins, bacteria, viruses, fungi and parasites are able to cause damage to living systems either humans, animals or plants. Optical techniques, in particular LIght Detection And Ranging (LIDAR), based on the transmission of laser pulses and analysis of the return signals, can be successfully applied to monitoring the release of biological agents into the atmosphere. It is well known that most of biological agents tend to emit specific fluorescence spectra, which in principle allow their detection and identification, if excited by light of the appropriate wavelength. For these reasons, the detection of the UVLight Induced Fluorescence (UV-LIF) emitted by BAs is particularly promising. On the other hand, the stand-off detection of BAs poses a series of challenging issues; one of the most severe is the automatic discrimination between various agents which emit very similar fluorescence spectra. In this paper, a new data analysis method, based on a combination of advanced filtering techniques and Support Vector Machines, is described. The proposed approach covers all the aspects of the data analysis process, from filtering and denoising to automatic recognition of the agents. A systematic series of numerical tests has been performed to assess the potential and limits of the proposed methodology. The first investigations of experimental data have already given very encouraging results.

  3. Polyacrylamide based ICG nanocarriers for enhanced fluorescence and photoacoustic imaging

    Science.gov (United States)

    Ray, Aniruddha; Yoon, Hyung Ki; Ryu, HeeJu; Koo Lee, Yong-Eun; Kim, Gwangseong; Wang, Xueding; Kopelman, Raoul

    2013-02-01

    Indocyanine green (ICG) is an FDA approved tricarbocyanine dye. This dye, with a strong absorbance in the near infrared (NIR) region, has been extensively used for fluorescence and photoacoustic imaging in vivo. ICG in its free form, however, has a few drawbacks that limit its in vivo applications, such as non-targetability, tendency to form aggregates which changes its optical properties, fast degradation, short plasma lifetime and reduced fluorescence at body temperature. In order to bypass these inherent drawbacks, we demonstrate a polyacrylamide based nanocarrier that was particularly designed to carry the negatively charged ICG molecules. These nanocarriers are biodegradable, biocompatible and can be specifically targeted to any cell or tissue. Using these nanocarriers we avoid all the problems associated with free ICG, such as degradation, aggregation and short plasma lifetime, and also enhance demonstrate its ability towards photoacoustics and fluorescence imaging.

  4. Fluorescence-Guided Probes of Aptamer-Targeted Gold Nanoparticles with Computed Tomography Imaging Accesses for in Vivo Tumor Resection

    Science.gov (United States)

    Li, Cheng-Hung; Kuo, Tsung-Rong; Su, Hsin-Jan; Lai, Wei-Yun; Yang, Pan-Chyr; Chen, Jinn-Shiun; Wang, Di-Yan; Wu, Yi-Chun; Chen, Chia-Chun

    2015-01-01

    Recent development of molecular imaging probes for fluorescence-guided surgery has shown great progresses for determining tumor margin to execute the tissue resection. Here we synthesize the fluorescent gold nanoparticles conjugated with diatrizoic acid and nucleolin-targeted AS1411 aptamer. The nanoparticle conjugates exhibit high water-solubility, good biocompatibility, visible fluorescence and strong X-ray attenuation for computed tomography (CT) contrast enhancement. The fluorescent nanoparticle conjugates are applied as a molecular contrast agent to reveal the tumor location in CL1-5 tumor-bearing mice by CT imaging. Furthermore, the orange-red fluorescence emitting from the conjugates in the CL1-5 tumor can be easily visualized by the naked eyes. After the resection, the IVIS measurements show that the fluorescence signal of the nanoparticle conjugates in the tumor is greatly enhanced in comparison to that in the controlled experiment. Our work has shown potential application of functionalized nanoparticles as a dual-function imaging agent in clinical fluorescence-guided surgery. PMID:26507179

  5. Deep UV Native Fluorescence Imaging of Antarctic Cryptoendolithic Communities

    Science.gov (United States)

    Storrie-Lombardi, M. C.; Douglas, S.; Sun, H.; McDonald, G. D.; Bhartia, R.; Nealson, K. H.; Hug, W. F.

    2001-01-01

    An interdisciplinary team at the Jet Propulsion Laboratory Center for Life Detection has embarked on a project to provide in situ chemical and morphological characterization of Antarctic cryptoendolithic microbial communities. We present here in situ deep ultraviolet (UV) native fluorescence and environmental scanning electron microscopy images transiting 8.5 mm into a sandstone sample from the Antarctic Dry Valleys. The deep ultraviolet imaging system employs 224.3, 248.6, and 325 nm lasers to elicit differential fluorescence and resonance Raman responses from biomolecules and minerals. The 224.3 and 248.6 nm lasers elicit a fluorescence response from the aromatic amino and nucleic acids. Excitation at 325 nm may elicit activity from a variety of biomolecules, but is more likely to elicit mineral fluorescence. The resultant fluorescence images provide in situ chemical and morphological maps of microorganisms and the associated organic matrix. Visible broadband reflectance images provide orientation against the mineral background. Environmental scanning electron micrographs provided detailed morphological information. The technique has made possible the construction of detailed fluorescent maps extending from the surface of an Antarctic sandstone sample to a depth of 8.5 mm. The images detect no evidence of microbial life in the superficial 0.2 mm crustal layer. The black lichen component between 0.3 and 0.5 mm deep absorbs all wavelengths of both laser and broadband illumination. Filamentous deep ultraviolet native fluorescent activity dominates in the white layer between 0.6 mm and 5.0 mm from the surface. These filamentous forms are fungi that continue into the red (iron-rich) region of the sample extending from 5.0 to 8.5 mm. Using differential image subtraction techniques it is possible to identify fungal nuclei. The ultraviolet response is markedly attenuated in this region, apparently from the absorption of ultraviolet light by iron-rich particles coating

  6. A matter of collection and detection for intraoperative and noninvasive near-infrared fluorescence molecular imaging: To see or not to see?

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Banghe; Rasmussen, John C.; Sevick-Muraca, Eva M., E-mail: Eva.Sevick@uth.tmc.edu [Center for Molecular Imaging, The Brown Foundation Institute of Molecular Medicine, The University of Texas Health Science Center, Houston, Texas 77030 (United States)

    2014-02-15

    Purpose: Although fluorescence molecular imaging is rapidly evolving as a new combinational drug/device technology platform for molecularly guided surgery and noninvasive imaging, there remains no performance standards for efficient translation of “first-in-humans” fluorescent imaging agents using these devices. Methods: The authors employed a stable, solid phantom designed to exaggerate the confounding effects of tissue light scattering and to mimic low concentrations (nM–pM) of near-infrared fluorescent dyes expected clinically for molecular imaging in order to evaluate and compare the commonly used charge coupled device (CCD) camera systems employed in preclinical studies and in human investigational studies. Results: The results show that intensified CCD systems offer greater contrast with larger signal-to-noise ratios in comparison to their unintensified CCD systems operated at clinically reasonable, subsecond acquisition times. Conclusions: Camera imaging performance could impact the success of future “first-in-humans” near-infrared fluorescence imaging agent studies.

  7. The Cyan Fluorescent Protein (CFP) Transgenic Mouse as a Model for Imaging Pancreatic Exocrine Cells

    Science.gov (United States)

    Cao, Hop S Tran; Kimura, Hiroaki; Kaushal, Sharmeela; Snyder, Cynthia S; Reynoso, Jose; Hoffman, Robert M; Bouvet, Michael

    2015-01-01

    Context The use of fluorescent proteins for in vivo imaging has opened many new areas of research. Among the important advances in the field have been the development of transgenic mice expressing various fluorescent proteins. Objective To report whole-body and organ-specific fluorescence imaging to characterize the transgenic cyan fluorescent protein mouse. Design Mice were imaged using two devices. Brightfield images were obtained with the OV100 Small Animal Imaging System (Olympus Corp., Tokyo, Japan). Fluorescence imaging was performed under the cyan fluorescent protein filter using the iBox Small Animal Imaging System (UVP, Upland, CA, USA). Intervention All animals were sacrificed immediately before imaging. They were imaged before and throughout multiple steps of a complete necropsy. Harvested organs were also imaged with both devices. Selected organs were then frozen and processed for histology, fluorescence microscopy, and H&E staining. Fluorescence microscopy was performed with an Olympus IMT-2 inverted fluorescence microscope. Main outcome measure Determination of fluorescence intensity of different organs. Results Surprisingly, we found that there is differential enhancement of fluorescence among organs; most notably, the pancreas stands out from the rest of the gastrointestinal tract, displaying the strongest fluorescence of all organs in the mouse. Fluorescence microscopy demonstrated that the cyan fluorescent protein fluorescence resided in the acinar cells of the pancreas and not the islet cells. Conclusions The cyan fluorescent protein mouse should lead to a deeper understanding of pancreatic function and pathology, including cancer. PMID:19287108

  8. The Cyan Fluorescent Protein (CFP Transgenic Mouse as a Model for Imaging Pancreatic Exocrine Cells

    Directory of Open Access Journals (Sweden)

    Hop S Tran Cao

    2009-03-01

    Full Text Available The use of fluorescent proteins for in vivo imaging has opened many new areas of research. Among the important advances in the field have been the development of transgenic mice expressing various fluorescent proteins. Objective To report whole-body and organ-specific fluorescence imaging to characterize the transgenic cyan fluorescent protein mouse. Design Mice were imaged using two devices. Brightfield images were obtained with the OV100 Small Animal Imaging System (Olympus Corp., Tokyo, Japan. Fluorescence imaging was performed under the cyan fluorescent protein filter using the iBox Small Animal Imaging System (UVP, Upland, CA, USA. Intervention All animals were sacrificed immediately before imaging. They were imaged before and throughout multiple steps of a complete necropsy. Harvested organs were also imaged with both devices. Selected organs were then frozen and processed for histology, fluorescence microscopy, and H&E staining. Fluorescence microscopy was performed with an Olympus IMT-2 inverted fluorescence microscope. Main outcome measure Determination of fluorescence intensity of different organs. Results Surprisingly, we found that there is differential enhancement of fluorescence among organs; most notably, the pancreas stands out from the rest of the gastrointestinal tract, displaying the strongest fluorescence of all organs in the mouse. Fluorescence microscopy demonstrated that the cyan fluorescent protein fluorescence resided in the acinar cells of the pancreas and not the islet cells. Conclusions The cyan fluorescent protein mouse should lead to a deeper understanding of pancreatic function and pathology, including cancer.

  9. Fluorescence Lifetime Imaging of Free and Protein-Bound NADH

    Science.gov (United States)

    Lakowicz, Joseph R.; Szmacinski, Henryk; Nowaczyk, Kazimierz; Johnson, Michael L.

    1992-02-01

    We introduce a methodology, fluorescence lifetime imaging (FLIM), in which the contrast depends on the fluorescence lifetime at each point in a two-dimensional image and not on the local concentration and/or intensity of the fluorophore. We used FLIM to create lifetime images of NADH when free in solution and when bound to malate dehydrogenase. This represents a challenging case for lifetime imaging because the NADH decay times are just 0.4 and 1.0 ns in the free and bound states, respectively. In the present apparatus, lifetime images are created from a series of phase-sensitive images obtained with a gain-modulated image intensifier and recorded with a charge-coupled device (CCD) camera. The intensifier gain is modulated at the light-modulation frequency or a harmonic thereof. A series of stationary phase-sensitive images, each obtained with various phase shifts of the gain-modulation signal, is used to determine the phase angle or modulation of the emission at each pixel, which is in essence the lifetime image. We also describe an imaging procedure that allows specific decay times to be suppressed, allowing in this case suppression of the emission from either free or bound NADH. Since the fluorescence lifetimes of probes are known to be sensitive to numerous chemical and physical factors such as pH, oxygen, temperature, cations, polarity, and binding to macromolecules, this method allows imaging of the chemical or property of interest in macroscopic and microscopic samples. The concept of FLIM appears to have numerous potential applications in the biosciences.

  10. Polyester Fabric's Fluorescent Dyeing in Supercritical Carbon Dioxide and its Fluorescence Imaging.

    Science.gov (United States)

    Xiong, Xiaoqing; Xu, Yanyan; Zheng, Laijiu; Yan, Jun; Zhao, Hongjuan; Zhang, Juan; Sun, Yanfeng

    2017-03-01

    As one of the most important coumarin-like dyes, disperse fluorescent Yellow 82 exhibits exceptionally large two-photon effects. Here, it was firstly introduced into the supercritical CO2 dyeing polyester fabrics in this work. Results of the present work showed that the dyeing parameters such as the dyeing time, pressure and temperature had remarkable influences on the color strength of fabrics. The optimized dyeing condition in supercritical CO2 dyeing has been proposed that the dyeing time was 60 min; the pressure was 25 MPa and the temperature was 120 °C. As a result, acceptable products were obtained with the wash and rub fastness rating at 5 or 4-5. The polyester fabrics dyed with fluorescent dyes can be satisfied for the requirement of manufacturing warning clothing. Importantly, the confocal microscopy imaging technology was successfully introduced into textile fields to observe the distribution and fluorescence intensity of disperse fluorescent Yellow 82 on polyester fabrics. As far as we know, this is the first report about supercritical CO2 dyeing polyester fabrics based on disperse fluorescent dyes. It will be very helpful for the further design of new fluorescent functional dyes suitable for supercritical CO2 dyeing technique.

  11. Two-photon fluorescence and fluorescence imaging of two styryl heterocyclic dyes combined with DNA.

    Science.gov (United States)

    Gao, Chao; Liu, Shu-yao; Zhang, Xian; Liu, Ying-kai; Qiao, Cong-de; Liu, Zhao-e

    2016-03-01

    Two new styryl heterocyclic two-photon (TP) materials, 4-[4-(N-methyl)styrene]-imidazo [4,5-f][1,10] phenanthroline-benzene iodated salt (probe-1) and 4,4-[4-(N-methyl)styrene]-benzene iodated salt (probe-2) were successfully synthesized and studied as potential fluorescent probes of DNA detection. The linear and nonlinear photophysical properties of two compounds in different solvents were investigated. The absorption, one- and two-photon fluorescent spectra of the free dye and dye-DNA complex were also examined to evaluate their photophysical properties. The binding constants of dye-DNA were obtained according to Scatchard equation with good values. The results showed that two probes could be used as fluorescent DNA probes by two-photon excitation, and TP fluorescent properties of probe-1 are superior to that of probe-2. The fluorescent method date indicated that the mechanisms of dye-DNA complex interaction may be groove binding for probe-1 and electrostatic interaction for probe-2, respectively. The MTT assay experiments showed two probes are low toxicity. Moreover, the TP fluorescence imaging of DNA detection in living cells at 800 nm indicated that the ability to locate in cell nuclei of probe-1 is better than that of probe-2.

  12. Fluorescence-lifetime identification of biological agents using deep ultraviolet light-emitting diodes

    Science.gov (United States)

    Vitta, P.; Kurilcik, N.; Jursenas, S.; Zukauskas, A.; Bakienė, E.; Zhang, J.; Katona, T.; Bilenko, Y.; Lunev, A.; Hu, X.; Deng, J.; Gaska, R.

    2005-10-01

    Recently developed deep-UV light-emitting diodes (LEDs) are already used in prototype fluorescence sensors for detection of hazardous biological agents. However, increasing of the sensor ability of discrimination against common interferents requires further development of measurement technique. In particular, LED-based fluorescence lifetime measurements are to be considered as a technique supplementary to fluorescence spectral and excitation measurements. Here we report on application of UVTOP® series deep-UV LEDs developed by Sensor Electronic Technology, Inc. for real-time measurements of fluorescence lifetime in the frequency domain. LEDs with the wavelengths of 280 nm (targeted to protein excitation) and 340 nm (for excitation of coenzymes NADH and flavins) were used. The output of the LEDs was harmonically modulated at frequencies up to 100 MHz and fluorescence lifetime on the nanosecond and subnanosecond scale was estimated by measuring the phase angle of the fluorescence signal in respect of the LED output. Dual-wavelength LED-based phase-resolved measurement technique was tested for discrimination of B. globigii against a variety of interferents such as diesel fuel, paper, cotton, dust, etc. We conclude that fluorescence phase measurements have potential to improve the discrimination ability of the "detect-to-warn" optical bioparticle sensors.

  13. Photon budget analysis for fluorescence lifetime imaging microscopy

    NARCIS (Netherlands)

    Zhao, Q.; Young, I.T.; De Jong, J.G.S.

    2011-01-01

    We have constructed a mathematical model to analyze the photon efficiency of frequency-domain fluorescence lifetime imaging microscopy (FLIM). The power of the light source needed for illumination in a FLIM system and the signal-to-noise ratio of the detector have led us to a photon “budget.” These

  14. Fluorescence lifetime imaging in biosciences: technologies and applications

    Institute of Scientific and Technical Information of China (English)

    Raluca NIESNER; Karl-Heinz GERICKE

    2008-01-01

    The biosciences require the development of methods that allow a non-invasive and rapid investigation of biological systems. In this aspect, high-end imaging tech-niques allow intravital microscopy in real-time, providing information on a molecular basis. Far-field fluorescence imaging techniques are some of the most adequate methods for such investigations. However, there are great differences between the common fluorescence imaging techniques, i.e., wide-field, confocal one-photon and two-photon microscopy, as far as their applicability in diverse bioscientific research areas is concerned. In the first part of this work, we briefly compare these techniques. Standard methods used in the biosciences, i.e., steady-state techniques based on the analy-sis of the total fluorescence signal originating from the sam-ple, can successfully be employed in the study of cell, tissue and organ morphology as well as in monitoring the macro-scopic tissue function. However, they are mostly inadequate for the quantitative investigation of the cellular function at the molecular level. The intrinsic disadvantages of steady-state techniques are countered by using time-resolved tech-niques. Among these fluorescence lifetime imaging (FLIM) is currently the most common. Different FLIM principles as well as applications of particular relevance for the biosci-ences, especially for fast intravital studies are discussed in this work.

  15. Colorful protein-based fluorescent probes for collagen imaging.

    Directory of Open Access Journals (Sweden)

    Stijn J A Aper

    Full Text Available Real-time visualization of collagen is important in studies on tissue formation and remodeling in the research fields of developmental biology and tissue engineering. Our group has previously reported on a fluorescent probe for the specific imaging of collagen in live tissue in situ, consisting of the native collagen binding protein CNA35 labeled with fluorescent dye Oregon Green 488 (CNA35-OG488. The CNA35-OG488 probe has become widely used for collagen imaging. To allow for the use of CNA35-based probes in a broader range of applications, we here present a toolbox of six genetically-encoded collagen probes which are fusions of CNA35 to fluorescent proteins that span the visible spectrum: mTurquoise2, EGFP, mAmetrine, LSSmOrange, tdTomato and mCherry. While CNA35-OG488 requires a chemical conjugation step for labeling with the fluorescent dye, these protein-based probes can be easily produced in high yields by expression in E. coli and purified in one step using Ni2+-affinity chromatography. The probes all bind specifically to collagen, both in vitro and in porcine pericardial tissue. Some first applications of the probes are shown in multicolor imaging of engineered tissue and two-photon imaging of collagen in human skin. The fully-genetic encoding of the new probes makes them easily accessible to all scientists interested in collagen formation and remodeling.

  16. A Review of Indocyanine Green Fluorescent Imaging in Surgery

    Directory of Open Access Journals (Sweden)

    Jarmo T. Alander

    2012-01-01

    Full Text Available The purpose of this paper is to give an overview of the recent surgical intraoperational applications of indocyanine green fluorescence imaging methods, the basics of the technology, and instrumentation used. Well over 200 papers describing this technique in clinical setting are reviewed. In addition to the surgical applications, other recent medical applications of ICG are briefly examined.

  17. Research Progress of Melanoma Imaging with Radionuclide Agents

    Directory of Open Access Journals (Sweden)

    YOU Qiang

    2015-11-01

    Full Text Available In comparison with CT, ultrasonography, MRI et al, scintigraphic imaging have the advantages of high specificity and recognizes lesions and structure-function of the receptor and metabolism in molecular level, which could diagnose melanoma and its metastasis earlier. With the emergence of more new imaging agents, this unique advantage was more obviously. This article discussed the latest research progress of melanoma imaging with radionuclide agents from aspects of single photon and positron.

  18. Fabrication of Indocyanine Green and 2H, 3H-perfluoropentane loaded microbubbles for fluorescence and ultrasound imaging

    Science.gov (United States)

    He, Yutong; Wu, Qiang; Ma, Rong; Chang, Shufang; Shao, Pengfei; Xu, Ronald

    2016-03-01

    As a near-infrared (NIR) fluorescence dye, Indocyanine Green (ICG) has not gained broader clinical applications, owing to its multiple limitations such as concentration-dependent aggregation, low fluorescence quantum yield, poor physicochemical stability and rapid elimination from the body. In the meanwhile, 2H,3H-perfluoropentane (H-PFP) has been widely studied in ultrasound imaging as a vehicle for targeted delivery of contrast agents and drugs. We synthesized a novel dual-modal fluorescence and ultrasound contrast agent by encapsulating ICG and H-PFP in lipid microbubbles using a liquid-driven coaxial flow focusing (LDCFF) process. Uniform microbubbles with the sizes ranging from 1-10um and great ICG loading efficiency was achieved by this method. Our benchtop experiments showed that ICG/H-PFP microbubbles exhibited less aggregation, increased fluorescence intensity and more stable photostability compared to free ICG aqueous solution. Our phantom experiments demonstrated that ICG/H-PFP microbubbles enhanced the imaging contrasts in fluorescence imaging and ultrasonography. Our animal experiments indicated that ICG/H-PFP microbubbles extended the ICG life time and facilitated dual mode fluorescence and ultrasound imaging in vivo.

  19. Fluorescence lifetime to image epidermal ionic concentrations

    Science.gov (United States)

    Behne, Martin J.; Barry, Nicholas P.; Moll, Ingrid; Gratton, Enrico; Mauro, Theodora M.

    2004-09-01

    Measurements of ionic concentrations in skin have traditionally been performed with an array of methods which either did not reveal detailed localization information, or only provided qualitative, not quantitative information. FLIM combines a number of advantages into a method ideally suited to visualize concentrations of ions such as H+ in intact, unperturbed epidermis and stratum corneum (SC). Fluorescence lifetime is dye concentration-independent, the method requires only low light intensities and is therefore not prone to photobleaching or phototoxic artifacts, and because multiphoton lasers of IR wavelength are used, light penetrates deep into intact tissue. The standard method to measure SC pH is the flat pH electrode, which provides reliable information only about surface pH changes, without further vertical or subcellular spatial resolution; i.e., specific microdomains such as the corneocyte interstices are not resolved, and the deeper SC is inaccessible without resorting to inherently disruptive stripping methods. Furthermore, the concept of a gradient of pH through the SC stems from such stripping experiments, but other confirmation for this concept is lacking. Our investigations into the SC pH distribution so far have revealed the crucial role of the Sodium/Hydrogen Antiporter NHE1 in generation of SC acidity, the colocalization of enzymatic lipid processing activity in the SC with acidic domains of the SC, and the timing and localization of emerging acidity in the SC of newborns. Together, these results have led to an improved understanding of the SC pH, its distribution, origin, and regulation. Future uses for this method include measurements of other ions important for epidermal processes, such as Ca2+, and a quantitative approach to topical drug penetration.

  20. A simple protocol for attenuating the auto-fluorescence of cyanobacteria for optimized fluorescence in situ hybridization (FISH) imaging.

    Science.gov (United States)

    Zeller, Perrine; Ploux, Olivier; Méjean, Annick

    2016-03-01

    Cyanobacteria contain pigments, which generate auto-fluorescence that interferes with fluorescence in situ hybridization (FISH) imaging of cyanobacteria. We describe simple chemical treatments using CuSO4 or H2O2 that significantly reduce the auto-fluorescence of Microcystis strains. These protocols were successfully applied in FISH experiments using 16S rRNA specific probes and filamentous cyanobacteria.

  1. Monitoring photosensitizer uptake using two photon fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Yeh, Shu-Chi Allison; Diamond, Kevin R; Patterson, Michael S; Nie, Zhaojun; Hayward, Joseph E; Fang, Qiyin

    2012-01-01

    Photodynamic Therapy (PDT) provides an opportunity for treatment of various invasive tumors by the use of a cancer targeting photosensitizing agent and light of specific wavelengths. However, real-time monitoring of drug localization is desirable because the induction of the phototoxic effect relies on interplay between the dosage of localized drug and light. Fluorescence emission in PDT may be used to monitor the uptake process but fluorescence intensity is subject to variability due to scattering and absorption; the addition of fluorescence lifetime may be beneficial to probe site-specific drug-molecular interactions and cell damage. We investigated the fluorescence lifetime changes of Photofrin(®) at various intracellular components in the Mat-LyLu (MLL) cell line. The fluorescence decays were analyzed using a bi-exponential model, followed by segmentation analysis of lifetime parameters. When Photofrin(®) was localized at the cell membrane, the slow lifetime component was found to be significantly shorter (4.3 ± 0.5 ns) compared to those at other locations (cytoplasm: 7.3 ± 0.3 ns; mitochondria: 7.0 ± 0.2 ns, p < 0.05).

  2. Monitoring Photosensitizer Uptake Using Two Photon Fluorescence Lifetime Imaging Microscopy

    Directory of Open Access Journals (Sweden)

    Shu-Chi Allison Yeh, Kevin R. Diamond, Michael S. Patterson, Zhaojun Nie, Joseph E. Hayward, Qiyin Fang

    2012-01-01

    Full Text Available Photodynamic Therapy (PDT provides an opportunity for treatment of various invasive tumors by the use of a cancer targeting photosensitizing agent and light of specific wavelengths. However, real-time monitoring of drug localization is desirable because the induction of the phototoxic effect relies on interplay between the dosage of localized drug and light. Fluorescence emission in PDT may be used to monitor the uptake process but fluorescence intensity is subject to variability due to scattering and absorption; the addition of fluorescence lifetime may be beneficial to probe site-specific drug-molecular interactions and cell damage. We investigated the fluorescence lifetime changes of Photofrin® at various intracellular components in the Mat-LyLu (MLL cell line. The fluorescence decays were analyzed using a bi-exponential model, followed by segmentation analysis of lifetime parameters. When Photofrin® was localized at the cell membrane, the slow lifetime component was found to be significantly shorter (4.3 ± 0.5 ns compared to those at other locations (cytoplasm: 7.3 ± 0.3 ns; mitochondria: 7.0 ± 0.2 ns, p < 0.05.

  3. Novel fluorescence molecular imaging of chemotherapy-induced intestinal apoptosis

    Science.gov (United States)

    Levin, Galit; Shirvan, Anat; Grimberg, Hagit; Reshef, Ayelet; Yogev-Falach, Merav; Cohen, Avi; Ziv, Ilan

    2009-09-01

    Chemotherapy-induced enteropathy (CIE) is one of the most serious complications of anticancer therapy, and tools for its early detection and monitoring are highly needed. We report on a novel fluorescence method for detection of CIE, based on molecular imaging of the related apoptotic process. The method comprises systemic intravenous administration of the ApoSense fluorescent biomarker (N,N'-didansyl-L-cystine DDC) in vivo and subsequent fluorescence imaging of the intestinal mucosa. In the reported proof-of-concept studies, mice were treated with either taxol+cyclophosphamide or doxil. DDC was administered in vivo at various time points after drug administration, and tracer uptake by ileum tissue was subsequently evaluated by ex vivo fluorescent microscopy. Chemotherapy caused marked and selective uptake of DDC in ileal epithelial cells, in correlation with other hallmarks of apoptosis (i.e., DNA fragmentation and Annexin-V binding). Induction of DDC uptake occurred early after chemotherapy, and its temporal profile was parallel to that of the apoptotic process, as assessed histologically. DDC may therefore serve as a useful tool for detection of CIE. Future potential integration of this method with fluorescent endoscopic techniques, or development of radio-labeled derivatives of DDC for emission tomography, may advance early diagnosis and monitoring of this severe adverse effect of chemotherapy.

  4. Wide-field imaging of fluorescent deoxy-glucose in ex vivo malignant and normal breast tissue

    Science.gov (United States)

    Langsner, R. J.; Middleton, L. P.; Sun, J.; Meric-Bernstam, F.; Hunt, K. K.; Drezek, R. A.; Yu, T. K.

    2011-01-01

    Rapid in situ determination of surgical resection margins during breast cancer surgery would reduce patient time under anesthesia. We present preliminary data supporting the use of a fluorescent glucose analog (2-NBDG) as an optical contrast agent to differentiate freshly excised breast tissue containing cancerous cells from normal breast tissue. Multi-spectral images of 14 breast cancer specimens acquired before and after incubation with 2-NBDG demonstrated increased fluorescent signal in all of the malignant tissue due to increased 2-NBDG consumption. We demonstrate that 2-NBDG has potential as an optical contrast agent to differentiate cancerous from non-cancerous tissue. PMID:21698015

  5. Infrared imaging of LED lighting tubes and fluorescent tubes

    Science.gov (United States)

    Siikanen, Sami; Kivi, Sini; Kauppinen, Timo; Juuti, Mikko

    2011-05-01

    The low energy efficiency of conventional light sources is mainly caused by generation of waste heat. We used infrared (IR) imaging in order to monitor the heating of both LED tube luminaires and ordinary T8 fluorescent tubes. The IR images showed clearly how the surface temperatures of the fluorescent tube ends quickly rose up to about +50...+70°C, whereas the highest surface temperatures seen on the LED tubes were only about +30...+40°C. The IR images demonstrated how the heat produced by the individual LED chips can be efficiently guided to the supporting structure in order to keep the LED emitters cool and hence maintain efficient operation. The consumed electrical power and produced illuminance were also recorded during 24 hour measurements. In order to assess the total luminous efficacy of the luminaires, separate luminous flux measurements were made in a large integrating sphere. The currently available LED tubes showed efficacies of up to 88 lm/W, whereas a standard "cool white" T8 fluorescent tube produced ca. 75 lm/W. Both lamp types gave ca. 110 - 130 lx right below the ceiling-mounted luminaire, but the LED tubes consume only 40 - 55% of the electric power compared to fluorescent tubes.

  6. Mueller matrix signature in advanced fluorescence microscopy imaging

    Science.gov (United States)

    Mazumder, Nirmal; Qiu, Jianjun; Kao, Fu-Jen; Diaspro, Alberto

    2017-02-01

    We have demonstrated the measurement and characterization of the polarization properties of a fluorescence signal using four-channel photon counting based Stokes-Mueller polarization microscopy. Thus, Lu-Chipman decomposition was applied to extract the critical polarization properties such as depolarization, linear retardance and the optical rotation of collagen type I fiber. We observed the spatial distribution of anisotropic and helical molecules of collagen from the reconstructed 2D Mueller images based on the fluorescence signal in a pixel-by-pixel manner.

  7. Dynamic fluorescence lifetime imaging based on acousto-optic deflectors

    Science.gov (United States)

    Yan, Wei; Peng, Xiao; Qi, Jing; Gao, Jian; Fan, Shunping; Wang, Qi; Qu, Junle; Niu, Hanben

    2014-11-01

    We report a dynamic fluorescence lifetime imaging (D-FLIM) system that is based on a pair of acousto-optic deflectors for the random regions of interest (ROI) study in the sample. The two-dimensional acousto-optic deflector devices are used to rapidly scan the femtosecond excitation laser beam across the sample, providing specific random access to the ROI. Our experimental results using standard fluorescent dyes in live cancer cells demonstrate that the D-FLIM system can dynamically monitor the changing process of the microenvironment in the ROI in live biological samples.

  8. Highly stable organic fluorescent nanorods for living- cell imaging

    Institute of Scientific and Technical Information of China (English)

    Minhuan Lan[1,3; Jinfeng Zhang[1,3; Xiaoyue Zhu[1; Pengfei Wang[2; Xianfeng Chen[1; Chun-Sing Lee[1; Wenjun Zhang[1

    2015-01-01

    Metal-free, organic-dye-based fluorescent nanorods were fabricated through a simple solvent-exchange procedure. The as-prepared nanorods exhibit low toxicity to living cells and excellent photostability. Furthermore, they are stable in solutions of various pHs and high ionic strength and in solutions with interfering metal ions. Compared with the free DPP-Br molecules in THF, these nanorods exhibit larger Stokes shift, broader absorption spectra, and greatly improved photostability. We successfully demonstrated the application of the nanorods, including their aforementioned beneficial characteristics, as a good fluorescence probe for bio-imaging.

  9. The study of blue LED to induce fluorescence spectroscopy and fluorescence imaging for oral carcinoma detection

    Science.gov (United States)

    Zheng, Longjiang; Hu, Yuanting

    2009-07-01

    Fluorescence spectroscopy and fluorescence imaging diagnosis of malignant lesions provides us with a new method to diagnose diseases in precancerous stage. Early diagnosis of disease has significant importance in cancer treatment, because most cancers can be cured well in precancerous, especially when the diffusion of cancer is limited in a restricted region. In this study, Golden hamster models were applied to 5% 9, 10 dimethyl-1, 2-benzanthracene (DMBA) to induce hamster buccal cheek pouch carcinoma three times a week. Rose Bengal, which has been used in clinican for years and avoids visible side-effect to human was chosen as photosensitizer. 405 nm blue LED was used to induce the fluorescence of photosensitizer. After topical application of photosensitizer, characteristic red emission fluorescence peak was observed around 600nm. Similar, normal oral cavity has special luminescence around 480nm. Fluorescence spectroscopy technology is based on analysing emission peaks of photosensitizer in the areas of oral carcinoma, moreover, red-to-green (IR/IG) intensity ratio is also applied as a diagnostic algorithm. A CCD which is connected with a computer is used to take pictures at carcinoma areas through different filters. Fluorescence images from normal hamster buccal cheek pouch are compared with those from carcinogen-induced models of carcinoma, and morphological differences between normal and lesion tissue can be distinguished. The pictures are analyzed by Matlab and shown on the screen of computer. This paper demonstrates that Rose Bengal could be used as photosensitizer to detect oral carcinoma, and blue LED as excitation source could not only have a good effect to diagnose oral carcinoma, but also decrease cost greatly.

  10. Nanoscale fluorescence lifetime imaging with a single diamond NV center

    CERN Document Server

    Beams, Ryan; Johnson, Timothy W; Oh, Sang-Hyun; Novotny, Lukas; Vamivakas, Nick

    2013-01-01

    Solid-state quantum emitters, such as artificially engineered quantum dots or naturally occurring defects in solids, are being investigated for applications ranging from quantum information science and optoelectronics to biomedical imaging. Recently, these same systems have also been studied from the perspective of nanoscale metrology. In this letter we study the near-field optical properties of a diamond nanocrystal hosting a single nitrogen vacancy center. We find that the nitrogen vacancy center is a sensitive probe of the surrounding electromagnetic mode structure. We exploit this sensitivity to demonstrate nanoscale fluorescence lifetime imaging microscopy (FLIM) with a single nitrogen vacancy center by imaging the local density of states of an optical antenna.

  11. A Pico Projector Source for Confocal Fluorescence and Ophthalmic Imaging.

    Science.gov (United States)

    Muller, Matthew S

    2012-09-02

    A Pico digital light projector has been implemented as an integrated illumination source and spatial light modulator for confocal imaging. The target is illuminated with a series of rapidly projected lines or points to simulate scanning. Light returning from the target is imaged onto a 2D rolling shutter CMOS sensor. By controlling the spatio-temporal relationship between the rolling shutter and illumination pattern, light returning from the target is spatially filtered. Confocal retinal, fluorescence, and Fourier-domain optical coherence tomography implementations of this novel imaging technique are presented.

  12. In Vivo Metal Ion Imaging Using Fluorescent Sensors.

    Science.gov (United States)

    Van de Bittner, Genevieve C; Hirayama, Tasuku

    2016-01-01

    In vivo imaging in living animals provides the ability to monitor alterations of signaling molecules, ions, and other biological components during various life stages and in disease. The data gained from in vivo imaging can be used for biological discovery or to determine elements of disease progression and can inform the development and translation of therapeutics. Herein, we present theories behind small-molecule, fluorescent, metal ion sensors as well as the methods for their successful application to in vivo metal ion imaging, including ex vivo validation.

  13. Fully integrated optical coherence tomography, ultrasound, and indocyanine green-based fluorescence tri-modality system for intravascular imaging.

    Science.gov (United States)

    Li, Yan; Jing, Joseph; Qu, Yueqiao; Miao, Yusi; Zhang, Buyun; Ma, Teng; Yu, Mingyue; Zhou, Qifa; Chen, Zhongping

    2017-02-01

    We present a tri-modality imaging system and fully integrated tri-modality probe for intravascular imaging. The tri-modality imaging system is able to simultaneously acquire optical coherence tomography (OCT), ultrasound (US), and fluorescence imaging. Moreover, for fluorescence imaging, we used the FDA-approved indocyanine green (ICG) dye as the contrast agent to target lipid-loaded macrophages. We conducted imaging from a male New Zealand white rabbit to evaluate the performance of the tri-modality system. In addition, tri-modality images of rabbit aortas were correlated with hematoxylin and eosin (H&E) histology to check the measurement accuracy. The fully integrated miniature tri-modality probe, together with the use of ICG dye suggest that the system is of great potential for providing a more accurate assessment of vulnerable plaques in clinical applications.

  14. Chlorophyll fluorescence imaging of cadmium-treated white cabbage plants

    Directory of Open Access Journals (Sweden)

    Borek M.

    2013-04-01

    Full Text Available The chlorophyll fluorescence imaging technique is a valuable tool to study the impact of heavy metal stress in plants. The aim of this paper was to investigate the influence of Cd on photosynthetic apparatus of white cabbage (Brassica oleracea subsp. capitata f. alba plants. Two cabbage cultivars ‘Ditmarska Najwcześniejsza’ (‘DN’; early and ‘Amager Polana’ (‘AP’; late were used. Cd was applied before planting seedlings (10 mg Cd kg−1 DM of soil.. Measurements were performed at the 3rd leaf after 2 weeks of planting. The level of Cd-induced stress to plants was estimated by chlorophyll (Chl content (photometrically and analyses of images and numeric values of the major fluorescence parameters of Chl (Chl fluorescence imaging system FluorCam. Cd negatively affected the chlorophyll content and Fv/Fm, Fv’/Fm’, Φ PSII and qP in leaves of early cultivar of white cabbage. However, in the case of late cv. we did not observe such distinct changes. It suggests that late cultivars. are more resistant to Cd than the early ones. Considering methodological aspect of the study, Chl fluorescence imaging can better reveal some alterations within the leaf, because numeric values of specific parameters, which are the averaged data collected from the whole leaf, cannot reflect the tissue specificity. Abbreviations: HM – heavy metal, Cd – cadmium, Chl – chlorophyll, Fv/Fm – photochemical efficiency of PSII in the dark-adapted state, F‘v’/F‘m’ – PSII maximum efficiency, Φ PSII – quantum efficiency of PSII electron transport, NPQ – nonphotochemical quenching of maximal Chl fluorescence, qP – photochemical quenching coefficient.

  15. Centimeter-deep tissue fluorescence microscopic imaging with high signal-to-noise ratio and picomole sensitivity

    CERN Document Server

    Cheng, Bingbing; Wei, Ming-Yuan; Pei, Yanbo; DSouza, Francis; Nguyen, Kytai T; Hong, Yi; Tang, Liping; Yuan, Baohong

    2015-01-01

    Fluorescence microscopic imaging in centimeter-deep tissue has been highly sought-after for many years because much interesting in vivo micro-information, such as microcirculation, tumor angiogenesis, and metastasis, may deeply locate in tissue. In this study, for the first time this goal has been achieved in 3-centimeter deep tissue with high signal-to-noise ratio (SNR) and picomole sensitivity under radiation safety thresholds. These results are demonstrated not only in tissue-mimic phantoms but also in actual tissues, such as porcine muscle, ex vivo mouse liver, ex vivo spleen, and in vivo mouse tissue. These results are achieved based on three unique technologies: excellent near infrared ultrasound-switchable fluorescence (USF) contrast agents, a sensitive USF imaging system, and an effective correlation method. Multiplex USF fluorescence imaging is also achieved. It is useful to simultaneously image multiple targets and observe their interactions. This work opens the door for future studies of centimeter...

  16. Fluorescent ligand for human progesterone receptor imaging in live cells.

    Science.gov (United States)

    Weinstain, Roy; Kanter, Joan; Friedman, Beth; Ellies, Lesley G; Baker, Michael E; Tsien, Roger Y

    2013-05-15

    We employed molecular modeling to design and then synthesize fluorescent ligands for the human progesterone receptor. Boron dipyrromethene (BODIPY) or tetramethylrhodamine were conjugated to the progesterone receptor antagonist RU486 (Mifepristone) through an extended hydrophilic linker. The fluorescent ligands demonstrated comparable bioactivity to the parent antagonist in live cells and triggered nuclear translocation of the receptor in a specific manner. The BODIPY labeled ligand was applied to investigate the dependency of progesterone receptor nuclear translocation on partner proteins and to show that functional heat shock protein 90 but not immunophilin FKBP52 activity is essential. A tissue distribution study indicated that the fluorescent ligand preferentially accumulates in tissues that express high levels of the receptor in vivo. The design and properties of the BODIPY-labeled RU486 make it a potential candidate for in vivo imaging of PR by positron emission tomography through incorporation of (18)F into the BODIPY core.

  17. Poly(Lactic-co-Glycolic) Acid as a Carrier for Imaging Contrast Agents

    Science.gov (United States)

    Doiron, Amber L.; Homan, Kimberly A.; Emelianov, Stanislav; Brannon-Peppas, Lisa

    2010-01-01

    Purpose With the broadening field of nanomedicine poised for future molecular level therapeutics, nano-and microparticles intended for the augmentation of either single- or multimodal imaging are created with PLGA as the chief constituent and carrier. Methods Emulsion techniques were used to encapsulate hydrophilic and hydrophobic imaging contrast agents in PLGA particles. The imaging contrast properties of these PLGA particles were further enhanced by reducing silver onto the PLGA surface, creating a silver cage around the polymeric core. Results The MRI contrast agent Gd-DTPA and the exogenous dye rhodamine 6G were both encapsulated in PLGA and shown to enhance MR and fluorescence contrast, respectively. The silver nanocage built around PLGA nanoparticles exhibited strong near infrared light absorbance properties, making it a suitable contrast agent for optical imaging strategies such as photoacoustic imaging. Conclusions The biodegradable polymer PLGA is an extremely versatile nano- and micro-carrier for several imaging contrast agents with the possibility of targeting diseased states at a molecular level. PMID:19034628

  18. Fluorescence imaging to study cancer burden on lymph nodes

    Science.gov (United States)

    D'Souza, Alisha V.; Elliott, Jonathan T.; Gunn, Jason R.; Samkoe, Kimberley S.; Tichauer, Kenneth M.; Pogue, Brian W.

    2015-03-01

    Morbidity and complexity involved in lymph node staging via surgical resection and biopsy calls for staging techniques that are less invasive. While visible blue dyes are commonly used in locating sentinel lymph nodes, since they follow tumor-draining lymphatic vessels, they do not provide a metric to evaluate presence of cancer. An area of active research is to use fluorescent dyes to assess tumor burden of sentinel and secondary lymph nodes. The goal of this work was to successfully deploy and test an intra-nodal cancer-cell injection model to enable planar fluorescence imaging of a clinically relevant blue dye, specifically methylene blue along with a cancer targeting tracer, Affibody labeled with IRDYE800CW and subsequently segregate tumor-bearing from normal lymph nodes. This direct-injection based tumor model was employed in athymic rats (6 normal, 4 controls, 6 cancer-bearing), where luciferase-expressing breast cancer cells were injected into axillary lymph nodes. Tumor presence in nodes was confirmed by bioluminescence imaging before and after fluorescence imaging. Lymphatic uptake from the injection site (intradermal on forepaw) to lymph node was imaged at approximately 2 frames/minute. Large variability was observed within each cohort.

  19. Chitosan-based formulations of drugs, imaging agents and biotherapeutics

    NARCIS (Netherlands)

    Amidi, M.; Hennink, W.E.

    2010-01-01

    This preface is part of the Advanced Drug Delivery Reviews theme issue on “Chitosan-Based Formulations of Drugs, Imaging Agents and Biotherapeutics”. This special Advanced Drug Delivery Reviews issue summarizes recent progress and different applications of chitosanbased formulations.

  20. Chitosan-based formulations of drugs, imaging agents and biotherapeutics

    NARCIS (Netherlands)

    Amidi, M.; Hennink, W.E.

    This preface is part of the Advanced Drug Delivery Reviews theme issue on “Chitosan-Based Formulations of Drugs, Imaging Agents and Biotherapeutics”. This special Advanced Drug Delivery Reviews issue summarizes recent progress and different applications of chitosanbased formulations.

  1. Stable J-aggregation enabled dual photoacoustic and fluorescence nanoparticles for intraoperative cancer imaging.

    Science.gov (United States)

    Shakiba, Mojdeh; Ng, Kenneth K; Huynh, Elizabeth; Chan, Harley; Charron, Danielle M; Chen, Juan; Muhanna, Nidal; Foster, F Stuart; Wilson, Brian C; Zheng, Gang

    2016-07-07

    J-aggregates display nanoscale optical properties which enable their use in fluorescence and photoacoustic imaging applications. However, control over their optical properties in an in vivo setting is hampered by the conformational lability of the J-aggregate structure in complex biological environments. J-aggregating nanoparticles (JNP) formed by self-assembly of bacteriopheophorbide-lipid (Bchl-lipid) in lipid nanovesicles represents a novel strategy to stabilize J-aggregates for in vivo bioimaging applications. We find that 15 mol% Bchl-lipid embedded within a saturated phospholipid bilayer vesicle was optimal in terms of maximizing Bchl-lipid dye loading, while maintaining a spherical nanoparticle morphology and retaining spectral properties characteristic of J-aggregates. The addition of cholesterol maintains the stability of the J-aggregate absorption band for up to 6 hours in the presence of 90% FBS. In a proof-of-concept experiment, we successfully applied JNPs as a fluorescence contrast agent for real-time intraoperative detection of metastatic lymph nodes in a rabbit head-and-neck cancer model. Lymph node metastasis delineation was further verified by visualizing the JNP within the excised lymph node using photoacoustic imaging. Using JNPs, we demonstrate the possibility of using J-aggregates as fluorescence and photoacoustic contrast agents and may potentially spur the development of other nanomaterials that can stably induce J-aggregation for in vivo cancer bioimaging applications.

  2. Contrast Agents for Photoacoustic and Thermoacoustic Imaging: A Review

    Directory of Open Access Journals (Sweden)

    Dan Wu

    2014-12-01

    Full Text Available Photoacoustic imaging (PAI and thermoacoustic imaging (TAI are two emerging biomedical imaging techniques that both utilize ultrasonic signals as an information carrier. Unique advantages of PAI and TAI are their abilities to provide high resolution functional information such as hemoglobin and blood oxygenation and tissue dielectric properties relevant to physiology and pathology. These two methods, however, may have a limited detection depth and lack of endogenous contrast. An exogenous contrast agent is often needed to effectively resolve these problems. Such agents are able to greatly enhance the imaging contrast and potentially break through the imaging depth limit. Furthermore, a receptor-targeted contrast agent could trace the molecular and cellular biological processes in tissues. Thus, photoacoustic and thermoacoustic molecular imaging can be outstanding tools for early diagnosis, precise lesion localization, and molecular typing of various diseases. The agents also could be used for therapy in conjugation with drugs or in photothermal therapy, where it functions as an enhancer for the integration of diagnosis and therapy. In this article, we present a detailed review about various exogenous contrast agents for photoacoustic and thermoacoustic molecular imaging. In addition, challenges and future directions of photoacoustic and thermoacoustic molecular imaging in the field of translational medicine are also discussed.

  3. Contrast agents for photoacoustic and thermoacoustic imaging: a review.

    Science.gov (United States)

    Wu, Dan; Huang, Lin; Jiang, Max S; Jiang, Huabei

    2014-12-18

    Photoacoustic imaging (PAI) and thermoacoustic imaging (TAI) are two emerging biomedical imaging techniques that both utilize ultrasonic signals as an information carrier. Unique advantages of PAI and TAI are their abilities to provide high resolution functional information such as hemoglobin and blood oxygenation and tissue dielectric properties relevant to physiology and pathology. These two methods, however, may have a limited detection depth and lack of endogenous contrast. An exogenous contrast agent is often needed to effectively resolve these problems. Such agents are able to greatly enhance the imaging contrast and potentially break through the imaging depth limit. Furthermore, a receptor-targeted contrast agent could trace the molecular and cellular biological processes in tissues. Thus, photoacoustic and thermoacoustic molecular imaging can be outstanding tools for early diagnosis, precise lesion localization, and molecular typing of various diseases. The agents also could be used for therapy in conjugation with drugs or in photothermal therapy, where it functions as an enhancer for the integration of diagnosis and therapy. In this article, we present a detailed review about various exogenous contrast agents for photoacoustic and thermoacoustic molecular imaging. In addition, challenges and future directions of photoacoustic and thermoacoustic molecular imaging in the field of translational medicine are also discussed.

  4. [Intraoperative graft assessment using fluorescent imaging system (SPY)].

    Science.gov (United States)

    Kawashima, T; Naraoka, S; Kakizaki, T

    2009-07-01

    We investigated the efficacy of intraoperative fluorescent imaging system for the assessment of coronary artery bypass grafting (CABG). We used SPY imaging system in 100 CABG (57 off-pump and 43 on-pump CABG), totalling 287 distal anastomoses. The total graft patency rate on postoperative angiography in this series was 96.2% (276/287). Graft revision was done in 10 cases (10.0%) and 13 anastomoses (4.5%) by SPY imaging, which all resulted in good patency at postoperative angiography. On the other hand, 7 distal anastomoses and 1 mammary graft (2.8%) appeared to be successful on intraoperative SPY imaging, but were revealed to be occluded by postoperative angiography. SPY imaging system is useful for graft validation, and may contribute to improvement of coronary bypass graft patency.

  5. Intraoperative near-infrared fluorescence imaging targeting folate receptors identifies lung cancer in a large-animal model.

    Science.gov (United States)

    Keating, Jane J; Runge, Jeffrey J; Singhal, Sunil; Nims, Sarah; Venegas, Ollin; Durham, Amy C; Swain, Gary; Nie, Shuming; Low, Philip S; Holt, David E

    2017-05-15

    Complete tumor resection is the most important predictor of patient survival with non-small cell lung cancer. Methods for intraoperative margin assessment after lung cancer excision are lacking. This study evaluated near-infrared (NIR) intraoperative imaging with a folate-targeted molecular contrast agent (OTL0038) for the localization of primary lung adenocarcinomas, lymph node sampling, and margin assessment. Ten dogs with lung cancer underwent either video-assisted thoracoscopic surgery or open thoracotomy and tumor excision after an intravenous injection of OTL0038. Lungs were imaged with an NIR imaging device both in vivo and ex vivo. The wound bed was re-imaged for retained fluorescence suspicious for positive tumor margins. The tumor signal-to-background ratio (SBR) was measured in all cases. Next, 3 human patients were enrolled in a proof-of-principle study. Tumor fluorescence was measured both in situ and ex vivo. All canine tumors fluoresced in situ (mean Fluoptics SBR, 5.2 [range, 2.7-8.1]; mean Karl Storz SBR 1.9 [range, 1.4-2.6]). In addition, the fluorescence was consistent with tumor margins on pathology. Three positive lymph nodes were discovered with NIR imaging. Also, a positive retained tumor margin was discovered upon NIR imaging of the wound bed. Human pulmonary adenocarcinomas were also fluorescent both in situ and ex vivo (mean SBR, > 2.0). NIR imaging can identify lung cancer in a large-animal model. In addition, NIR imaging can discriminate lymph nodes harboring cancer cells and also bring attention to a positive tumor margin. In humans, pulmonary adenocarcinomas fluoresce after the injection of the targeted contrast agent. Cancer 2017;123:1051-60. © 2016 American Cancer Society. © 2016 American Cancer Society.

  6. Phenotypic charactheristics of fluorescent pseudomonss, biological control agent of lincat disease of temanggung tobacco

    Directory of Open Access Journals (Sweden)

    NINING NURUL AZIZAH

    2007-04-01

    Full Text Available Fluorescent pseudomonass isolated from local plants-rishosphere in temanggung controlled lincat disease of tobacco. This report describe phenotypic charactheristics of the bacteria in order to be used as a base for the development of the bacteria as a biological control agent of lincat disease. Phenotypic charactheristics of six isolates of fluorescent Pseudomonass which controlled lincat disease in the field were determined in the laboratory of Plant Bacteriology, Faculty of Agriculture, Gadjah Mada University. Plant pathogenicity tests were conducted by hypersensitive reaction into tobacco leaf and inoculation to tobacco plants. Antagonism test between fluorescent Pseudomonass and other candidate of biological control agents were also conducted. The results indicated that the bacteria were rod shape, Gram negative, positive reaction in catalase and oxidase tests. Nitrate reduce to nitrite, arginine was hydrolysed, fluorescent pigment were produced on King’s B medium, levan formation positive and all bacteria denitrifiy. The bacteria used urea, tween 80 and amylum were not hydrolised, poly--hydroxybutyrate was not accumulated in the cells. Negative reactions were observed for lysine decarboxylation, indol production, VP/MR reaction, and gelatn liquefation. Some compounds could be used as solely carbon sources. All isolates grew on the medium containing 2% NaCl. The best pH for growth was 6-7 and all isolates grew at 20-41C. Negative result were obtained for hypersensitive reaction and pathogenicity tests.

  7. Proton-induced x-ray fluorescence CT imaging.

    Science.gov (United States)

    Bazalova-Carter, Magdalena; Ahmad, Moiz; Matsuura, Taeko; Takao, Seishin; Matsuo, Yuto; Fahrig, Rebecca; Shirato, Hiroki; Umegaki, Kikuo; Xing, Lei

    2015-02-01

    To demonstrate the feasibility of proton-induced x-ray fluorescence CT (pXFCT) imaging of gold in a small animal sized object by means of experiments and Monte Carlo (MC) simulations. First, proton-induced gold x-ray fluorescence (pXRF) was measured as a function of gold concentration. Vials of 2.2 cm in diameter filled with 0%-5% Au solutions were irradiated with a 220 MeV proton beam and x-ray fluorescence induced by the interaction of protons, and Au was detected with a 3 × 3 mm(2) CdTe detector placed at 90° with respect to the incident proton beam at a distance of 45 cm from the vials. Second, a 7-cm diameter water phantom containing three 2.2-diameter vials with 3%-5% Au solutions was imaged with a 7-mm FWHM 220 MeV proton beam in a first generation CT scanning geometry. X-rays scattered perpendicular to the incident proton beam were acquired with the CdTe detector placed at 45 cm from the phantom positioned on a translation/rotation stage. Twenty one translational steps spaced by 3 mm at each of 36 projection angles spaced by 10° were acquired, and pXFCT images of the phantom were reconstructed with filtered back projection. A simplified geometry of the experimental data acquisition setup was modeled with the MC TOPAS code, and simulation results were compared to the experimental data. A linear relationship between gold pXRF and gold concentration was observed in both experimental and MC simulation data (R(2) > 0.99). All Au vials were apparent in the experimental and simulated pXFCT images. Specifically, the 3% Au vial was detectable in the experimental [contrast-to-noise ratio (CNR) = 5.8] and simulated (CNR = 11.5) pXFCT image. Due to fluorescence x-ray attenuation in the higher concentration vials, the 4% and 5% Au contrast were underestimated by 10% and 15%, respectively, in both the experimental and simulated pXFCT images. Proton-induced x-ray fluorescence CT imaging of 3%-5% gold solutions in a small animal sized water phantom has been demonstrated

  8. Proton-induced x-ray fluorescence CT imaging

    Energy Technology Data Exchange (ETDEWEB)

    Bazalova-Carter, Magdalena, E-mail: bazalova@stanford.edu; Xing, Lei [Department of Radiation Oncology, Stanford University, Stanford, California 94305-5847 and Global Station for Quantum Medical Science and Engineering, Global Institution for Collaborative Research and Education (GI-CoRE), Hokkaido University, Sapporo 060-8648 (Japan); Ahmad, Moiz [Department of Radiation Oncology, Stanford University, Stanford, California 94305-5847 (United States); Matsuura, Taeko; Takao, Seishin; Shirato, Hiroki; Umegaki, Kikuo [Department of Medical Physics, Proton Beam Therapy Center, Hokkaido University Hospital, Sapporo 060-8648, Japan and Global Station for Quantum Medical Science and Engineering, Global Institution for Collaborative Research and Education (GI-CoRE), Hokkaido University, Sapporo 060-8648 (Japan); Matsuo, Yuto [Department of Medical Physics, Proton Beam Therapy Center, Hokkaido University Hospital, Sapporo 060-8648 (Japan); Fahrig, Rebecca [Department of Radiology, Stanford University, Stanford, California 94305 (United States)

    2015-02-15

    Purpose: To demonstrate the feasibility of proton-induced x-ray fluorescence CT (pXFCT) imaging of gold in a small animal sized object by means of experiments and Monte Carlo (MC) simulations. Methods: First, proton-induced gold x-ray fluorescence (pXRF) was measured as a function of gold concentration. Vials of 2.2 cm in diameter filled with 0%–5% Au solutions were irradiated with a 220 MeV proton beam and x-ray fluorescence induced by the interaction of protons, and Au was detected with a 3 × 3 mm{sup 2} CdTe detector placed at 90° with respect to the incident proton beam at a distance of 45 cm from the vials. Second, a 7-cm diameter water phantom containing three 2.2-diameter vials with 3%–5% Au solutions was imaged with a 7-mm FWHM 220 MeV proton beam in a first generation CT scanning geometry. X-rays scattered perpendicular to the incident proton beam were acquired with the CdTe detector placed at 45 cm from the phantom positioned on a translation/rotation stage. Twenty one translational steps spaced by 3 mm at each of 36 projection angles spaced by 10° were acquired, and pXFCT images of the phantom were reconstructed with filtered back projection. A simplified geometry of the experimental data acquisition setup was modeled with the MC TOPAS code, and simulation results were compared to the experimental data. Results: A linear relationship between gold pXRF and gold concentration was observed in both experimental and MC simulation data (R{sup 2} > 0.99). All Au vials were apparent in the experimental and simulated pXFCT images. Specifically, the 3% Au vial was detectable in the experimental [contrast-to-noise ratio (CNR) = 5.8] and simulated (CNR = 11.5) pXFCT image. Due to fluorescence x-ray attenuation in the higher concentration vials, the 4% and 5% Au contrast were underestimated by 10% and 15%, respectively, in both the experimental and simulated pXFCT images. Conclusions: Proton-induced x-ray fluorescence CT imaging of 3%–5% gold solutions in a

  9. Directional bilateral filters for smoothing fluorescence microscopy images

    Directory of Open Access Journals (Sweden)

    Manasij Venkatesh

    2015-08-01

    Full Text Available Images obtained through fluorescence microscopy at low numerical aperture (NA are noisy and have poor resolution. Images of specimens such as F-actin filaments obtained using confocal or widefield fluorescence microscopes contain directional information and it is important that an image smoothing or filtering technique preserve the directionality. F-actin filaments are widely studied in pathology because the abnormalities in actin dynamics play a key role in diagnosis of cancer, cardiac diseases, vascular diseases, myofibrillar myopathies, neurological disorders, etc. We develop the directional bilateral filter as a means of filtering out the noise in the image without significantly altering the directionality of the F-actin filaments. The bilateral filter is anisotropic to start with, but we add an additional degree of anisotropy by employing an oriented domain kernel for smoothing. The orientation is locally adapted using a structure tensor and the parameters of the bilateral filter are optimized for within the framework of statistical risk minimization. We show that the directional bilateral filter has better denoising performance than the traditional Gaussian bilateral filter and other denoising techniques such as SURE-LET, non-local means, and guided image filtering at various noise levels in terms of peak signal-to-noise ratio (PSNR. We also show quantitative improvements in low NA images of F-actin filaments.

  10. Increasing precision of lifetime determination in fluorescence lifetime imaging

    Science.gov (United States)

    Chang, Ching-Wei; Mycek, Mary-Ann

    2010-02-01

    The interest in fluorescence lifetime imaging microscopy (FLIM) is increasing, as commercial FLIM modules become available for confocal and multi-photon microscopy. In biological FLIM applications, low fluorescence signals from samples can be a challenge, and this causes poor precision in lifetime. In this study, for the first time, we applied wavelet-based denoising methods in time-domain FLIM, and compared them with our previously developed total variation (TV) denoising methods. They were first tested using artificial FLIM images. We then applied them to lowlight live-cell images. The results demonstrated that our TV methods could improve lifetime precision multi-fold in FLIM images and preserve the overall lifetime and pre-exponential term values when improving local lifetime fitting, while wavelet-based methods were faster. The results here can enhance the precision of FLIM, especially for low-light and / or fast video-rate imaging, to improve current and rapidly emerging new applications of FLIM such as live-cell, in vivo whole-animal, or endoscopic imaging.

  11. Fluorescence imaging of dendritic spines of Golgi-Cox-stained neurons using brightening background

    Science.gov (United States)

    Ai, Min; Xiong, Hanqing; Yang, Tao; Shang, Zhenhua; Chen, Muqing; Liu, Xiuli; Zeng, Shaoqun

    2015-01-01

    We report a novel fluorescence imaging approach to imaging nonfluorescence-labeled biological tissue samples. The method was demonstrated by imaging neurons in Golgi-Cox-stained and epoxy-resin-embedded samples through the excitation of the background fluorescence of the specimens. The dark neurons stood out clearly against background fluorescence in the images, enabling the tracing of a single dendritic spine using both confocal and wide-field fluorescence microscopy. The results suggest that the reported fluorescence imaging method would provide an effective alternative solution to image nonfluorescence-labeled samples, and it allows tracing the dendritic spine structure of neurons.

  12. Fluorescence lifetime imaging microscopy of nanodiamonds in vivo

    Science.gov (United States)

    Kuo, Yung; Hsu, Tsung-Yuan; Wu, Yi-Chun; Hsu, Jui-Hung; Chang, Huan-Cheng

    2013-03-01

    The negatively charged nitrogen-vacancy (NV-) center in bulk diamond is a photostable fluorophore with a radiative lifetime of 11.6 ns at room temperature. The lifetime substantially increases to ~20 ns for diamond nanoparticles (size ~ 100 nm) suspended in water due to the change in refractive index of the surrounding medium of the NV- centers. This fluorescence decay time is much longer than that (typically 1 - 4 ns) of endogenous and exogenous fluorophores commonly used in biological imaging, making it possible to detect NV--containing nanodiamonds in vivo at the single particle level by fluorescence lifetime imaging microscopy (FLIM). We demonstrate the feasibility of this approach using Caenorhabditis elegans (C. elegans) as a model organism.

  13. Novel fluorescent carbonic nanomaterials for sensing and imaging

    Science.gov (United States)

    Demchenko, Alexander P.; Dekaliuk, Mariia O.

    2013-12-01

    Small brightly fluorescent carbon nanoparticles have emerged as a new class of materials important for sensing and imaging applications. We analyze comparatively the properties of nanodiamonds, graphene and graphene oxide ‘dots’, of modified carbon nanotubes and of diverse carbon nanoparticles known as ‘C-dots’ obtained by different methods. The mechanisms of their light absorption and luminescence emission are still unresolved and the arguments are presented for their common origin. Regarding present and potential applications, we provide critical comparison with the other types of fluorescence reporters, such as organic dyes and semiconductor quantum dots. Their most prospective applications in sensing (based on the changes of intensity, FRET and lifetime) and in imaging technologies on the level of living cells and whole bodies are overviewed. The possibilities for design on their basis of multifunctional nanocomposites on a broader scale of theranostics are outlined.

  14. Multifunctional ultrasound contrast agents for imaging guided photothermal therapy.

    Science.gov (United States)

    Guo, Caixin; Jin, Yushen; Dai, Zhifei

    2014-05-21

    Among all the imaging techniques, ultrasound imaging has a unique advantage due to its features of real-time, low cost, high safety, and portability. Ultrasound contrast agents (UCAs) have been widely used to enhance ultrasonic signals. One of the most exciting features of UCAs for use in biomedicine is the possibility of easily putting new combinations of functional molecules into microbubbles (MBs), which are the most routinely used UCAs. Various therapeutic agents and medical nanoparticles (quantum dots, gold, Fe3O4, etc.) can be loaded into ultrasound-responsive MBs. Hence, UCAs can be developed as multifunctional agents that integrate capabilities for early detection and diagnosis and for imaging guided therapy of various diseases. The current review will focus on such state-of-the-art UCA platforms that have been exploited for multimodal imaging and for imaging guided photothermal therapy.

  15. Modulated electron-multiplied fluorescence lifetime imaging microscope: all-solid-state camera for fluorescence lifetime imaging.

    Science.gov (United States)

    Zhao, Qiaole; Schelen, Ben; Schouten, Raymond; van den Oever, Rein; Leenen, René; van Kuijk, Harry; Peters, Inge; Polderdijk, Frank; Bosiers, Jan; Raspe, Marcel; Jalink, Kees; Geert Sander de Jong, Jan; van Geest, Bert; Stoop, Karel; Young, Ian Ted

    2012-12-01

    We have built an all-solid-state camera that is directly modulated at the pixel level for frequency-domain fluorescence lifetime imaging microscopy (FLIM) measurements. This novel camera eliminates the need for an image intensifier through the use of an application-specific charge coupled device design in a frequency-domain FLIM system. The first stage of evaluation for the camera has been carried out. Camera characteristics such as noise distribution, dark current influence, camera gain, sampling density, sensitivity, linearity of photometric response, and optical transfer function have been studied through experiments. We are able to do lifetime measurement using our modulated, electron-multiplied fluorescence lifetime imaging microscope (MEM-FLIM) camera for various objects, e.g., fluorescein solution, fixed green fluorescent protein (GFP) cells, and GFP-actin stained live cells. A detailed comparison of a conventional microchannel plate (MCP)-based FLIM system and the MEM-FLIM system is presented. The MEM-FLIM camera shows higher resolution and a better image quality. The MEM-FLIM camera provides a new opportunity for performing frequency-domain FLIM.

  16. Neural imaging in songbirds using fiber optic fluorescence microscopy

    Science.gov (United States)

    Nooshabadi, Fatemeh; Hearn, Gentry; Lints, Thierry; Maitland, Kristen C.

    2012-02-01

    The song control system of juvenile songbirds is an important model for studying the developmental acquisition and generation of complex learned vocal motor sequences, two processes that are fundamental to human speech and language. To understand the neural mechanisms underlying song production, it is critical to characterize the activity of identified neurons in the song control system when the bird is singing. Neural imaging in unrestrained singing birds, although technically challenging, will advance our understanding of neural ensemble coding mechanisms in this system. We are exploring the use of a fiber optic microscope for functional imaging in the brain of behaving and singing birds in order to better understand the contribution of a key brain nucleus (high vocal center nucleus; HVC) to temporal aspects of song motor control. We have constructed a fluorescence microscope with LED illumination, a fiber bundle for transmission of fluorescence excitation and emission light, a ~2x GRIN lens, and a CCD for image acquisition. The system has 2 μm resolution, 375 μm field of view, 200 μm working distance, and 1 mm outer diameter. As an initial characterization of this setup, neurons in HVC were imaged using the fiber optic microscope after injection of quantum dots or fluorescent retrograde tracers into different song nuclei. A Lucid Vivascope confocal microscope was used to confirm the imaging results. Long-term imaging of the activity of these neurons in juvenile birds during singing may lead us to a better understanding of the central motor codes for song and the central mechanism by which auditory experience modifies song motor commands to enable vocal learning and imitation.

  17. Photodegradation-based detection of fluorescent whitening agents in a mountain river.

    Science.gov (United States)

    Assaad, Aziz; Pontvianne, Steve; Pons, Marie-Noëlle

    2014-04-01

    Fluorescent whitening agents (FWAs) are highly soluble and poorly biodegradable ingredients used in laundry detergents and in industries (paper, textile, plastic manufacturing). They are likely to pass through biological wastewater treatment systems. The presence of FWAs in a mountain river was detected by monitoring the decay of synchronous fluorescence intensity at λ(ex)=360 nm after exposing samples to ultraviolet (UV) light (365 nm), for mimicking sunlight, for 15 min. The method was first validated on four commercial FWAs (DAS-1, FB28, DMA-X and CBS-X) in different water matrices (deionized water and pristine river water in the presence of humic acid and dyes). A 40% decay was observed after 15 min for the least photosensitive FWA (CBS-X). A field application was then performed on samples collected along a mountain river in which impacts of FWAs from domestic sources (laundry greywater) and industrial sources (paper and textile mills) were suspected. Variations of fluorescence decay at λ(ex)=360 nm could be explained by these potential sources of pollution. It is suggested that the fluorescence decay at λ(ex)=280 nm also be considered as an indicator, as some FWAs can exhibit fluorescence at that excitation wavelength.

  18. In Vivo Dual Fluorescence Imaging to Detect Joint Destruction.

    Science.gov (United States)

    Cho, Hongsik; Bhatti, Fazal-Ur-Rehman; Lee, Sangmin; Brand, David D; Yi, Ae-Kyung; Hasty, Karen A

    2016-10-01

    Diagnosis of cartilage damage in early stages of arthritis is vital to impede the progression of disease. In this regard, considerable progress has been made in near-infrared fluorescence (NIRF) optical imaging technique. Arthritis can develop due to various mechanisms but one of the main contributors is the production of matrix metalloproteinases (MMPs), enzymes that can degrade components of the extracellular matrix. Especially, MMP-1 and MMP-13 have main roles in rheumatoid arthritis and osteoarthritis because they enhance collagen degradation in the process of arthritis. We present here a novel NIRF imaging strategy that can be used to determine the activity of MMPs and cartilage damage simultaneously by detection of exposed type II collagen in cartilage tissue. In this study, retro-orbital injection of mixed fluorescent dyes, MMPSense 750 FAST (MMP750) dye and Alexa Fluor 680 conjugated monoclonal mouse antibody immune-reactive to type II collagen, was administered in the arthritic mice. Both dyes were detected with different intensity according to degree of joint destruction in the animal. Thus, our dual fluorescence imaging method can be used to detect cartilage damage as well as MMP activity simultaneously in early stage arthritis.

  19. Fluorescent quantum dots: synthesis, biomedical optical imaging, and biosafety assessment.

    Science.gov (United States)

    Ji, Xiaoyuan; Peng, Fei; Zhong, Yiling; Su, Yuanyuan; He, Yao

    2014-12-01

    The marriage of nanomaterials with biology has significantly promoted advancement of biological techniques, profoundly facilitating basic research and practical applications in biological and biomedical fields. Taking advantages of unique optical properties (e.g., strong fluorescence, robust photostability, size-tunable emission wavelengths, etc.), fluorescent quantum dots (QDs), appearing as high-performance biological fluorescent nanoprobes, have been extensively explored for a variety of biomedical optical imaging applications. In this review, we present representative synthetic strategies for preparation of QDs and their applications in biomedical optical imaging, as well as risk assessments in vitro and in vivo. Briefly, we first summarize recent progress in fabrication of QDs via two rudimentary approaches, i.e., organometallic route and aqueous synthesis. Next we present representative achievement in QDs-based in vitro and in vivo biomedical optical imaging applications. We further discuss the toxicity assessment of QDs, ranging from cell studies to animal models. In the final section, we discuss challenges and perspectives for the QDs-relative bioapplications in the future.

  20. All-optically integrated multimodality imaging system: combined photoacoustic microscopy, optical coherence tomography, and fluorescence imaging

    Science.gov (United States)

    Chen, Zhongjiang; Yang, Sihua; Xing, Da

    2016-10-01

    We have developed a multimodality imaging system by optically integrating all-optical photoacoustic microscopy (AOPAM), optical coherence tomography (OCT) and fluorescence microscopy (FLM) to provide complementary information including optical absorption, optical back-scattering and fluorescence contrast of biological tissue. By sharing the same low-coherence Michelson interferometer, AOPAM and OCT could be organically optically combined to obtain the absorption and scattering information of the biological tissues. Also, owing to using the same laser source and objective lens, intrinsically registered photoacoustic and fluorescence signals are obtained to present the radiative and nonradiative transition process of absorption. Simultaneously photoacoustic angiography, tissue structure and fluorescence molecular in vivo images of mouse ear were acquired to demonstrate the capabilities of the optically integrated trimodality imaging system, which can present more information to study tumor angiogenesis, vasculature, anatomical structure and microenvironments in vivo.

  1. Refractive index sensing of green fluorescent proteins in living cells using fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    van Manen, Henk-Jan; Verkuijlen, Paul; Wittendorp, Paul; Subramaniam, Vinod; van den Berg, Timo K; Roos, Dirk; Otto, Cees

    2008-04-15

    We show that fluorescence lifetime imaging microscopy (FLIM) of green fluorescent protein (GFP) molecules in cells can be used to report on the local refractive index of intracellular GFP. We expressed GFP fusion constructs of Rac2 and gp91(phox), which are both subunits of the phagocyte NADPH oxidase enzyme, in human myeloid PLB-985 cells and showed by high-resolution confocal fluorescence microscopy that GFP-Rac2 and GFP-gp91(phox) are targeted to the cytosol and to membranes, respectively. Frequency-domain FLIM experiments on these PLB-985 cells resulted in average fluorescence lifetimes of 2.70 ns for cytosolic GFP-Rac2 and 2.31 ns for membrane-bound GFP-gp91(phox). By comparing these lifetimes with a calibration curve obtained by measuring GFP lifetimes in PBS/glycerol mixtures of known refractive index, we found that the local refractive indices of cytosolic GFP-Rac2 and membrane-targeted GFP-gp91(phox) are approximately 1.38 and approximately 1.46, respectively, which is in good correspondence with reported values for the cytosol and plasma membrane measured by other techniques. The ability to measure the local refractive index of proteins in living cells by FLIM may be important in revealing intracellular spatial heterogeneities within organelles such as the plasma and phagosomal membrane.

  2. Activatable iRGD-based peptide monolith: Targeting, internalization, and fluorescence activation for precise tumor imaging.

    Science.gov (United States)

    Cho, Hong-Jun; Lee, Sung-Jin; Park, Sung-Jun; Paik, Chang H; Lee, Sang-Myung; Kim, Sehoon; Lee, Yoon-Sik

    2016-09-10

    A disulfide-bridged cyclic RGD peptide, named iRGD (internalizing RGD, c(CRGDK/RGPD/EC)), is known to facilitate tumor targeting as well as tissue penetration. After the RGD motif-induced targeting on αv integrins expressed near tumor tissue, iRGD encounters proteolytic cleavage to expose the CendR motif that promotes penetration into cancer cells via the interaction with neuropilin-1. Based on these proteolytic cleavage and internalization mechanism, we designed an iRGD-based monolithic imaging probe that integrates multiple functions (cancer-specific targeting, internalization and fluorescence activation) within a small peptide framework. To provide the capability of activatable fluorescence signaling, we conjugated a fluorescent dye to the N-terminal of iRGD, which was linked to the internalizing sequence (CendR motif), and a quencher to the opposite C-terminal. It turned out that fluorescence activation of the dye/quencher-conjugated monolithic peptide probe requires dual (reductive and proteolytic) cleavages on both disulfide and amide bond of iRGD peptide. Furthermore, the cleavage of the iRGD peptide leading to fluorescence recovery was indeed operative depending on the tumor-related angiogenic receptors (αvβ3 integrin and neuropilin-1) in vitro as well as in vivo. Compared to an 'always fluorescent' iRGD control probe without quencher conjugation, the dye/quencher-conjugated activatable monolithic peptide probe visualized tumor regions more precisely with lower background noise after intravenous injection, owing to the multifunctional responses specific to tumor microenvironment. All these results, along with minimal in vitro and in vivo toxicity profiles, suggest potential of the iRGD-based activatable monolithic peptide probe as a promising imaging agent for precise tumor diagnosis.

  3. Two photon fluorescence imaging of lipid membrane domains and potentials using advanced fluorescent probes

    Science.gov (United States)

    Kilin, Vasyl; Darwich, Zeinab; Richert, Ludovic; Didier, Pascal; Klymchenko, Andrey; Mély, Yves

    2013-02-01

    Biomembranes are ordered and dynamic nanoscale structures critical for cell functions. The biological functions of the membranes strongly depend on their physicochemical properties, such as electrostatics, phase state, viscosity, polarity and hydration. These properties are essential for the membrane structure and the proper folding and function of membrane proteins. To monitor these properties, fluorescence techniques and notably, two-photon microscopy appear highly suited due to their exquisite sensitivity and their capability to operate in complex biological systems, such as living cells and tissues. In this context, we have developed multiparametric environment-sensitive fluorescent probes tailored for precise location in the membrane bilayer. We notably developed probes of the 3-hydroxychromone family, characterized by an excited state intramolecular proton transfer reaction, which generates two tautomeric emissive species with well-separated emission bands. As a consequence, the response of these probes to changes in their environment could be monitored through changes in the ratios of the two bands, as well as through changes in the fluorescence lifetimes. Using two-photon ratiometric imaging and FLIM, these probes were used to monitor the surface membrane potential, and were applied to detect apoptotic cells and image membrane domains.

  4. Differential structured illumination microendoscopy for in vivo imaging of molecular contrast agents

    Science.gov (United States)

    Keahey, Pelham; Ramalingam, Preetha; Schmeler, Kathleen

    2016-01-01

    Fiber optic microendoscopy has shown promise for visualization of molecular contrast agents used to study disease in vivo. However, fiber optic microendoscopes have limited optical sectioning capability, and image contrast is limited by out-of-focus light generated in highly scattering tissue. Optical sectioning techniques have been used in microendoscopes to remove out-of-focus light but reduce imaging speed or rely on bulky optical elements that prevent in vivo imaging. Here, we present differential structured illumination microendoscopy (DSIMe), a fiber optic system that can perform structured illumination in real time for optical sectioning without any opto-mechanical components attached to the distal tip of the fiber bundle. We demonstrate the use of DSIMe during in vivo fluorescence imaging in patients undergoing surgery for cervical adenocarcinoma in situ. Images acquired using DSIMe show greater contrast than standard microendoscopy, improving the ability to detect cellular atypia associated with neoplasia. PMID:27621464

  5. 64Cu loaded liposomes as positron emission tomography imaging agents

    DEFF Research Database (Denmark)

    Petersen, Anncatrine Luisa; Binderup, Tina; Rasmussen, Palle

    2011-01-01

    We have developed a highly efficient method for utilizing liposomes as imaging agents for positron emission tomography (PET) giving high resolution images and allowing direct quantification of tissue distribution and blood clearance. Our approach is based on remote loading of a copper-radionuclid...

  6. Nanogels as imaging agents for modalities spanning the electromagnetic spectrum.

    Science.gov (United States)

    Chan, Minnie; Almutairi, Adah

    2016-01-21

    In the past few decades, advances in imaging equipment and protocols have expanded the role of imaging in in vivo diagnosis and disease management, especially in cancer. Traditional imaging agents have rapid clearance and low specificity for disease detection. To improve accuracy in disease identification, localization and assessment, novel nanomaterials are frequently explored as imaging agents to achieve high detection specificity and sensitivity. A promising material for this purpose are hydrogel nanoparticles, whose high hydrophilicity, biocompatibility, and tunable size in the nanometer range make them ideal for imaging. These nanogels (10 to 200 nm) can circumvent uptake by the reticuloendothelial system, allowing longer circulation times than small molecules. In addition, their size/surface properties can be further tailored to optimize their pharmacokinetics for imaging of a particular disease. Herein, we provide a comprehensive review of nanogels as imaging agents in various modalities with sources of signal spanning the electromagnetic spectrum, including MRI, NIR, UV-vis, and PET. Many materials and formulation methods will be reviewed to highlight the versatility of nanogels as imaging agents.

  7. Folate receptor-targeted fluorescent paramagnetic bimodal liposomes for tumor imaging

    Directory of Open Access Journals (Sweden)

    Ding N

    2011-10-01

    Full Text Available Nan Ding1,2, Yao Lu1, Robert J Lee3, Chang Yang1, Lei Huang1, Jian Liu1, Guangya Xiang1,41School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People’s Republic of China; 2Department of Pharmacy, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People’s Republic of China; 3Division of Pharmaceutics, College of Pharmacy, The Ohio State University, Columbus, Ohio, USA; 4Hubei Key Laboratory of Natural Medicinal Chemistry and Resource Evaluation, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People’s Republic of China The first three authors contributed equally to this work. Rationale and objective: Receptor-targeted delivery of imaging and therapeutic agents can lead to enhanced efficacy for both. Multimodality imaging offers unique advantages over traditional single modality imaging. Tumor marker folate receptor (FR-targeted fluorescent paramagnetic bimodal liposomes were synthesized to co-deliver paramagnetic and fluorescence agents for magnetic resonance (MR and optical bimodal imaging contrast enhancement. Materials and methods: Fluorescent and paramagnetic bimodal liposomes were synthesized with a mean diameter of 136 nm and a low polydispersity index. The liposomes incorporated folate-PEG3350-CHEMS for FR targeting, Gd(III[N,N-Bis-stearylamidomethyl-N’-amidomethyl]diethylenetriamine tetraacetic acid (Gd-DTPA-BSA for MR contrast, and calcein for fluorescence. To determine the specificity and efficiency of delivery, the liposomes were evaluated in FR-positive KB and HeLa cells and FR-negative A549 cells, which were analyzed by fluorescence microscopy, magnetic resonance imaging (MRI, and flow cytometry (FCM. Results: FR-specific and efficient cellular uptake of the FR-targeted bimodal liposomes was confirmed by fluorescence microscopy and by FCM. The mean fluorescence intensity (MFI of KB cells treated with FR

  8. Photouncaging nanoparticles for MRI and fluorescence imaging in vitro and in vivo.

    Science.gov (United States)

    Shibu, Edakkattuparambil S; Ono, Kenji; Sugino, Sakiko; Nishioka, Ayami; Yasuda, Akikazu; Shigeri, Yasushi; Wakida, Shin-Ichi; Sawada, Makoto; Biju, Vasudevanpillai

    2013-11-26

    Multimodal and multifunctional nanomaterials are promising candidates for bioimaging and therapeutic applications in the nanomedicine settings. Here we report the preparation of photouncaging nanoparticles with fluorescence and magnetic modalities and evaluation of their potentials for in vitro and in vivo bioimaging. Photoactivation of such bimodal nanoparticles prepared using photouncaging ligands, CdSe/ZnS quantum dots, and super paramagnetic iron oxide nanoparticles results in the systematic uncaging of the particles, which is correlated with continuous changes in the absorption, mass and NMR spectra of the ligands. Fluorescence and magnetic components of the bimodal nanoparticles are characterized using scanning electron microscopy (SEM), transmission electron microscopy (TEM), and elemental analyses using energy dispersive X-ray (EDX) spectroscopy and X-ray photoelectron spectroscopy (XPS). Bioconjugation of the nanoparticles with peptide hormones renders them with biocompatibility and efficient intracellular transport as seen in the fluorescence and MRI images of mouse melanoma cells (B16) or human lung epithelial adenocarcinoma cells (H1650). Biocompatibility of the nanoparticles is evaluated using MTT cytotoxicity assays, which show cell viability over 90%. Further, we combine MRI and NIR fluorescence imaging in C57BL/6 (B6) mice subcutaneously or intravenously injected with the photouncaging nanoparticles and follow the in vivo fate of the nanoparticles. Interestingly, the intravenously injected nanoparticles initially accumulate in the liver within 30 min post injection and subsequently clear by the renal excretion within 48 h as seen in the time-dependent MRI and fluorescence images of the liver, urinary bladder, and urine samples. Photouncaging ligands such as the ones reported in this article are promising candidates for not only the site-specific delivery of nanomaterials-based contrast agents and drugs but also the systematic uncaging and renal

  9. Normalized fluorescence lifetime imaging for tumor identification and margin delineation

    Science.gov (United States)

    Sherman, Adria J.; Papour, Asael; Bhargava, Siddharth; Taylor, Zach; Grundfest, Warren S.; Stafsudd, Oscar M.

    2013-03-01

    Fluorescence lifetime imaging microscopy (FLIM) is a technique that has been proven to produce quantitative and qualitative differentiation and identification of substances with good specificity and sensitivity based on lifetime extracted information. This technique has shown the ability to also differentiate between a wide range of tissue types to identify malignant from benign tissue in vivo and ex vivo. However, the complexity, long duration and effort required to generate this information has limited the adoption of these techniques in a clinical setting. Our group has developed a time-resolved imaging system (patent pending) that does not require the extraction of lifetimes or use of complex curve fitting algorithms to display the needed information. The technique, entitled Lifetime Fluorescence Imaging (LFI, or NoFYI), converts fluorescence lifetime decay information directly into visual contrast. Initial studies using Fluorescein and Rhodamine-B demonstrated the feasibility of this approach. Subsequent studies demonstrated the ability to separate collagen and elastin powders. The technique uses nanosecond pulsed UV LEDs at 375 nm for average illumination intensities of ~4.5 μW on the tissue surface with detection by a gated CCD camera. To date, we have imaged 11 surgical head and neck squamous cell carcinoma and brain cancer biopsy specimens including 5 normal and 6 malignant samples. Images at multiple wavelengths clearly demonstrate differentiation between benign and malignant tissue, which was later confirmed by histology. Contrast was obtained between fluorophores with 35 μm spatial resolution and an SNR of ~30 dB allowing us to clearly define tumor margins in these highly invasive cancers. This method is capable of providing both anatomical and chemical information for the pathologist and the surgeon. These results suggest that this technology has a possible role in identifying tumors in tissue specimens and detecting tumor margins during procedures.

  10. Intracellular temperature mapping with a fluorescent polymeric thermometer and fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Okabe, Kohki; Inada, Noriko; Gota, Chie; Harada, Yoshie; Funatsu, Takashi; Uchiyama, Seiichi

    2012-02-28

    Cellular functions are fundamentally regulated by intracellular temperature, which influences biochemical reactions inside a cell. Despite the important contributions to biological and medical applications that it would offer, intracellular temperature mapping has not been achieved. Here we demonstrate the first intracellular temperature mapping based on a fluorescent polymeric thermometer and fluorescence lifetime imaging microscopy. The spatial and temperature resolutions of our thermometry were at the diffraction limited level (200 nm) and 0.18-0.58 °C. The intracellular temperature distribution we observed indicated that the nucleus and centrosome of a COS7 cell, both showed a significantly higher temperature than the cytoplasm and that the temperature gap between the nucleus and the cytoplasm differed depending on the cell cycle. The heat production from mitochondria was also observed as a proximal local temperature increase. These results showed that our new intracellular thermometry could determine an intrinsic relationship between the temperature and organelle function.

  11. Simultaneous Fluorescence and Phosphorescence Lifetime Imaging Microscopy in Living Cells.

    Science.gov (United States)

    Jahn, Karolina; Buschmann, Volker; Hille, Carsten

    2015-09-22

    In living cells, there are always a plethora of processes taking place at the same time. Their precise regulation is the basis of cellular functions, since small failures can lead to severe dysfunctions. For a comprehensive understanding of intracellular homeostasis, simultaneous multiparameter detection is a versatile tool for revealing the spatial and temporal interactions of intracellular parameters. Here, a recently developed time-correlated single-photon counting (TCSPC) board was evaluated for simultaneous fluorescence and phosphorescence lifetime imaging microscopy (FLIM/PLIM). Therefore, the metabolic activity in insect salivary glands was investigated by recording ns-decaying intrinsic cellular fluorescence, mainly related to oxidized flavin adenine dinucleotide (FAD) and the μs-decaying phosphorescence of the oxygen-sensitive ruthenium-complex Kr341. Due to dopamine stimulation, the metabolic activity of salivary glands increased, causing a higher pericellular oxygen consumption and a resulting increase in Kr341 phosphorescence decay time. Furthermore, FAD fluorescence decay time decreased, presumably due to protein binding, thus inducing a quenching of FAD fluorescence decay time. Through application of the metabolic drugs antimycin and FCCP, the recorded signals could be assigned to a mitochondrial origin. The dopamine-induced changes could be observed in sequential FLIM and PLIM recordings, as well as in simultaneous FLIM/PLIM recordings using an intermediate TCSPC timing resolution.

  12. Red Fluorescent Carbon Nanoparticle-Based Cell Imaging Probe.

    Science.gov (United States)

    Ali, Haydar; Bhunia, Susanta Kumar; Dalal, Chumki; Jana, Nikhil R

    2016-04-13

    Fluorescent carbon nanoparticle-based probes with tunable visible emission are biocompatible, environment friendly and most suitable for various biomedical applications. However, synthesis of red fluorescent carbon nanoparticles and their transformation into functional nanoparticles are very challenging. Here we report red fluorescent carbon nanoparticle-based nanobioconjugates of nanoparticles are synthesized via high temperature colloid-chemical approach and transformed into water-soluble functional nanoparticles via coating with amphiphilic polymer followed by covalent linking with desired biomolecules. Following this approach, carbon nanoparticles are functionalized with polyethylene glycol, primary amine, glucose, arginine, histidine, biotin and folic acid. These functional nanoparticles can be excited with blue/green light (i.e., 400-550 nm) to capture their emission spanning from 550 to 750 nm. Arginine and folic acid functionalized nanoparticles have been demonstrated as fluorescent cell labels where blue and green excitation has been used for imaging of labeled cells. The presented method can be extended for the development of carbon nanoparticle-based other bioimaging probes.

  13. Development of Fluorescence Imaging Lidar for Boat-Based Coral Observation

    Directory of Open Access Journals (Sweden)

    Sasano Masahiko

    2016-01-01

    Full Text Available A fluorescence imaging lidar system installed in a boat-towable buoy has been developed for the observation of reef-building corals. Long-range fluorescent images of the sea bed can be recorded in the daytime with this system. The viability of corals is clear in these fluorescent images because of the innate fluorescent proteins. In this study, the specifications and performance of the system are shown.

  14. FISHji: New ImageJ macros for the quantification of fluorescence in epifluorescence images

    DEFF Research Database (Denmark)

    Fontenete, Sílvia; Carvalho, Daniel R; Lourenço, Anália

    2016-01-01

    and tools has been trying to overcome this problem, however, the determination of fluorescent intensity in microscopy images still has issues due to the lack of precision in the results and the complexity of existing software. This work presents FISHji, a set of new ImageJ methods for automated...

  15. Synthesis, and Fluorescence Properties of Coumarin and Benzocoumarin Derivatives Conjugated Pyrimidine Scaffolds for Biological Imaging Applications.

    Science.gov (United States)

    Al-Masoudi, Najim A; Al-Salihi, Niran J; Marich, Yossra A; Markus, Timo

    2015-11-01

    Series of coumarin and 5,6-benzomcomarin substituted pyrimidine derivatives 11-15 and 22-25 were synthesized, aiming to develop new imaging fluorescent agents. Analogously, treatment of 4-chloropyrimidine analog 16 with coumarin 3-carbohyrazide 5 under MWI condition followed by boiling with NH4OAc in HOAc furnished coumarin-1,2,4-triazolo-pyrimidine analog 18. The fluorescence property was investigated spectrophotometrically in MeOH with Rhodamine 6G as standard dye. All the compounds showed emission in the region between 331 and 495 nm. The quantum yield of all the compounds were found to be weak, except methyl benzocoumarin 3-carboxylate 22 which showed (ΦF = 0.98) in comparison to Rhodamine 6G as standard (ΦF = 0.95).

  16. Single camera imaging system for color and near-infrared fluorescence image guided surgery.

    Science.gov (United States)

    Chen, Zhenyue; Zhu, Nan; Pacheco, Shaun; Wang, Xia; Liang, Rongguang

    2014-08-01

    Near-infrared (NIR) fluorescence imaging systems have been developed for image guided surgery in recent years. However, current systems are typically bulky and work only when surgical light in the operating room (OR) is off. We propose a single camera imaging system that is capable of capturing NIR fluorescence and color images under normal surgical lighting illumination. Using a new RGB-NIR sensor and synchronized NIR excitation illumination, we have demonstrated that the system can acquire both color information and fluorescence signal with high sensitivity under normal surgical lighting illumination. The experimental results show that ICG sample with concentration of 0.13 μM can be detected when the excitation irradiance is 3.92 mW/cm(2) at an exposure time of 10 ms.

  17. Ultrafast superresolution fluorescence imaging with spinning disk confocal microscope optics.

    Science.gov (United States)

    Hayashi, Shinichi; Okada, Yasushi

    2015-05-01

    Most current superresolution (SR) microscope techniques surpass the diffraction limit at the expense of temporal resolution, compromising their applications to live-cell imaging. Here we describe a new SR fluorescence microscope based on confocal microscope optics, which we name the spinning disk superresolution microscope (SDSRM). Theoretically, the SDSRM is equivalent to a structured illumination microscope (SIM) and achieves a spatial resolution of 120 nm, double that of the diffraction limit of wide-field fluorescence microscopy. However, the SDSRM is 10 times faster than a conventional SIM because SR signals are recovered by optical demodulation through the stripe pattern of the disk. Therefore a single SR image requires only a single averaged image through the rotating disk. On the basis of this theory, we modified a commercial spinning disk confocal microscope. The improved resolution around 120 nm was confirmed with biological samples. The rapid dynamics of micro-tubules, mitochondria, lysosomes, and endosomes were observed with temporal resolutions of 30-100 frames/s. Because our method requires only small optical modifications, it will enable an easy upgrade from an existing spinning disk confocal to a SR microscope for live-cell imaging. © 2015 Hayashi and Okada. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  18. Selective Detection of Neurotransmitters by Fluorescence and Chemiluminescence Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Ziqiang Wang; Edward S. Yeung

    2001-08-06

    In recent years, luminescence imaging has been widely employed in neurochemical analysis. It has a number of advantages for the study of neuronal and other biological cells: (1) a particular molecular species or cellular constituent can be selectively visualized in the presence of a large excess of other species in a heterogeneous environment; (2) low concentration detection limits can be achieved because of the inherent sensitivity associated with fluorescence and chemiluminescence; (3) low excitation intensities can be used so that long-term observation can be realized while the viability of the specimen is preserved; and (4) excellent spatial resolution can be obtained with the light microscope so subcellular compartments can be identified. With good sensitivity, temporal and spatial resolution, the flux of ions and molecules and the distribution and dynamics of intracellular species can be measured in real time with specific luminescence probes, substrates, or with native fluorescence. A noninvasive detection scheme based on glutamate dehydrogenase (GDH) enzymatic assay combined with microscopy was developed to measure the glutamate release in cultured cells from the central nervous system (CNS). The enzyme reaction is very specific and sensitive. The detection limit with CCD imaging is down to {micro}M levels of glutamate with reasonable response time. They also found that chemiluminescence associated with the ATP-dependent reaction between luciferase and luciferin can be used to image ATP at levels down to 10 nM in the millisecond time scale. Similar imaging experiments should be feasible in a broad spectrum of biological systems.

  19. Rapid fluorescence determination of diquat herbicide in food grains using quantum dots as new reducing agent

    Energy Technology Data Exchange (ETDEWEB)

    Carrillo-Carrion, Carolina; Simonet, Bartolome M. [Department of Analytical Chemistry, University of Cordoba, E-14071 Cordoba (Spain); Valcarcel, Miguel, E-mail: qa1meobj@uco.es [Department of Analytical Chemistry, University of Cordoba, E-14071 Cordoba (Spain)

    2011-04-29

    CdSe/ZnS QDs have demonstrated capacity to act as reducing agent in organic media such as acetonitrile and ethanol. By using fluorescence and Raman spectroscopy, it has been demonstrated that QDs reduce diquat herbicide to its monocation radical. The reaction is characterized to present a high reaction rate making possible to perform the reaction by simple filtration of the solution containing the herbicide through a QDs modified filter. The monocation radical presents a high fluorescence emission spectrum which was selected as the analytical signal to quantify the diquat herbicide. The method described here for the analysis of diquat herbicide in oat grains is simple and fast allowing the analysis of trace level of herbicide in only 6 min. The excellent sensitivity and reproducibility of the methods indicate that the reaction is favoured from both thermodynamic and kinetic point of view. The results presented open up the possibility to use QDs as redox agent. The sensitivity of the method expressed as detection limit was only of 0.01 mg kg{sup -1}.The lineal range was between 0.05 and 0.5 mg kg{sup -1}. The time of analysis per sample, including extraction, reaction and fluorescent measurement was only of 6 min.

  20. Nanoparticles and nanocomposites for fluorescence sensing and imaging

    Science.gov (United States)

    Demchenko, Alexander P.

    2013-06-01

    The assortment of fluorescence reporters is changing dramatically. Traditionally explored intrinsic fluorescence of biological macromolecules and cellular pigments and of externally introduced organic dyes are presently in strong competition with new nanomaterials. Among them are conjugated polymers, semiconductor nanocrystals (quantum dots), up-converting nanocrystals, magic-size clusters of silver and gold, nanodiamonds and carbon dots. They demonstrate diverse photophysical behavior and allow one to obtain diverse information when used in analytical tools or when they form images in biological systems. Based on them, functional nanocomposites displaying a variety of useful features, thus extending dramatically the information content of output data, can be constructed. We describe their properties and compare them with those of small-molecular emitters, such as organic dyes. With their aid, one can modulate over a wide range the wavelengths of excitation and emission, the lifetimes and anisotropies and design the systems with ‘superenhancement’ and ‘superquenching’. Such unlimited possibilities are offered by combining different types of luminophores based on electronic conjugation, plasmonic effects or excited-state resonance energy transfer. This tutorial review provides a comparative analysis of the properties of new nanoscale materials and of their hybrid nanocomposites for applications in fluorescence sensing and imaging.

  1. Photostable and photoswitching fluorescent dyes for super-resolution imaging.

    Science.gov (United States)

    Minoshima, Masafumi; Kikuchi, Kazuya

    2017-01-12

    Super-resolution fluorescence microscopy is a recently developed imaging tool for biological researches. Several methods have been developed for detection of fluorescence signals from molecules in a subdiffraction-limited area, breaking the diffraction limit of the conventional optical microscopies and allowing visualization of detailed macromolecular structures in cells. As objectives are exposed to intense laser in the optical systems, fluorophores for super-resolution microscopy must be tolerated even under severe light irradiation conditions. The fluorophores must also be photoactivatable and photoswitchable for single-molecule localization-based super-resolution microscopy, because the number of active fluorophores must be controlled by light irradiation. This has led to growing interest in these properties in the development of fluorophores. In this mini-review, we focus on the development of photostable and photoswitching fluorescent dyes for super-resolution microscopy. We introduce recent efforts, including improvement of fluorophore photostability and control of photoswitching behaviors of fluorophores based on photochemical and photophysical processes. Understanding and manipulation of chemical reactions in excited fluorophores can develop highly photostable and efficiently photoswitchable fluorophores that are suitable for super-resolution imaging applications.

  2. Portable multispectral fluorescence imaging system for food safety applications

    Science.gov (United States)

    Lefcourt, Alan M.; Kim, Moon S.; Chen, Yud-Ren

    2004-03-01

    Fluorescence can be a sensitive method for detecting food contaminants. Of particular interest is detection of fecal contamination as feces is the source of many pathogenic organisms. Feces generally contain chlorophyll a and related compounds due to ingestion of plant materials, and these compounds can readily be detected using fluorescence techniques. Described is a fluorescence-imaging system consisting primarily of a UV light source, an intensified camera with a six-position filter wheel, and software for controlling the system and automatically analyzing the resulting images. To validate the system, orchard apples artificially contaminated with dairy feces were used in a "hands-on" public demonstration. The contamination sites were easily identified using automated edge detection and threshold detection algorithms. In addition, by applying feces to apples and then washing sets of apples at hourly intervals, it was determined that five h was the minimum contact time that allowed identification of the contamination site after the apples were washed. There are many potential uses for this system, including studying the efficacy of apple washing systems.

  3. RNA Imaging with Multiplexed Error Robust Fluorescence in situ Hybridization

    Science.gov (United States)

    Moffitt, Jeffrey R.; Zhuang, Xiaowei

    2016-01-01

    Quantitative measurements of both the copy number and spatial distribution of large fractions of the transcriptome in single-cells could revolutionize our understanding of a variety of cellular and tissue behaviors in both healthy and diseased states. Single-molecule Fluorescence In Situ Hybridization (smFISH)—an approach where individual RNAs are labeled with fluorescent probes and imaged in their native cellular and tissue context—provides both the copy number and spatial context of RNAs but has been limited in the number of RNA species that can be measured simultaneously. Here we describe Multiplexed Error Robust Fluorescence In Situ Hybridization (MERFISH), a massively parallelized form of smFISH that can image and identify hundreds to thousands of different RNA species simultaneously with high accuracy in individual cells in their native spatial context. We provide detailed protocols on all aspects of MERFISH, including probe design, data collection, and data analysis to allow interested laboratories to perform MERFISH measurements themselves. PMID:27241748

  4. Fluorescence and image guided resection in high grade glioma.

    Science.gov (United States)

    Panciani, Pier Paolo; Fontanella, Marco; Schatlo, Bawarjan; Garbossa, Diego; Agnoletti, Alessandro; Ducati, Alessandro; Lanotte, Michele

    2012-01-01

    The extent of resection in high grade glioma is increasingly been shown to positively effect survival. Nevertheless, heterogeneity and migratory behavior of glioma cells make gross total resection very challenging. Several techniques were used in order to improve the detection of residual tumor. Aim of this study was to analyze advantages and limitations of fluorescence and image guided resection. A multicentric prospective study was designed to evaluate the accuracy of each method. Furthermore, the role of 5-aminolevulinc acid and neuronavigation were reviewed. Twenty-three patients harboring suspected high grade glioma, amenable to complete resection, were enrolled. Fluorescence and image guides were used to perform surgery. Multiple samples were obtained from the resection cavity of each lesion according to 5-ALA staining positivity and boundaries as delineated by neuronavigation. All samples were analyzed by a pathologist blinded to the intra-operative labeling. Decision-making based on fluorescence showed a sensitivity of 91.1% and a specificity of 89.4% (pimage-guided resection accuracy was low (sensitivity: 57.8%; specificity: 57.4%; p=0.346). We observed that the sensitivity of 5-ALA can be improved by the combined use of neuronavigation, but this leads to a significant reduction in specificity. Thus, the use of auxiliary techniques should always be subject to critical skills of the surgeon. We advocate a large-scale study to further improve the assessment of multimodal approaches.

  5. Image registration and averaging of low laser power two-photon fluorescence images of mouse retina.

    Science.gov (United States)

    Alexander, Nathan S; Palczewska, Grazyna; Stremplewski, Patrycjusz; Wojtkowski, Maciej; Kern, Timothy S; Palczewski, Krzysztof

    2016-07-01

    Two-photon fluorescence microscopy (TPM) is now being used routinely to image live cells for extended periods deep within tissues, including the retina and other structures within the eye . However, very low laser power is a requirement to obtain TPM images of the retina safely. Unfortunately, a reduction in laser power also reduces the signal-to-noise ratio of collected images, making it difficult to visualize structural details. Here, image registration and averaging methods applied to TPM images of the eye in living animals (without the need for auxiliary hardware) demonstrate the structural information obtained with laser power down to 1 mW. Image registration provided between 1.4% and 13.0% improvement in image quality compared to averaging images without registrations when using a high-fluorescence template, and between 0.2% and 12.0% when employing the average of collected images as the template. Also, a diminishing return on image quality when more images were used to obtain the averaged image is shown. This work provides a foundation for obtaining informative TPM images with laser powers of 1 mW, compared to previous levels for imaging mice ranging between 6.3 mW [Palczewska G., Nat Med.20, 785 (2014) Sharma R., Biomed. Opt. Express4, 1285 (2013)].

  6. Fluorescence Imaging In Vivo up to 1700 nm

    CERN Document Server

    Diao, Shuo; Hong, Guosong; Antaris, Alexander L; Chang, Junlei; Wu, Justin Z; Zhang, Bo; Kuo, Calvin J; Dai, Hongjie

    2015-01-01

    Compared to visible and near-infrared regions below ~ 900 nm, imaging in the second near-infrared window beyond 1000 nm (NIR-II, 1000-1700 nm) is promising for deep-tissue high-resolution optical imaging in vivo owing to reduced scattering of photons traversing through tissues. Here, we succeeded fluorescence imaging in vivo in the long 1500-1700 nm (NIR-IIb) region using a novel, chemical separation enriched large-diameter semiconducting single-walled carbon nanotube material. Imaging in the 1500-1700 nm window resolved 3-4 um wide capillary blood vessels at ~ 3 millimeters depth through the intact body and brain of mice with the ability of blood-flow speed mapping in individual capillary vessels. Further, non-invasive single fluorophore imaging inside the tumor of a live mouse was achieved in the 1500-1700 nm window. NIR-IIb imaging can be generalized to a wide range of fluorophores emitting up to 1700 nm for a new paradigm of high performance in vivo optical imaging.

  7. NOVEL AMPHIPHILIC FLUORESCENT GRAFT COPOLYMER: SYNTHESIS,CHARACTERIZATION AND ENCAPSULATION OF A HYDROPHOBIC AGENT

    Institute of Scientific and Technical Information of China (English)

    Zhao-qiang Wu; Shu Yang; Wen-yan Liao; Ling-zhi Meng

    2006-01-01

    Novel amphiphilic fluorescent graft copolymer (PVP-PyAHy) was successfully synthesized by the free radical copolymerization of hydrophobic monomer N-(1-pyrenebutyryl)-N'-acryloyl hydrazide (PyAHy) with hydrophilic precursor polymers of vinyl-functionalized poly(N-vinylpyrrolidone) (PVP) in DMF. The copolymer is amphiphilic and has intrinsic fluorescence. FT-IR, 1H-NMR, TEM, gel permeation chromatography-multi-angle laser light scattering, UV-Vis spectroscopy and fluorescence spectroscopy were used to characterize this copolymer. The TEM observation shows that the copolymer PVP-PyAHy forms micelles in aqueous solution. Results of fluorometric measurements illustrate that the critical micelle concentration (CMC) value of PVP-PyAHy in aqueous solution is about 0.90 mg/mL. To examine the encapsulation ability of the copolymer in aqueous media, methyl yellow was employed as a model hydrophobic agent. The loading level of the polymer to methyl yellow is 8.8 mg/g. The cytotoxicity assays for Madin Darby Canine Kidney (MDCK) cells shows good biocompatibility of PVP-PyAHy in vitro. These results suggest the potential of this copolymer PVP-PyAHy as drugs delivery carrier and fluorescent tracer.

  8. Near-Infrared Fluorescent NanoGUMBOS for Biomedical Imaging

    Energy Technology Data Exchange (ETDEWEB)

    Bwambok, David [Louisiana State University; El-Zahab, Bilal [Lousianna State University; Challa, Santhosh [Louisiana State University; Li, Min [Lousianna State University; Chandler, Lin [Horiba Jobin Yvon Inc.; Baker, Gary A [ORNL; Warner, Isiah M [ORNL

    2009-01-01

    Herein, we report on near-infrared (NIR) fluorescent nanoparticles generated from an emergent class of materials we refer to as a Group of Uniform Materials Based on Organic Salts (GUMBOS). GUMBOS are largely frozen ionic liquids, although the concept is more general and is also easily applied to solid ionic materials with melting points in excess of 100 C. Nanoparticles based on GUMBOS (nanoGUMBOS) derived from a NIR fluorophore are prepared using a reprecipitation method and evaluated for in vivo fluorescence imaging. Due to their uniformity, single-step preparation, and composite nature, nanoGUMBOS help to resolve issues with dye leakage problems innate to alternate cellular stains and unlock a myriad of applications for these materials, highlighting exciting possibilities for multifunctional nanoGUMBOS.

  9. Development of temperature imaging using two-line atomic fluorescence.

    Science.gov (United States)

    Medwell, Paul R; Chan, Qing N; Kalt, Peter A M; Alwahabi, Zeyad T; Dally, Bassam B; Nathan, Graham J

    2009-02-20

    This work aims to advance understanding of the coupling between temperature and soot. The ability to image temperature using the two-line atomic fluorescence (TLAF) technique is demonstrated. Previous TLAF theory is extended from linear excitation into the nonlinear fluence regime. Nonlinear regime two-line atomic fluorescence (NTLAF) provides superior signal and reduces single-shot uncertainty from 250 K for conventional TLAF down to 100 K. NTLAF is shown to resolve the temperature profile across the stoichiometric envelope for hydrogen, ethylene, and natural gas flames, with deviation from thermocouple measurements not exceeding 100 K, and typically ≲30 K. Measurements in flames containing soot demonstrate good capacity of NTLAF to exclude interferences that hamper most two-dimensional thermometry techniques.

  10. Simultaneous x-ray fluorescence and K-edge CT imaging with photon-counting detectors

    Science.gov (United States)

    Li, Liang; Li, Ruizhe; Zhang, Siyuan; Chen, Zhiqiang

    2016-10-01

    Rapid development of the X-ray phonon-counting detection technology brings tremendous research and application opportunities. In addition to improvements in conventional X-ray imaging performance such as radiation dose utilization and beam hardening correction, photon-counting detectors allows significantly more efficient X-ray fluorescence (XRF) and K-edge imaging, and promises a great potential of X-ray functional, cellular and molecular imaging. XRF is the characteristic emission of secondary X-ray photons from a material excited by initial X-rays. The phenomenon is widely used for chemical and elemental analysis. K-edge imaging identifies a material based on its chemically-specific absorption discontinuity over X-ray photon energy. In this paper, we try to combine XRF and K-edge signals from the contrast agents (e.g., iodine, gadolinium, gold nanoparticles) to simultaneously realize XFCT and K-edge CT imaging for superior image performance. As a prerequisite for this dual-modality imaging, the accurate energy calibration of multi-energy-bin photon-counting detectors is critically important. With the measured XRF data of different materials, we characterize the energy response function of a CZT detector for energy calibration and spectrum reconstruction, which can effectively improve the energy resolution and decrease the inconsistence of the photon counting detectors. Then, a simultaneous K-edge and X-ray fluorescence CT imaging (SKYFI) experimental setup is designed which includes a cone-beam X-ray tube, two separate photon counting detector arrays, a pin-hole collimator and a rotation stage. With a phantom containing gold nanoparticles the two types of XFCT and K-edge CT datasets are collected simultaneously. Then, XFCT and K-edge CT images are synergistically reconstructed in a same framework. Simulation results are presented and quantitative analyzed and compared with the separate XFCT and K-edge CT results.

  11. Defining a superlens operating regime for imaging fluorescent molecules.

    Directory of Open Access Journals (Sweden)

    Kareem Elsayad

    Full Text Available It has been shown that thin metal-based films can at certain frequencies act as planar near-field lenses for certain polarization components. A desirable property of such "lenses" is that they can also enhance and focus some large transverse spatial frequency components which contain sub-diffraction limit details. Over the last decade there has been much work in optimizing designs to reduce effects (such as material losses and surface roughness that are detrimental to image reconstruction. One design that can reduce some of these undesirable effects, and which has received a fair amount of attention recently, is the stacked metal-dielectric superlens. Here we theoretically explore the imaging ability of such a design for the specific purpose of imaging a fluorescent dye (the common bio-marker GFP in the vicinity of the superlens surface. Our calculations take into consideration the interaction (damping of an oscillating electric dipole with the metallic layers in the superlens. We also assume a Gaussian frequency distribution spectrum for the dipole. We treat the metallic-alloy and dielectric-alloy layers separately using an appropriate effective medium theory. The transmission properties are evaluated via Transfer matrix (-matrix calculations that were performed in the MatLab and MathCad environments. Our study shows that it is in principle possible to image fluorescent molecules using a simple bilayer planar superlens. We find that optimal parameters for such a superlens occur when the peak dipole emission-frequency is slightly offset from the Surface Plasmon resonance frequency of the metal-dielectric interfaces. The best resolution is obtained when the fluorescent molecules are not too close (>/ approximately 10 nm or too far (>/approximately 30 nm from the superlens surface. The realization and application of a superlens with the specified design is possible using current nanofabrication techniques. When combined with e.g. a sub

  12. Materials characterization using micro-x-ray fluorescence elemental imaging.

    Energy Technology Data Exchange (ETDEWEB)

    Havrilla, G. J. (George J.); Miller, T. C. (Thomasin C.); Joseph, M. R. (Martha R.)

    2002-01-01

    Materials characterization continues to be a key challenge in a variety of programs. Although bulk elemental composition provides overall concentration of both major and trace elements, the distribution of these elements both on micro and macro scales can determine the performance and ultimately the physical properties of the materials. Hence elemental imaging can provide a new level of information for major and in some cases bulk trace concentrations of elements. Micro X-ray fluorescence (MXRF) offers unique capabilities in terms of elemental imaging. This approach is based on a meso scale level of resolution around 50 micrometer X-ray spot size. When coupled with a moveable stage, specimens several inches on a side can be imaged with surprising detail. In most instances, qualitative images are sufficient to illustrate the elemental heterogeneity. This information can then be used to determine if the material meets the desired physical characteristics and whether this is due to the observed heterogeneity or in spite of it. Several examples of elemental imaging will be presented. These will include the aging of polymers and the effects of residual organotin catalyst. The tin can be imaged using MXRF and has been show to be mobile within the polymeric material over time. Corrosion is a serious issue throughout the industrial world. A specific example of chloride attack on a metal, which creates problems in waste storage. Finally, MXRF used in high throughput screening in the development of novel peptide receptors will be shown. The advantage of MXRF is that no fluorescent tags need be added to the target molecules. This insures the unhindered interaction of the target molecules and allows for additional characterization using molecular spectroscopic techniques.

  13. Synthesis of polymeric fluorescent brightener based on coumarin and its performances on paper as light stabilizer, fluorescent brightener and surface sizing agent

    Science.gov (United States)

    Zhang, Guanghua; Zheng, Hua; Guo, Mingyuan; Du, Lun; Liu, Guojun; Wang, Peng

    2016-03-01

    In this work, a novel polymeric fluorescent brightener based on coumarin (PFBC) was synthesized, using three-step synthetic route, from 7-amino-4-methylcoumarin, coumarin monomer (FBC), Acrylamide (AM) and methacrylatoethyl trimethyl ammonium chloride (DMC). The structure of PFBC was characterized by FT-IR, 1HNMR and GPC. PFBC was applied to paper fiber as light stabilizer, fluorescent brightener and surface sizing agent and its performances were evaluated by measuring the UV-vis, fluorescence, thermal stability, the cationic degree, surface strength and smoothness of paper, the brightness degree of paper and the PC value of paper. Results showed that PFBC had better solubility in water than that of FBC, by measuring the optical properties. Through the surface sizing experiment and UV aging experiment, PFBC not only enhanced the surface strength and smoothness of paper as a surface sizing agent, but also had better effect on anti-UV aging than that of FBC as light stabilizer and fluorescent brightener.

  14. Multispectral fluorescence imaging technique for discrimination of cucumber (Cucumis Sativus) seed viability

    Science.gov (United States)

    In this study, we developed a nondestructive method for discriminating viable cucumber (Cucumis sativus) seeds based on hyperspectral fluorescence imaging. The fluorescence spectra of cucumber seeds in the 420–700 nm range were extracted from hyperspectral fluorescence images obtained using 365 nm u...

  15. Fluorescence and fluorescence-lifetime imaging microscopy (FLIM) to characterize yeast strains by autofluorescence

    Science.gov (United States)

    Bhatta, H.; Goldys, E. M.; Ma, J.

    2006-02-01

    We characterised populations of wild type baking and brewing yeast cells using intrinsic fluorescence and fluorescence lifetime microscopy, in order to obtain quantitative identifiers of different strains. The cell autofluorescence was excited at 405 nm and observed within 440-540 nm range where strong cell to cell variability was observed. The images were analyzed using customised public domain software, which provided information on cell size, intensity and texture-related features. In light of significant diversity of the data, statistical methods were utilized to assess the validity of the proposed quantitative identifiers for strain differentiation. The Kolmogorov-Smirnov test was applied to confirm that empirical distribution functions for size, intensity and entropy for different strains were statistically different. These characteristics were followed with culture age of 24, 48 and 72 h, (the latter corresponding to a stationary growth phase) and size, and to some extent entropy, were found to be independent of age. The fluorescence intensity presented a distinctive evolution with age, different for each of the examined strains. The lifetime analysis revealed a short decay time component of 1.4 ns and a second, longer one with the average value of 3.5 ns and a broad distribution. High variability of lifetime values within cells was observed however a lifetime texture feature in the studied strains was statistically different.

  16. Contrast Agent in Magnetic Resonance Imaging

    DEFF Research Database (Denmark)

    Vu-Quang, Hieu

    2015-01-01

    formulated in order to suppress inflamed cytokine expression by siRNA transfection as well as following the migration of macrophage using MRI and NIR bio-imaging. The nano-complexes could inhibit 50 % mRNA expression and 39 % protein expression. In the in vivo cell tracking NIR bio-imaging and MRI...... for chemotherapy. The nanoparticles were 150 nm in size with spherical shape, which contained PFOB in the inner core and Dox and ICG in the polymeric shell. More importantly, they could target folate receptor expressing cancer cells, which provide positive in vitro and in vivo NIR and 19F MRI results. In project...... stem cells and Raw 264.7 macrophages were chitosan-to-particles weight ratios of w0.1 and w0.01, respectively. In vivo 19F MRI results showed the possibility of capturing labeled cells indicating the potential use of PLGA PFOB in future research involving such as cell migration. . In regard of magnetic...

  17. Fluorescence Quenching Nanoprobes Dedicated to In Vivo Photoacoustic Imaging and High-Efficient Tumor Therapy in Deep-Seated Tissue.

    Science.gov (United States)

    Qin, Huan; Zhou, Ting; Yang, Sihua; Xing, Da

    2015-06-10

    Photoacoustic imaging (PAI) and photoacoustic (PA) therapy have promising applications for treating tumors. It is known that the utilization of high-absorption-coefficient probes can selectively enhance the PAI target contrast and PA tumor therapy efficiency in deep-seated tissue. Here, the design of a probe with the highest availability of optical-thermo conversion by using graphene oxide (GO) and dyes via π-π stacking interactions is reported. The GO serves as a base material for loading dyes and quenching dye fluorescence via fluorescence resonance energy transfer (FRET), with the one purpose of maximum of PA efficiency. Experiments verify that the designed fluorescence quenching nanoprobes can produce stronger PA signals than the sum of the separate signals generated in the dye and the GO. Potential applications of the fluorescence quenching nanoprobes are demonstrated, dedicating to enhance PA contrast of targets in deep-seated tissues and tumors in living mice. PA therapy efficiency both in vitro and in vivo by using the fluorescence quenching nanoprobes is found to be higher than with the commonly used PA therapy agents. Taken together, quenching dye fluorescence via FRET will provide a valid means for developing high-efficiency PA probes. Fluorescence quenching nanoprobes are likely to become a promising candidate for deep-seated tumor imaging and therapy.

  18. Fluorescence lifetime imaging for the characterization of the biochemical composition of atherosclerotic plaques

    Science.gov (United States)

    Phipps, Jennifer; Sun, Yinghua; Saroufeem, Ramez; Hatami, Nisa; Fishbein, Michael C.; Marcu, Laura

    2011-09-01

    This study investigates the ability of a flexible fiberoptic-based fluorescence lifetime imaging microscopy (FLIM) technique to resolve biochemical features in plaque fibrotic cap associated with plaque instability and based solely on fluorescence decay characteristics. Autofluorescence of atherosclerotic human aorta (11 autopsy samples) was measured at 48 locations through two filters, F377: 377/50 and F460: 460/60 nm (center wavelength/bandwidth). The fluorescence decay dynamic was described by average lifetime (τ) and four Laguerre coefficients (LECs) retrieved through a Laguerre deconvolution technique. FLIM-derived parameters discriminated between four groups [elastin-rich (ER), elastin and macrophage-rich (E+M), collagen-rich (CR), and lipid-rich (LR)]. For example, τF377 discriminated ER from CR (R = 0.84); τF460 discriminated E+M from CR and ER (R = 0.60 and 0.54, respectively); LEC-1F377 discriminated CR from LR and E+M (R = 0.69 and 0.77, respectively); P 87% (all cases) and sensitivity as high as 86%. Current results demonstrate for the first time that clinically relevant features (e.g., ratios of lipid versus collagen versus elastin) can be evaluated with a flexible-fiber based FLIM technique without the need for fluorescence intensity information or contrast agents.

  19. Facile synthesis of CdTe@GdS fluorescent-magnetic nanoparticles for tumor-targeted dual-modal imaging.

    Science.gov (United States)

    Zhang, Fei; Kong, Xiu-Qi; Li, Qiong; Sun, Ting-Ting; Chai, Chao; Shen, Wen; Hong, Zhang-Yong; He, Xi-Wen; Li, Wen-You; Zhang, Yu-Kui

    2016-01-01

    Multimodal imaging has made great contribution for diagnosis and therapy of disease since it can provide more effective and complementary information in comparison to any single imaging modality. The design and fabrication of fluorescent-magnetic nanoparticles for multimodal imaging has rapidly developed over the years. Herein, we demonstrate the facile synthesis of GdS coated CdTe nanoparticles (CdTe@GdS NPs) as multimodal agents for fluorescence (FL) and T1-weighted magnetic resonance (MR) imaging. These nanoparticles obtain both prominent fluorescent and paramagnetic properties by coating the GdS shell on the surface of CdTe core via a simple room-temperature route in aqueous solution directly. It is shown that the as-prepared CdTe@GdS NPs have high quantum yield (QY) value of 12% and outstanding longitudinal relaxation rate (r1) of 11.25 mM s(-1), which allow them to be employed as FL/MR dual-modal imaging contrast agents. They also exhibit small particle size of 5 nm, excellent colloidal stability and low cellular toxicity for concentrations up to 750 μg mL(-1). In addition, with the conjugation of folic acid, the nanoparticles were successfully used for tumor-targeted FL/MR dual-modal imaging in vitro and in vivo.

  20. SIMA: Python software for analysis of dynamic fluorescence imaging data

    Directory of Open Access Journals (Sweden)

    Patrick eKaifosh

    2014-09-01

    Full Text Available Fluorescence imaging is a powerful method for monitoring dynamic signals in the nervous system. However, analysis of dynamic fluorescence imaging data remains burdensome, in part due to the shortage of available software tools. To address this need, we have developed SIMA, an open source Python package that facilitates common analysis tasks related to fluorescence imaging. Functionality of this package includes correction of motion artifacts occurring during in vivo imaging with laser-scanning microscopy, segmentation of imaged fields into regions of interest (ROIs, and extraction of signals from the segmented ROIs. We have also developed a graphical user interface (GUI for manual editing of the automatically segmented ROIs and automated registration of ROIs across multiple imaging datasets. This software has been designed with flexibility in mind to allow for future extension with different analysis methods and potential integration with other packages. Software, documentation, and source code for the SIMA package and ROI Buddy GUI are freely available at http://www.losonczylab.org/sima/.

  1. Breast cancer margin delineation with fluorescence lifetime imaging (Conference Presentation)

    Science.gov (United States)

    Phipps, Jennifer E.; Gorpas, Dimitris; Darrow, Morgan; Unger, Jakob; Bold, Richard; Marcu, Laura

    2017-02-01

    The current standard of care for early stages of breast cancer is breast-conserving surgery (BCS). BCS involves a lumpectomy procedure, during which the tumor is removed with a rim of normal tissue-if cancer cells found in that rim of tissue, it is called a positive margin and means part of the tumor remains in the breast. Currently there is no method to determine if cancer cells exist at the margins of lumpectomy specimens aside from time-intensive histology methods that result in reoperations in up to 38% of cases. We used fluorescence lifetime imaging (FLIm) to measure time-resolved autofluorescence from N=13 ex vivo human breast cancer specimens (N=10 patients undergoing lumpectomy or mastectomy) and compared our results to histology. Tumor (both invasive and ductal carcinoma in situ), fibrous tissue, fat and fat necrosis have unique fluorescence signatures. For instance, between 500-580 nm, fluorescence lifetime of tumor was shortest (4.7 +/- 0.4 ns) compared to fibrous tissue (5.5 +/- 0.7 ns) and fat (7.0 +/- 0.1 ns), P<0.05 (ANOVA). These differences are due to the biochemical properties of lipid, nicotineamide adenine dinucleotide (NADH) and collagen fibers in the fat, tumor and fibrous tissue, respectively. Additionally, the FLIm data is augmented to video of the breast tissue with image processing algorithms that track a blue (450 nm) aiming beam used in parallel with the 355 nm excitation beam. This allows for accurate histologic co-registration and in the future will allow for three-dimensional lumpectomy surfaces to be imaged for cancer margin delineation.

  2. FIZICS: fluorescent imaging zone identification system, a novel macro imaging system.

    Science.gov (United States)

    Skwish, Stephen; Asensio, Francisco; King, Greg; Clarke, Glenn; Kath, Gary; Salvatore, Michael J; Dufresne, Claude

    2004-12-01

    Constantly improving biological assay development continues to drive technological requirements. Recently, a specification was defined for capturing white light and fluorescent images of agar plates ranging in size from the NUNC Omni tray (96-well footprint, 128 x 85 mm) to the NUNC Bio Assay Dish (245 x 245 mm). An evaluation of commercially available products failed to identify any system capable of fluorescent macroimaging with discrete wavelength selection. To address the lack of a commercially available system, a custom imaging system was designed and constructed. This system provides the same capabilities of many commercially available systems with the added ability to fluorescently image up to a 245 x 245 mm area using wavelengths in the visible light spectrum.

  3. Gadolinium-based contrast agents in pediatric magnetic resonance imaging

    Energy Technology Data Exchange (ETDEWEB)

    Gale, Eric M.; Caravan, Peter [Massachusetts General Hospital, Harvard Medical School, Department of Radiology, The Martinos Center for Biomedical Imaging, Boston, MA (United States); Rao, Anil G. [Medical University of South Carolina, Department of Radiology and Radiological Science, Charleston, SC (United States); McDonald, Robert J. [College of Medicine, Mayo Clinic, Department of Radiology, Rochester, MN (United States); Winfeld, Matthew [University of Pennsylvania Perelman School of Medicine, Philadelphia, PA (United States); Fleck, Robert J. [Cincinnati Children' s Hospital Medical Center, Department of Pediatric Radiology, Cincinnati, OH (United States); Gee, Michael S. [MassGeneral Hospital for Children, Harvard Medical School, Division of Pediatric Imaging, Department of Radiology, Boston, MA (United States)

    2017-05-15

    Gadolinium-based contrast agents can increase the accuracy and expediency of an MRI examination. However the benefits of a contrast-enhanced scan must be carefully weighed against the well-documented risks associated with administration of exogenous contrast media. The purpose of this review is to discuss commercially available gadolinium-based contrast agents (GBCAs) in the context of pediatric radiology. We discuss the chemistry, regulatory status, safety and clinical applications, with particular emphasis on imaging of the blood vessels, heart, hepatobiliary tree and central nervous system. We also discuss non-GBCA MRI contrast agents that are less frequently used or not commercially available. (orig.)

  4. Cyanine-based probe\\tag-peptide pair fluorescence protein imaging and fluorescence protein imaging methods

    Science.gov (United States)

    Mayer-Cumblidge, M. Uljana; Cao, Haishi

    2013-01-15

    A molecular probe comprises two arsenic atoms and at least one cyanine based moiety. A method of producing a molecular probe includes providing a molecule having a first formula, treating the molecule with HgOAc, and subsequently transmetallizing with AsCl.sub.3. The As is liganded to ethanedithiol to produce a probe having a second formula. A method of labeling a peptide includes providing a peptide comprising a tag sequence and contacting the peptide with a biarsenical molecular probe. A complex is formed comprising the tag sequence and the molecular probe. A method of studying a peptide includes providing a mixture containing a peptide comprising a peptide tag sequence, adding a biarsenical probe to the mixture, and monitoring the fluorescence of the mixture.

  5. Radioiodinated agents for imaging multidrug resistant tumors.

    Science.gov (United States)

    Kortylewicz, Zbigniew P; Augustine, Ann M; Nearman, Jessica; McGarry, Jonathon; Baranowska-Kortylewicz, Janina

    2009-03-01

    Diagnostic agents enabling characterization of multidrug resistance (MDR) in tumors can aid in the selection of chemotherapy regimens. We report here synthesis and evaluation of radiopharmaceuticals based on the second-generation MDR-reversing drug MS-209. 5-[3-{4-(2-Phenyl-2-(4'-[(125)I]iodo-phenyl)acetyl)piperazin-1-yl}-2-hydroxypropoxy]quino-line (17) was prepared from the 4'-tributylstannyl precursor (16) in >95% radiochemical yield. (16) was synthesized in a six-step process with the overall yield of 25%. In vitro studies were conducted in MES-SA (drug-sensitive) and MES-SA/Dx5 (MDR) human uterine sarcoma cell lines. In vivo studies were performed in athymic mice bearing MES-SA and MES-SA/Dx5 xenografts. The uptake of (17) is higher in MES-SA than MES-SA/Dx5 cells. The uptake and efflux of (17) depend on temperature and concentration, and indicate active transport mechanism(s). Incubation of drug sensitive MES-SA cells with verapamil or (15), a nonradioactive analog of (17), alters the cellular retention of radioactivity only marginally. However, MES-SA/Dx5 cells retain approximately 12% more of (17) when incubated with 10 muM verapamil. The addition of (15) or high concentrations of (17) also increase the uptake of (17) in MES-SA/Dx5 up to 200%, depending on the concentration and temperature. The dependence of (17) uptake on the MDR status is also evident in the ex vivo binding studies. In vivo tests in mice xenografted simultaneously with both tumor cell lines indicate distinct pharmacokinetics for each tumor. The absorption half-life in MES-SA/Dx5 xenograft is approximately 10x shorter and the mean residence time approximately 50% shorter compared to MES-SA xenograft in the same mouse. Radioiodinated derivatives of MS-209 appear to be good indicators of multidrug resistance.

  6. Image reconstruction for diagnosis and prognosis of breast cancer using fluorescence measurements: phantom studies

    Science.gov (United States)

    Roy, R.; Godavarty, A.; Thompson, A. B., Jr.; Sevick-Muraca, E. M.

    2005-04-01

    Fluorescence-enhance optical tomography is performed using (i) point illumination and point collection and (ii) area illumination and area collection geometrics in 3D. In both measurement techniques, an image-intensified charge-coupled (ICCD) imaging system is used in the frequency-domain to image near-infrared contrast agents. The experimental measurements are compared to diffusion model predictions in least squares form in the inverse problem. For image recovery for both area and point illumination geometries, an efficient gradient-based optimization technique based on the Penalty/modified barrier function (PMBF) method and the constrained truncated Newton with trust region (CONTN) method is developed. Targets in 3D were reconstructed from experimental data under two conditions of (i) perfect uptake (1:0, target to background ratio) and (ii) imperfect uptake (100:1, target to background ratio). Parameters of absorption cross section due to fluorophore and lifetimes are reconstructed. The present work demonstrates that 3D fluorescence enhanced optical tomography reconstructions can be successfully performed from both point/area illumination and collection experimental measurements of the time-dependent light propagation on clinically relevant tissue phantoms using a gain-modulated ICCD camera.

  7. An (125)I-labeled octavalent peptide fluorescent nanoprobe for tumor-homing imaging in vivo.

    Science.gov (United States)

    Luo, Haiming; Shi, Jiyun; Jin, Honglin; Fan, Di; Lu, Lisen; Wang, Fan; Zhang, Zhihong

    2012-06-01

    Targeting radiopeptides are promising agents for radio-theranostics. However, in vivo evaluation of their targeting specificity is often obscured by their short biologic half-lives and low binding affinities. Here, we report an approach to efficiently examine targeting radiopeptides with a new class of octavalent peptide fluorescent nanoprobe (Octa-FNP) platform, which is composed of candidate targeting peptides and a tetrameric far-red fluorescent protein (tfRFP) scaffold. To shed light on this process, (125)I-Octa-FNP, (125)I-tfRFP and (125)I-peptide were synthesized, and their targeting functionalities were compared. Both fluorescence imaging and radioactive quantification results confirmed that (125)I-Octa-FNP had a significantly higher cellular binding capability than (125)I-tfRFP. In vivo biodistribution studies show that at 6 h post-injection, (125)I-Octa-FNP had 2-fold and 30-fold higher tumor uptake than that of (125)I-tfRFP and (125)I-peptide, respectively. Moreover, γ-imaging at 24 h post-injection revealed a remarkable accumulation of (125)I-Octa-FNP in the tumor while maintaining an extremely low background contrast, which was further confirmed by immunofluorescence analysis. These data suggested that, as an engineered and multivalent platform, Octa-FNP could enhance the tumor targeting of a designed peptide and provide excellent contrast radioimaging, making it a valuable tool for the evaluation of the targeting ability of specifically designed radiopeptides for cancer theranostics.

  8. 4-haloethenylphenyl tropane:serotonin transporter imaging agents

    Science.gov (United States)

    Goodman, Mark M.; Martarello, Laurent

    2005-01-18

    A series of compounds in the 4-fluoroalkyl-3-halophenyl nortropanes and 4-haloethenylphenyl tropane families are described as diagnostic and therapeutic agents for diseases associated with serotonin transporter dysfunction. These compounds bind to serotonin transporter protein with high affinity and selectivity. The invention provides methods of synthesis which incorporate radioisotopic halogens at a last step which permit high radiochemical yield and maximum usable product life. The radiolabeled compounds of the invention are useful as imaging agents for visualizing the location and density of serotonin transporter by PET and SPECT imaging.

  9. Modelling of microcracks image treated with fluorescent dye

    Science.gov (United States)

    Glebov, Victor; Lashmanov, Oleg U.

    2015-06-01

    The main reasons of catastrophes and accidents are high level of wear of equipment and violation of the production technology. The methods of nondestructive testing are designed to find out defects timely and to prevent break down of aggregates. These methods allow determining compliance of object parameters with technical requirements without destroying it. This work will discuss dye penetrant inspection or liquid penetrant inspection (DPI or LPI) methods and computer model of microcracks image treated with fluorescent dye. Usually cracks on image look like broken extended lines with small width (about 1 to 10 pixels) and ragged edges. The used method of inspection allows to detect microcracks with depth about 10 or more micrometers. During the work the mathematical model of image of randomly located microcracks treated with fluorescent dye was created in MATLAB environment. Background noises and distortions introduced by the optical systems are considered in the model. The factors that have influence on the image are listed below: 1. Background noise. Background noise is caused by the bright light from external sources and it reduces contrast on the objects edges. 2. Noises on the image sensor. Digital noise manifests itself in the form of randomly located points that are differing in their brightness and color. 3. Distortions caused by aberrations of optical system. After passing through the real optical system the homocentricity of the bundle of rays is violated or homocentricity remains but rays intersect at the point that doesn't coincide with the point of the ideal image. The stronger the influence of the above-listed factors, the worse the image quality and therefore the analysis of the image for control of the item finds difficulty. The mathematical model is created using the following algorithm: at the beginning the number of cracks that will be modeled is entered from keyboard. Then the point with random position is choosing on the matrix whose size is

  10. Carbon nanotubes allow capture of krypton, barium and lead for multichannel biological X-ray fluorescence imaging

    Science.gov (United States)

    Serpell, Christopher J.; Rutte, Reida N.; Geraki, Kalotina; Pach, Elzbieta; Martincic, Markus; Kierkowicz, Magdalena; de Munari, Sonia; Wals, Kim; Raj, Ritu; Ballesteros, Belén; Tobias, Gerard; Anthony, Daniel C.; Davis, Benjamin G.

    2016-10-01

    The desire to study biology in situ has been aided by many imaging techniques. Among these, X-ray fluorescence (XRF) mapping permits observation of elemental distributions in a multichannel manner. However, XRF imaging is underused, in part, because of the difficulty in interpreting maps without an underlying cellular `blueprint' this could be supplied using contrast agents. Carbon nanotubes (CNTs) can be filled with a wide range of inorganic materials, and thus can be used as `contrast agents' if biologically absent elements are encapsulated. Here we show that sealed single-walled CNTs filled with lead, barium and even krypton can be produced, and externally decorated with peptides to provide affinity for sub-cellular targets. The agents are able to highlight specific organelles in multiplexed XRF mapping, and are, in principle, a general and versatile tool for this, and other modes of biological imaging.

  11. Fluorescence-Doped Particles for Simultaneous Temperature and Velocity Imaging

    Science.gov (United States)

    Danehy, Paul M.; Tiemsin, Pacita I.; Wohl, Chrostopher J.; Verkamp, Max; Lowe, T.; Maisto, P.; Byun, G.; Simpson, R.

    2012-01-01

    Polystyrene latex microspheres (PSLs) have been used for particle image velocimetry (PIV) and laser Doppler velocimetry (LDV) measurements for several decades. With advances in laser technologies, instrumentation, and data processing, the capability to collect more information about fluid flow beyond velocity is possible using new seed materials. To provide additional measurement capability, PSLs were synthesized with temperature-sensitive fluorescent dyes incorporated within the particle. These multifunctional PSLs would have the greatest impact if they could be used in large scale facilities with minimal modification to the facilities or the existing instrumentation. Consequently, several potential dyes were identified that were amenable to existing laser systems currently utilized in wind tunnels at NASA Langley Research Center as well as other wind and fluid (water) tunnels. PSLs incorporated with Rhodamine B, dichlorofluorescein (DCF, also known as fluorescein 548 or fluorescein 27) and other dyes were synthesized and characterized for morphology and spectral properties. The resulting particles were demonstrated to exhibit fluorescent emission, which would enable determination of both fluid velocity and temperature. They also would allow near-wall velocity measurements whereas laser scatter from surfaces currently prevents near-wall measurements using undoped seed materials. Preliminary results in a wind tunnel facility located at Virginia Polytechnic Institute and State University (Virginia Tech) have verified fluorescent signal detection and temperature sensitivity of fluorophore-doped PSLs.

  12. Red emitting neutral fluorescent glycoconjugates for membrane optical imaging.

    Science.gov (United States)

    Redon, Sébastien; Massin, Julien; Pouvreau, Sandrine; De Meulenaere, Evelien; Clays, Koen; Queneau, Yves; Andraud, Chantal; Girard-Egrot, Agnès; Bretonnière, Yann; Chambert, Stéphane

    2014-04-16

    A family of neutral fluorescent probes was developed, mimicking the overall structure of natural glycolipids in order to optimize their membrane affinity. Nonreducing commercially available di- or trisaccharidic structures were connected to a push-pull chromophore based on dicyanoisophorone electron-accepting group, which proved to fluoresce in the red region with a very large Stokes shift. This straightforward synthetic strategy brought structural variations to a series of probes, which were studied for their optical, biophysical, and biological properties. The insertion properties of the different probes into membranes were evaluated on a model system using the Langmuir monolayer balance technique. Confocal fluorescence microscopy performed on muscle cells showed completely different localizations and loading efficiencies depending on the structure of the probes. When compared to the commercially available ANEPPS, a family of commonly used membrane imaging dyes, the most efficient probes showed a similar brightness, but a sharper pattern was observed. According to this study, compounds bearing one chromophore, a limited size of the carbohydrate moiety, and an overall rod-like shape gave the best results.

  13. Iodinated silica/porphyrin hybrid nanoparticles for X-ray computed tomography/fluorescence dual-modal imaging of tumors

    Directory of Open Access Journals (Sweden)

    Koichiro Hayashi

    2014-12-01

    Full Text Available Silica nanoparticles containing covalently linked iodine and a near-infrared (NIR fluorescence dye, namely porphyrin, have been synthesized through a one-pot sol–gel reaction. These particles are called iodinated silica/porphyrin hybrid nanoparticles (ISP HNPs. The ISP HNPs have both high X-ray absorption coefficient and NIR fluorescence. The ISP HNPs modified with folic acid (FA and polyethylene glycol (PEG, denoted as FA-PEG-ISP HNPs, enabled the successful visualization of tumors in mice by both X-ray computed tomography (CT and fluorescence imaging (FI. Thus, the FA-PEG-ISP HNPs are useful as contrast agents or probes for CT/FI dual-modal imaging.

  14. Software Agent with Reinforcement Learning Approach for Medical Image Segmentation

    Institute of Scientific and Technical Information of China (English)

    Mahsa Chitsaz; Chaw Seng Woo

    2011-01-01

    Many image segmentation solutions are problem-based. Medical images have very similar grey level and texture among the interested objects. Therefore, medical image segmentation requires improvements although there have been researches done since the last few decades. We design a self-learning framework to extract several objects of interest simultaneously from Computed Tomography (CT) images. Our segmentation method has a learning phase that is based on reinforcement learning (RL) system. Each RL agent works on a particular sub-image of an input image to find a suitable value for each object in it. The RL system is define by state, action and reward. We defined some actions for each state in the sub-image. A reward function computes reward for each action of the RL agent. Finally, the valuable information, from discovering all states of the interest objects, will be stored in a Q-matrix and the final result can be applied in segmentation of similar images. The experimental results for cranial CT images demonstrated segmentation accuracy above 95%.

  15. Fluorescence-tagged amphiphilic brush copolymer encapsulated Gd2O3 core-shell nanostructures for enhanced T 1 contrast effect and fluorescent imaging

    Science.gov (United States)

    Wang, Fenghe; Peng, Erwin; Liu, Feng; Li, Pingjing; Fong Yau Li, Sam; Xue, Jun Min

    2016-10-01

    To obtain suitable T 1 contrast agents for magnetic resonance imaging (MRI) application, aqueous Gd2O3 nanoparticles (NPs) with high longitudinal relativity (r 1) are demanded. High quality Gd2O3 NPs are usually synthesized through a non-hydrolytic route which requires post-synthetic modification to render the NPs water soluble. The current challenge is to obtain aqueous Gd2O3 NPs with high colloidal stability and enhanced r 1 relaxivity. To overcome this challenge, fluorescence-tagged amphiphilic brush copolymer (AFCP) encapsulated Gd2O3 NPs were proposed as suitable T 1 contrast agents. Such a coating layer provided (i) superior aqueous stability, (ii) biocompatibility, as well as (iii) multi-modality (conjugation with fluorescence dye). The polymeric coating layer thickness was simply adjusted by varying the phase-transfer parameters. By reducing the coating thickness, i.e. the distance between the paramagnetic centre and surrounding water protons, the r 1 relaxivity could be enhanced. In contrast, a thicker polymeric layer coating prevents Gd3+ ions leakage, thus improving its biocompatibility. Therefore, it is important to strike a balance between the biocompatibility and the r 1 relaxivity behaviour. Lastly, by conjugating fluorescence moiety, an additional imaging modality was enabled, as demonstrated from the cell-labelling experiment.

  16. Fluorescence imaging of chromosomal DNA using click chemistry

    Science.gov (United States)

    Ishizuka, Takumi; Liu, Hong Shan; Ito, Kenichiro; Xu, Yan

    2016-09-01

    Chromosome visualization is essential for chromosome analysis and genetic diagnostics. Here, we developed a click chemistry approach for multicolor imaging of chromosomal DNA instead of the traditional dye method. We first demonstrated that the commercially available reagents allow for the multicolor staining of chromosomes. We then prepared two pro-fluorophore moieties that served as light-up reporters to stain chromosomal DNA based on click reaction and visualized the clear chromosomes in multicolor. We applied this strategy in fluorescence in situ hybridization (FISH) and identified, with high sensitivity and specificity, telomere DNA at the end of the chromosome. We further extended this approach to observe several basic stages of cell division. We found that the click reaction enables direct visualization of the chromosome behavior in cell division. These results suggest that the technique can be broadly used for imaging chromosomes and may serve as a new approach for chromosome analysis and genetic diagnostics.

  17. Fluorescent cyanine probe for DNA detection and cellular imaging

    Science.gov (United States)

    Zheng, Yong-Chao; Zheng, Mei-Ling; Zhao, Zhen-Sheng; Duan, Xuan-Ming

    2014-03-01

    In our study, two carbazole-based cyanines, 3,6-bis[2-(1-methylpyridinium)vinyl]-9-methyl carbazole diiodide (A) and 6,6'-bis[2-(1-methylpyridinium)vinyl]-bis(9-methyl-carbazol-3yl)methane diiodide (B) were synthesized and employed as light-up probes for DNA and cell imaging. Both of the cyanine probes possess a symmetric structure and bis-cationic center. The obvious induced circular dichroism signals in circular dichroism spectra reveal that the molecules can specifically interact with DNA. Strong fluorescence enhancement is observed when these two cyanines are bound to DNA. These cyanine probes show high binding affinity to oligonucleotides but different binding preferences to various secondary structures. Confocal microscopy images of fixed cell stained by the probes exhibit strong brightness and high contrast in nucleus with a very low cytoplasmic background.

  18. Fluorescence imaging of chromosomal DNA using click chemistry

    Science.gov (United States)

    Ishizuka, Takumi; Liu, Hong Shan; Ito, Kenichiro; Xu, Yan

    2016-01-01

    Chromosome visualization is essential for chromosome analysis and genetic diagnostics. Here, we developed a click chemistry approach for multicolor imaging of chromosomal DNA instead of the traditional dye method. We first demonstrated that the commercially available reagents allow for the multicolor staining of chromosomes. We then prepared two pro-fluorophore moieties that served as light-up reporters to stain chromosomal DNA based on click reaction and visualized the clear chromosomes in multicolor. We applied this strategy in fluorescence in situ hybridization (FISH) and identified, with high sensitivity and specificity, telomere DNA at the end of the chromosome. We further extended this approach to observe several basic stages of cell division. We found that the click reaction enables direct visualization of the chromosome behavior in cell division. These results suggest that the technique can be broadly used for imaging chromosomes and may serve as a new approach for chromosome analysis and genetic diagnostics. PMID:27620982

  19. Dynamic fluorescence imaging for multiparametric measurement of tumor vasculature

    Science.gov (United States)

    Choi, Myunghwan; Choi, Kyungsun; Ryu, Seung-Wook; Lee, Jungwhoi; Choi, Chulhee

    2011-04-01

    Angiogenesis is essential for tumor growth and a promising target for cancer therapy. Blood vessel monitoring is an indispensable tool for evaluation and development of anti-angiogenic drugs. Here, we report a new noninvasive in vivo imaging tool, named dynamic fluorescence imaging (DyFI), for the simultaneous measurement of multiple vascular parameters including vascular density, perfusion rate, and permeability using spatiotemporal profiles of indocyanine green. Using DyFI in a tumor xenograft model, we quantitatively measured multiple vascular parameters in tumors and normal tissues with high spatial resolution. The multimodality of this method allowed us to find negative spatial correlations between perfusion and permeability. Moreover, DyFI was effective for revealing the early effects of an anti-angiogenic drug. We suggest that DyFI could be a useful tool for the preclinical development of anti-angiogenic drugs.

  20. Sentinel lymph node imaging by a fluorescently labeled DNA tetrahedron.

    Science.gov (United States)

    Kim, Kyoung-Ran; Lee, Yong-Deok; Lee, Taemin; Kim, Byeong-Su; Kim, Sehoon; Ahn, Dae-Ro

    2013-07-01

    Sentinel lymph nodes (SLNs) are the first lymph nodes which cancer cells reach after traveling through lymphatic vessels from the primary tumor. Evaluating the nodal status is crucial in accurate staging of human cancers and accordingly determines prognosis and the most appropriate treatment. The commonly used methods for SLN identification in clinics are based on employment of a colloid of radionuclide or injection of a small dye. Although these methods have certainly contributed to improve surgical practice, new imaging materials are still required to overcome drawbacks of the techniques such as inconvenience of handling radioactive materials and short retention time of small dyes in SLNs. Here, we prepare a fluorescence-labeled DNA tetrahedron and perform SLN imaging by using the DNA nanoconstruct. With a successful identification of SLNs by the DNA nanoconstruct, we suggest that DNA tetrahedron hold great promises for clinical applications.

  1. A Quantitative Method for Microtubule Analysis in Fluorescence Images.

    Science.gov (United States)

    Lan, Xiaodong; Li, Lingfei; Hu, Jiongyu; Zhang, Qiong; Dang, Yongming; Huang, Yuesheng

    2015-12-01

    Microtubule analysis is of significant value for a better understanding of normal and pathological cellular processes. Although immunofluorescence microscopic techniques have proven useful in the study of microtubules, comparative results commonly rely on a descriptive and subjective visual analysis. We developed an objective and quantitative method based on image processing and analysis of fluorescently labeled microtubular patterns in cultured cells. We used a multi-parameter approach by analyzing four quantifiable characteristics to compose our quantitative feature set. Then we interpreted specific changes in the parameters and revealed the contribution of each feature set using principal component analysis. In addition, we verified that different treatment groups could be clearly discriminated using principal components of the multi-parameter model. High predictive accuracy of four commonly used multi-classification methods confirmed our method. These results demonstrated the effectiveness and efficiency of our method in the analysis of microtubules in fluorescence images. Application of the analytical methods presented here provides information concerning the organization and modification of microtubules, and could aid in the further understanding of structural and functional aspects of microtubules under normal and pathological conditions.

  2. Dual-frequency transducer for nonlinear contrast agent imaging.

    Science.gov (United States)

    Guiroy, Axel; Novell, Anthony; Ringgaard, Erling; Lou-Moeller, Rasmus; Grégoire, Jean-Marc; Abellard, André-Pierre; Zawada, Tomasz; Bouakaz, Ayache; Levassort, Franck

    2013-12-01

    Detection of high-order nonlinear components issued from microbubbles has emerged as a sensitive method for contrast agent imaging. Nevertheless, the detection of these high-frequency components, including the third, fourth, and fifth harmonics, remains challenging because of the lack of transducer sensitivity and bandwidth. In this context, we propose a new design of imaging transducer based on a simple fabrication process for high-frequency nonlinear imaging. The transducer is composed of two elements: the outer low-frequency (LF) element was centered at 4 MHz and used in transmit mode, whereas the inner high-frequency (HF) element centered at 14 MHz was used in receive mode. The center element was pad-printed using a lead zirconate titanate (PZT) paste. The outer element was molded using a commercial PZT, and curved porous unpoled PZT was used as backing. Each piezoelectric element was characterized to determine the electromechanical performance with thickness coupling factor around 45%. After the assembly of the two transducer elements, hydrophone measurements (electroacoustic responses and radiation patterns) were carried out and demonstrated a large bandwidth (70% at -3 dB) of the HF transducer. Finally, the transducer was evaluated for contrast agent imaging using contrast agent microbubbles. The results showed that harmonic components (up to the sixth harmonic) of the microbubbles were successfully detected. Moreover, images from a flow phantom were acquired and demonstrated the potential of the transducer for high-frequency nonlinear contrast imaging.

  3. Metal-organic frameworks as sensory materials and imaging agents.

    Science.gov (United States)

    Liu, Demin; Lu, Kuangda; Poon, Christopher; Lin, Wenbin

    2014-02-17

    Metal-organic frameworks (MOFs) are a class of hybrid materials self-assembled from organic bridging ligands and metal ion/cluster connecting points. The combination of a variety of organic linkers, metal ions/clusters, and structural motifs can lead to an infinite array of new materials with interesting properties for many applications. In this Forum Article, we discuss the design and applications of MOFs in chemical sensing and biological imaging. The first half of this article focuses on the development of MOFs as chemical sensors by highlighting how unique attributes of MOFs can be utilized to enhance sensitivity and selectivity. We also discuss some of the issues that need to be addressed in order to develop practically useful MOF sensors. The second half of this article focuses on the design and applications of nanoscale MOFs (NMOFs) as imaging contrast agents. NMOFs possess several interesting attributes, such as high cargo loading capacity, ease of postmodification, tunable size and shape, and intrinsic biodegradability, to make them excellent candidates as imaging contrast agents. We discuss the use of representative NMOFs in magnetic resonance imaging (MRI), X-ray computed tomography (CT), and optical imaging. Although still in their infancy, we believe that the compositional tunability and mild synthetic conditions of NMOF imaging agents should greatly facilitate their further development for clinical translation.

  4. Fluorescence lifetime imaging of membrane lipid order with a ratiometric fluorescent probe.

    Science.gov (United States)

    Kilin, Vasyl; Glushonkov, Oleksandr; Herdly, Lucas; Klymchenko, Andrey; Richert, Ludovic; Mely, Yves

    2015-05-19

    To monitor the lateral segregation of lipids into liquid-ordered (Lo) and -disordered (Ld) phases in lipid membranes, environment-sensitive dyes that partition in both phases but stain them differently have been developed. Of particular interest is the dual-color F2N12S probe, which can discriminate the two phases through the ratio of its two emission bands. These bands are associated with the normal (N(∗)) and tautomer (T(∗)) excited-state species that result from an excited-state intramolecular proton transfer. In this work, we investigated the potency of the time-resolved fluorescence parameters of F2N12S to discriminate lipid phases in model and cell membranes. Both the long and mean lifetime values of the T(∗) form of F2N12S were found to differ by twofold between Ld and Lo phases as a result of the restriction in the relative motions of the two aromatic moieties of F2N12S imposed by the highly packed Lo phase. This differed from the changes in the ratio of the two emission bands between the two phases, which mainly resulted from the decreased hydration of the N(∗) form in the Lo phase. Importantly, the strong difference in lifetimes between the two phases was preserved when cholesterol was added to the Ld phase. The two phases could be imaged with high contrast by fluorescence lifetime imaging microscopy (FLIM) on giant unilamellar vesicles. FLIM images of F2N12S-labeled live HeLa cells confirmed that the plasma membrane was mainly in the Lo-like phase. Furthermore, the two phases were found to be homogeneously distributed all over the plasma membrane, indicating that they are highly mixed at the spatiotemporal resolution of the FLIM setup. Finally, FLIM could also be used to sensitively monitor the change in lipid phase upon cholesterol depletion and apoptosis.

  5. Prussian Blue Modified PLA Microcapsules Containing R6G for Ultrasonic/Fluorescent Bimodal Imaging Guided Photothermal Tumor Therapy.

    Science.gov (United States)

    Feng, Shanshan; Wang, Jinrui; Ma, Fang; Liang, Xiaolong; Li, Xiaoda; Xing, Sen; Yue, Xiuli

    2016-03-01

    A theranostic agent has been successfully constructed for fluorescence/ultrasound dual-modal imaging guided photothermal therapy by loading the fluorescent dye R6G into polylactide microcapsules (PLA MCs) followed by deposition of Prussian blue nanoparticles (PB NPs) into the surface of PLA MCs. It was proved that the obtained microcapsules of R6G@PLA/PB MCs could serve as an efficient probe to simultaneously enhance fluorescence imaging and ultrasound imaging greatly in vivo. R6G@PLA/PB MCs exhibited significant photothermal cytotoxicity. Cancer cells could be killed efficiently through photothermal effects of R6G@PLA/PB MCs due to the strong absorption of PB NPs in the near infrared region under laser irradiation. In a word, R6G@PLA/PB MCs integrate multiple capabilities for effective tumor imaging and therapy. Such a single agent provides us a possibility to interpret accurately the obtained images, identify the size and location of the tumor, as well as guide and monitor the photothermal therapy.

  6. Stable J-aggregation enabled dual photoacoustic and fluorescence nanoparticles for intraoperative cancer imaging

    Science.gov (United States)

    Shakiba, Mojdeh; Ng, Kenneth K.; Huynh, Elizabeth; Chan, Harley; Charron, Danielle M.; Chen, Juan; Muhanna, Nidal; Foster, F. Stuart; Wilson, Brian C.; Zheng, Gang

    2016-06-01

    J-aggregates display nanoscale optical properties which enable their use in fluorescence and photoacoustic imaging applications. However, control over their optical properties in an in vivo setting is hampered by the conformational lability of the J-aggregate structure in complex biological environments. J-aggregating nanoparticles (JNP) formed by self-assembly of bacteriopheophorbide-lipid (Bchl-lipid) in lipid nanovesicles represents a novel strategy to stabilize J-aggregates for in vivo bioimaging applications. We find that 15 mol% Bchl-lipid embedded within a saturated phospholipid bilayer vesicle was optimal in terms of maximizing Bchl-lipid dye loading, while maintaining a spherical nanoparticle morphology and retaining spectral properties characteristic of J-aggregates. The addition of cholesterol maintains the stability of the J-aggregate absorption band for up to 6 hours in the presence of 90% FBS. In a proof-of-concept experiment, we successfully applied JNPs as a fluorescence contrast agent for real-time intraoperative detection of metastatic lymph nodes in a rabbit head-and-neck cancer model. Lymph node metastasis delineation was further verified by visualizing the JNP within the excised lymph node using photoacoustic imaging. Using JNPs, we demonstrate the possibility of using J-aggregates as fluorescence and photoacoustic contrast agents and may potentially spur the development of other nanomaterials that can stably induce J-aggregation for in vivo cancer bioimaging applications.J-aggregates display nanoscale optical properties which enable their use in fluorescence and photoacoustic imaging applications. However, control over their optical properties in an in vivo setting is hampered by the conformational lability of the J-aggregate structure in complex biological environments. J-aggregating nanoparticles (JNP) formed by self-assembly of bacteriopheophorbide-lipid (Bchl-lipid) in lipid nanovesicles represents a novel strategy to stabilize J

  7. Estrogen receptor-targeted optical imaging of breast cancer cells with near-infrared fluorescent dye

    Science.gov (United States)

    Jose, Iven; Deodhar, Kodand; Chiplunkar, Shuba V.; Patkar, Meena

    2010-02-01

    Molecular imaging provides the in vivo characterization of cellular molecular events involved in normal and pathologic processes. With the advent of optical molecular imaging, specific molecules, proteins and genes may be tagged with a luminescent reporter and visualized in small animals. This powerful new tool has pushed in vivo optical imaging to the forefront as it allows for direct determination of drug bio-distribution and uptake kinetics as well as an indicator of biochemical activity and drug efficacy. Although optical imaging encompasses diverse techniques and makes use of various wavelengths of light, a great deal of excitement in molecular research lies in the use of tomographic and fluorescence techniques to image living tissues with near-infrared (NIR) light. Nonionizing, noninvasive near-infrared optical imaging has great potential to become promising alternative for breast cancer detection. Fluorescence spectroscopy studies of human tissue suggest that a variety of lesions show distinct fluorescence spectra compared to those of normal tissue. It has also been shown that exogenous dyes exhibit selective uptake in neoplastic lesions and may offer the best contrast for optical imaging. Use of exogenous agents would provide fluorescent markers, which could serve to detect embedded tumors in the breast. In particular, the ability to monitor the fluorescent yield and lifetime may also enable biochemical specificity if the fluorophore is sensitive to a specific metabolite, such as oxygen. As a first step, we have synthesized and characterized one such NIR fluorescent dye conjugate, which could potentially be used to detect estrogen receptors (ER)[2] . The conjugate was synthesized by ester formation between 17-β estradiol and a hydrophilic derivative of indocyanine green (ICG) cyanine dye, bis-1, 1-(4-sulfobutyl) indotricarbocyanine-5- carboxylic acid, sodium salt. The ester formed was found to have an extra binding ability with the receptor cites as

  8. Ultrasound imaging beyond the vasculature with new generation contrast agents.

    Science.gov (United States)

    Perera, Reshani H; Hernandez, Christopher; Zhou, Haoyan; Kota, Pavan; Burke, Alan; Exner, Agata A

    2015-01-01

    Current commercially available ultrasound contrast agents are gas-filled, lipid- or protein-stabilized microbubbles larger than 1 µm in diameter. Because the signal generated by these agents is highly dependent on their size, small yet highly echogenic particles have been historically difficult to produce. This has limited the molecular imaging applications of ultrasound to the blood pool. In the area of cancer imaging, microbubble applications have been constrained to imaging molecular signatures of tumor vasculature and drug delivery enabled by ultrasound-modulated bubble destruction. Recently, with the rise of sophisticated advancements in nanomedicine, ultrasound contrast agents, which are an order of magnitude smaller (100-500 nm) than their currently utilized counterparts, have been undergoing rapid development. These agents are poised to greatly expand the capabilities of ultrasound in the field of targeted cancer detection and therapy by taking advantage of the enhanced permeability and retention phenomenon of many tumors and can extravasate beyond the leaky tumor vasculature. Agent extravasation facilitates highly sensitive detection of cell surface or microenvironment biomarkers, which could advance early cancer detection. Likewise, when combined with appropriate therapeutic agents and ultrasound-mediated deployment on demand, directly at the tumor site, these nanoparticles have been shown to contribute to improved therapeutic outcomes. Ultrasound's safety profile, broad accessibility and relatively low cost make it an ideal modality for the changing face of healthcare today. Aided by the multifaceted nano-sized contrast agents and targeted theranostic moieties described herein, ultrasound can considerably broaden its reach in future applications focused on the diagnosis and staging of cancer.

  9. Recent advances in the development of amyloid imaging agents.

    Science.gov (United States)

    Furumoto, Shozo; Okamura, Nobuyuki; Iwata, Ren; Yanai, Kazuhiko; Arai, Hiroyuki; Kudo, Yukitsuka

    2007-01-01

    Excessive amyloid-beta (Abeta) deposition in the brain is one of the most crucial events in the early pathological stage of Alzheimer's disease (AD). Therefore, Abeta deposits have enough potential to become a useful biomarker for not only an early diagnosis of AD, but also for the assessment of the clinical efficacy of anti-Abeta therapies, if they can be measured non-invasively and reliably in living patients. As a potent candidate technique to measure this biomarker, PET amyloid imaging using a radioligand for Abeta deposits has received much attention. A large number of Abeta ligands have been synthesized and evaluated as candidates for amyloid imaging agents. These can be classified into six categories of derivatives: Congo-red, Thioflavine T, stilbene, vinylbenzoxazole, DDNP, and miscellaneous. Many of these derivatives exhibit high binding affinities to Abeta fibrils (below 20 nM) and some of them also show excellent brain pharmacokinetic profiles. The concept of amyloid imaging is currently being tested in human PET studies using optimized amyloid imaging agents. Despite the small number of subjects, these studies have demonstrated sufficiently promising results. This review article provides an overview of recent advances in the development of amyloid imaging agents, and includes: a summary of the fundamental basis and clinical significance of amyloid imaging; lists of binding affinity data for 135 compounds classified into 12 molecular frameworks; a comprehensive discussion of the in vitro and in vivo features of representative Abeta ligands; and a discussion of the current state of clinical evaluation of these amyloid imaging agents (PIB, SB-13, BF-227, and FDDNP).

  10. Improved detection of fluorescently labeled microspheres and vessel architecture with an imaging cryomicrotome

    NARCIS (Netherlands)

    P. van Horssen; M. Siebes; I. Hoefer; J.A.E. Spaan; J.P.H.M. van den Wijngaard

    2010-01-01

    Due to spectral overlap, the number of fluorescent labels for imaging cryomicrotome detection was limited to 4. The aim of this study was to increase the separation of fluorescent labels. In the new imaging cryomicrotome, the sample is cut in slices of 40 mu m. Six images are taken for each cutting

  11. A modified phasor approach for analyzing time-gated fluorescence lifetime images

    NARCIS (Netherlands)

    Fereidouni, F.; Esposito, A.; Blab, G.; Gerritsen, H.C.

    2011-01-01

    Fluorescence lifetime imaging is a versatile tool that permits mapping the biochemical environment in the cell. Among various fluorescence lifetime imaging techniques, timecorrelated single photon counting and time-gating methods have been demonstrated to be very efficient and robust for the imaging

  12. Fluorescence imaging and chlorophyll fluorescence to evaluate the role of EDU in UV-B protection in cucumber

    Science.gov (United States)

    Sandhu, Ravinder K.; Kim, Moon S.; Krizek, Donald T.; Middleton, Elizabeth M.

    1997-07-01

    A fluorescence imaging system and chlorophyll fluorescence emissions were used to evaluate whether EDU, N-[2-(2-oxo-1- imidazolidinyl) ethyl]-N'-phenylurea, provided protection against ultraviolet-B (UV-B) irradiation (290 - 320 nm) in cucumber (Cucumis sativus L.) leaves. Plants were grown in growth chambers illuminated for 14 h per day with 400 W high pressure sodium and metal halide lamps. Photosynthetically active radiation (PAR) for 1 hr at the beginning and end of each cycle was provided at 270 micrometers ol m-2 s-1 PAR; during the other 12 hr of the photoperiod, the plants received 840 micrometers ol m-2 s-1 PAR. Beginning on the twelfth day, the plants were exposed to UV-B radiation (0.2 & 18.0 kJ m-2d-1) for 2 days at 8 h per day centered in the photoperiod. Rapidly acquired (less than 1 s), high spatial resolution (less than 1 mm2) images were obtained for whole adaxial leaf surfaces using a fluorescence imaging system. The steady-state fluorescence images were acquired in four spectral regions: blue (F450 nm), green (F550 nm), red (F680 nm), and far-red (F740 nm). Fluorescence emission spectra for leaf pigments extracted in dimethyl sulfoxide (DMSO) were obtained by excitation at 280 and 380 nm (280EX 300 - 530 nm; 380EX 400 - 800 nm). Both UV-B and EDU induced stress responses in cucumber leaves that altered the fluorescence emissions obtained from extracts. In the fluorescence images only UV-B induced stress responses were observed but this damage was detected before it was visually apparent. There was no evidence that EDU afforded protection against UV-B irradiation. Use of fluorescence imaging may provide an early stress detection capability for helping to assess damage to the photosynthetic apparatus of plants.

  13. Zwitterion functionalized gold nanoclusters for multimodal near infrared fluorescence and photoacoustic imaging

    Directory of Open Access Journals (Sweden)

    Danjin Shen

    2017-05-01

    Full Text Available Gold nanoclusters (Au NCs are an emerging type of theranostic agents combining therapeutic and imaging features with reduced toxicity. Au NCs stabilized by a zwitterion ligand with a fine control of the metal core size and the ligand coverage were synthesized by wet chemistry. Intense fluorescence signal is reported for the highest ligand coverage, whereas photoacoustic signal is stronger for the largest metal core. The best Au NC candidate with an average molecular weight of 17 kDa could be detected with high sensitivity on a 2D-near-infrared imaging instrument (limit of detection (LOD = 2.3 μM and by photoacoustic imaging. In vitro and in vivo experiments demonstrate an efficient cell uptake in U87 cell lines, a fast renal clearance (t1/2α = 6.5 ± 1.3 min, and a good correlation between near infrared fluorescence and photoacoustic measurements to follow the early uptake of Au NCs in liver.

  14. Fluorescence Imaging of the Cytoskeleton in Plant Roots.

    Science.gov (United States)

    Dyachok, Julia; Paez-Garcia, Ana; Yoo, Cheol-Min; Palanichelvam, Karuppaiah; Blancaflor, Elison B

    2016-01-01

    During the past two decades the use of live cytoskeletal probes has increased dramatically due to the introduction of the green fluorescent protein. However, to make full use of these live cell reporters it is necessary to implement simple methods to maintain plant specimens in optimal growing conditions during imaging. To image the cytoskeleton in living Arabidopsis roots, we rely on a system involving coverslips coated with nutrient supplemented agar where the seeds are directly germinated. This coverslip system can be conveniently transferred to the stage of a confocal microscope with minimal disturbance to the growth of the seedling. For roots with a larger diameter such as Medicago truncatula, seeds are first germinated in moist paper, grown vertically in between plastic trays, and roots mounted on glass slides for confocal imaging. Parallel with our live cell imaging approaches, we routinely process fixed plant material via indirect immunofluorescence. For these methods we typically use non-embedded vibratome-sectioned and whole mount permeabilized root tissue. The clearly defined developmental regions of the root provide us with an elegant system to further understand the cytoskeletal basis of plant development.

  15. Nonnegative matrix factorization: a blind spectra separation method for in vivo fluorescent optical imaging.

    Science.gov (United States)

    Montcuquet, Anne-Sophie; Hervé, Lionel; Navarro, Fabrice; Dinten, Jean-Marc; Mars, Jérôme I

    2010-01-01

    Fluorescence imaging in diffusive media is an emerging imaging modality for medical applications that uses injected fluorescent markers that bind to specific targets, e.g., carcinoma. The region of interest is illuminated with near-IR light and the emitted back fluorescence is analyzed to localize the fluorescence sources. To investigate a thick medium, as the fluorescence signal decreases with the light travel distance, any disturbing signal, such as biological tissues intrinsic fluorescence (called autofluorescence) is a limiting factor. Several specific markers may also be simultaneously injected to bind to different molecules, and one may want to isolate each specific fluorescent signal from the others. To remove the unwanted fluorescence contributions or separate different specific markers, a spectroscopic approach is explored. The nonnegative matrix factorization (NMF) is the blind positive source separation method we chose. We run an original regularized NMF algorithm we developed on experimental data, and successfully obtain separated in vivo fluorescence spectra.

  16. Intraoperative near-infrared fluorescent imaging during robotic operations.

    Science.gov (United States)

    Macedo, Antonio Luiz de Vasconcellos; Schraibman, Vladimir

    2016-01-01

    The intraoperative identification of certain anatomical structures because they are small or visually occult may be challenging. The development of minimally invasive surgery brought additional difficulties to identify these structures due to the lack of complete tactile sensitivity. A number of different forms of intraoperative mapping have been tried. Recently, the near-infrared fluorescence imaging technology with indocyanine green has been added to robotic platforms. In addition, this technology has been tested in several types of operations, and has advantages such as safety, low cost and good results. Disadvantages are linked to contrast distribution in certain clinical scenarios. The intraoperative near-infrared fluorescent imaging is new and promising addition to robotic surgery. Several reports show the utility of this technology in several different procedures. The ideal dose, time and site for dye injection are not well defined. No high quality evidence-based comparative studies and long-term follow-up outcomes have been published so far. Initial results, however, are good and safe. RESUMO A identificação intraoperatória de certas estruturas anatômicas, por seu tamanho ou por elas serem ocultas à visão, pode ser desafiadora. O desenvolvimento da cirurgia minimamente invasiva trouxe dificuldades adicionais, pela falta da sensibilidade tátil completa. Diversas formas de detecção intraoperatória destas estruturas têm sido tentadas. Recentemente, a tecnologia de fluorescência infravermelha com verde de indocianina foi associada às plataformas robóticas. Além disso, essa tecnologia tem sido testada em uma variedade de cirurgias, e suas vantagens parecem estar ligadas a baixo custo, segurança e bons resultados. As desvantagens estão associadas à má distribuição do contraste em determinados cenários. A imagem intraoperatória por fluorescência infravermelha é uma nova e promissora adição à cirurgia robótica. Diversas séries mostram

  17. High resolution fluorescent bio-imaging with electron beam excitation.

    Science.gov (United States)

    Kawata, Yoshimasa; Nawa, Yasunori; Inami, Wataru

    2014-11-01

    We have developed electron beam excitation assisted (EXA) optical microscope[1-3], and demonstrated its resolution higher than 50 nm. In the microscope, a light source in a few nanometers size is excited by focused electron beam in a luminescent film. The microscope makes it possible to observe dynamic behavior of living biological specimens in various surroundings, such as air or liquids. Scan speed of the nanometric light source is faster than that in conventional near-field scanning optical microscopes. The microscope enables to observe optical constants such as absorption, refractive index, polarization, and their dynamic behavior on a nanometric scale. The microscope opens new microscopy applications in nano-technology and nano-science.Figure 1(a) shows schematic diagram of the proposed EXA microscope. An electron beam is focused on a luminescent film. A specimen is put on the luminescent film directly. The inset in Fig. 1(a) shows magnified image of the luminescent film and the specimen. Nanometric light source is excited in the luminescent film by the focused electron beam. The nanometric light source illuminates the specimen, and the scattered or transmitted radiation is detected with a photomultiplier tube (PMT). The light source is scanned by scanning of the focused electron beam in order to construct on image. Figure 1(b) shows a luminescence image of the cells acquired with the EXA microscope, and Fig. 1(c) shows a phase contrast microscope image. Cells were observed in culture solution without any treatments, such as fixation and drying. The shape of each cell was clearly recognized and some bright spots were observed in cells. We believe that the bright spots indicated with arrows were auto-fluorescence of intracellular granules and light- grey regions were auto-fluorescence of cell membranes. It is clearly demonstrated that the EXA microscope is useful tool for observation of living biological cells in physiological conditions.jmicro;63/suppl_1/i

  18. Facile and green approach to prepare fluorescent carbon dots: Emergent nanomaterial for cell imaging and detection of vitamin B2.

    Science.gov (United States)

    Kundu, Aniruddha; Nandi, Sudipta; Das, Pradip; Nandi, Arun K

    2016-04-15

    Carbon dots (CDs) are a new representative in carbonaceous family and have initiated remarkable research interests over the past one decade in a large variety of fields. Herein, we have utilized a facile, one-step carbonization method to prepare fluorescent carbon dots using poly(vinyl alcohol) (PVA) both as a carbon source and as a surface passivating agent. The as prepared CDs emit bright blue fluorescence under ultraviolet illumination. The structure and optical properties of the CDs are thoroughly investigated by several methods such as high-resolution transmission electron microscopy; dynamic light scattering; UV-vis, fluorescence and Fourier transform infrared spectroscopy. The CDs exhibit excellent water solubility and demonstrate average hydrodynamic diameter of 11.3 nm, holding great promise for biological applications. The biocompatibility evaluation and in vitro imaging study reveals that the synthesized CDs can be used as effective fluorescent probes in bio-imaging without noticeable cytotoxicity. In addition, a unique sensor for the detection of vitamin B2 in aqueous solution is proposed on the basis of spontaneous fluorescence resonance energy transfer from CD to vitamin B2. These findings therefore suggest that the CDs can find potential applications in cellular imaging along with sensing of vitamin B2.

  19. In vivo tomographic imaging of lung colonization of tumour in mouse with simultaneous fluorescence and X-ray CT.

    Science.gov (United States)

    Zhang, Bin; Gao, Fuping; Wang, Mengjiao; Cao, Xu; Liu, Fei; Wang, Xin; Luo, Jianwen; Wang, Guangzhi; Bai, Jing

    2014-01-01

    Non-invasive in vivo imaging of diffuse and wide-spread colonization within the lungs, rather than distinct solid primary tumors, is still a challenging work. In this work, a lung colonization mouse model bearing A549 human lung tumor was simultaneously scanned by a dual-modality fluorescence molecular tomography (FMT) and X-ray computed tomography (CT) system in vivo. A two steps method which incorporates CT structural information into the FMT reconstruction procedure is employed to provide concurrent anatomical and functional information. By using the target-specific fluorescence agent, the fluorescence tomographic results show elevated fluorescence intensity deep within the lungs which is colonized with diffuse and wide-spread tumors. The results were confirmed with ex vivo fluorescence reflectance imaging and histological examination of the lung tissues. With FMT reconstruction combined with the CT information, the dual-modality FMT/micro-CT system is expected to offer sensitive and noninvasive imaging of diffuse tumor colonization within the lungs in vivo. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Robust overlay schemes for the fusion of fluorescence and color channels in biological imaging.

    Science.gov (United States)

    Glatz, Jürgen; Symvoulidis, Panagiotis; Garcia-Allende, P Beatriz; Ntziachristos, Vasilis

    2014-04-01

    Molecular fluorescence imaging is a commonly used method in various biomedical fields and is undergoing rapid translation toward clinical applications. Color images are commonly superimposed with fluorescence measurements to provide orientation, anatomical information, and molecular tissue properties in a single image. New adaptive methods that produce a more robust composite image than conventional lime green alpha blending are presented and demonstrated herein. Moreover, visualization through temporal changes is showcased as an alternative for real-time imaging systems.

  1. In Vivo Follow-up of Brain Tumor Growth via Bioluminescence Imaging and Fluorescence Tomography

    Directory of Open Access Journals (Sweden)

    Coralie Genevois

    2016-10-01

    Full Text Available Reporter gene-based strategies are widely used in experimental oncology. Bioluminescence imaging (BLI using the firefly luciferase (Fluc as a reporter gene and d-luciferin as a substrate is currently the most widely employed technique. The present paper compares the performances of BLI imaging with fluorescence imaging using the near infrared fluorescent protein (iRFP to monitor brain tumor growth in mice. Fluorescence imaging includes fluorescence reflectance imaging (FRI, fluorescence diffuse optical tomography (fDOT, and fluorescence molecular Imaging (FMT®. A U87 cell line was genetically modified for constitutive expression of both the encoding Fluc and iRFP reporter genes and assayed for cell, subcutaneous tumor and brain tumor imaging. On cultured cells, BLI was more sensitive than FRI; in vivo, tumors were first detected by BLI. Fluorescence of iRFP provided convenient tools such as flux cytometry, direct detection of the fluorescent protein on histological slices, and fluorescent tomography that allowed for 3D localization and absolute quantification of the fluorescent signal in brain tumors.

  2. Enhanced live cell imaging via photonic crystal enhanced fluorescence microscopy.

    Science.gov (United States)

    Chen, Weili; Long, Kenneth D; Yu, Hojeong; Tan, Yafang; Choi, Ji Sun; Harley, Brendan A; Cunningham, Brian T

    2014-11-21

    We demonstrate photonic crystal enhanced fluorescence (PCEF) microscopy as a surface-specific fluorescence imaging technique to study the adhesion of live cells by visualizing variations in cell-substrate gap distance. This approach utilizes a photonic crystal surface incorporated into a standard microscope slide as the substrate for cell adhesion, and a microscope integrated with a custom illumination source as the detection instrument. When illuminated with a monochromatic light source, angle-specific optical resonances supported by the photonic crystal enable efficient excitation of surface-confined and amplified electromagnetic fields when excited at an on-resonance condition, while no field enhancement occurs when the same photonic crystal is illuminated in an off-resonance state. By mapping the fluorescence enhancement factor for fluorophore-tagged cellular components between on- and off-resonance states and comparing the results to numerical calculations, the vertical distance of labelled cellular components from the photonic crystal substrate can be estimated, providing critical and quantitative information regarding the spatial distribution of the specific components of cells attaching to a surface. As an initial demonstration of the concept, 3T3 fibroblast cells were grown on fibronectin-coated photonic crystals with fluorophore-labelled plasma membrane or nucleus. We demonstrate that PCEF microscopy is capable of providing information about the spatial distribution of cell-surface interactions at the single-cell level that is not available from other existing forms of microscopy, and that the approach is amenable to large fields of view, without the need for coupling prisms, coupling fluids, or special microscope objectives.

  3. Fluorescent Cy5 silica nanoparticles for cancer cell imaging

    Science.gov (United States)

    O'Connell, Claire; Nooney, Robert I.; Glynn, MacDara; Ducree, Jens; McDonagh, Colette

    2015-08-01

    Cancer is a leading cause of death worldwide, with metastasis responsible for the majority of cancer-related deaths. Circulating tumour cells (CTCs) play a central role in metastasis. Fluorescent silica particles (NPs), of diameter ~50 nm which contain a large concentration of Cy5 dye molecules and are extremely bright, have been developed to detect these rare CTCs. Due to this brightness, the particles have superior performance compared to single Cy5 dye molecule labels, for detecting cancer cells. Fluorescence measurements show that the NPs are almost 100 times brighter than the free dye. They do not photo bleach as readily and, due to the biocompatible silica surface, they can be chemically modified, layer-by-layer, in order to bind to cells. The choice of these chemical layers, in particular the NP to antibody linker, along with the incubation period and type of media used in the incubation, has a strong influence on the specific binding abilities of the NPs. In this work, NPs have been shown to selectively bind to the MCF-7 cell line by targeting epithelial cellular adhesion molecule (EpCAM) present on the MCF-7 cell membrane by conjugating anti-EpCAM antibody to the NP surface. Results have shown a high signal to noise ratio for this cell line in comparison to a HeLa control line. NP attachment to cells was verified qualitatively with the use of fluorescence microscopy and quantitatively using image analysis methods. Once the system has been optimised, other dyes will be doped into the silica NPs and their use in multiplexing will be investigated.

  4. Analysis of human aorta using fluorescence lifetime imaging microscopy (FLIM)

    Science.gov (United States)

    Vieira-Damiani, Gislaine; Adur, J.; Ferro, D. P.; Adam, R. L.; Pelegati, V.; Thomáz, A.; Cesar, C. L.; Metze, K.

    2012-03-01

    The use of photonics has improved our understanding of biologic phenomena. For the study of the normal and pathologic architecture of the aorta the use of Two-Photon Excited Fluorescence (TPEF) and Second Harmonic Generation showed interesting details of morphologic changes of the elastin-collagen architecture during aging or development of hypertension in previous studies. In this investigation we tried to apply fluorescence lifetime imaging (FLIM) for the morphologic analysis of human aortas. The aim of our study was to use FLIM in non-stained formalin-fixed and paraffin-embedded samples of the aorta ascendants in hypertensive and normotensive patients of various ages, examining two different topographical regions. The FLIM-spectra of collagen and elastic fibers were clearly distinguishable, thus permitting an exact analysis of unstained material on the microscopic level. Moreover the FLIM spectrum of elastic fibers revealed variations between individual cases, which indicate modifications on a molecular level and might be related to FLIM age or diseases states and reflect modifications on a molecular level.

  5. Application of hyperspectral fluorescence lifetime imaging to tissue autofluorescence: arthritis

    Science.gov (United States)

    Talbot, C. B.; Benninger, R. K. P.; de Beule, P.; Requejo-Isidro, J.; Elson, D. S.; Dunsby, C.; Munro, I.; Neil, M. A.; Sandison, A.; Sofat, N.; Nagase, H.; French, P. M. W.; Lever, M. J.

    2005-08-01

    Tissue contains many natural fluorophores and therefore by exploiting autofluorescence, we can obtain information from tissue with less interference than conventional histological techniques. However, conventional intensity imaging is prone to artifacts since it is an absolute measurement. Fluorescence lifetime and spectral measurements are relative measurements and therefore allow for better measurements. We have applied FLIM and hyperspectral FLIM to the study of articular cartilage and its disease arthritis. We have analyzed normal human articular cartilage and cartilage which was in the early stages of disease. In this case, it was found that FLIM was able to detect changes in the diseased tissue that were not detectable with the conventional diagnosis. Specifically, the fluorescence lifetimes (FL) of the cells were different between the two samples. We have also applied hyperspectral FLIM to degraded cartilage through treatment with interleukin-1. In this case, it was found that there was a shift in the emission spectrum with treatment and that the lifetime had also increased. We also showed that there was greater contrast between the cells and the extracellular matrix (ECM) at longer wavelengths.

  6. Near-infrared (NIR) fluorescence imaging of head and neck squamous cell carcinoma for fluorescence-guided surgery (Conference Presentation)

    Science.gov (United States)

    Moore, Lindsay; Warram, Jason M.; de Boer, Esther; Carroll, William R.; Morlandt, Anthony; Withrow, Kirk P.; Rosenthal, Eben L.

    2016-03-01

    During fluorescence-guided surgery, a cancer-specific optical probe is injected and visualized using a compatible device intraoperatively to provide visual contrast between diseased and normal tissues to maximize resection of cancer and minimize the resection of precious adjacent normal tissues. Six patients with squamous cell carcinomas of the head and neck region (oral cavity (n=4) or cutaneous (n=2)) were injected with an EGFR-targeting antibody (Cetuximab) conjugated to a near-infrared (NIR) fluorescent dye (IRDye800) 3, 4, or 7 days prior to surgical resection of the cancer. Each patient's tumor was then imaged using a commercially available, open-field NIR fluorescence imaging device each day prior to surgery, intraoperatively, and post-operatively. The mean fluorescence intensity (MFI) of the tumor was calculated for each specimen at each imaging time point. Adjacent normal tissue served as an internal anatomic control for each patient to establish a patient-matched "background" fluorescence. Resected tissues were also imaged using a closed-field NIR imaging device. Tumor to background ratios (TBRs) were calculated for each patient using both devices. Fluorescence histology was correlated with traditional pathology assessment to verify the specificity of antibody-dye conjugate binding. Peak TBRs using the open-field device ranged from 2.2 to 11.3, with an average TBR of 4.9. Peak TBRs were achieved between days 1 and 4. This study demonstrated that a commercially available NIR imaging device suited for intraoperative and clinical use can successfully be used with a fluorescently-labeled dye to delineate between diseased and normal tissue in this single cohort human study, illuminated the potential for its use in fluoresence-guided surgery.

  7. Single aflatoxin contaminated corn kernel analysis with fluorescence hyperspectral image

    Science.gov (United States)

    Yao, Haibo; Hruska, Zuzana; Kincaid, Russell; Ononye, Ambrose; Brown, Robert L.; Cleveland, Thomas E.

    2010-04-01

    Aflatoxins are toxic secondary metabolites of the fungi Aspergillus flavus and Aspergillus parasiticus, among others. Aflatoxin contaminated corn is toxic to domestic animals when ingested in feed and is a known carcinogen associated with liver and lung cancer in humans. Consequently, aflatoxin levels in food and feed are regulated by the Food and Drug Administration (FDA) in the US, allowing 20 ppb (parts per billion) limits in food and 100 ppb in feed for interstate commerce. Currently, aflatoxin detection and quantification methods are based on analytical tests including thin-layer chromatography (TCL) and high performance liquid chromatography (HPLC). These analytical tests require the destruction of samples, and are costly and time consuming. Thus, the ability to detect aflatoxin in a rapid, nondestructive way is crucial to the grain industry, particularly to corn industry. Hyperspectral imaging technology offers a non-invasive approach toward screening for food safety inspection and quality control based on its spectral signature. The focus of this paper is to classify aflatoxin contaminated single corn kernels using fluorescence hyperspectral imagery. Field inoculated corn kernels were used in the study. Contaminated and control kernels under long wavelength ultraviolet excitation were imaged using a visible near-infrared (VNIR) hyperspectral camera. The imaged kernels were chemically analyzed to provide reference information for image analysis. This paper describes a procedure to process corn kernels located in different images for statistical training and classification. Two classification algorithms, Maximum Likelihood and Binary Encoding, were used to classify each corn kernel into "control" or "contaminated" through pixel classification. The Binary Encoding approach had a slightly better performance with accuracy equals to 87% or 88% when 20 ppb or 100 ppb was used as classification threshold, respectively.

  8. A 3-D fluorescence imaging system incorporating structured illumination technology

    Science.gov (United States)

    Antos, L.; Emord, P.; Luquette, B.; McGee, B.; Nguyen, D.; Phipps, A.; Phillips, D.; Helguera, M.

    2010-02-01

    A currently available 2-D high-resolution, optical molecular imaging system was modified by the addition of a structured illumination source, OptigridTM, to investigate the feasibility of providing depth resolution along the optical axis. The modification involved the insertion of the OptigridTM and a lens in the path between the light source and the image plane, as well as control and signal processing software. Projection of the OptigridTM onto the imaging surface at an angle, was resolved applying the Scheimpflug principle. The illumination system implements modulation of the light source and provides a framework for capturing depth resolved mages. The system is capable of in-focus projection of the OptigridTM at different spatial frequencies, and supports the use of different lenses. A calibration process was developed for the system to achieve consistent phase shifts of the OptigridTM. Post-processing extracted depth information using depth modulation analysis using a phantom block with fluorescent sheets at different depths. An important aspect of this effort was that it was carried out by a multidisciplinary team of engineering and science students as part of a capstone senior design program. The disciplines represented are mechanical engineering, electrical engineering and imaging science. The project was sponsored by a financial grant from New York State with equipment support from two industrial concerns. The students were provided with a basic imaging concept and charged with developing, implementing, testing and validating a feasible proof-of-concept prototype system that was returned to the originator of the concept for further evaluation and characterization.

  9. Monomeric red fluorescent protein variants used for imaging studies in different species

    NARCIS (Netherlands)

    Mueller-Taubenberger, Annette; Vos, Michel J.; Boettger, Angelika; Lasi, Margherita; Lai, Frank P. L.; Fischer, Markus; Rottner, Klemens

    2006-01-01

    Fluorescent proteins have proven to be excellent tools for live-cell imaging studies. In addition to green fluorescent protein (GFP) and its variants, recent progress was achieved in the development of monomeric red fluorescent proteins (mRFPs) that show improved properties in respect to maturation

  10. Applying two-photon excitation fluorescence lifetime imaging microscopy to study photosynthesis in plant leaves

    NARCIS (Netherlands)

    Broess, K.; Borst, J.W.; Amerongen, van H.

    2009-01-01

    This study investigates to which extent two-photon excitation (TPE) fluorescence lifetime imaging microscopy can be applied to study picosecond fluorescence kinetics of individual chloroplasts in leaves. Using femtosecond 860 nm excitation pulses, fluorescence lifetimes can be measured in leaves of

  11. Inspection of fecal contamination on strawberries using fluorescence imaging

    Science.gov (United States)

    Chuang, Yung-Kun; Yang, Chun-Chieh; Kim, Moon S.; Delwiche, Stephen R.; Lo, Y. Martin; Chen, Suming; Chan, Diane E.

    2013-05-01

    Fecal contamination of produce is a food safety issue associated with pathogens such as Escherichia coli that can easily pollute agricultural products via animal and human fecal matters. Outbreaks of foodborne illnesses associated with consuming raw fruits and vegetables have occurred more frequently in recent years in the United States. Among fruits, strawberry is one high-potential vector of fecal contamination and foodborne illnesses since the fruit is often consumed raw and with minimal processing. In the present study, line-scan LED-induced fluorescence imaging techniques were applied for inspection of fecal material on strawberries, and the spectral characteristics and specific wavebands of strawberries were determined by detection algorithms. The results would improve the safety and quality of produce consumed by the public.

  12. Aptamer-assembled nanomaterials for fluorescent sensing and imaging

    Directory of Open Access Journals (Sweden)

    Lu Danqing

    2017-01-01

    Full Text Available Aptamers, which are selected in vitro by a technology known as the systematic evolution of ligands by exponential enrichment (SELEX, represent a crucial recognition element in molecular sensing. With advantages such as good biocompatibility, facile functionalization, and special optical and physical properties, various nanomaterials can protect aptamers from enzymatic degradation and nonspecific binding in living systems and thus provide a preeminent platform for biochemical applications. Coupling aptamers with various nanomaterials offers many opportunities for developing highly sensitive and selective sensing systems. Here, we focus on the recent applications of aptamer-assembled nanomaterials in fluorescent sensing and imaging. Different types of nanomaterials are examined along with their advantages and disadvantages. Finally, we look toward the future of aptamer-assembled nanomaterials.

  13. Aptamer-assembled nanomaterials for fluorescent sensing and imaging

    Science.gov (United States)

    Lu, Danqing; He, Lei; Zhang, Ge; Lv, Aiping; Wang, Ruowen; Zhang, Xiaobing; Tan, Weihong

    2017-01-01

    Aptamers, which are selected in vitro by a technology known as the systematic evolution of ligands by exponential enrichment (SELEX), represent a crucial recognition element in molecular sensing. With advantages such as good biocompatibility, facile functionalization, and special optical and physical properties, various nanomaterials can protect aptamers from enzymatic degradation and nonspecific binding in living systems and thus provide a preeminent platform for biochemical applications. Coupling aptamers with various nanomaterials offers many opportunities for developing highly sensitive and selective sensing systems. Here, we focus on the recent applications of aptamer-assembled nanomaterials in fluorescent sensing and imaging. Different types of nanomaterials are examined along with their advantages and disadvantages. Finally, we look toward the future of aptamer-assembled nanomaterials.

  14. Dual-modality, fluorescent, PLGA encapsulated bismuth nanoparticles for molecular and cellular fluorescence imaging and computed tomography

    Science.gov (United States)

    Swy, Eric R.; Schwartz-Duval, Aaron S.; Shuboni, Dorela D.; Latourette, Matthew T.; Mallet, Christiane L.; Parys, Maciej; Cormode, David P.; Shapiro, Erik M.

    2014-10-01

    Reports of molecular and cellular imaging using computed tomography (CT) are rapidly increasing. Many of these reports use gold nanoparticles. Bismuth has similar CT contrast properties to gold while being approximately 1000-fold less expensive. Herein we report the design, fabrication, characterization, and CT and fluorescence imaging properties of a novel, dual modality, fluorescent, polymer encapsulated bismuth nanoparticle construct for computed tomography and fluorescence imaging. We also report on cellular internalization and preliminary in vitro and in vivo toxicity effects of these constructs. 40 nm bismuth(0) nanocrystals were synthesized and encapsulated within 120 nm Poly(dl-lactic-co-glycolic acid) (PLGA) nanoparticles by oil-in-water emulsion methodologies. Coumarin-6 was co-encapsulated to impart fluorescence. High encapsulation efficiency was achieved ~70% bismuth w/w. Particles were shown to internalize within cells following incubation in culture. Bismuth nanocrystals and PLGA encapsulated bismuth nanoparticles exhibited >90% and >70% degradation, respectively, within 24 hours in acidic, lysosomal environment mimicking media and both remained nearly 100% stable in cytosolic/extracellular fluid mimicking media. μCT and clinical CT imaging was performed at multiple X-ray tube voltages to measure concentration dependent attenuation rates as well as to establish the ability to detect the nanoparticles in an ex vivo biological sample. Dual fluorescence and CT imaging is demonstrated as well. In vivo toxicity studies in rats revealed neither clinically apparent side effects nor major alterations in serum chemistry and hematology parameters. Calculations on minimal detection requirements for in vivo targeted imaging using these nanoparticles are presented. Indeed, our results indicate that these nanoparticles may serve as a platform for sensitive and specific targeted molecular CT and fluorescence imaging.Reports of molecular and cellular imaging using

  15. Self-Assembled Polyelectrolyte Nanoparticles as Fluorophore-Free Contrast Agents for Multicolor Optical Imaging

    Directory of Open Access Journals (Sweden)

    Da Hye Shin

    2015-03-01

    Full Text Available In this work, we describe the fabrication of self-assembled polyelectrolyte nanoparticles that provide a multicolor optical imaging modality. Poly(γ-glutamic acid(γ-PGA formed self-assembled nanoparticles through electrostatic interactions with two different cationic polymers: poly(L-lysine(PLL and chitosan. The self-assembled γ-PGA/PLL and γ-PGA/chitosan nanoparticles were crosslinked by glutaraldehyde. Crosslinking of the ionic self-assembled nanoparticles with glutaraldehyde not only stabilized the nanoparticles but also generated a strong autofluorescence signal. Fluorescent Schiff base bonds (C=N and double bonds (C=C were generated simultaneously by crosslinking of the amine moiety of the cationic polyelectrolytes with monomeric glutaraldehyde or with polymeric glutaraldehyde. The unique optical properties of the nanoparticles that resulted from the crosslinking by glutaraldehyde were analyzed using UV/Vis and fluorescence spectroscopy. We observed that the fluorescence intensity of the nanoparticles could be regulated by adjusting the crosslinker concentration and the reaction time. The nanoparticles also exhibited high performance in the labeling and monitoring of therapeutic immune cells (macrophages and dendritic cells. These self-assembled nanoparticles are expected to be a promising multicolor optical imaging contrast agent for the labeling, detection, and monitoring of cells.

  16. Study of adsorption kinetics for fluorescent whitening agent on fiber surfaces

    Institute of Scientific and Technical Information of China (English)

    HUANG XianNan; HUANG YanGui; CHAI XinSheng; WEI Wei

    2008-01-01

    Adsorption kinetics for a stilbene derivative type fluorescent whitening agent (FWA) on fiber surfaces was studied based on a real-time spectroscopic measurement system. Results showed that the time-dependent behavior of FWA agrees to that of a mono-molecular adsorption layer on fiber surfaces, as characterized in Langmuir-type expression. The adsorption has two distinguishable stages include-ing initial fast phase with the primary constant of 1.51×1014 and the later near-equilibrium phase with the secondary constant of 4.96×10-4, The maximum amount of FWA adsorbed on fiber surfaces is 1.67×10-4g (per dry weight based fiber) in the initial phase. A mathematical model of adsorption kinetics was therefore established and evaluated, This model is important for the optimization of FWA applica-tion in papermaking.

  17. Study of adsorption kinetics for fluorescent whitening agent on fiber surfaces

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Adsorption kinetics for a stilbene derivative type fluorescent whitening agent (FWA) on fiber surfaces was studied based on a real-time spectroscopic measurement system. Results showed that the time-dependent behavior of FWA agrees to that of a mono-molecular adsorption layer on fiber surfaces,as characterized in Langmuir-type expression. The adsorption has two distinguishable stages includ-ing initial fast phase with the primary constant of 1.51×1014 and the later near-equilibrium phase with the secondary constant of 4.96×10-4. The maximum amount of FWA adsorbed on fiber surfaces is 1.67×10-4g (per dry weight based fiber) in the initial phase. A mathematical model of adsorption kinetics was therefore established and evaluated. This model is important for the optimization of FWA applica-tion in papermaking.

  18. Near-infrared fluorescence imaging with a mobile phone (Conference Presentation)

    Science.gov (United States)

    Ghassemi, Pejhman; Wang, Bohan; Wang, Jianting; Wang, Quanzeng; Chen, Yu; Pfefer, T. Joshua

    2017-03-01

    Mobile phone cameras employ sensors with near-infrared (NIR) sensitivity, yet this capability has not been exploited for biomedical purposes. Removing the IR-blocking filter from a phone-based camera opens the door to a wide range of techniques and applications for inexpensive, point-of-care biophotonic imaging and sensing. This study provides proof of principle for one of these modalities - phone-based NIR fluorescence imaging. An imaging system was assembled using a 780 nm light source along with excitation and emission filters with 800 nm and 825 nm cut-off wavelengths, respectively. Indocyanine green (ICG) was used as an NIR fluorescence contrast agent in an ex vivo rodent model, a resolution test target and a 3D-printed, tissue-simulating vascular phantom. Raw and processed images for red, green and blue pixel channels were analyzed for quantitative evaluation of fundamental performance characteristics including spectral sensitivity, detection linearity and spatial resolution. Mobile phone results were compared with a scientific CCD. The spatial resolution of CCD system was consistently superior to the phone, and green phone camera pixels showed better resolution than blue or green channels. The CCD exhibited similar sensitivity as processed red and blue pixels channels, yet a greater degree of detection linearity. Raw phone pixel data showed lower sensitivity but greater linearity than processed data. Overall, both qualitative and quantitative results provided strong evidence of the potential of phone-based NIR imaging, which may lead to a wide range of applications from cancer detection to glucose sensing.

  19. Application of fluorescent tracer agent technology to point-of-care gastrointestinal permeability measurement

    Science.gov (United States)

    Dorshow, Richard B.; Shieh, Jeng-Jong; Rogers, Thomas E.; Hall-Moore, Carla; Shaikh, Nurmohammad; Talcott, Michael; Tarr, Phillip I.

    2016-03-01

    Gut dysfunction, often accompanied by increased mucosal permeability to gut contents, frequently accompanies a variety of human intestinal inflammatory conditions. These disorders include inflammatory bowel diseases (e.g., Crohn's Disease) and environmental enteropathy and enteric dysfunction, a condition strongly associated with childhood malnutrition and stunting in resource poor areas of the world. The most widely used diagnostic assay for gastrointestinal permeability is the lactulose to mannitol ratio (L:M) measurement. These sugars are administered orally, differentially absorbed by the gut, and then cleared from the body by glomerular filtration in the kidney. The amount of each sugar excreted in the urine is measured. The larger sugar, lactulose, is minimally absorbed through a healthy gut. The smaller sugar, mannitol, in contrast, is readily absorbed through both a healthy and injured gut. Thus a higher ratio of lactulose to mannitol reflects increased intestinal permeability. However, several issues prevent widespread use of the L:M ratio in clinical practice. Urine needs to be collected over time intervals of several hours, the specimen then needs to be transported to an analytical laboratory, and sophisticated equipment is required to measure the concentration of each sugar in the urine. In this presentation we show that fluorescent tracer agents with molecular weights similar to those of the sugars, selected from our portfolio of biocompatible renally cleared fluorophores, mimic the L:M ratio test for gut permeability. This fluorescent tracer agent detection technology can be used to overcome the limitations of the L:M assay, and is amenable to point-of-care clinical use.

  20. Screening system of blocking agents of the receptor for advanced glycation endproducts in cells using fluorescence.

    Science.gov (United States)

    Jung, Dong Ho; Kim, Young Sook; Kim, Jin Sook

    2012-01-01

    Activation of the receptor for advanced glycation endproducts (RAGE) triggers cellular responses implicated in the pathogenesis of diabetic complications; blockade of RAGE has been shown to inhibit the development of diabetic complications. To develop a screening system to identify novel disruptors of advanced glycation endproducts (AGE)-RAGE binding, we used an AGE-RAGE binding system in RAGE-overexpressing cells; test compounds were screened using this system. To construct human RAGE-overexpressing cells, mouse mesangial cells (MMCs) were stably transfected with the pcDNA-human RAGE (hRAGE) vector and selected under 1 mg/mL gentamicin (G418). RAGE expression in hRAGE-overexpressing MMCs was analyzed by Western blotting with specific RAGE antibody. To identify novel disruptors of AGE-RAGE binding, 50 single compounds and AGE-bovine serum albumin (BSA)-Alexa 488 (AGE-BSA labeled with Alexa 488) were treated to the hRAGE-overexpressing MMCs. Nonbinding AGE-BSA-Alexa 488 was washed and fluorescence measured by microtiter plate reader (excitation wavelength, 485 nm; emission wavelength, 528 nm). In hRAGE-overexpressing cells, only treatment with AGE-BSA-Alexa 488 significantly increased fluorescence intensity in a dose-dependent manner. Of 50 compounds tested, genistein disrupted AGE-RAGE binding in a dose-dependent manner. This AGE-RAGE binding system using AGE-BSA-Alexa 488 in hRAGE-overexpressing cells was suitable for screening of agents that disrupt AGE-hRAGE binding.

  1. Deep-red polymer dots with bright two-photon fluorescence and high biocompatibility for in vivo mouse brain imaging

    Science.gov (United States)

    Alifu, Nuernisha; Sun, Zezhou; Zebibula, Abudureheman; Zhu, Zhenggang; Zhao, Xinyuan; Wu, Changfeng; Wang, Yalun; Qian, Jun

    2017-09-01

    With high contrast and deep penetration, two-photon fluorescence (2PF) imaging has become one of the most promising in vivo fluorescence imaging techniques. To obtain good imaging contrast, fluorescent nanoprobes with good 2PF properties are highly needed. In this work, bright 2PF polymer dots (P dots) were applied for in vivo mouse brain imaging. Deep-red emissive P dots with PFBT as the donor and PFDBT5 as the acceptor were synthesized and used as a contrast agent. P dots were further encapsulated by poly(styrene-co-maleic anhydride) (PSMA) and grafted with poly(ethylene glycol) (PEG). The P dots-PEG exhibit large two-photon absorption (2PA) cross-sections (δ≥8500 g), good water dispersibility, and high biocompatibility. P dots-PEG was further utilized first time for in vivo vascular imaging of mouse ear and brain, under 690-900 nm femtosecond (fs) laser excitation. Due to the large 2PA cross-section and deep-red emission, a large imaging depth ( 720 μm) was achieved.

  2. Development of [F-18]-Labeled Amyloid Imaging Agents for PET

    Energy Technology Data Exchange (ETDEWEB)

    Mathis, CA

    2007-05-09

    The applicant proposes to design and synthesize a series of fluorine-18-labeled radiopharmaceuticals to be used as amyloid imaging agents for positron emission tomography (PET). The investigators will conduct comprehensive iterative in vitro and in vivo studies based upon well defined acceptance criteria in order to identify lead agents suitable for human studies. The long term goals are to apply the selected radiotracers as potential diagnostic agents of Alzheimer's disease (AD), as surrogate markers of amyloid in the brain to determine the efficacy of anti-amyloid therapeutic drugs, and as tools to help address basic scientific questions regarding the progression of the neuropathology of AD, such as testing the "amyloid cascade hypothesis" which holds that amyloid accumulation is the primary cause of AD.

  3. Integrated Lymphography using Fluorescence Imaging and Magnetic Resonance Imaging in Intact Mice

    Directory of Open Access Journals (Sweden)

    Yusuke Inoue

    2011-09-01

    Full Text Available We assessed lymph drainage in living mice by an integrated imaging method using fluorescence imaging (FLI and magnetic resonance imaging (MRI. Mice were subcutaneously injected with quantum dots and gadofluorine 8 into the right rear footpad. They were fixed on a transparent flat plate and underwent FLI and MRI successively. Small markers were attached to the mouse surface for spatial coregistration, and image fusion of FLIs and MRIs was performed. Two-dimensional fluorescence reflectance imaging was used for FLI. FLI and MRI provided generally consistent results and demonstrated lymphatic flow to the popliteal, sacral, and iliac lymph nodes in most mice and to the renal, inguinal, and lumbar-aortic lymph nodes in some mice. On the fusion images, the locations of the lymph nodes in the mouse trunk were in good agreement between FLI and MRI, indicating successful spatial registration even for the deep structures. The popliteal node tended to be visualized a little farther caudally in FLI than in MRI, presumably because the overlying tissues were thicker in the cranial portion. Integrated FLI/MRI lymphography with image fusion appears to be a useful tool for analysis of the murine lymphatic system.

  4. The potential of a fluorescent-based approach for bioassay of antifungal agents against chili anthracnose disease in Thailand.

    Science.gov (United States)

    Chutrakul, Chanikul; Khaokhajorn, Pratoomporn; Auncharoen, Patchanee; Boonruengprapa, Tanapong; Mongkolporn, Orarat

    2013-01-01

    Severe chili anthracnose disease in Thailand is caused by Colletotrichum gloeosporioides and C. capsici. To discover anti-anthracnose substances we developed an efficient dual-fluorescent labeling bioassay based on a microdilution approach. Indicator strains used in the assay were constructed by integrating synthetic green fluorescent protein (sGFP) and Discosoma sp. red fluorescent protein (DsRedExp) genes into the genomes of C. gloeosporioides or C. capsici respectively. Survival of co-spore cultures in the presence of inhibitors was determined by the expression levels of these fluorescent proteins. This developed assay has high potential for utilization in the investigation of selective inhibition activity to either one of the pathogens as well as the broad-range inhibitory effect against both pathogens. The value of using the dual-fluorescent assay is rapid, reliable, and consistent identification of anti-anthracnose agents. Most of all, the assay enables the identification of specific inhibitors under the co-cultivation condition.

  5. Two-photon fluorescence and confocal reflected light imaging of thick tissue structures

    Science.gov (United States)

    Kim, Ki H.; So, Peter T. C.; Kochevar, Irene E.; Masters, Barry R.; Gratton, Enrico

    1998-04-01

    The technology of two-photon excitation has opened a window of opportunity for developing non-invasive medical diagnostic tools capable of monitoring thick tissue biochemical states. Using cellular endogenous chromophores, (beta) -nicotinamide- adenine dinucleotide phosphate [NAD(P)H], the cellular metabolic rates in living human skin were determined. Although important functional information can be obtained from the fluorescence spectroscopy of endogenous chromophores, these chromophores are rather poor contrast enhancing agent for mapping cellular morphology. First, most endogenous chromophores are confined to the cellular cytoplasm which prevents the visualization of other cellular organelles. Second, there is significant variability in the distribution and the quantum yield of endogenous chromophores which depends on tissue biochemistry but prevents consistent comparison of cellular morphology. On the other hand, the deep tissue cellular morphology has been imaged with excellent resolution using reflected light confocal microscopy. In reflected light microscopy, the image contrast originates from the index of refraction differences of the cellular structures. The organelle boundaries with significant index differences such as the plasma membrane and the nucleus envelope can be consistently visualized. A combination of morphological and functional information is required for a thorough tissue study. This presentation describes the development of a new microscope which is capable of simultaneously collecting both two-photon fluorescence and confocal reflected light signals. Promising biomedical applications include the non-invasive diagnosis of skin cancer and the study of wound healing.

  6. In vivo imaging agents: an international market report

    Energy Technology Data Exchange (ETDEWEB)

    1990-03-01

    The purpose of this study is to provide a global perspective of the in vivo imaging agents business to market planning executives who are working for companies that develop, produce and distribute various types of in vivo imaging agents. Others that could find this study useful include investment bankers, regulatory and governmental authorities and purchasers of these products. The study attempts to diligently provide market data by type for important geographic markets - Western Europe, the U.S.A., and Japan. A competitive intelligence section which discusses companies involved in these markets constitutes the last part of this study. These profiles are not intended to extensively evaluate each company's marketing strengths or strategies but to provide a general idea of the market presence and prospects. A combination of primary and secondary research is used for all findings. (author).

  7. High contrast imaging of reversibly switchable fluorescent proteins via temporally unmixed Multispectral Optoacoustic Tomography (tuMSOT)

    CERN Document Server

    Stiel, Andre C; Jiang, Yuanyuan; Ntziachristos, Vasilis; Razansky, Daniel; Westmeyer, Gil G

    2014-01-01

    Photocontrol of reversibly switchable fluorescent proteins (RSFPs) was used to program optoacoustic signal time courses that were temporally unmixed to increase the proteins contrast-to-noise-ratios (CNRs) in optoacoustic imaging. In this way, two variants of the RSFP Dronpa with very similar optoacoustic spectra could be readily discriminated in the presence of highly absorbing blood. Addition of temporal unmixing to multispectral optoacoustic tomography (tuMSOT) in conjunction with synthetic or genetically controlled photochromic contrast agents and customized photoswitching schedules can increase the performance of multiplexed and high contrast molecular optoacoustic imaging.

  8. Perfluorocarbon Nanoparticles: Evolution of a Multimodality and Multifunctional Imaging Agent

    Directory of Open Access Journals (Sweden)

    Patrick M. Winter

    2014-01-01

    Full Text Available Perfluorocarbon nanoparticles offer a biologically inert, highly stable, and nontoxic platform that can be specifically designed to accomplish a range of molecular imaging and drug delivery functions in vivo. The particle surface can be decorated with targeting ligands to direct the agent to a variety of biomarkers that are associated with diseases such as cancer, cardiovascular disease, obesity, and thrombosis. The surface can also carry a high payload of imaging agents, ranging from paramagnetic metals for MRI, radionuclides for nuclear imaging, iodine for CT, and florescent tags for histology, allowing high sensitivity mapping of cellular receptors that may be expressed at very low levels in the body. In addition to these diagnostic imaging applications, the particles can be engineered to carry highly potent drugs and specifically deposit them into cell populations that display biosignatures of a variety of diseases. The highly flexible and robust nature of this combined molecular imaging and drug delivery vehicle has been exploited in a variety of animal models to demonstrate its potential impact on the care and treatment of patients suffering from some of the most debilitating diseases.

  9. Cellular uptake of modified oligonucleotides enhanced by porphyrins studied by time-resolved microspectrofluorimetry and fluorescence imaging techniques

    Science.gov (United States)

    Praus, P.; Kočišová, E.; Mojzeš, P.; Štěpánek, J.; Turpin, P.-Y.; Sureau, F.

    2011-05-01

    Fluorescence microimaging and homodyne phase-resolved confocal microspectrofluorimetry were used to monitor the transport of antisense oligonucleotide into 3T3 living cells and its subsequent intracellular distribution. Phosphorothioate analog of 15-mer oligothymidylate labeled by ATTO 425 was complexed with 5,10,15,20-tetrakis (1-methyl-4-pyridyl) porphyrin (H 2TMPyP4) as an uptake-mediating agent. High frequency (up to 180 MHz) analog modulation of both exciting diode laser and the detector image intensifier gain was used to record time-resolved fluorescence spectra. Fluorescence lifetime data within a broad spectral range have revealed preservation of oligonucleotide/porphyrin complex integrity and binding properties of both components inside the cell.

  10. Development of a Multifaceted Ovarian Cancer Imaging Agent

    Science.gov (United States)

    2010-04-01

    method for a recombinant disintegrin vicrostatin (VN), whose structure is based on the snake venom disintegrin contortrostatin (CN), and the use of the...TITLE: Development of a Multifaceted Ovarian Cancer Imaging Agent PRINCIPAL INVESTIGATOR: Francis S . Markland Ph.D...distribution unlimited The views, opinions and/or findings contained in this report are those of the author( s ) and should not be construed

  11. Synthesis of serotonin transporter imaging agent [125I]ADAM

    Institute of Scientific and Technical Information of China (English)

    LU Chun-Xiong; WU Chun-Ying; JIANG Quan-Fu; CHEN Zheng-Ping; ZHANG Tong-Xing; LI Xiao-Ming; WANG Song-Pei

    2005-01-01

    The synthesis of serotonin transporter imaging agent [125I] -2-((2-((dimethylamino)methyl)phenyl)thio)-5-iodophenylamine([125I] ADAM) was reported. The chemical structure of the labeling precursor 5- (tributylstannyl) -2-((2-((dimethylamino)methyl)phenyl)thio)phenylamine and all its intermediates were verified by IR,1HNMR and MS. The .radioiodinated compound was prepared using iododestannylation reaction by hydrogen peroxide. Final radiochemical purity was above 95% determined by TLC.

  12. Radiolabelled D2 agonists as prolactinoma imaging agents

    Energy Technology Data Exchange (ETDEWEB)

    Otto, C.A.

    1991-12-31

    Research conducted in this terminal year of support centered on three distinct areas: mAChR ligand localization in pancreas and the effect of Ca{sup +2} on localization, continuation of assessment of quaternized and neutral mAChR ligands for possible use as PET myocardial imaging agents, and initiation of a study to determine the relationship of the nAChR receptor to the cellular receptor for measles virus. Several tables and figures illustrating the results are included.

  13. Segmentation of fluorescence microscopy cell images using unsupervised mining.

    Science.gov (United States)

    Du, Xian; Dua, Sumeet

    2010-05-28

    The accurate measurement of cell and nuclei contours are critical for the sensitive and specific detection of changes in normal cells in several medical informatics disciplines. Within microscopy, this task is facilitated using fluorescence cell stains, and segmentation is often the first step in such approaches. Due to the complex nature of cell issues and problems inherent to microscopy, unsupervised mining approaches of clustering can be incorporated in the segmentation of cells. In this study, we have developed and evaluated the performance of multiple unsupervised data mining techniques in cell image segmentation. We adapt four distinctive, yet complementary, methods for unsupervised learning, including those based on k-means clustering, EM, Otsu's threshold, and GMAC. Validation measures are defined, and the performance of the techniques is evaluated both quantitatively and qualitatively using synthetic and recently published real data. Experimental results demonstrate that k-means, Otsu's threshold, and GMAC perform similarly, and have more precise segmentation results than EM. We report that EM has higher recall values and lower precision results from under-segmentation due to its Gaussian model assumption. We also demonstrate that these methods need spatial information to segment complex real cell images with a high degree of efficacy, as expected in many medical informatics applications.

  14. In vivo imaging of Lactococcus lactis, Lactobacillus plantarum and Escherichiacoli expressing infrared fluorescent protein in mice

    OpenAIRE

    Berlec, Aleš; Štrukelj, Borut; Završnik, Janja; Turk, Boris; Butinar, Miha

    2016-01-01

    Background In vivo imaging of orally administered lactic acid bacteria (LAB) and commensal bacteria in mice is shown to provide information on the spatial and temporal distribution of bacteria in the gastrointestinal tract. The bacteria can be detected and monitored using bioluminescence or near-infrared fluorescence. Results Fluorescence imaging of bacteria was established by expressing the infrared fluorescent protein IRFP713 in Lactococcus lactis, Lactobacillus plantarum and Escherichia co...

  15. A FRET-based ratiometric fluorescent and colorimetric probe for the facile detection of organophosphonate nerve agent mimic DCP.

    Science.gov (United States)

    Xuan, Weimin; Cao, Yanting; Zhou, Jiahong; Wang, Wei

    2013-11-18

    A FRET ratiometric fluorescent probe enabling a fast and highly sensitive response to OP nerve agent mimic DCP within 1 min and with as low as 0.17 ppm concentration detection limit has been developed. Moreover, the probe exhibits noticeable color changes under UV light and even with the naked eye. It is also demonstrated that it can detect both liquid and gas nerve agents.

  16. In vivo multiphoton tomography and fluorescence lifetime imaging of human brain tumor tissue.

    Science.gov (United States)

    Kantelhardt, Sven R; Kalasauskas, Darius; König, Karsten; Kim, Ella; Weinigel, Martin; Uchugonova, Aisada; Giese, Alf

    2016-05-01

    High resolution multiphoton tomography and fluorescence lifetime imaging differentiates glioma from adjacent brain in native tissue samples ex vivo. Presently, multiphoton tomography is applied in clinical dermatology and experimentally. We here present the first application of multiphoton and fluorescence lifetime imaging for in vivo imaging on humans during a neurosurgical procedure. We used a MPTflex™ Multiphoton Laser Tomograph (JenLab, Germany). We examined cultured glioma cells in an orthotopic mouse tumor model and native human tissue samples. Finally the multiphoton tomograph was applied to provide optical biopsies during resection of a clinical case of glioblastoma. All tissues imaged by multiphoton tomography were sampled and processed for conventional histopathology. The multiphoton tomograph allowed fluorescence intensity- and fluorescence lifetime imaging with submicron spatial resolution and 200 picosecond temporal resolution. Morphological fluorescence intensity imaging and fluorescence lifetime imaging of tumor-bearing mouse brains and native human tissue samples clearly differentiated tumor and adjacent brain tissue. Intraoperative imaging was found to be technically feasible. Intraoperative image quality was comparable to ex vivo examinations. To our knowledge we here present the first intraoperative application of high resolution multiphoton tomography and fluorescence lifetime imaging of human brain tumors in situ. It allowed in vivo identification and determination of cell density of tumor tissue on a cellular and subcellular level within seconds. The technology shows the potential of rapid intraoperative identification of native glioma tissue without need for tissue processing or staining.

  17. Statistical image segmentation for the detection of skin lesion borders in UV fluorescence excitation

    Science.gov (United States)

    Ortega-Martinez, Antonio; Padilla-Martinez, Juan Pablo; Franco, Walfre

    2016-04-01

    The skin contains several fluorescent molecules or fluorophores that serve as markers of structure, function and composition. UV fluorescence excitation photography is a simple and effective way to image specific intrinsic fluorophores, such as the one ascribed to tryptophan which emits at a wavelength of 345 nm upon excitation at 295 nm, and is a marker of cellular proliferation. Earlier, we built a clinical UV photography system to image cellular proliferation. In some samples, the naturally low intensity of the fluorescence can make it difficult to separate the fluorescence of cells in higher proliferation states from background fluorescence and other imaging artifacts -- like electronic noise. In this work, we describe a statistical image segmentation method to separate the fluorescence of interest. Statistical image segmentation is based on image averaging, background subtraction and pixel statistics. This method allows to better quantify the intensity and surface distributions of fluorescence, which in turn simplify the detection of borders. Using this method we delineated the borders of highly-proliferative skin conditions and diseases, in particular, allergic contact dermatitis, psoriatic lesions and basal cell carcinoma. Segmented images clearly define lesion borders. UV fluorescence excitation photography along with statistical image segmentation may serve as a quick and simple diagnostic tool for clinicians.

  18. Combination of a spinning disc confocal unit with frequency-domain fluorescence lifetime imaging microscopy.

    NARCIS (Netherlands)

    van Munster, E.B.; Goedhart, J.; Kremers, G.J.; Manders, E.M.M.; Gadella, Th.W.J.

    2007-01-01

    BACKGROUND: Wide-field frequency-domain fluorescence lifetime imaging microscopy (FLIM) is an established technique to determine fluorescence lifetimes. Disadvantage of wide-field imaging is that measurements are compromised by out-of-focus blur. Conventional scanning confocal typically means long

  19. Accurate study of FosPeg® distribution in a mouse model using fluorescence imaging technique and fluorescence white monte carlo simulations

    DEFF Research Database (Denmark)

    Xie, Haiyan; Liu, Haichun; Svenmarker, Pontus

    2010-01-01

    Fluorescence imaging is used for quantitative in vivo assessment of drug concentration. Light attenuation in tissue is compensated for through Monte-Carlo simulations. The intrinsic fluorescence intensity, directly proportional to the drug concentration, could be obtained....

  20. Magnetic resonance imaging with hyperpolarized agents: methods and applications.

    Science.gov (United States)

    Adamson, Erin B; Ludwig, Kai D; Mummy, David G; Fain, Sean B

    2017-07-07

    In the past decade, hyperpolarized (HP) contrast agents have been under active development for MRI applications to address the twin challenges of functional and quantitative imaging. Both HP helium ((3)He) and xenon ((129)Xe) gases have reached the stage where they are under study in clinical research. HP (129)Xe, in particular, is poised for larger scale clinical research to investigate asthma, chronic obstructive pulmonary disease, and fibrotic lung diseases. With advances in polarizer technology and unique capabilities for imaging of (129)Xe gas exchange into lung tissue and blood, HP (129)Xe MRI is attracting new attention. In parallel, HP (13)C and (15)N MRI methods have steadily advanced in a wide range of pre-clinical research applications for imaging metabolism in various cancers and cardiac disease. The HP [1-(13)C] pyruvate MRI technique, in particular, has undergone phase I trials in prostate cancer and is poised for investigational new drug trials at multiple institutions in cancer and cardiac applications. This review treats the methodology behind both HP gases and HP (13)C and (15)N liquid state agents. Gas and liquid phase HP agents share similar technologies for achieving non-equilibrium polarization outside the field of the MRI scanner, strategies for image data acquisition, and translational challenges in moving from pre-clinical to clinical research. To cover the wide array of methods and applications, this review is organized by numerical section into (1) a brief introduction, (2) the physical and biological properties of the most common polarized agents with a brief summary of applications and methods of polarization, (3) methods for image acquisition and reconstruction specific to improving data acquisition efficiency for HP MRI, (4) the main physical properties that enable unique measures of physiology or metabolic pathways, followed by a more detailed review of the literature describing the use of HP agents to study: (5) metabolic pathways

  1. Magnetic resonance imaging with hyperpolarized agents: methods and applications

    Science.gov (United States)

    Adamson, Erin B.; Ludwig, Kai D.; Mummy, David G.; Fain, Sean B.

    2017-07-01

    In the past decade, hyperpolarized (HP) contrast agents have been under active development for MRI applications to address the twin challenges of functional and quantitative imaging. Both HP helium (3He) and xenon (129Xe) gases have reached the stage where they are under study in clinical research. HP 129Xe, in particular, is poised for larger scale clinical research to investigate asthma, chronic obstructive pulmonary disease, and fibrotic lung diseases. With advances in polarizer technology and unique capabilities for imaging of 129Xe gas exchange into lung tissue and blood, HP 129Xe MRI is attracting new attention. In parallel, HP 13C and 15N MRI methods have steadily advanced in a wide range of pre-clinical research applications for imaging metabolism in various cancers and cardiac disease. The HP [1-13C] pyruvate MRI technique, in particular, has undergone phase I trials in prostate cancer and is poised for investigational new drug trials at multiple institutions in cancer and cardiac applications. This review treats the methodology behind both HP gases and HP 13C and 15N liquid state agents. Gas and liquid phase HP agents share similar technologies for achieving non-equilibrium polarization outside the field of the MRI scanner, strategies for image data acquisition, and translational challenges in moving from pre-clinical to clinical research. To cover the wide array of methods and applications, this review is organized by numerical section into (1) a brief introduction, (2) the physical and biological properties of the most common polarized agents with a brief summary of applications and methods of polarization, (3) methods for image acquisition and reconstruction specific to improving data acquisition efficiency for HP MRI, (4) the main physical properties that enable unique measures of physiology or metabolic pathways, followed by a more detailed review of the literature describing the use of HP agents to study: (5) metabolic pathways in cancer and cardiac

  2. Hyperspectral instrumentation to image and characterize the fluorescence of materials

    Science.gov (United States)

    Bourcier, Frédéric; Walter, Philippe; Pedetti, Silvia; Faye, Delphine; Spezzigu, Piero; Infante, Fulvio; Le Nouy, Patrice; Zedda, Edoardo

    2016-09-01

    Optical instruments for space applications with improved performances (smaller pixels and spectral range extension) are becoming more and more sensitive to chemical contamination and particle sedimentation. Outgassing under vacuum conditions causes dramatic flux losses, especially in the UV bandwidth. Furthermore, it is difficult to perform physicochemical analyses of contaminated surfaces on flight models, in a clean room. Conventional analytical techniques such as FTIR (Fourier Transform Infrared interferometer) need the tool to be in contact with the studied area, which is forbidden when working on satellites. In addition, it does not give any information about the distribution of the contaminants in the field of view. The probed area is large, mono-pixel, and the sensitivity of the instrument is too low for hundred nanometer thin film deposits. A first study has shown that we could benefit from using the UV/visible fluorescence spectra to partially identify contaminants and polymer materials. The shape of the fluorescence spectra of adhesives, paints and varnishes have specific signatures that could be recorded into a designated reference database. The location of the presence of these contaminants on such sensitive optics is also relevant. To acquire both these parameters, we designed a specific compact hyperspectral instrument to remotely acquire cube images (500x500 pixels) in a 5 degree field of view, and on a wide range of continuous wavelengths from UV at 320 nm up to the near infrared at 1000 nm. This paper will present the chosen trade-off between different critical optics for a new portable version of this instrument. It is dedicated to space and cultural heritage applications and the first results on an engineering prototype will be shown.

  3. Novel fluorescence nanobubbles for contrast-enhanced ultrasound imaging in rabbit VX2 hepatocellular carcinoma model

    Science.gov (United States)

    Yu, Houqiang; Wang, Wei; He, Xiaoling; Zhou, Qibing; Ding, Mingyue

    2017-03-01

    Ultrasound contrast agents (UCAs) such as SonoVue or Optison have been used widely in clinic for contrast-enhanced vascular imaging. However, microbubbles UCAs display limitations in tumor-targeted imaging due to the large sizes, nanoscaled UCAs has consequently attracted increasing attentions. In this work, we synthesized nanobubbles (NBs) by ultrasonic cavitation method, then a fluorescent marker of Alexa Fluor 680 was conjugated to the shell in order to observe the localization of NBs in tumor tissue. Measurement of fundamental characteristics showed that the NBs had homogeneous distribution of mean diameter of 267.9 +/- 19.2 nm and polydispersity index of 0.410 +/- 0.056. To assess in vivo tumor-selectivity of NBs, we established the rabbits VX2 hepatocellular carcinoma model though surgical implantation method. After the rabbits were intravenous administered of NBs, contrast-enhanced sonograms was observed in the surrounding of VX2 tumor, which showed there are rich capillaries in the tumor periphery. We additionally investigated the toxic of the NBs by hematoxylin-eosin staining. The results indicated that the NBs is a biocompatible non-toxic lipid system. Furthermore, the VX2 tumors and major organs were analyzed using ex vivo fluorescence imaging to confirm the targeted selectivity of NBs, and the results verified that the NBs were capable of targeting VX2 tumor. Confocal laser scanning microscopy examination showed that the NBs can traverse the VX2 tumor capillaries and target to the hepatocellular carcinoma tumor cells. All these results suggested that the newly prepared NBs have a potential application in molecular imaging and tumor-targeting therapy.

  4. Photoacoustic imaging of a near-infrared fluorescent marker based on dual wavelength pump-probe excitation

    Science.gov (United States)

    Märk, Julia; Theiss, Christoph; Schmitt, Franz-Josef; Laufer, Jan

    2014-03-01

    Photoacoustic imaging has been used to determine the spatial distribution of fluorophores, such as exogenous dyes and genetically expressed proteins, from images acquired in phantoms and in vivo. Most methods involve the acquisition of multiwavelength images and rely on differences in the absorption spectra of the tissue chromophores to estimate the spatial distribution and abundance of the latter using spectral decomposition techniques, such as model based inversion schemes. However, the inversion of 3-D images can be computationally expensive. Experimental approaches to localising contrast agents may therefore be useful, especially if quantification is not essential. This work aims to develop a method for determining the spatial distribution of a near-infrared fluorescent cell marker from images acquired using dual wavelength excitation. The excitation wavelengths coincided with the absorption and emission spectrum of the fluorophore. The contrast mechanism relies on reducing the excited state lifetime of the fluorophore by inducing stimulated emission. This changes the amount of energy thermalized by the fluorophore, and hence the photoacoustic signal amplitude. Since this is not observed in endogenous chromophores, the background may be removed by subtracting two images acquired with and without pulse delay between the pump and probe pulses. To characterise the fluorophore, the signal amplitude is measured in a cuvette as a function of pulse delay, concentration, and fluence. The spatial distribution of the fluorophore is determined from images acquired in realistic tissue phantoms. This method may be suitable for in vivo applications, such as imaging of exogenous or genetically expressed fluorescent cell markers.

  5. Ultrasound contrast agent imaging: Real-time imaging of the superharmonics

    NARCIS (Netherlands)

    Peruzzini, D.; Viti, J.; Tortoli, P.; Verweij, M.D.; De Jong, N.; Vos, H.J.

    2015-01-01

    Currently, in medical ultrasound contrast agent (UCA) imaging the second harmonic scattering of the microbubbles is regularly used. This scattering is in competition with the signal that is caused by nonlinear wave propagation in tissue. It was reported that UCA imaging based on the third or higher

  6. Ultrasound contrast agent imaging: Real-time imaging of the superharmonics

    NARCIS (Netherlands)

    Peruzzini, D.; J. Viti (Jacopo); P. Tortoli (Piero); M.D. Verweij (Martin D.); N. de Jong (Nico); H.J. Vos (Rik)

    2015-01-01

    textabstractCurrently, in medical ultrasound contrast agent (UCA) imaging the second harmonic scattering of the microbubbles is regularly used. This scattering is in competition with the signal that is caused by nonlinear wave propagation in tissue. It was reported that UCA imaging based on the

  7. Fluorescence guided lymph node biopsy in large animals using direct image projection device

    Science.gov (United States)

    Ringhausen, Elizabeth; Wang, Tylon; Pitts, Jonathan; Akers, Walter J.

    2016-03-01

    The use of fluorescence imaging for aiding oncologic surgery is a fast growing field in biomedical imaging, revolutionizing open and minimally invasive surgery practices. We have designed, constructed, and tested a system for fluorescence image acquisition and direct display on the surgical field for fluorescence guided surgery. The system uses a near-infrared sensitive CMOS camera for image acquisition, a near-infra LED light source for excitation, and DLP digital projector for projection of fluorescence image data onto the operating field in real time. Instrument control was implemented in Matlab for image capture, processing of acquired data and alignment of image parameters with the projected pattern. Accuracy of alignment was evaluated statistically to demonstrate sensitivity to small objects and alignment throughout the imaging field. After verification of accurate alignment, feasibility for clinical application was demonstrated in large animal models of sentinel lymph node biopsy. Indocyanine green was injected subcutaneously in Yorkshire pigs at various locations to model sentinel lymph node biopsy in gynecologic cancers, head and neck cancer, and melanoma. Fluorescence was detected by the camera system during operations and projected onto the imaging field, accurately identifying tissues containing the fluorescent tracer at up to 15 frames per second. Fluorescence information was projected as binary green regions after thresholding and denoising raw intensity data. Promising results with this initial clinical scale prototype provided encouraging results for the feasibility of optical projection of acquired luminescence during open oncologic surgeries.

  8. A paramagnetic CEST agent for imaging glucose by MRI.

    Science.gov (United States)

    Zhang, Shanrong; Trokowski, Robert; Sherry, A Dean

    2003-12-17

    The europium(III) complex of a DOTA-tetraamide ligand (DOTA = 1,4,7,10-tetraazacyclododecane-N,N',N' ',N' ''-tetraacetic acids) containing two phenyl boronate pendent arms binds glucose reversibly with an association constant of 383 M-1 at pH 7. Glucose binding results in slowing of water exchange between a single Eu(III)-bound water molecule and bulk water, and this can be imaged by MRI using chemical exchange saturation transfer (CEST) imaging sequence. This metabolite-responsive paramagnetic CEST agent responds to changes in glucose over the physiologically important range (0-20 mM), and thus it offers the possibility of high-sensitivity MR imaging glucose in tissues using bulk water protons as antenna.

  9. NADH fluorescence imaging of isolated biventricular working rabbit hearts.

    Science.gov (United States)

    Asfour, Huda; Wengrowski, Anastasia M; Jaimes, Rafael; Swift, Luther M; Kay, Matthew W

    2012-07-24

    Since its inception by Langendorff(1), the isolated perfused heart remains a prominent tool for studying cardiac physiology(2). However, it is not well-suited for studies of cardiac metabolism, which require the heart to perform work within the context of physiologic preload and afterload pressures. Neely introduced modifications to the Langendorff technique to establish appropriate left ventricular (LV) preload and afterload pressures(3). The model is known as the isolated LV working heart model and has been used extensively to study LV performance and metabolism(4-6). This model, however, does not provide a properly loaded right ventricle (RV). Demmy et al. first reported a biventricular model as a modification of the LV working heart model(7, 8). They found that stroke volume, cardiac output, and pressure development improved in hearts converted from working LV mode to biventricular working mode(8). A properly loaded RV also diminishes abnormal pressure gradients across the septum to improve septal function. Biventricular working hearts have been shown to maintain aortic output, pulmonary flow, mean aortic pressure, heart rate, and myocardial ATP levels for up to 3 hours(8). When studying the metabolic effects of myocardial injury, such as ischemia, it is often necessary to identify the location of the affected tissue. This can be done by imaging the fluorescence of NADH (the reduced form of nicotinamide adenine dinucleotide)(9-11), a coenzyme found in large quantities in the mitochondria. NADH fluorescence (fNADH) displays a near linearly inverse relationship with local oxygen concentration(12) and provides a measure of mitochondrial redox state(13). fNADH imaging during hypoxic and ischemic conditions has been used as a dye-free method to identify hypoxic regions(14, 15) and to monitor the progression of hypoxic conditions over time(10). The objective of the method is to monitor the mitochondrial redox state of biventricular working hearts during protocols

  10. [Studies on laticifers and milk of greater celandine (Chelidonium majus L.) with fluorescence imaging and fluorescence spectroscopic methods].

    Science.gov (United States)

    Póczi, Dorottya; Böddi, Béla

    2010-01-01

    Using fluorescence imaging and fluorescence spectroscopic methods, the localisation of the laticifers and the native spectral properties of the milk were studied in various organs of greater celandine (Chelidonium majus L.). Direct measurements on tissue pieces (without the extraction and the separation of the components) provided information about the complexity of the milk and the various ratios of the alkaloid contents in the tissues. Whole plant were studied in a gel documentation system using ultraviolet light source, while the localisation of the laticifers was observed along the leaf veins in fluorescence microscope, using blue excitation light. Measuring different tissue pieces, fluorescence spectroscopic studies showed that the greater celandine alkaloids have emission bands at 469, 530-531, 553, 572-575 and 592 nm and excitation bands at 365, 370, 386 is 400 nm. These results give a possibility for conclusions about the alkaloid contents and composition or ratios of the alkaloid components in various tissue pieces directly, via comparisons with alkaloid standards.

  11. Cryo-imaging of fluorescently labeled single cells in a mouse

    Science.gov (United States)

    Steyer, Grant J.; Roy, Debashish; Salvado, Olivier; Stone, Meredith E.; Wilson, David L.

    2009-02-01

    We developed a cryo-imaging system to provide single-cell detection of fluorescently labeled cells in mouse, with particular applicability to stem cells and metastatic cancer. The Case cryoimaging system consists of a fluorescence microscope, robotic imaging positioner, customized cryostat, PC-based control system, and visualization/analysis software. The system alternates between sectioning (10-40 μm) and imaging, collecting color brightfield and fluorescent blockface image volumes >60GB. In mouse experiments, we imaged quantum-dot labeled stem cells, GFP-labeled cancer and stem cells, and cell-size fluorescent microspheres. To remove subsurface fluorescence, we used a simplified model of light-tissue interaction whereby the next image was scaled, blurred, and subtracted from the current image. We estimated scaling and blurring parameters by minimizing entropy of subtracted images. Tissue specific attenuation parameters were found [uT : heart (267 +/- 47.6 μm), liver (218 +/- 27.1 μm), brain (161 +/- 27.4 μm)] to be within the range of estimates in the literature. "Next image" processing removed subsurface fluorescence equally well across multiple tissues (brain, kidney, liver, adipose tissue, etc.), and analysis of 200 microsphere images in the brain gave 97+/-2% reduction of subsurface fluorescence. Fluorescent signals were determined to arise from single cells based upon geometric and integrated intensity measurements. Next image processing greatly improved axial resolution, enabled high quality 3D volume renderings, and improved enumeration of single cells with connected component analysis by up to 24%. Analysis of image volumes identified metastatic cancer sites, found homing of stem cells to injury sites, and showed microsphere distribution correlated with blood flow patterns. We developed and evaluated cryo-imaging to provide single-cell detection of fluorescently labeled cells in mouse. Our cryo-imaging system provides extreme (>60GB), micron

  12. Laser-induced fluorescence imaging of subsurface tissue structures with a volume holographic spatial-spectral imaging system.

    Science.gov (United States)

    Luo, Yuan; Gelsinger-Austin, Paul J; Watson, Jonathan M; Barbastathis, George; Barton, Jennifer K; Kostuk, Raymond K

    2008-09-15

    A three-dimensional imaging system incorporating multiplexed holographic gratings to visualize fluorescence tissue structures is presented. Holographic gratings formed in volume recording materials such as a phenanthrenquinone poly(methyl methacrylate) photopolymer have narrowband angular and spectral transmittance filtering properties that enable obtaining spatial-spectral information within an object. We demonstrate this imaging system's ability to obtain multiple depth-resolved fluorescence images simultaneously.

  13. Iodinated oil-loaded, fluorescent mesoporous silica-coated iron oxide nanoparticles for magnetic resonance imaging/computed tomography/fluorescence trimodal imaging

    Science.gov (United States)

    Xue, Sihan; Wang, Yao; Wang, Mengxing; Zhang, Lu; Du, Xiaoxia; Gu, Hongchen; Zhang, Chunfu

    2014-01-01

    In this study, a novel magnetic resonance imaging (MRI)/computed tomography (CT)/fluorescence trifunctional probe was prepared by loading iodinated oil into fluorescent mesoporous silica-coated superparamagnetic iron oxide nanoparticles (i-fmSiO4@SPIONs). Fluorescent mesoporous silica-coated superparamagnetic iron oxide nanoparticles (fmSiO4@SPIONs) were prepared by growing fluorescent dye-doped silica onto superparamagnetic iron oxide nanoparticles (SPIONs) directed by a cetyltrimethylammonium bromide template. As prepared, fmSiO4@SPIONs had a uniform size, a large surface area, and a large pore volume, which demonstrated high efficiency for iodinated oil loading. Iodinated oil loading did not change the sizes of fmSiO4@SPIONs, but they reduced the MRI T2 relaxivity (r2) markedly. I-fmSiO4@SPIONs were stable in their physical condition and did not demonstrate cytotoxic effects under the conditions investigated. In vitro studies indicated that the contrast enhancement of MRI and CT, and the fluorescence signal intensity of i-fmSiO4@SPION aqueous suspensions and macrophages, were intensified with increased i-fmSiO4@SPION concentrations in suspension and cell culture media. Moreover, for the in vivo study, the accumulation of i-fmSiO4@SPIONs in the liver could also be detected by MRI, CT, and fluorescence imaging. Our study demonstrated that i-fmSiO4@SPIONs had great potential for MRI/CT/fluorescence trimodal imaging. PMID:24904212

  14. Precise diagnosis in different scenarios using photoacoustic and fluorescence imaging with dual-modality nanoparticles

    Science.gov (United States)

    Peng, Dong; Du, Yang; Shi, Yiwen; Mao, Duo; Jia, Xiaohua; Li, Hui; Zhu, Yukun; Wang, Kun; Tian, Jie

    2016-07-01

    Photoacoustic imaging and fluorescence molecular imaging are emerging as important research tools for biomedical studies. Photoacoustic imaging offers both strong optical absorption contrast and high ultrasonic resolution, and fluorescence molecular imaging provides excellent superficial resolution, high sensitivity, high throughput, and the ability for real-time imaging. Therefore, combining the imaging information of both modalities can provide comprehensive in vivo physiological and pathological information. However, currently there are limited probes available that can realize both fluorescence and photoacoustic imaging, and advanced biomedical applications for applying this dual-modality imaging approach remain underexplored. In this study, we developed a dual-modality photoacoustic-fluorescence imaging nanoprobe, ICG-loaded Au@SiO2, which was uniquely designed, consisting of gold nanorod cores and indocyanine green with silica shell spacer layers to overcome fluorophore quenching. This nanoprobe was examined by both PAI and FMI for in vivo imaging on tumor and ischemia mouse models. Our results demonstrated that the nanoparticles can specifically accumulate at the tumor and ischemic areas and be detected by both imaging modalities. Moreover, this dual-modality imaging strategy exhibited superior advantages for a precise diagnosis in different scenarios. The new nanoprobe with the dual-modality imaging approach holds great potential for diagnosis and stage classification of tumor and ischemia related diseases.Photoacoustic imaging and fluorescence molecular imaging are emerging as important research tools for biomedical studies. Photoacoustic imaging offers both strong optical absorption contrast and high ultrasonic resolution, and fluorescence molecular imaging provides excellent superficial resolution, high sensitivity, high throughput, and the ability for real-time imaging. Therefore, combining the imaging information of both modalities can provide

  15. A CMOS image sensor with draining only modulation pixels for fluorescence lifetime imaging

    Science.gov (United States)

    Li, Zhuo; Yasutomi, Keita; Takasawa, Taishi; Itoh, Shinya; Kawahito, Shoji

    2011-03-01

    Fluorescence lifetime imaging is becoming a powerful tool in biology. A charge-domain CMOS Fluorescence Lifetime Imaging Microscopy (FLIM) chip using a pinned photo diode (PPD) and the pinned storage diode (PSD) with different depth of potential wells has been previously developed by the authors. However, a transfer gate between PPD and PSD causes charge transfer noise due to traps at the channel surface. This paper presents a time-resolved CMOS image sensor with draining only modulation pixels for fluorescence lifetime imaging, which removes the transfer gate between PPD and PSD. The time windowing is done by draining with a draining gate only, which is attached along the carrier path from PPD to PSD. This allows us to realize a trapping less charge transfer between PPD and PSD, leading to a very low-noise time-resolved signal detection. A video-rate CMOS FLIM chip has been fabricated using 0.18μm standard CMOS pinned diode image sensor process. The pixel consists of a PPD, a PSD, a charge draining gate (TD), a readout transfer gate (TX) between the PSD and the floating diffusion (FD), a reset transistor and a source follower amplifier transistor. The pixel array has 200(Row) x 256(Column) pixels and the pixel pitch is 7.5μm. Fundamental characteristics of the implemented CMOS FLIM chip are measured. The signal intensity of the PSD as a function of the TD gate voltage is also measured. The ratio of the signal for the TD off to the signal for the TD on is 212 : 1.

  16. Near-Infrared Fluorescence Enhanced (NIR-FE) Molecular Imaging of Live Cells on Gold Substrates

    CERN Document Server

    Hong, Guosong; Welsher, Kevin; Chen, Zhuo; Robinson, Joshua T; Wang, Hailiang; Zhang, Bo; Dai, Hongjie

    2011-01-01

    Low quantum yields of near infrared (NIR) fluorophores have limited their capabilities as imaging probes in a transparent, low background imaging window. Here for the first time we reported near-infrared fluorescence enhance (NIR-FE) cell imaging using nanostructured Au substrate, which was employed as a general platform for both single-walled carbon nanotubes (SWNTs) and organic fluorescent labels in the NIR region. Fluorescence intensity, as well as cell targeting specificity, was greatly improved by this novel imaging technique. With NIR-FE imaging, we were able to image SWNT-stained cells at short exposure time of 300ms, and push the detectable limit of SWNT staining of cells down to an ultralow concentration of ~50 pM. Further, different degrees of fluorescence enhancement for endocytosed, intracellular SWNTs vs. nanotubes on the cell membrane at the cell/gold interface were observed, suggesting the possibility of using this technique to track the transmembrane behavior of NIR fluorophores.

  17. Deep-brain imaging via epi-fluorescence Computational Cannula Microscopy

    Science.gov (United States)

    Kim, Ganghun; Nagarajan, Naveen; Pastuzyn, Elissa; Jenks, Kyle; Capecchi, Mario; Shepherd, Jason; Menon, Rajesh

    2017-03-01

    Here we demonstrate widefield (field diameter = 200 μm) fluorescence microscopy and video imaging inside the rodent brain at a depth of 2 mm using a simple surgical glass needle (cannula) of diameter 0.22 mm as the primary optical element. The cannula guides excitation light into the brain and the fluorescence signal out of the brain. Concomitant image-processing algorithms are utilized to convert the spatially scrambled images into fluorescent images and video. The small size of the cannula enables minimally invasive imaging, while the long length (>2 mm) allow for deep-brain imaging with no additional complexity in the optical system. Since no scanning is involved, widefield fluorescence video at the native frame rate of the camera can be achieved.

  18. Segmenting Intracellular Distribution Images Derived by Fluorescent Dyes Using a Potts Model Hamiltonian

    CERN Document Server

    Hu, Dandan; Ronhovde, Peter; Bloch, Sharon; Achilefu, Samuel; Nussinov, Zohar

    2012-01-01

    We apply a multiresolution community detection algorithm to perform unsupervised segmentation of complex intracellular signals derived using fluorescent dyes. In our earlier work, when applying our method to benchmarks, our algorithm was shown to be one of the best and to be especially suited to the detection of camouflage images. In the current manuscript, we have explored this algorithm in a more complex scenario. The current image processing problem is framed as identifying clusters with respective average fluorescent lifetimes (FLTs) against a background or "solvent" in fluorescence lifetime imaging microscopy (FLIM) images derived using NIR fluorescent dyes. We have identified significant multiresolution structures using replica correlations in these images, where such correlations are manifested by information theoretic overlaps of the independent solutions ("replicas") attained using the proposed algorithm from different starting points. Our method is more efficient than a well-known image segmentation...

  19. Deep-brain imaging via epi-fluorescence Computational Cannula Microscopy

    Science.gov (United States)

    Kim, Ganghun; Nagarajan, Naveen; Pastuzyn, Elissa; Jenks, Kyle; Capecchi, Mario; Shepherd, Jason; Menon, Rajesh

    2017-01-01

    Here we demonstrate widefield (field diameter = 200 μm) fluorescence microscopy and video imaging inside the rodent brain at a depth of 2 mm using a simple surgical glass needle (cannula) of diameter 0.22 mm as the primary optical element. The cannula guides excitation light into the brain and the fluorescence signal out of the brain. Concomitant image-processing algorithms are utilized to convert the spatially scrambled images into fluorescent images and video. The small size of the cannula enables minimally invasive imaging, while the long length (>2 mm) allow for deep-brain imaging with no additional complexity in the optical system. Since no scanning is involved, widefield fluorescence video at the native frame rate of the camera can be achieved. PMID:28317915

  20. 1,2-hydroxypyridonates as contrast agents for magnetic resonance imaging: TREN-1,2-HOPO.

    Science.gov (United States)

    Jocher, Christoph J; Moore, Evan G; Xu, Jide; Avedano, Stefano; Botta, Mauro; Aime, Silvio; Raymond, Kenneth N

    2007-10-29

    1,2-Hydroxypyridinones (1,2-HOPO) form very stable lanthanide complexes that may be useful as contrast agents for magnetic resonance imaging (MRI). X-ray diffraction of single crystals established that the solid-state structures of the Eu(III) and the previously reported [Inorg. Chem. 2004, 43, 5452] Gd(III) complex are identical. The recently discovered sensitizing properties of 1,2-HOPO chelates for Eu(III) luminescence [J. Am. Chem. Soc. 2006, 128, 10 067] allow for direct measurement of the number of water molecules coordinated to the metal center. Fluorescence measurements of the Eu(III) complex corroborate that, in solution, two water molecules coordinate the lanthanide (q = 2) as proposed from the analysis of NMRD profiles. In addition, fluorescence measurements have verified the anion binding interactions of lanthanide TREN-1,2-HOPO complexes in solution, studied by relaxivity, revealing only very weak oxalate binding (KA = 82.7 +/- 6.5 M-1). Solution thermodynamic studies of the metal complex and free ligand have been carried out using potentiometry, spectrophotometry, and fluorescence spectroscopy. The metal ion selectivity of TREN-1,2-HOPO supports the feasibility of using 1,2-HOPO ligands for selective lanthanide binding [pGd = 19.3 (2), pZn = 15.2 (2), pCa = 8.8 (3)].

  1. Comparison of Folate Receptor Targeted Optical Contrast Agents for Intraoperative Molecular Imaging

    Directory of Open Access Journals (Sweden)

    Elizabeth De Jesus

    2015-01-01

    Full Text Available Background. Intraoperative imaging can identify cancer cells in order to improve resection; thus fluorescent contrast agents have emerged. Our objective was to do a preclinical comparison of two fluorescent dyes, EC17 and OTL38, which both target folate receptor but have different fluorochromes. Materials. HeLa and KB cells lines were used for in vitro and in vivo comparisons of EC17 and OTL38 brightness, sensitivity, pharmacokinetics, and biodistribution. In vivo experiments were then performed in mice. Results. The peak excitation and emission wavelengths of EC17 and OTL38 were 470/520 nm and 774/794 nm, respectively. In vitro, OTL38 required increased incubation time compared to EC17 for maximum fluorescence; however, peak signal-to-background ratio (SBR was 1.4-fold higher compared to EC17 within 60 minutes (p<0.001. Additionally, the SBR for detecting smaller quantity of cells was improved with OTL38. In vivo, the mean improvement in SBR of tumors visualized using OTL38 compared to EC17 was 3.3 fold (range 1.48–5.43. Neither dye caused noticeable toxicity in animal studies. Conclusions. In preclinical testing, OTL38 appears to have superior sensitivity and brightness compared to EC17. This coincides with the accepted belief that near infrared (NIR dyes tend to have less autofluorescence and scattering issues than visible wavelength fluorochromes.

  2. Prolactin receptor-mediated internalization of imaging agents detects epithelial ovarian cancer

    Science.gov (United States)

    Sundaram, Karthik M.

    Epithelial ovarian cancer (EOC) has the highest mortality rate of all gynecologic malignant tumors. Diagnosis of epithelial ovarian cancer (EOC) presents two main challenges. The first challenge is detecting low volume (prolactin receptor (PRLR) - a cell surface tyrosine kinase receptor that is over-expressed in moderate to high levels on > 98% of epithelial ovarian cancers. Upon binding of native ligands to PRLR, the ligand:PRLR complex is internalized by cells. By conjugating gadolinium-chelates, molecules normally used as contrast agents diagnostically, to human placental lactogen (hPL), a native ligand of PRLR, we show that MRI becomes highly sensitive and specific for detecting PRLR (+) tumors in a nude mouse model of EOC. We further establish the adaptability of this approach for fluorescence-based imaging techniques using an hPL conjugated Cy5.5 dye. We conclude that molecular imaging of PRLR with hPL-conjugated imaging agents can address the current challenges that limit EOC diagnosis.

  3. Enhancing in vivo tumor boundary delineation with structured illumination fluorescence molecular imaging and spatial gradient mapping

    Science.gov (United States)

    Sun, Jessica; Miller, Jessica P.; Hathi, Deep; Zhou, Haiying; Achilefu, Samuel; Shokeen, Monica; Akers, Walter J.

    2016-08-01

    Fluorescence imaging, in combination with tumor-avid near-infrared (NIR) fluorescent molecular probes, provides high specificity and sensitivity for cancer detection in preclinical animal models, and more recently, assistance during oncologic surgery. However, conventional camera-based fluorescence imaging techniques are heavily surface-weighted such that surface reflection from skin or other nontumor tissue and nonspecific fluorescence signals dominate, obscuring true cancer-specific signals and blurring tumor boundaries. To address this challenge, we applied structured illumination fluorescence molecular imaging (SIFMI) in live animals for automated subtraction of nonspecific surface signals to better delineate accumulation of an NIR fluorescent probe targeting α4β1 integrin in mice bearing subcutaneous plasma cell xenografts. SIFMI demonstrated a fivefold improvement in tumor-to-background contrast when compared with other full-field fluorescence imaging methods and required significantly reduced scanning time compared with diffuse optical spectroscopy imaging. Furthermore, the spatial gradient mapping enhanced highlighting of tumor boundaries. Through the relatively simple hardware and software modifications described, SIFMI can be integrated with clinical fluorescence imaging systems, enhancing intraoperative tumor boundary delineation from the uninvolved tissue.

  4. Cell Permeable Ratiometric Fluorescent Sensors for Imaging Phosphoinositides.

    Science.gov (United States)

    Mondal, Samsuzzoha; Rakshit, Ananya; Pal, Suranjana; Datta, Ankona

    2016-07-15

    Phosphoinositides are critical cell-signal mediators present on the plasma membrane. The dynamic change of phosphoinositide concentrations on the membrane including clustering and declustering mediates signal transduction. The importance of phosphoinositides is scored by the fact that they participate in almost all cell-signaling events, and a defect in phosphoinositide metabolism is linked to multiple diseases including cancer, bipolar disorder, and type-2 diabetes. Optical sensors for visualizing phosphoinositide distribution can provide information on phosphoinositide dynamics. This exercise will ultimately afford a handle into understanding and manipulating cell-signaling processes. The major requirement in phosphoinositide sensor development is a selective, cell permeable probe that can quantify phosphoinositides. To address this requirement, we have developed short peptide-based ratiometric fluorescent sensors for imaging phosphoinositides. The sensors afford a selective response toward two crucial signaling phosphoinositides, phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2) and phosphatidylinositol-4-phosphate (PI4P), over other anionic membrane phospholipids and soluble inositol phosphates. Dissociation constant values indicate up to 4 times higher probe affinity toward PI(4,5)P2 when compared to PI4P. Significantly, the sensors are readily cell-permeable and enter cells within 15 min of incubation as indicated by multiphoton excitation confocal microscopy. Furthermore, the sensors light up signaling phosphoinositides present both on the cell membrane and on organelle membranes near the perinuclear space, opening avenues for quantifying and monitoring phosphoinositide signaling.

  5. (Non-targeted) radioactive/fluorescent nanoparticles and their potential in combined pre- and intraoperative imaging during sentinel lymph node resection

    Energy Technology Data Exchange (ETDEWEB)

    Buckle, Tessa; Chin, Patrick T K; Van Leeuwen, Fijs W B, E-mail: fw.v.leeuwen@nki.nl [Departments of Radiology and Nuclear Medicine, Division of Diagnostic Oncology at the Netherlands Cancer Institute-Antoni van Leeuwenhoek Hospital, 1066 CX Amsterdam (Netherlands)

    2010-12-03

    One clinical precedent for the use of nanosized imaging agents is the localization of the tumor draining sentinel lymph nodes. In this application, radiocolloids such as {sup 99m}Tc-NanoColl are currently used to plan the surgical procedure and to provide acoustic guidance during the intervention. Additional injections of dyes are common to provide optical surgical guidance. Bimodal imaging agents, which are both radioactive and fluorescent, have the potential to be used for both surgical planning and intraoperative fluorescence guidance towards the sentinel lymph nodes. This review provides an overview of the radioactive, fluorescent, and size properties of (non-targeted) bimodal nanoparticles, and their (potential) value in sentinel lymph node detection. (topical review)

  6. Electron beam dispersion measurements in nitrogen using two-dimensional imaging of N2(+) fluorescence

    Science.gov (United States)

    Clapp, L. H.; Twiss, R. G.; Cattolica, R. J.

    Experimental results are presented related to the radial spread of fluorescence excited by 10 and 20 KeV electron beams passing through nonflowing rarefied nitrogen at 293 K. An imaging technique for obtaining species distributions from measured beam-excited fluorescence is described, based on a signal inversion scheme mathematically equivalent to the inversion of the Abel integral equation. From fluorescence image data, measurements of beam radius, integrated signal intensity, and spatially resolved distributions of N2(+) first-negative-band fluorescence-emitting species have been made. Data are compared with earlier measurements and with an heuristic beam spread model.

  7. A fast image registration approach of neural activities in light-sheet fluorescence microscopy images

    Science.gov (United States)

    Meng, Hui; Hui, Hui; Hu, Chaoen; Yang, Xin; Tian, Jie

    2017-03-01

    The ability of fast and single-neuron resolution imaging of neural activities enables light-sheet fluorescence microscopy (LSFM) as a powerful imaging technique in functional neural connection applications. The state-of-art LSFM imaging system can record the neuronal activities of entire brain for small animal, such as zebrafish or C. elegans at single-neuron resolution. However, the stimulated and spontaneous movements in animal brain result in inconsistent neuron positions during recording process. It is time consuming to register the acquired large-scale images with conventional method. In this work, we address the problem of fast registration of neural positions in stacks of LSFM images. This is necessary to register brain structures and activities. To achieve fast registration of neural activities, we present a rigid registration architecture by implementation of Graphics Processing Unit (GPU). In this approach, the image stacks were preprocessed on GPU by mean stretching to reduce the computation effort. The present image was registered to the previous image stack that considered as reference. A fast Fourier transform (FFT) algorithm was used for calculating the shift of the image stack. The calculations for image registration were performed in different threads while the preparation functionality was refactored and called only once by the master thread. We implemented our registration algorithm on NVIDIA Quadro K4200 GPU under Compute Unified Device Architecture (CUDA) programming environment. The experimental results showed that the registration computation can speed-up to 550ms for a full high-resolution brain image. Our approach also has potential to be used for other dynamic image registrations in biomedical applications.

  8. The use of contrast agent for imaging biological samples

    Energy Technology Data Exchange (ETDEWEB)

    Dammer, J; Sopko, V; Jakubek, J [Institute of Experimental and Applied Physics, Czech Technical University in Prague, Horska 3a/22, CZ 12800 Prague 2 (Czech Republic); Weyda, F, E-mail: jiri.dammer@utef.cvut.cz [Biological center of the Academy of Sciences of the Czech Republic, Institute of Entomology, Branisovska 31, CZ-37005 Ceske Budejovice (Czech Republic)

    2011-01-15

    The technique of X-ray transmission imaging has been available for over a century and is still among the fastest and easiest approaches to the studies of internal structure of biological samples. Recent advances in semiconductor technology have led to the development of new types of X-ray detectors with direct conversion of interacting X-ray photon to an electric signal. Semiconductor pixel detectors seem to be specially promising; compared to the film technique, they provide single-quantum and real-time digital information about the objects being studied. We describe the recently developed radiographic apparatus, equipped with Medipix2 semiconductor pixel detector. The detector is used as an imager that counts individual photons of ionizing radiation, emitted by an X-ray tube (micro- or nano-focus FeinFocus). Thanks to the wide dynamic range of the Medipix2 detector and its high spatial resolution better than 1{mu}m, the setup is particularly suitable for radiographic imaging of small biological samples, including in-vivo observations with contrast agent (Optiray). Along with the description of the apparatus we provide examples of the use iodine contrast agent as a tracer in various insects as model organisms. The motivation of our work is to develop our imaging techniques as non-destructive and non-invasive. Microradiographic imaging helps detect organisms living in a not visible environment, visualize the internal biological processes and also to resolve the details of their body (morphology). Tiny live insects are an ideal object for our studies.

  9. AAZTA: an ideal chelating agent for the development of {sup 44}Sc PET imaging agents

    Energy Technology Data Exchange (ETDEWEB)

    Nagy, Gabor; Szikra, Dezso; Trencsenyi, Gyoergy [Scanomed Ltd., Debrecen (Hungary); University of Debrecen, Medical Imaging Clinic (Hungary); Fekete, Aniko [University of Debrecen, Medical Imaging Clinic (Hungary); Garai, Ildiko [Scanomed Ltd., Debrecen (Hungary); Giani, Arianna M.; Negri, Roberto [Dipartimento di Scienze del Farmaco, Universita del Piemonte Orientale, Novara (Italy); Masciocchi, Norberto [Dipartimento di Scienza e Alta Tecnologia e To.Sca.Lab, Universita degli Studi dell' Insubria, Como (Italy); Maiocchi, Alessandro; Uggeri, Fulvio [Bracco Imaging spa, Bracco Research Centre, Colleretto Giacosa (Italy); Toth, Imre [Department of Inorganic and Analytical Chemistry, University of Debrecen (Hungary); Aime, Silvio [Dipartimento di Biotecnologie Molecolari e Scienze della Salute, Centro di Imaging Molecolare e Preclinico, Universita degli Studi di Torino (Italy); Giovenzana, Giovanni B. [Dipartimento di Scienze del Farmaco, Universita del Piemonte Orientale, Novara (Italy); CAGE Chemicals srl, Novara (Italy); Baranyai, Zsolt [Bracco Imaging spa, Bracco Research Centre, Colleretto Giacosa (Italy); Department of Inorganic and Analytical Chemistry, University of Debrecen (Hungary)

    2017-02-13

    Unprecedented fast and efficient complexation of Sc{sup III} was demonstrated with the chelating agent AAZTA (AAZTA=1,4-bis(carboxymethyl)-6-[bis(carboxymethyl)] amino-6-methylperhydro-1,4-d iazepine) under mild experimental conditions. The robustness of the {sup 44}Sc(AAZTA){sup -} chelate and conjugated biomolecules thereof is further shown by in vivo PET imaging in healthy and tumor mice models. The new results pave the way towards development of efficient Sc-based radiopharmaceuticals using the AAZTA chelator. (copyright 2017 Wiley-VCH Verlag GmbH and Co. KGaA, Weinheim)

  10. Early detection of breast cancer: a molecular optical imaging approach using novel estrogen conjugate fluorescent dye

    Science.gov (United States)

    Bhattacharjee, Shubhadeep; Jose, Iven

    2011-02-01

    Estrogen induced proliferation of mutant cells is widely understood to be the one of major risk determining factor in the development of breast cancer. Hence determination of the Estrogen Receptor[ER] status is of paramount importance if cancer pathogenesis is to be detected and rectified at an early stage. Near Infrared Fluorescence [NIRf] Molecular Optical Imaging is emerging as a powerful tool to monitor bio-molecular changes in living subjects. We discuss pre-clinical results in our efforts to develop an optical imaging diagnostic modality for the early detection of breast cancer. We have successfully carried out the synthesis and characterization of a novel target-specific NIRf dye conjugate aimed at measuring Estrogen Receptor[ER] status. The conjugate was synthesized by ester formation between 17-β estradiol and a hydrophilic derivative of Indocyanine Green (ICG) cyanine dye, bis-1,1-(4-sulfobutyl) indotricarbocyanine-5-carboxylic acid, sodium salt. In-vitro studies regarding specific binding and endocytocis of the dye performed on ER+ve [MCF-7] and control [MDA-MB-231] adenocarcinoma breast cancer cell lines clearly indicated nuclear localization of the dye for MCF-7 as compared to plasma level staining for MDA-MB-231. Furthermore, MCF-7 cells showed ~4.5-fold increase in fluorescence signal intensity compared to MDA-MB-231. A 3-D mesh model mimicking the human breast placed in a parallel-plate DOT Scanner is created to examine the in-vivo efficacy of the dye before proceeding with clinical trials. Photon migration and florescence flux intensity is modeled using the finite-element method with the coefficients (quantum yield, molar extinction co-efficient etc.) pertaining to the dye as obtained from photo-physical and in-vitro studies. We conclude by stating that this lipophilic dye can be potentially used as a target specific exogenous contrast agent in molecular optical imaging for early detection of breast cancer.

  11. Photoactivation and imaging of optical highlighter fluorescent proteins.

    Science.gov (United States)

    Patterson, George H

    2011-07-01

    A major advance in the microscopic study of cells and tissues is the introduction of photoactivatable fluorescent proteins, which can specifically mark proteins of interest within a living cell. Fluorescent proteins are now available that allow a pool of molecules to be "turned on" by photoactivation. This unit discusses technical aspects for the general use of photoactivatable fluorescent proteins and introduces some specific applications in the concluding remarks.

  12. Functional surface engineering of quantum dot hydrogels for selective fluorescence imaging of extracellular lactate release.

    Science.gov (United States)

    Zhang, Xiaomeng; Ding, Shushu; Cao, Sumei; Zhu, Anwei; Shi, Guoyue

    2016-06-15

    Selective and sensitive detection of extracellular lactate is of fundamental significance for studying the metabolic alterations in tumor progression. Here we report the rational design and synthesis of a quantum-dot-hydrogel-based fluorescent probe for biosensing and bioimaging the extracellular lactate. By surface engineering the destabilized quantum dot sol with Nile Blue, the destabilized Nile-Blue-functionalized quantum dot sol cannot only self-assemble forming quantum dot hydrogel but also monitor lactate in the presence of nicotinamide adenine dinucleotide cofactor and lactate dehydrogenase through fluorescence resonance energy transfer. Notably, the surface engineered quantum dot hydrogel show high selectivity toward lactate over common metal ions, amino acids and other small molecules that widely coexist in biological system. Moreover, the destabilized Nile-Blue-functionalized quantum dots can encapsulate isolated cancer cells when self-assembled into a hydrogel and thus specifically detect and image the extracellular lactate metabolism. By virtue of these properties, the functionalized quantum dot hydrogel was further successfully applied to monitor the effect of metabolic agents.

  13. Semiautomatic Landmark-Based Two-Dimensional—Three-Dimensional Image Fusion in Living Mice: Correlation of Near-Infrared Fluorescence Imaging of Cy5.5-Labeled Antibodies with Flat-Panel Volume Computed Tomography

    Directory of Open Access Journals (Sweden)

    Christian Dullin

    2009-01-01

    Full Text Available Connecting fluorescence signals with anatomic structures enhances our ability to monitor biologic processes in mice. Here, we present a semiautomated approach to correlate two-dimensional (2D noninvasive near-infrared fluorescence (NIRF imaging with three-dimensional (3D, high-resolution, flat-panel volume computed tomography (fpVCT. We developed an algorithm to colocalize fluorescence signals of NIRF-labeled antibodies directed against matriptase and urokinase plasminogen activator receptor (uPAR to orthotopic carcinomas in mice visualized by fpVCT. For this purpose, mice were anesthetized and fixed on a multimodality animal bed containing fiducial markers filled with iodine-containing contrast agent and fluorescent dye. After intravenous administration of contrast agent and Cy5.5-labeled antibodies, NIRF and fpVCT images were obtained, without repositioning the mice. Binding of Cy5.5-labeled matriptase-specific antibody to pancreatic tumors and Cy5.5-labeled uPAR-specific antibody to mammary carcinomas was assessed by time-domain NIRF imaging measuring the location of fluorescence intensity and its lifetime. In summary, we developed a novel 2D-3D registration technique for image fusion with NIRF imaging and fpVCT to provide complementary information in tumor models on the in vivo association of functional information with anatomic structures. The combination of fpVCT with NIRF imaging will now allow targeted and effective monitoring of preclinical tumor therapies.

  14. Paired-agent imaging for resection during surgery (PAIRS) of head and neck squamous cell carcinomas (Conference Presentation)

    Science.gov (United States)

    Samkoe, Kimberley S.; Tichauer, Kenneth M.; Chen, Eunice; Gunn, Jason R.; Hoopes, P. Jack; Wells, Wendy A.; Hasan, Tayyaba; Pogue, Brian W.

    2016-03-01

    Ninety percent of patients with head and neck squamous cell carcinomas (HNSCC) have overexpression of epidermal growth factor receptor (EGFR), which is correlated with poor prognosis. Complete surgical resection of HNSCC tumors has a large impact on patient survival, where detection of tumor at or close to surgical margins increases the risk of death at 5-years by 90%. In addition, large surgical margins can greatly increase the morbidity experienced by the patient due to functional and cosmetic damage of oral and facial structures. Single fluorescence targeting agents are often used for tumor detection in in vivo pre-clinical imaging; however, the arising signal is qualitative at best because it is a complex mixture of vascular perfusion, vascular leakage, inhibited lymphatic clearance, and receptor binding. In vivo ratiometric receptor concentration imaging (RCI) allows quantification of receptor expression (hence identification of cancerous tissue) by utilizing co-administered paired-agents consisting of a targeted agent and non-targeted perfusion agent to reference the plasma delivery and leakage. A panel of HNSCC tumors with varying levels of EGFR expression (SCC-15 >SCC-25 > SCC-09) have been imaged using ABY-029, a clinically relevant anti-EGFR affibody labeled with IRDye 800CW, and affibody control imaging agent labeled with IRDye 680RD. RCI maps of in vivo tissue have been created and are spatially correlated with EGFR and CD31 immunohistochemistry and basic H and E staining. The RCI threshold parameters for distinguishing tumor from normal tissues (skin and muscle) and the accuracy of margin detection in these tumors will be presented. RCI surgical resection will be further developed using a novel multi-channel, gated fluorescence-guided surgery (FGS) imaging system that is capable of performing RCI in normal room light.

  15. Study of the hydrogen peroxide bleaching agent effects on bovine enamel using X-ray fluorescence

    Energy Technology Data Exchange (ETDEWEB)

    Moreira, Ruda F.; Calazans, Fernanda S.; Miranda, Mauro S.; Santos, Ramon S.; Anjos, Marcelino J.; Assis, Joaquim T. [Universidade do Estado do Rio de Janeiro (UERJ), Rio de Janeiro, RJ (Brazil)

    2013-07-01

    Hydrogen Peroxide's a bleaching agent capable of oxidizing a wide range of colored organic, causing discoloration and hence bleaching of the substrate, but some authors related the occurrence of side effects related to bleaching of the tooth structure, such as changes in morphology superficial. It was used 6 bovine incisors, each tooth was initially evaluated six times in different areas to obtain the count of elements phosphorus and calcium using X-Ray Fluorescence. The teeth were randomly divided in two groups: both groups were submitted to bleaching in office with hydrogen peroxide 38%, once a week during three weeks. Group 1 was stored in distilled water and group 2 in artificial saliva, between the sessions. The measurements were repeated every seven days before the bleaching treatment. Besides that, changes in mineral levels were always assessed in the same area and using the same procedure. It was observed that the bleaching was not able to demineralize the tooth enamel studied. (author)

  16. Fluorescence-enhanced imaging using a novel hand-held based optical imager: phantom studies

    Science.gov (United States)

    Ge, Jiajia; Zhu, Banghe; Regalado, Steven; Godavarty, Anuradha

    2008-02-01

    Near-infrared (NIR) optical imaging is an emerging noninvasive modality for breast cancer diagnosis. The currently available optical imaging systems towards tomography studies are limited either by instrument portability, patient comfort, or flexibility to image any given tissue volume. Hence, a novel hand-held probe based gain modulated intensified CCD camera imaging system is developed such that it can possibly overcome some of the above limitations. The unique features of this hand-held probe based optical imaging system are: (i) to perform simultaneous multiple point illumination and detection, thus decreasing the total imaging time and improving overall signal strength; (ii) to adapt to the tissue contours, thus decreasing the light leakage at contact surface; and (iii) to obtain trans-illumination measurements apart from reflectance measurements, thus improving the depth information. Phantom studies are performed to demonstrate the feasibility of performing fluorescence optical imaging under different target depths using cubical phantoms (10×6.5×10 cc). The effect of simultaneous multiple point illumination over sequential single point illumination is demonstrated from experimental phantom studies.

  17. Influence of angle's ranges for recording an X-ray fluorescence hologram on reconstructed atomic images

    Institute of Scientific and Technical Information of China (English)

    XIE Hong-Lan; CHEN Jian-Wen; GAO Hong-Yi; ZHU Hua-Feng; LI Ru-Xin; XU Zhi-Zhan

    2004-01-01

    X-ray fluorescence holography (XFH) is a novel method for three-dimensional (3D) imaging of atomic structure. Theoretically, in an XFH experiment, one has to measure the fluorescence energy on a spherical surface to get well-resolved 3D images of atoms. But in practice, the experimental system arrangement does not allow the measurement of the fluorescent intensity oscillations in the full sphere. The holographic information losses because of the limited sampling range (less than 4π) will directly result in defective reconstructed atomic images. In this work, the atomic image of a Fe single crystal (001) was reconstructed by numerically simulating X-ray fluorescence holograms of the crystal at different recording angle's ranges and step lengths. Influences of the ranges of azimuth angles and polar angles and the step length of polar angles on the reconstructed atomic images were discussed.

  18. Two-photon excited fluorescence microendoscopic imaging using a GRIN lens

    Science.gov (United States)

    Yan, Wei; Peng, Xiao; Lin, Danying; Wang, Qi; Gao, Jian; Zhou, Jie; Ye, Tong; Qu, Junle; Niu, Hanben

    2015-03-01

    With the rapid development of life sciences, there is an increasing demand for intravital fluorescence imaging of small animals. However, large dimensions and limited working distances of objective lenses in traditional fluorescence microscopes have limited the imaging applications mostly to superficial tissues. To overcome this disadvantage, researchers have developed the graded-index (GRIN) probes with small diameters for imaging internal organs of small animals in a minimally invasive fashion. Here, we present the development of a fluorescence endoscopic imaging system based on a GRIN lens using two-photon excitation. Experimental results showed that this system could perform dynamic fluorescence microendoscopic imaging and monitor the blood flow in anesthetized living mice using two-photon excitation.

  19. A detection instrument for enhanced-fluorescence and label-free imaging on photonic crystal surfaces.

    Science.gov (United States)

    Block, Ian D; Mathias, Patrick C; Ganesh, Nikhil; Jones, Sarah I; Dorvel, Brian R; Chaudhery, Vikram; Vodkin, Lila O; Bashir, Rashid; Cunningham, Brian T

    2009-07-20

    We report on the design and demonstration of an optical imaging system capable of exciting surface-bound fluorophores within the resonant evanescent electric field of a photonic crystal surface and gathering fluorescence emission that is directed toward the imaging objective by the photonic crystal. The system also has the ability to quantify shifts in the local resonance angle induced by the adsorption of biomolecules on the photonic crystal surface for label-free biomolecular imaging. With these two capabilities combined within a single detection system, we demonstrate label-free images self-registered to enhanced fluorescence images with 328x more sensitive fluorescence detection relative to a glass surface. This technique is applied to a DNA microarray where label-free quantification of immobilized capture DNA enables improved quality control and subsequent enhanced fluorescence detection of dye-tagged hybridized DNA yields 3x more genes to be detected versus commercially available microarray substrates.

  20. First X-ray fluorescence CT experimental results at the SSRF X-ray imaging beamline

    Institute of Scientific and Technical Information of China (English)

    DENG Biao; YANG Qun; XIE Hong-Lan; DU Guo-Hao; XIAO Wi-Qiao

    2011-01-01

    X-ray fluorescence CT is a non-destructive technique for detecting elemental composition and distribution inside a specimen. In this paper, the first experimental results of X-ray fluorescence CT obtained at the SSRF X-ray imaging beamline (BL13W1) are described. The test samples were investigated and the 2D elemental image was reconstructed using a filtered back-projection algorithm. In the sample the element Cd was observed. Up to now, the X-ray fluorescence CT could be carried out at the SSRF X-ray imaging beamline.

  1. Fluorescence tomographic imaging of sentinel lymph node using near-infrared emitting bioreducible dextran nanogels

    Directory of Open Access Journals (Sweden)

    Li J

    2014-12-01

    Full Text Available Jiejing Li,1* Beiqi Jiang,1* Chao Lin,2 Zhigang Zhuang1 1Department of Breast Surgery, Shanghai First Maternity and Infant Hospital, 2The Institute for Biomedical Engineering and Nanoscience, Tongji University School of Medicine, Tongji University, Shanghai, People’s Republic of China *These authors contributed equally to this work Abstract: Sentinel lymph node (SLN mapping is a critical procedure for SLN biopsy and its diagnosis as tumor metastasis in clinical practice. However, SLN mapping agents used in the clinic frequently cause side effects and complications in the patients. Here, we report the development of a near-infrared (NIR emitting polymeric nanogel with hydrodynamic diameter of ~28 nm – which is the optimal size for SLN uptake – for noninvasive fluorescence mapping of SLN in a mouse. This polymeric nanogel was obtained by coupling Cy7, an NIR dye, to the self-assembled nanogel from disulfide-linked dextran-deoxycholic acid conjugate with the dextran of 10 kDa, denoted as Dex–Cy7. Fluorescence imaging analysis showed that Dex–Cy7 nanogels had an enhanced photostability when compared to Cy7 alone. After intradermal injection of Dex–Cy7 nanogel into the front paw of a mouse, the nanogels were able to migrate into the mouse’s axillary lymph node, exhibiting longer retention time and higher fluorescence intensity in the node when compared to Cy7 alone. An immunohistofluorescence assay revealed that the nanogels were localized in the central region of lymph node and that the uptake was largely by the macrophages. In vitro and in vivo toxicity results indicated that the dextran-based nanogels were of low cytotoxicity at a polymer concentration up to 1,000 µg/mL and harmless to normal liver and kidney organs in mice at an intravenous dose of 1.25 mg/kg. The results of this study suggest that NIR-emitting polymeric nanogels based on bioreducible dextran-deoxycholic acid conjugates show high potential as fluorescence

  2. Liposomal encapsulation of a near-infrared fluorophore enhances fluorescence quenching and reliable whole body optical imaging upon activation in vivo.

    Science.gov (United States)

    Tansi, Felista L; Rüger, Ronny; Rabenhold, Markus; Steiniger, Frank; Fahr, Alfred; Kaiser, Werner A; Hilger, Ingrid

    2013-11-11

    In the past decade, there has been significant progress in the development of water soluble near-infrared fluorochromes for use in a wide range of imaging applications. Fluorochromes with high photo and thermal stability, sensitivity, adequate pharmacological properties and absorption/emission maxima within the near infrared window (650-900 nm) are highly desired for in vivo imaging, since biological tissues show very low absorption and auto-fluorescence at this spectrum window. Taking these properties into consideration, a myriad of promising near infrared fluorescent probes has been developed recently. However, a hallmark of most of these probes is a rapid clearance in vivo, which hampers their application. It is hypothesized that encapsulation of the near infrared fluorescent dye DY-676-COOH, which undergoes fluorescence quenching at high concentrations, in the aqueous interior of liposomes will result in protection and fluorescence quenching, which upon degradation by phagocytes in vivo will lead to fluorescence activation and enable imaging of inflammation. Liposomes prepared with high concentrations of DY-676-COOH reveal strong fluorescence quenching. It is demonstrated that the non-targeted PEGylated fluorescence-activatable liposomes are taken up predominantly by phagocytosis and degraded in lysosomes. Furthermore, in zymosan-induced edema models in mice, the liposomes are taken up by monocytes and macrophages which migrate to the sites of inflammation. Opposed to free DY-676-COOH, prolonged stability and retention of liposomal-DY-676-COOH is reflected in a significant increase in fluorescence intensity of edema. Thus, protected delivery and fluorescence quenching make the DY-676-COOH-loaded liposomes a highly promising contrast agent for in vivo optical imaging of inflammatory diseases.

  3. Detection of fecal residue on poultry carcasses by laser-induced fluorescence imaging.

    Science.gov (United States)

    Cho, B; Kim, M S; Chao, K; Lawrence, K; Park, B; Kim, K

    2009-04-01

    Feasibility of fluorescence imaging technique for the detection of diluted fecal matters from various parts of the digestive tract, including colon, ceca, small intestine, and duodenum, on poultry carcasses was investigated. One of the challenges for using fluorescence imaging for inspection of agricultural material is the low fluorescence yield in that fluorescence can be masked by ambient light. A laser-induced fluorescence imaging system (LIFIS) developed by our group allowed acquisition of fluorescence from feces-contaminated poultry carcasses in ambient light. Fluorescence emission images at 630 nm were captured with 415-nm laser excitation. Image processing algorithms including threshold and image erosion were used to identify fecal spots diluted up to 1: 10 by weight with double distilled water. Feces spots on the carcasses, without dilution and up to 1: 5 dilutions, could be detected with 100% accuracy regardless of feces type. Detection accuracy for fecal matters diluted up to 1: 10 was 96.6%. The results demonstrated good potential of the LIFIS for detection of diluted poultry fecal matter, which can harbor pathogens, on poultry carcasses.

  4. Evaluation of chemical fluorescent dyes as a protein conjugation partner for live cell imaging.

    Directory of Open Access Journals (Sweden)

    Yoko Hayashi-Takanaka

    Full Text Available To optimize live cell fluorescence imaging, the choice of fluorescent substrate is a critical factor. Although genetically encoded fluorescent proteins have been used widely, chemical fluorescent dyes are still useful when conjugated to proteins or ligands. However, little information is available for the suitability of different fluorescent dyes for live imaging. We here systematically analyzed the property of a number of commercial fluorescent dyes when conjugated with antigen-binding (Fab fragments directed against specific histone modifications, in particular, phosphorylated H3S28 (H3S28ph and acetylated H3K9 (H3K9ac. These Fab fragments were conjugated with a fluorescent dye and loaded into living HeLa cells. H3S28ph-specific Fab fragments were expected to be enriched in condensed chromosomes, as H3S28 is phosphorylated during mitosis. However, the degree of Fab fragment enrichment on mitotic chromosomes varied depending on the conjugated dye. In general, green fluorescent dyes showed higher enrichment, compared to red and far-red fluorescent dyes, even when dye:protein conjugation ratios were similar. These differences are partly explained by an altered affinity of Fab fragment after dye-conjugation; some dyes have less effect on the affinity, while others can affect it more. Moreover, red and far-red fluorescent dyes tended to form aggregates in the cytoplasm. Similar results were observed when H3K9ac-specific Fab fragments were used, suggesting that the properties of each dye affect different Fab fragments similarly. According to our analysis, conjugation with green fluorescent dyes, like Alexa Fluor 488 and Dylight 488, has the least effect on Fab affinity and is the best for live cell imaging, although these dyes are less photostable than red fluorescent dyes. When multicolor imaging is required, we recommend the following dye combinations for optimal results: Alexa Fluor 488 (green, Cy3 (red, and Cy5 or CF640 (far-red.

  5. Green synthesis of peptide-templated fluorescent copper nanoclusters for temperature sensing and cellular imaging.

    Science.gov (United States)

    Huang, Hong; Li, Hua; Wang, Ai-Jun; Zhong, Shu-Xian; Fang, Ke-Ming; Feng, Jiu-Ju

    2014-12-21

    A simple and green approach was developed for the preparation of fluorescent Cu nanoclusters (NCs) using the artificial peptide CLEDNN as a template. The as-synthesized Cu NCs exhibited a high fluorescence quantum yield (7.3%) and good stability, along with excitation and temperature dependent fluorescent properties, which could be employed for temperature sensing. Further investigations demonstrated low toxicity of Cu NCs for cellular imaging.

  6. Development of Tc-99m Imaging Agents for Abeta Plaques

    Energy Technology Data Exchange (ETDEWEB)

    Zhi-Ping, Zhuang; Mei-Ping Kung; Catherihne Hou; Hank F. Kung

    2008-09-26

    Development of SPECT imaging agents based on Tc-99m targeting Aβ plaques is useful for diagnosis of Alzheimer’s disease (AD). A stilbene derivative, [11C]SB-13, showing promise in detecting senile plaques present in AD patients has been reported previously1,2. Based on the 4’-amino-stilbene core structure we have added substituted groups through which a chelating group, N2S2, was conjugated. We report herein a series of Tc-99m labeled stilbene derivative conjugated with a TcO[N2S2] core. The syntheses of stilbenes containing a N2S2 chelating ligand are achieved by a scheme shown. Lipophilic 99mTc stilbene complexes were successfully prepared and purified through HPLC. Preliminary results of in vitro labeling of brain sections from transgenic mice showed very promising plaque labeling. These 99mTc stilbene derivatives are warranted for further evaluations as potential imaging agents targeting amyloid plaques.

  7. Precise diagnosis in different scenarios using photoacoustic and fluorescence imaging with dual-modality nanoparticles.

    Science.gov (United States)

    Peng, Dong; Du, Yang; Shi, Yiwen; Mao, Duo; Jia, Xiaohua; Li, Hui; Zhu, Yukun; Wang, Kun; Tian, Jie

    2016-08-14

    Photoacoustic imaging and fluorescence molecular imaging are emerging as important research tools for biomedical studies. Photoacoustic imaging offers both strong optical absorption contrast and high ultrasonic resolution, and fluorescence molecular imaging provides excellent superficial resolution, high sensitivity, high throughput, and the ability for real-time imaging. Therefore, combining the imaging information of both modalities can provide comprehensive in vivo physiological and pathological information. However, currently there are limited probes available that can realize both fluorescence and photoacoustic imaging, and advanced biomedical applications for applying this dual-modality imaging approach remain underexplored. In this study, we developed a dual-modality photoacoustic-fluorescence imaging nanoprobe, ICG-loaded Au@SiO2, which was uniquely designed, consisting of gold nanorod cores and indocyanine green with silica shell spacer layers to overcome fluorophore quenching. This nanoprobe was examined by both PAI and FMI for in vivo imaging on tumor and ischemia mouse models. Our results demonstrated that the nanoparticles can specifically accumulate at the tumor and ischemic areas and be detected by both imaging modalities. Moreover, this dual-modality imaging strategy exhibited superior advantages for a precise diagnosis in different scenarios. The new nanoprobe with the dual-modality imaging approach holds great potential for diagnosis and stage classification of tumor and ischemia related diseases.

  8. Wavefront sensorless approaches to adaptive optics for in vivo fluorescence imaging of mouse retina

    Science.gov (United States)

    Wahl, Daniel J.; Bonora, Stefano; Mata, Oscar S.; Haunerland, Bengt K.; Zawadzki, Robert J.; Sarunic, Marinko V.; Jian, Yifan

    2016-03-01

    Adaptive optics (AO) is necessary to correct aberrations when imaging the mouse eye with high numerical aperture. In order to obtain cellular resolution, we have implemented wavefront sensorless adaptive optics for in vivo fluorescence imaging of mouse retina. Our approach includes a lens-based system and MEMS deformable mirror for aberration correction. The AO system was constructed with a reflectance channel for structural images and fluorescence channel for functional images. The structural imaging was used in real-time for navigation on the retina using landmarks such as blood vessels. We have also implemented a tunable liquid lens to select the retinal layer of interest at which to perform the optimization. At the desired location on the mouse retina, the optimization algorithm used the fluorescence image data to drive a modal hill-climbing algorithm using an intensity or sharpness image quality metric. The optimization requires ~30 seconds to complete a search up to the 20th Zernike mode. In this report, we have demonstrated the AO performance for high-resolution images of the capillaries in a fluorescence angiography. We have also made progress on an approach to AO with pupil segmentation as a possible sensorless technique suitable for small animal retinal imaging. Pupil segmentation AO was implemented on the same ophthalmic system and imaging performance was demonstrated on fluorescent beads with induced aberrations.

  9. Multiphoton fluorescence and second harmonic generation microscopy for imaging keratoconus

    Science.gov (United States)

    Sun, Yen; Lo, Wen; Lin, Sung-Jan; Lin, Wei-Chou; Jee, Shiou-Hwa; Tan, Hsin-Yuan; Dong, Chen-Yuan

    2006-02-01

    The purpose of this study is to assess the possible application of multiphoton fluorescence and second harmonic generation (SHG) microscopy for imaging the structural features of keratoconus cornea and to evaluate its potential as being a clinical in vivo monitoring technique. Using the near-infrared excitation source from a titanium-sapphire laser pumped by a diode-pumped, solid state (DPSS) laser system, we can induce and simultaneously acquire multiphoton autofluorescence and SHG signals from the cornea specimens with keratoconus. A home-modified commercial microscope system with specified optical components is used for optimal signal detection. Keratoconus cornea button from patient with typical clinical presentation of keratoconus was obtained at the time of penetrating keratoplasty. The specimen was also sent for the histological examination as comparison. In all samples of keratoconus, destruction of lamellar structure with altered collagen fiber orientation was observed within whole layer of the diseased stromal area. In addition, the orientation of the altered collagen fibers within the cone area shows a trend directing toward the apex of the cone, which might implicate the biomechanical response of the keratoconus stroma to the intraocular pressure. Moreover, increased autofluorescent cells were also found in the cone area, with increased density as one approaches the apical area. In conclusion, multiphoton autofluorescence and SHG microscopy non-invasively demonstrated the morphological features of keratoconus cornea, especially the structural alternations of the stromal lamellae. We believe that in the future the multiphoton microscopy can be applied in vivo as an effective, non-invasive diagnostic and monitoring technique for keratoconus.

  10. Submicron hard X-ray fluorescence imaging of synthetic elements

    Energy Technology Data Exchange (ETDEWEB)

    Jensen, Mark P., E-mail: mjensen@anl.gov [Chemical Sciences and Engineering Division, Argonne National Laboratory, Argonne, IL 60439 (United States); Aryal, Baikuntha P. [Chemical Sciences and Engineering Division, Argonne National Laboratory, Argonne, IL 60439 (United States); Department of Chemistry, University of Chicago, Chicago, IL 60637 (United States); Gorman-Lewis, Drew [Chemical Sciences and Engineering Division, Argonne National Laboratory, Argonne, IL 60439 (United States); Paunesku, Tatjana [Department of Radiation Oncology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States); Department of Radiology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States); Lai, Barry; Vogt, Stefan [X-ray Science Division, Advanced Photon Source, Argonne National Laboratory, Argonne, IL 60439 (United States); Woloschak, Gayle E. [Department of Radiation Oncology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States); Department of Radiology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611 (United States)

    2012-04-13

    Highlights: Black-Right-Pointing-Pointer Actinide elements are mapped with L-edge X-rays and better than 400 nm resolution. Black-Right-Pointing-Pointer A typical detection limit was 2.9 Multiplication-Sign 10{sup -20} moles Pu {mu}m{sup -2}. Black-Right-Pointing-Pointer XANES measurements provide chemical information in 0.1 {mu}m{sup 2} spots. Black-Right-Pointing-Pointer Selection of materials for encapsulation is important for avoiding interferences. - Abstract: Synchrotron-based X-ray fluorescence microscopy (XFM) using hard X-rays focused into sub-micron spots is a powerful technique for elemental quantification and mapping, as well as microspectroscopic measurements such as {mu}-XANES (X-ray absorption near edge structure). We have used XFM to image and simultaneously quantify the transuranic element plutonium at the L{sub 3} or L{sub 2}-edge as well as Th and lighter biologically essential elements in individual rat pheochromocytoma (PC12) cells after exposure to the long-lived plutonium isotope {sup 242}Pu. Elemental maps demonstrate that plutonium localizes principally in the cytoplasm of the cells and avoids the cell nucleus, which is marked by the highest concentrations of phosphorus and zinc, under the conditions of our experiments. The minimum detection limit under typical acquisition conditions with an incident X-ray energy of 18 keV for an average 202 {mu}m{sup 2} cell is 1.4 fg Pu or 2.9 Multiplication-Sign 10{sup -20} moles Pu {mu}m{sup -2}, which is similar to the detection limit of K-edge XFM of transition metals at 10 keV. Copper electron microscopy grids were used to avoid interference from gold X-ray emissions, but traces of strontium present in naturally occurring calcium can still interfere with plutonium detection using its L{sub {alpha}} X-ray emission.

  11. Highly stable polymer coated nano-clustered silver plates: a multimodal optical contrast agent for biomedical imaging.

    Science.gov (United States)

    Ray, Aniruddha; Mukundan, Ananya; Xie, Zhixing; Karamchand, Leshern; Wang, Xueding; Kopelman, Raoul

    2014-11-07

    Here, we present a new optical contrast agent based on silver nanoplate clusters embedded inside of a polymer nano matrix. Unlike nanosphere clusters, which have been well studied, nanoplate clusters have unique properties due to the different possible orientations of interaction between the individual plates, resulting in a significant broadening of the absorption spectra. These nanoclusters were immobilized inside of a polymer cladding so as to maintain their stability and optical properties under in vivo conditions. The polymer-coated silver nanoplate clusters show a lower toxicity compared to the uncoated nanoparticles. At high nanoparticle concentrations, cell death occurs mostly due to apoptosis. These nanoparticles were used for targeted fluorescence imaging in a rat glioma cell line by incorporating a fluorescent dye into the matrix, followed by conjugation of a tumor targeting an F3 peptide. We further used these nanoparticles as photoacoustic contrast agents in vivo to enhance the contrast of the vasculature structures in a rat ear model. We observed a contrast enhancement of over 90% following the nanoparticle injection. It is also shown that these NPs can serve as efficient contrast agents, with specific targeting abilities for broadband multimodal imaging that are usable for diagnostic applications and that extend into use as therapeutic agents as well.

  12. Highly stable polymer coated nano-clustered silver plates: a multimodal optical contrast agent for biomedical imaging

    Science.gov (United States)

    Ray, Aniruddha; Mukundan, Ananya; Xie, Zhixing; Karamchand, Leshern; Wang, Xueding; Kopelman, Raoul

    2014-11-01

    Here, we present a new optical contrast agent based on silver nanoplate clusters embedded inside of a polymer nano matrix. Unlike nanosphere clusters, which have been well studied, nanoplate clusters have unique properties due to the different possible orientations of interaction between the individual plates, resulting in a significant broadening of the absorption spectra. These nanoclusters were immobilized inside of a polymer cladding so as to maintain their stability and optical properties under in vivo conditions. The polymer-coated silver nanoplate clusters show a lower toxicity compared to the uncoated nanoparticles. At high nanoparticle concentrations, cell death occurs mostly due to apoptosis. These nanoparticles were used for targeted fluorescence imaging in a rat glioma cell line by incorporating a fluorescent dye into the matrix, followed by conjugation of a tumor targeting an F3 peptide. We further used these nanoparticles as photoacoustic contrast agents in vivo to enhance the contrast of the vasculature structures in a rat ear model. We observed a contrast enhancement of over 90% following the nanoparticle injection. It is also shown that these NPs can serve as efficient contrast agents, with specific targeting abilities for broadband multimodal imaging that are usable for diagnostic applications and that extend into use as therapeutic agents as well.

  13. In vivo magnetic resonance and fluorescence dual imaging of tumor sites by using dye-doped silica-coated iron oxide nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Jang, Haeyun; Lee, Chaedong [Seoul National University, Program in Nano Science and Technology, Graduate School of Convergence Science and Technology (Korea, Republic of); Nam, Gi-Eun [University of Ulsan College of Medicine, Department of Radiology, Asan Medical Center (Korea, Republic of); Quan, Bo [Seoul National University, Program in Nano Science and Technology, Graduate School of Convergence Science and Technology (Korea, Republic of); Choi, Hyuck Jae [University of Ulsan College of Medicine, Department of Radiology, Asan Medical Center (Korea, Republic of); Yoo, Jung Sun [Seoul National University, Department of Transdisciplinary Studies, Graduate School of Convergence Science and Technology, Smart Humanity Convergence Center (Korea, Republic of); Piao, Yuanzhe, E-mail: parkat9@snu.ac.kr [Seoul National University, Program in Nano Science and Technology, Graduate School of Convergence Science and Technology (Korea, Republic of)

    2016-02-15

    The difficulty in delineating tumor is a major obstacle for better outcomes in cancer treatment of patients. The use of single-imaging modality is often limited by inadequate sensitivity and resolution. Here, we present the synthesis and the use of monodisperse iron oxide nanoparticles coated with fluorescent silica nano-shells for fluorescence and magnetic resonance dual imaging of tumor. The as-synthesized core–shell nanoparticles were designed to improve the accuracy of diagnosis via simultaneous tumor imaging with dual imaging modalities by a single injection of contrast agent. The iron oxide nanocrystals (∼11 nm) were coated with Rhodamine B isothiocyanate-doped silica shells via reverse microemulsion method. Then, the core–shell nanoparticles (∼54 nm) were analyzed to confirm their size distribution by transmission electron microscopy and dynamic laser scattering. Photoluminescence spectroscopy was used to characterize the fluorescent property of the dye-doped silica shell-coated nanoparticles. The cellular compatibility of the as-prepared nanoparticles was confirmed by a trypan blue dye exclusion assay and the potential as a dual-imaging contrast agent was verified by in vivo fluorescence and magnetic resonance imaging. The experimental results show that the uniform-sized core–shell nanoparticles are highly water dispersible and the cellular toxicity of the nanoparticles is negligible. In vivo fluorescence imaging demonstrates the capability of the developed nanoparticles to selectively target tumors by the enhanced permeability and retention effects and ex vivo tissue analysis was corroborated this. Through in vitro phantom test, the core/shell nanoparticles showed a T2 relaxation time comparable to Feridex{sup ®} with smaller size, indicating that the as-made nanoparticles are suitable for imaging tumor. This new dual-modality-nanoparticle approach has promised for enabling more accurate tumor imaging.

  14. In vivo magnetic resonance and fluorescence dual imaging of tumor sites by using dye-doped silica-coated iron oxide nanoparticles

    Science.gov (United States)

    Jang, Haeyun; Lee, Chaedong; Nam, Gi-Eun; Quan, Bo; Choi, Hyuck Jae; Yoo, Jung Sun; Piao, Yuanzhe

    2016-02-01

    The difficulty in delineating tumor is a major obstacle for better outcomes in cancer treatment of patients. The use of single-imaging modality is often limited by inadequate sensitivity and resolution. Here, we present the synthesis and the use of monodisperse iron oxide nanoparticles coated with fluorescent silica nano-shells for fluorescence and magnetic resonance dual imaging of tumor. The as-synthesized core-shell nanoparticles were designed to improve the accuracy of diagnosis via simultaneous tumor imaging with dual imaging modalities by a single injection of contrast agent. The iron oxide nanocrystals ( 11 nm) were coated with Rhodamine B isothiocyanate-doped silica shells via reverse microemulsion method. Then, the core-shell nanoparticles ( 54 nm) were analyzed to confirm their size distribution by transmission electron microscopy and dynamic laser scattering. Photoluminescence spectroscopy was used to characterize the fluorescent property of the dye-doped silica shell-coated nanoparticles. The cellular compatibility of the as-prepared nanoparticles was confirmed by a trypan blue dye exclusion assay and the potential as a dual-imaging contrast agent was verified by in vivo fluorescence and magnetic resonance imaging. The experimental results show that the uniform-sized core-shell nanoparticles are highly water dispersible and the cellular toxicity of the nanoparticles is negligible. In vivo fluorescence imaging demonstrates the capability of the developed nanoparticles to selectively target tumors by the enhanced permeability and retention effects and ex vivo tissue analysis was corroborated this. Through in vitro phantom test, the core/shell nanoparticles showed a T2 relaxation time comparable to Feridex® with smaller size, indicating that the as-made nanoparticles are suitable for imaging tumor. This new dual-modality-nanoparticle approach has promised for enabling more accurate tumor imaging.

  15. Time-Resolved Fluorescence Spectroscopy and Fluorescence Lifetime Imaging Microscopy for Characterization of Dendritic Polymer Nanoparticles and Applications in Nanomedicine

    Directory of Open Access Journals (Sweden)

    Alexander Boreham

    2016-12-01

    Full Text Available The emerging field of nanomedicine provides new approaches for the diagnosis and treatment of diseases, for symptom relief and for monitoring of disease progression. One route of realizing this approach is through carefully constructed nanoparticles. Due to the small size inherent to the nanoparticles a proper characterization is not trivial. This review highlights the application of time-resolved fluorescence spectroscopy and fluorescence lifetime imaging microscopy (FLIM for the analysis of nanoparticles, covering aspects ranging from molecular properties to particle detection in tissue samples. The latter technique is particularly important as FLIM allows for distinguishing of target molecules from the autofluorescent background and, due to the environmental sensitivity of the fluorescence lifetime, also offers insights into the local environment of the nanoparticle or its interactions with other biomolecules. Thus, these techniques offer highly suitable tools in the fields of particle development, such as organic chemistry, and in the fields of particle application, such as in experimental dermatology or pharmaceutical research.

  16. A fluorescent probe for imaging p53-MDM2 protein-protein interaction.

    Science.gov (United States)

    Liu, Zhenzhen; Miao, Zhenyuan; Li, Jin; Fang, Kun; Zhuang, Chunlin; Du, Lupei; Sheng, Chunquan; Li, Minyong

    2015-04-01

    In this article, we describe a no-wash small-molecule fluorescent probe for detecting and imaging p53-MDM2 protein-protein interaction based on an environment-sensitive fluorescent turn-on mechanism. After extensive biological examination, this probe L1 exhibited practical activity and selectivity in vitro and in cellulo.

  17. Evaluation of intraoperative fluorescence imaging-guided surgery in cancer-bearing dogs: a prospective proof-of-concept phase II study in 9 cases.

    Science.gov (United States)

    Cabon, Quentin; Sayag, David; Texier, Isabelle; Navarro, Fabrice; Boisgard, Raphaël; Virieux-Watrelot, Dorothée; Ponce, Frédérique; Carozzo, Claude

    2016-04-01

    The objective was to prospectively evaluate the application of intraoperative fluorescence imaging (IOFI) in the surgical excision of malignant masses in dogs, using a novel lipid nanoparticle contrast agent. Dogs presenting with spontaneous soft-tissue sarcoma or subcutaneous tumors were prospectively enrolled. Clinical staging and whole-body computed tomography (CT) were performed. All the dogs received an intravenous injection of dye-loaded lipid nanoparticles, LipImage 815. Wide or radical resection was realized after CT examination. Real-time IOFI was performed before skin incision and after tumor excision. In cases of radical resection, the lymph nodes (LNs) were imaged. The margin/healthy tissues fluorescence ratio or LN/healthy tissues fluorescence ratio was measured and compared with the histologic margins or LN status. Nine dogs were included. Limb amputation was performed in 3 dogs, and wide resection in 6. No adverse effect was noted. Fluorescence was observed in all 9 of the tumors. The margins were clean in 5 of 6 dogs after wide surgical resection, and the margin/healthy tissues fluorescence ratio was close to 1.0 in all these dogs. Infiltrated margins were observed in 1 case, with a margin/healthy tissues fluorescence ratio of 3.2. Metastasis was confirmed in 2 of 3 LNs, associated with LN/healthy tissues fluorescence ratios of 2.1 and 4.2, whereas nonmetastatic LN was associated with a ratio of 1.0. LipImage 815 used as a contrast agent during IOFI seemed to allow for good discrimination between tumoral and healthy tissues. Future studies are scheduled to evaluate the sensitivity and specificity of IOFI using LipImage 815 as a tracer.

  18. Delivery of imaging and therapeutic agents to tumor using pHLIP

    Science.gov (United States)

    Wijesinghe, Dayanjali; Moshnikova, Anna; Rossi, Bethany; Engelman, Donald; Andreev, Oleg; Reshetnyak, Yana

    2012-02-01

    We are developing a novel technology for selective delivery of imaging probes and membrane-impermeable molecules to cancer cells. It is based on action of water-soluble membrane peptide, pHLIP^ (pH [Low] Insertion Peptide), which has ability to insert and fold in cellular membrane at slightly acidic environment, which is a characteristic for various pathological states including cancer. The insertion of the peptide is unidirectional: C-terminus moves inside the cell across membrane, while N-terminus flags outside. Thus pHLIP possess dual delivery capability. Imaging agents (fluorescent, PET, SPECT or MRI) could be attached to the N-terminus of the peptide to mark tumor mass and tumor margins with high precision. At the same time, therapeutic molecules attached to the C-inserting end, could be moved across membrane to reach cytoplasmic target. Among translocated molecules are synthetic cyclic peptides, gene regulation agent (peptide nucleic acid) and phalla- and amanita toxins with hydrophobicity tuned by attachment of fatty acids for optimum delivery. Currently we have family of pHLIP peptides for various applications. The work is supported by NIH grants CA133890 to OAA, DME, YRK.

  19. Fluorescence imaging of single Kinesin motors on immobilized microtubules.

    Science.gov (United States)

    Korten, Till; Nitzsche, Bert; Gell, Chris; Ruhnow, Felix; Leduc, Cécile; Diez, Stefan

    2011-01-01

    Recent developments in optical microscopy and nanometer tracking have greatly improved our understanding of cytoskeletal motor proteins. Using fluorescence microscopy, dynamic interactions are now routinely observed in vitro on the level of single molecules mainly using a geometry, where fluorescently labeled motors move on surface-immobilized filaments. In this chapter, we review recent methods related to single-molecule kinesin motility assays. In particular, we aim to provide practical advice on: how to set up the assays, how to acquire high-precision data from fluorescently labeled kinesin motors and attached quantum dots, and how to analyze data by nanometer tracking.

  20. Spectral phasor analysis allows rapid and reliable unmixing of fluorescence microscopy spectral images

    NARCIS (Netherlands)

    Fereidouni, F.; Bader, A.N.; Gerritsen, H.C.

    2012-01-01

    A new global analysis algorithm to analyse (hyper-) spectral images is presented. It is based on the phasor representation that has been demonstrated to be very powerful for the analysis of lifetime imaging data. In spectral phasor analysis the fluorescence spectrum of each pixel in the image is Fou

  1. Near-infrared fluorescence imaging of cancer mediated by tumor hypoxia and HIF1α/OATPs signaling axis.

    Science.gov (United States)

    Wu, Jason Boyang; Shao, Chen; Li, Xiangyan; Shi, Changhong; Li, Qinlong; Hu, Peizhen; Chen, Yi-Ting; Dou, Xiaoliang; Sahu, Divya; Li, Wei; Harada, Hiroshi; Zhang, Yi; Wang, Ruoxiang; Zhau, Haiyen E; Chung, Leland W K

    2014-09-01

    Near-infrared fluorescence (NIRF) imaging agents are promising tools for noninvasive cancer imaging. Here, we explored the mechanistic properties of a specific group of NIR heptamethine carbocyanines including MHI-148 dye we identified and synthesized, and demonstrated these dyes to achieve cancer-specific imaging and targeting via a hypoxia-mediated mechanism. We found that cancer cells and tumor xenografts exhibited hypoxia-dependent MHI-148 dye uptake in vitro and in vivo, which was directly mediated by hypoxia-inducible factor 1α (HIF1α). Microarray analysis and dye uptake assay further revealed a group of hypoxia-inducible organic anion-transporting polypeptides (OATPs) responsible for dye uptake, and the correlation between OATPs and HIF1α was manifested in progressive clinical cancer specimens. Finally, we demonstrated increased uptake of MHI-148 dye in situ in perfused clinical tumor samples with activated HIF1α/OATPs signaling. Our results establish these NIRF dyes as potential tumor hypoxia-dependent cancer-targeting agents and provide a mechanistic rationale for continued development of NIRF imaging agents for improved cancer detection, prognosis and therapy.

  2. Evaluating the use of fluorescent imaging for the quantification of dental fluorosis.

    Science.gov (United States)

    McGrady, Michael G; Ellwood, Roger P; Taylor, Andrew; Maguire, Anne; Goodwin, Michaela; Boothman, Nicola; Pretty, Iain A

    2012-11-01

    The quantification of fluorosis using fluorescence imaging (QLF) hardware and stain analysis software has been demonstrated in selected populations with good correlation between fluorescent image metrics and TF Index scores from photographs. The aim of this study was to evaluate the ability of QLF to quantify fluorosis in a population of subjects (aged 11-13) participating in an epidemiological caries and fluorosis survey in fluoridated and non-fluoridated communities in Northern England. Fluorescent images of the maxillary incisors were captured together with standardized photographs were scored blind for fluorosis using the TF Index. Subjects were excluded from the analysis if there were restorations or caries on the maxillary central incisors. Data were available for 1774 subjects (n=905 Newcastle, n=869 Manchester). The data from the fluorescence method demonstrated a significant correlation with TF Index scores from photographs (Kendall's tau = 0.332 pfluorosis or at low levels of fluorosis severity had an adverse impact on tooth fluorescence and hence the outcome variable. This in conjunction with an uneven distribution of subjects across the range of fluorosis presentations may have resulted in the lower than anticipated correlations between the fluorescent imaging metrics and the photographic fluorosis scores. Nevertheless, the fluorescence imaging technique was able to discriminate between a fluoridated and non-fluoridated population (pfluorosis when used adjunctively with photographic scoring.

  3. Determination of the modulation transfer function for a time-gated fluorescence imaging system.

    Science.gov (United States)

    Gundy, Sarah; Van der Putten, Wil; Shearer, Andy; Buckton, Daniel; Ryder, Alan G

    2004-01-01

    The use of fluorescence for cancer detection is currently under investigation. Presently, steady-state fluorescence detection methods are in use, but have limitations due to poor contrast between the fluorescence of the tumor and background autofluorescence. Improved contrast can be obtained with time-resolved techniques because of the differing lifetimes between autofluorescence and exogenous photosensitizers that selectively accumulate within tumor tissue. An imaging system is constructed using a fast-gated (200-ps) charge-coupled device (CCD) camera and a pulsed 635-nm laser diode. To characterize the ability of the system to transfer object contrast to an image, the modulation transfer function (MTF) of the system is acquired by employing an extended knife-edge technique. A knife-edge target is assembled by drilling a rectangular well into a block of polymethyl methacrylate (PMMA). The imaging system records images of the photosensitizer, chloroaluminum phthalocyanine tetrasulfonate (AlPcTS), within the well. AlPcTS was chosen to test the system because of its strong absorption of 635-nm, high fluorescence yield, and relatively long fluorescence lifetime (approximately 7.5 ns). The results show that the system is capable of resolving 10(-4) M AlPcTS fluorescence as small as 1 mm. The findings of this study contribute to the development of a time-gated imaging system using fluorescence lifetimes. Copyright 2004 Society of Photo-Optical Instrumentation Engineers.

  4. B-Spline potential function for maximum a-posteriori image reconstruction in fluorescence microscopy

    Directory of Open Access Journals (Sweden)

    Shilpa Dilipkumar

    2015-03-01

    Full Text Available An iterative image reconstruction technique employing B-Spline potential function in a Bayesian framework is proposed for fluorescence microscopy images. B-splines are piecewise polynomials with smooth transition, compact support and are the shortest polynomial splines. Incorporation of the B-spline potential function in the maximum-a-posteriori reconstruction technique resulted in improved contrast, enhanced resolution and substantial background reduction. The proposed technique is validated on simulated data as well as on the images acquired from fluorescence microscopes (widefield, confocal laser scanning fluorescence and super-resolution 4Pi microscopy. A comparative study of the proposed technique with the state-of-art maximum likelihood (ML and maximum-a-posteriori (MAP with quadratic potential function shows its superiority over the others. B-Spline MAP technique can find applications in several imaging modalities of fluorescence microscopy like selective plane illumination microscopy, localization microscopy and STED.

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

    NARCIS (Netherlands)

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

    2014-01-01

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

  6. Endoscopic image-guided thermal therapy using targeted near infrared fluorescent gold nanorods (Conference Presentation)

    Science.gov (United States)

    Elson, Daniel S.

    2016-09-01

    We present an in vivo study of endoscopic fluorescence image-guided photothermal therapy of human oesophageal adenocarcinoma in a murine xenograft model, using intratumoural or intravenous gold nanorods functionalised with Cy5.5 and EGFR.

  7. Fluorescence cell imaging and manipulation using conventional halogen lamp microscopy.

    Directory of Open Access Journals (Sweden)

    Kazuo Yamagata

    Full Text Available Technologies for vitally labeling cells with fluorescent dyes have advanced remarkably. However, to excite fluorescent dyes currently requires powerful illumination, which can cause phototoxic damage to the cells and increases the cost of microscopy. We have developed a filter system to excite fluorescent dyes using a conventional transmission microscope equipped with a halogen lamp. This method allows us to observe previously invisible cell organelles, such as the metaphase spindle of oocytes, without causing phototoxicity. Cells remain healthy even after intensive manipulation under fluorescence observation, such as during bovine, porcine and mouse somatic cell cloning using nuclear transfer. This method does not require expensive epifluorescence equipment and so could help to reduce the science gap between developed and developing countries.

  8. [Removal of fluorescent whitening agent by hydrogen peroxide oxidation catalyzed by activated carbon].

    Science.gov (United States)

    Liu, Hai-Long; Zhang, Zhong-Min; Zhao, Xia; Jiao, Ru-Yuan

    2014-06-01

    Degradation of fluorescent whitening agent VBL in the processes of activated carbon (AC) and activated carbon modified (ACM) adsorptions, hydrogen peroxide (H2O2) oxidation, and hydrogen peroxide oxidation catalyzed by activated carbon were studied. Mechanism of the above catalytic oxidation was also investigated by adding tert-Butyl alcohol (TBA), the free radical scavenger, and detecting the released gases. The results showed that: the activated carbon modified by Fe (NO3)3 (ACM)exhibited better adsorption removal than AC. Catalytic oxidation showed efficient removal of VBL, and the catalytic removal of AC (up to 95%) was significantly higher than that of ACM (58% only). Catalytic oxidation was inhibited by TBA, which indicates that the above reaction involved *OH radicals and atom oxygen generated by hydrogen peroxide with the presence of AC. The results of H2O2 decomposition and released gases detection involved in the process showed that activated carbon enhanced the decomposition of H2O2 which released oxygen and heat. More O2 was produced and higher temperature of the reactor was achieved, which indicated that H2O2 decomposition catalyzed by ACM was significantly faster than that of AC. Combining the results of VBL removal, it could be concluded that the rate of active intermediates (*OH radicals and atom oxygen) production by ACM catalytic reaction was faster than that of AC. These intermediates consumed themselves and produced O2 instead of degrading VBL. It seemed that the improper mutual matching of the forming rate of activating intermediates and the supply rate of reactants was an important reason for the lower efficiency of ACM catalytic reaction comparing with AC.

  9. Live imaging of Tribolium castaneum embryonic development using light-sheet-based fluorescence microscopy.

    Science.gov (United States)

    Strobl, Frederic; Schmitz, Alexander; Stelzer, Ernst H K

    2015-10-01

    Tribolium castaneum has become an important insect model organism for evolutionary developmental biology, genetics and biotechnology. However, few protocols for live fluorescence imaging of Tribolium have been reported, and little image data is available. Here we provide a protocol for recording the development of Tribolium embryos with light-sheet-based fluorescence microscopy. The protocol can be completed in 4-7 d and provides procedural details for: embryo collection, microscope configuration, embryo preparation and mounting, noninvasive live imaging for up to 120 h along multiple directions, retrieval of the live embryo once imaging is completed, and image data processing, for which exemplary data is provided. Stringent quality control criteria for developmental biology studies are also discussed. Light-sheet-based fluorescence microscopy complements existing toolkits used to study Tribolium development, can be adapted to other insect species, and requires no advanced imaging or sample preparation skills.

  10. Enhanced image reconstruction of three-dimensional fluorescent assays by subtractive structured-light illumination microscopy.

    Science.gov (United States)

    Choi, Jong-ryul; Kim, Donghyun

    2012-10-01

    We investigate improved image reconstruction of structured light illumination for high-resolution imaging of three-dimensional (3D) cell-based assays. For proof of concept, an in situ fluorescence optical detection system was built with a digital micromirror device as a spatial light modulator, for which phase and tilting angle in a grid pattern were varied to implement specific image reconstruction schemes. Subtractive reconstruction algorithms based on structured light illumination were used to acquire images of fluorescent microbeads deposited as a two-dimensional monolayer or in 3D alginate matrix. We have confirmed that an optical subtraction algorithm improves axial and lateral resolution by effectively removing out-of-focus fluorescence. The results suggest that subtractive image reconstruction can be useful for structured illumination microscopy of broad types of cell-based assays with high image resolution.

  11. Fabry-Perot-based Fourier-transform hyperspectral imaging allows multi-labeled fluorescence analysis.

    Science.gov (United States)

    Pisani, Marco; Zucco, Massimo

    2014-05-10

    We demonstrate the ability of our hyperspectral imaging device, based on a scanning Fabry-Perot interferometer, to obtain a single hyper-image of a sample marked with different fluorescent molecules, and to unambiguously discriminate them by observing their spectral fingerprints. An experiment carried out with cyanines, fluorescein, and quantum dots emitting in the yellow-orange region, demonstrates the feasibility of multi-labeled fluorescence microscopy without the use of multiple filter sets or dispersive means.

  12. Comparison of two detection algorithms for spot tracking in fluorescence microscopy images

    CSIR Research Space (South Africa)

    Mabaso, M

    2014-11-01

    Full Text Available for spot tracking in fluorescence microscopy images Matsilele Mabaso∗, Daniel Withey‡, Bhekisipho Twala† ∗ ‡MDS(MIAS) Council for Scientific and Industrial Research Pretoria, South Africa, Email: ∗MMabaso@csir.co.za †Department of Electrical Engineering.... The quantitative comparative results demonstrated the importance of spot detection in tracking contexts. I. INTRODUCTION In recent years, the field of fluorescence microscopy has been improved and automated, and a large volume of image data are being generated...

  13. MULTIPHOTON MICROSCOPIC IMAGING OF MOUSE INTESTINAL MUCOSA BASED ON TWO-PHOTON EXCITED FLUORESCENCE AND SECOND HARMONIC GENERATION

    Directory of Open Access Journals (Sweden)

    REN'AN XU

    2013-01-01

    Full Text Available Multiphoton microscopy (MPM, based on two-photon excited fluorescence and second harmonic generation, enables direct noninvasive visualization of tissue architecture and cell morphology in live tissues without the administration of exogenous contrast agents. In this paper, we used MPM to image the microstructures of the mucosa in fresh, unfixed, and unstained intestinal tissue of mouse. The morphology and distribution of the main components in mucosa layer such as columnar cells, goblet cells, intestinal glands, and a little collagen fibers were clearly observed in MPM images, and then compared with standard H&E images from paired specimens. Our results indicate that MPM combined with endoscopy and miniaturization probes has the potential application in the clinical diagnosis and in vivo monitoring of early intestinal cancer.

  14. Green Synthesis of Bifunctional Fluorescent Carbon Dots from Garlic for Cellular Imaging and Free Radical Scavenging.

    Science.gov (United States)

    Zhao, Shaojing; Lan, Minhuan; Zhu, Xiaoyue; Xue, Hongtao; Ng, Tsz-Wai; Meng, Xiangmin; Lee, Chun-Sing; Wang, Pengfei; Zhang, Wenjun

    2015-08-12

    Nitrogen and sulfur codoped carbon dots (CDs) were prepared from garlic by a hydrothermal method. The as-prepared CDs possess good water dispersibility, strong blue fluorescence emission with a fluorescent quantum yield of 17.5%, and excellent photo and pH stabilities. It is also demonstrated that the fluorescence of CDs are resistant to the interference of metal ions, biomolecules, and high ionic strength environments. Combining with low cytotoxicity properties, CDs could be used as an excellent fluorescent probe for cellular multicolor imaging. Moreover, the CDs were also demonstrated to exhibit favorable radical scavenging activity.

  15. Fluorescence imaging of tryptophan and collagen cross-links to evaluate wound closure ex vivo

    Science.gov (United States)

    Wang, Ying; Ortega-Martinez, Antonio; Farinelli, Bill; Anderson, R. R.; Franco, Walfre

    2016-02-01

    Wound size is a key parameter in monitoring healing. Current methods to measure wound size are often subjective, time-consuming and marginally invasive. Recently, we developed a non-invasive, non-contact, fast and simple but robust fluorescence imaging (u-FEI) method to monitor the healing of skin wounds. This method exploits the fluorescence of native molecules to tissue as functional and structural markers. The objective of the present study is to demonstrate the feasibility of using variations in the fluorescence intensity of tryptophan and cross-links of collagen to evaluate proliferation of keratinocyte cells and quantitate size of wound during healing, respectively. Circular dermal wounds were created in ex vivo human skin and cultured in different media. Two serial fluorescence images of tryptophan and collagen cross-links were acquired every two days. Histology and immunohistology were used to validate correlation between fluorescence and epithelialization. Images of collagen cross-links show fluorescence of the exposed dermis and, hence, are a measure of wound area. Images of tryptophan show higher fluorescence intensity of proliferating keratinocytes forming new epithelium, as compared to surrounding keratinocytes not involved in epithelialization. These images are complementary since collagen cross-links report on structure while tryptophan reports on function. HE and immunohistology show that tryptophan fluorescence correlates with newly formed epidermis. We have established a fluorescence imaging method for studying epithelialization processes during wound healing in a skin organ culture model, our approach has the potential to provide a non-invasive, non-contact, quick, objective and direct method for quantitative measurements in wound healing in vivo.

  16. Fluorescent imaging of single nanoparticles and viruses on a smart phone.

    Science.gov (United States)

    Wei, Qingshan; Qi, Hangfei; Luo, Wei; Tseng, Derek; Ki, So Jung; Wan, Zhe; Göröcs, Zoltán; Bentolila, Laurent A; Wu, Ting-Ting; Sun, Ren; Ozcan, Aydogan

    2013-10-22

    Optical imaging of nanoscale objects, whether it is based on scattering or fluorescence, is a challenging task due to reduced detection signal-to-noise ratio and contrast at subwavelength dimensions. Here, we report a field-portable fluorescence microscopy platform installed on a smart phone for imaging of individual nanoparticles as well as viruses using a lightweight and compact opto-mechanical attachment to the existing camera module of the cell phone. This hand-held fluorescent imaging device utilizes (i) a compact 450 nm laser diode that creates oblique excitation on the sample plane with an incidence angle of ~75°, (ii) a long-pass thin-film interference filter to reject the scattered excitation light, (iii) an external lens creating 2× optical magnification, and (iv) a translation stage for focus adjustment. We tested the imaging performance of this smart-phone-enabled microscopy platform by detecting isolated 100 nm fluorescent particles as well as individual human cytomegaloviruses that are fluorescently labeled. The size of each detected nano-object on the cell phone platform was validated using scanning electron microscopy images of the same samples. This field-portable fluorescence microscopy attachment to the cell phone, weighing only ~186 g, could be used for specific and sensitive imaging of subwavelength objects including various bacteria and viruses and, therefore, could provide a valuable platform for the practice of nanotechnology in field settings and for conducting viral load measurements and other biomedical tests even in remote and resource-limited environments.

  17. In-vivo optical detection of cancer using chlorin e6 – polyvinylpyrrolidone induced fluorescence imaging and spectroscopy

    Directory of Open Access Journals (Sweden)

    Soo Khee

    2009-01-01

    Full Text Available Abstract Background Photosensitizer based fluorescence imaging and spectroscopy is fast becoming a promising approach for cancer detection. The purpose of this study was to examine the use of the photosensitizer chlorin e6 (Ce6 formulated in polyvinylpyrrolidone (PVP as a potential exogenous fluorophore for fluorescence imaging and spectroscopic detection of human cancer tissue xenografted in preclinical models as well as in a patient. Methods Fluorescence imaging was performed on MGH human bladder tumor xenografted on both the chick chorioallantoic membrane (CAM and the murine model using a fluorescence endoscopy imaging system. In addition, fiber optic based fluorescence spectroscopy was performed on tumors and various normal organs in the same mice to validate the macroscopic images. In one patient, fluorescence imaging was performed on angiosarcoma lesions and normal skin in conjunction with fluorescence spectroscopy to validate Ce6-PVP induced fluorescence visual assessment of the lesions. Results Margins of tumor xenografts in the CAM model were clearly outlined under fluorescence imaging. Ce6-PVP-induced fluorescence imaging yielded a specificity of 83% on the CAM model. In mice, fluorescence intensity of Ce6-PVP was higher in bladder tumor compared to adjacent muscle and normal bladder. Clinical results confirmed that fluorescence imaging clearly captured the fluorescence of Ce6-PVP in angiosarcoma lesions and good correlation was found between fluorescence imaging and spectral measurement in the patient. Conclusion Combination of Ce6-PVP induced fluorescence imaging and spectroscopy could allow for optical detection and discrimination between cancer and the surrounding normal tissues. Ce6-PVP seems to be a promising fluorophore for fluorescence diagnosis of cancer.

  18. High-throughput Identification of Phage-derived Imaging Agents

    Directory of Open Access Journals (Sweden)

    Kimberly A. Kelly

    2006-01-01

    Full Text Available The use of phage-displayed peptide libraries is a powerful method for selecting peptides with desired binding properties. However, the validation and prioritization of “hits” obtained from this screening approach remains challenging. Here, we describe the development and testing of a new analysis method to identify and display hits from phage-display experiments and high-throughput enzyme-linked immunosorbent assay screens. We test the method using a phage screen against activated macrophages to develop imaging agents with higher specificity for active disease processes. The new methodology should be useful in identifying phage hits and is extendable to other library screening methods such as small-molecule and nanoparticle libraries.

  19. Non-fused phospholes as fluorescent probes for imaging of lipid droplets in living cells

    Science.gov (United States)

    Öberg, Elisabet; Appelqvist, Hanna; Nilsson, K. Peter R.

    2017-04-01

    Molecular tools for fluorescent imaging of specific compartments in cells are essential for understanding the function and activity of cells. Here, we report the synthesis of a series of pyridyl- and thienyl-substituted phospholes and the evaluation of these dyes for fluorescent imaging of cells. The thienyl-substituted phospholes proved to be successful for staining of cultured normal and malignant cells due to their fluorescent properties and low toxicity. Co-staining experiments demonstrated that these probes target lipid droplets, which are, lipid-storage organelles found in the cytosol of nearly all cell types. Our findings confirm that thienyl-substituted phospholes can be utilized as fluorescent tools for vital staining of cells, and we foresee that these fluorescent dyes might be used in studies to unravel the roles that lipid droplets play in cellular physiology and their role in diseases.

  20. In Vivo Imaging of Far-red Fluorescent Proteins after DNA Electrotransfer to Muscle Tissue

    Directory of Open Access Journals (Sweden)

    Hojman Pernille

    2009-04-01

    Full Text Available Abstract DNA electrotransfer to muscle tissue yields long-term, high levels of gene expression; showing great promise for future gene therapy. We want to characterize the novel far-red fluorescent protein Katushka as a marker for gene expression using time domain fluorescence in vivo imaging. Highly efficient transgenic expression was observed after DNA electrotransfer with 100-fold increase in fluorescent intensity. The fluorescent signal peaked 1 week after transfection and returned to background level within 4 weeks. Katushka expression was not as stable as GFP expression, which was detectable for 8 weeks. Depth and 3D analysis proved that the expression was located in the target muscle. In vivo bio-imaging using the novel Katushka fluorescent protein enables excellent evaluation of the transfection efficacy, and spatial distribution, but lacks long-term stability.

  1. Silica nanocapsules of fluorescent conjugated polymers and superparamagnetic nanocrystals for dual-mode cellular imaging.

    Science.gov (United States)

    Tan, Happy; Wang, Miao; Yang, Chang-Tong; Pant, Shilpa; Bhakoo, Kishore Kumar; Wong, Siew Yee; Chen, Zhi-Kuan; Li, Xu; Wang, John

    2011-06-01

    We describe here a facile and benign synthetic strategy to integrate the fluorescent behavior of conjugated polymers and superparamagnetic properties of iron oxide nanocrystals into silica nanocapsules, forming a new type of bifunctional magnetic fluorescent silica nanocapsule (BMFSN). The resultant BMFSNs are uniform, colloidally stable in aqueous medium, and exhibit the desired dual functionality of fluorescence and superparamagnetism in a single entity. Four conjugated polymers with different emissions were used to demonstrate the versatility of employing this class of fluorescent materials for the preparation of BMFSNs. The applicability of BMFSNs in cellular imaging was studied by incubating them with human liver cancer cells, the result of which demonstrated that the cells could be visualized by dual-mode fluorescence and magnetic resonance imaging. Furthermore, the superparamagnetic behavior of the BMFSNs was exploited for in vitro magnetic-guided delivery of the nanocapsules into the cancer cells, thereby highlighting their potential for targeting biomedical applications.

  2. In Vivo Imaging of Far-red Fluorescent Proteins after DNA Electrotransfer to Muscle Tissue

    DEFF Research Database (Denmark)

    Hojman, Pernille; Eriksen, Jens; Gehl, Julie

    2009-01-01

    DNA electrotransfer to muscle tissue yields long-term, high levels of gene expression; showing great promise for future gene therapy. We want to characterize the novel far-red fluorescent protein Katushka as a marker for gene expression using time domain fluorescence in vivo imaging. Highly...... weeks. Depth and 3D analysis proved that the expression was located in the target muscle. In vivo bio-imaging using the novel Katushka fluorescent protein enables excellent evaluation of the transfection efficacy, and spatial distribution, but lacks long-term stability....

  3. A Novel Molecular Fluorescent Technique for Imaging the Somatostatin Receptor 2, Using a DOTATOC Lanthanide Conjugate

    DEFF Research Database (Denmark)

    Andersen, Rune Wiik; Prakash, Vineet; Stensballe, Allan

    to easily obtain commercial receptor antibodies. We propose an alternative with the novel use of lanthanide fluorescent DOTATOC imaging.Purpose is to prove that it is feasible to combine the fluorescent nuclear imaging of neuroendocrine tumors with histopathological correlates using the same bio......-functional DOTATOC component.                       METHOD AND MATERIALS            The chelation of Europium and Samarium to DOTATOC was proven using MALDI-TOF Mass Spectrometry. The rise in quantum yield between unchelated lanthanides and those bound by DOTATOC was examined using fluorescence spectroscopy...

  4. Characterization of nanoparticle-based contrast agents for molecular magnetic resonance imaging

    Science.gov (United States)

    Shan, Liang; Chopra, Arvind; Leung, Kam; Eckelman, William C.; Menkens, Anne E.

    2012-09-01

    The development of molecular imaging agents is currently undergoing a dramatic expansion. As of October 2011, 4,800 newly developed agents have been synthesized and characterized in vitro and in animal models of human disease. Despite this rapid progress, the transfer of these agents to clinical practice is rather slow. To address this issue, the National Institutes of Health launched the Molecular Imaging and Contrast Agents Database (MICAD) in 2005 to provide freely accessible online information regarding molecular imaging probes and contrast agents for the imaging community. While compiling information regarding imaging agents published in peer-reviewed journals, the MICAD editors have observed that some important information regarding the characterization of a contrast agent is not consistently reported. This makes it difficult for investigators to evaluate and meta-analyze data generated from different studies of imaging agents, especially for the agents based on nanoparticles. This article is intended to serve as a guideline for new investigators for the characterization of preclinical studies performed with nanoparticle-based MRI contrast agents. The common characterization parameters are summarized into seven categories: contrast agent designation, physicochemical properties, magnetic properties, in vitro studies, animal studies, MRI studies, and toxicity. Although no single set of parameters is suitable to define the properties of the various types of contrast agents, it is essential to ensure that these agents meet certain quality control parameters at the preclinical stage, so that they can be used without delay for clinical studies.

  5. Bright field microscopy as an alternative to whole cell fluorescence in automated analysis of macrophage images.

    Directory of Open Access Journals (Sweden)

    Jyrki Selinummi

    Full Text Available BACKGROUND: Fluorescence microscopy is the standard tool for detection and analysis of cellular phenomena. This technique, however, has a number of drawbacks such as the limited number of available fluorescent channels in microscopes, overlapping excitation and emission spectra of the stains, and phototoxicity. METHODOLOGY: We here present and validate a method to automatically detect cell population outlines directly from bright field images. By imaging samples with several focus levels forming a bright field -stack, and by measuring the intensity variations of this stack over the -dimension, we construct a new two dimensional projection image of increased contrast. With additional information for locations of each cell, such as stained nuclei, this bright field projection image can be used instead of whole cell fluorescence to locate borders of individual cells, separating touching cells, and enabling single cell analysis. Using the popular CellProfiler freeware cell image analysis software mainly targeted for fluorescence microscopy, we validate our method by automatically segmenting low contrast and rather complex shaped murine macrophage cells. SIGNIFICANCE: The proposed approach frees up a fluorescence channel, which can be used for subcellular studies. It also facilitates cell shape measurement in experiments where whole cell fluorescent staining is either not available, or is dependent on a particular experimental condition. We show that whole cell area detection results using our projected bright field images match closely to the standard approach where cell areas are localized using fluorescence, and conclude that the high contrast bright field projection image can directly replace one fluorescent channel in whole cell quantification. Matlab code for calculating the projections can be downloaded from the supplementary site: http://sites.google.com/site/brightfieldorstaining.

  6. Applying fluorescence lifetime imaging microscopy to evaluate the efficacy of anticancer drugs

    Science.gov (United States)

    Kawanabe, Satoshi; Araki, Yoshie; Uchimura, Tomohiro; Imasaka, Totaro

    2015-06-01

    Fluorescence lifetime imaging microscopy was applied to evaluate the efficacy of anticancer drugs. A decrease in the fluorescence lifetime of the nucleus in apoptotic cancer cells stained by SYTO 13 dye was detected after treatment with antitumor antibiotics such as doxorubicin or epirubicin. It was confirmed that the change in fluorescence lifetime occurred earlier than morphological changes in the cells. We found that the fluorescence lifetime of the nucleus in the cells treated with epirubicin decreased more rapidly than that of the cells treated with doxorubicin. This implies that epirubicin was more efficacious than doxorubicin in the treatment of cancer cells. The change in fluorescence lifetime was, however, not indicated when the cells were treated with cyclophosphamide. The decrease in fluorescence lifetime was associated with the processes involving caspase activation and chromatin condensation. Therefore, this technique would provide useful information about apoptotic cells, particularly in the early stages.

  7. An endoscopic fluorescence imaging system for simultaneous visual examination and photodetection of cancers

    Science.gov (United States)

    Wagnières, Georges A.; Studzinski, André P.; van den Bergh, Hubert E.

    1997-01-01

    We describe the design and performance tested during six years of clinical trials of a fluorescence endoscope for the detection and delineation of cancers in several hollow organs. The apparatus is based on the imaging of the laser-induced fluorescence that differs between a tumor and its surrounding normal tissue. The tests are carried out in the upper aerodigestive tract, the tracheobronchial tree, the esophagus, and the colon. In the three former cases an exogenous dye is used (Photofrin II), whereas in the latter case fluorescein molecules conjugated with monoclonal antibodies directed against carcinoembryonic antigen are injected. The decrease of native tissue autofluorescence observed in early cancers is also used for detecting lesions in the tracheobronchial tree. The fluorescence contrast between the tumor and surrounding normal tissue is enhanced by real time image processing. This is done by simultaneously recording the fluorescence image in two spectral domains, after which these two images are digitized and manipulated with a mathematical operator (look-up table) at video frequency. Moreover, the device that is described below allows for an immediate observation of the endoscopic area under white light illumination during fluorescence detection in order to localize the origin of the "positive" fluorescence signals. Typical results obtained in the tracheobronchial tree and in the colon are presented and the sources of false positives and false negatives are evaluated in terms of the fluorescent dye, tissue optical properties, and illumination optics.

  8. The Impact of “Omic” and Imaging Technologies on Assessing the Host Immune Response to Biodefence Agents

    Directory of Open Access Journals (Sweden)

    Julia A. Tree

    2014-01-01

    Full Text Available Understanding the interactions between host and pathogen is important for the development and assessment of medical countermeasures to infectious agents, including potential biodefence pathogens such as Bacillus anthracis, Ebola virus, and Francisella tularensis. This review focuses on technological advances which allow this interaction to be studied in much greater detail. Namely, the use of “omic” technologies (next generation sequencing, DNA, and protein microarrays for dissecting the underlying host response to infection at the molecular level; optical imaging techniques (flow cytometry and fluorescence microscopy for assessing cellular responses to infection; and biophotonic imaging for visualising the infectious disease process. All of these technologies hold great promise for important breakthroughs in the rational development of vaccines and therapeutics for biodefence agents.

  9. Imaging Membrane Potential with Two Types of Genetically Encoded Fluorescent Voltage Sensors.

    Science.gov (United States)

    Lee, Sungmoo; Piao, Hong Hua; Sepheri-Rad, Masoud; Jung, Arong; Sung, Uhna; Song, Yoon-Kyu; Baker, Bradley J

    2016-02-04

    Genetically encoded voltage indicators (GEVIs) have improved to the point where they are beginning to be useful for in vivo recordings. While the ultimate goal is to image neuronal activity in vivo, one must be able to image activity of a single cell to ensure successful in vivo preparations. This procedure will describe how to image membrane potential in a single cell to provide a foundation to eventually image in vivo. Here we describe methods for imaging GEVIs consisting of a voltage-sensing domain fused to either a single fluorescent protein (FP) or two fluorescent proteins capable of Förster resonance energy transfer (FRET) in vitro. Using an image splitter enables the projection of images created by two different wavelengths onto the same charge-coupled device (CCD) camera simultaneously. The image splitter positions a second filter cube in the light path. This second filter cube consists of a dichroic and two emission filters to separate the donor and acceptor fluorescent wavelengths depending on the FPs of the GEVI. This setup enables the simultaneous recording of both the acceptor and donor fluorescent partners while the membrane potential is manipulated via whole cell patch clamp configuration. When using a GEVI consisting of a single FP, the second filter cube can be removed allowing the mirrors in the image splitter to project a single image onto the CCD camera.

  10. Fluorescence detection and imaging of amino-functionalized organic monolayer

    Energy Technology Data Exchange (ETDEWEB)

    Shirahata, Naoto [National Institute for Materials Science (NIMS), 1-2-1 Sengen, Tsukuba, Ibaraki 305-0047 (Japan)], E-mail: SHIRAHATA.naoto@nims.go.jp; Furumi, Seiichi [National Institute for Materials Science (NIMS), 1-2-1 Sengen, Tsukuba, Ibaraki 305-0047 (Japan); Masuda, Yoshitake; Hozumi, Atsushi [National Institute of Advanced Industrial Science and Technology (AIST), 2266-98 Anagahora, Shimo-shidami, Moriyama, Nagoya 463-8560 (Japan); Sakka, Yoshio [National Institute for Materials Science (NIMS), 1-2-1 Sengen, Tsukuba, Ibaraki 305-0047 (Japan)

    2008-03-03

    Amino-terminated organic monolayer formed on silicon covered with native oxide (SiO{sub 2}/Si) was directly visualized under observation with fluorescent microscopy. This successful fluorescence visualization was achieved by a combination of fluorescamine method and photopatterning of the amino-terminated surface. As a typical example, an amino-terminated self-assembled monolayer (SAM) was formed on SiO{sub 2}/Si substrate in a vapor of 12.5 vol.% solution of N-(6-aminohexyl)-3-aminopropyltrimethoxysilane [H{sub 2}N(CH{sub 2}){sub 6}NH(CH{sub 2}){sub 3}Si(OCH{sub 3}){sub 3}, AHAPS] diluted with absolute toluene. A micropattern of AHAPS-SAM was photolithographycally prepared using 172 nm vacuum ultraviolet (VUV) light under a reduced pressure of 10 Pa for 30 min through a photomask. The resultant micropattern composed of AHAPS- and SiOH-covered regions was provided to fluorescamine method. Due to a nonluminescence of fluorescamine itself under UV/visible irradiation, a fluorescent emission could not be observed on SiOH regions of the micropattern. In contrast, fluorescamine reacted with the outermost amino group of the AHAPS-SAM to give a fluorescent emission. A comprehensible fluorescence method for verifying formation of an amino-terminated organic monolayer has been developed.

  11. Functional brain fluorescence plurimetry in rat by implantable concatenated CMOS imaging system.

    Science.gov (United States)

    Kobayashi, Takuma; Masuda, Hiroyuki; Kitsumoto, Chikara; Haruta, Makito; Motoyama, Mayumi; Ohta, Yasumi; Noda, Toshihiko; Sasagawa, Kiyotaka; Tokuda, Takashi; Shiosaka, Sadao; Ohta, Jun

    2014-03-15

    Measurement of brain activity in multiple areas simultaneously by minimally invasive methods contributes to the study of neuroscience and development of brain machine interfaces. However, this requires compact wearable instruments that do not inhibit natural movements. Application of optical potentiometry with voltage-sensitive fluorescent dye using an implantable image sensor is also useful. However, the increasing number of leads required for the multiple wired sensors to measure larger domains inhibits natural behavior. For imaging broad areas by numerous sensors without excessive wiring, a web-like sensor that can wrap the brain was developed. Kaleidoscopic potentiometry is possible using the imaging system with concatenated sensors by changing the alignment of the sensors. This paper describes organization of the system, evaluation of the system by a fluorescence imaging, and finally, functional brain fluorescence plurimetry by the sensor. The recorded data in rat somatosensory cortex using the developed multiple-area imaging system compared well with electrophysiology results.

  12. Removal of Out-of-Plane Fluorescence for Single Cell Visualization and Quantification in Cryo-Imaging

    OpenAIRE

    Steyer, Grant J.; Roy, Debashish; Salvado, Olivier; Stone, Meredith E.; Wilson, David L.

    2009-01-01

    We developed a cryo-imaging system, which alternates between sectioning (10–40 μm) and imaging bright field and fluorescence block-face image volumes with micronscale-resolution. For applications requiring single-cell detection of fluorescently labeled cells anywhere in a mouse, we are developing software for reduction of out-of-plane fluorescence. In mouse experiments, we imaged GFP-labeled cancer and stem cells, and cell-sized fluorescent microspheres. To remove out-of-plane fluorescence, w...

  13. Compact multispectral fluorescence imaging system with spectral multiplexed volume holographic grating

    Science.gov (United States)

    Lv, Yanlu; Cai, Chuangjian; Bai, Jing; Luo, Jianwen

    2016-12-01

    Traditional spectral imaging systems mainly rely on spatial scanning or spectral scanning methods to acquire spatial and spectral features. The acquisition is time-consuming and cannot fully satisfy the need of monitoring dynamic phenomenon and observing different structures of the specimen simultaneously. To overcome these barriers, we develop a video-rate simultaneous multispectral imaging system built with a spectral multiplexed volume holographic grating (VHG) and few optical components. Four spectral multiplexed volume holograms optimized for four discrete spectral bands (centered at 488 nm, 530 nm, 590 nm and 620 nm) are recorded into an 8×12 mm photo-thermal refractive glass. The diffraction efficiencies of all the holograms within the multiplexed VHG are greater than 80%. With the high throughout multiplexed VHG, the system can work with both reflection and fluorescence modes and allow simultaneous acquisition of spectral and spatial information with a single exposure. Imaging experiments demonstrate that the multispectral images of the target illuminated with white light source can be obtained. Fluorescence images of multiple fluorescence objects (two glass beads filled with 20 uL 1.0 mg/mL quantum dots solutions that emit 530 +/- 15 nm and 620 +/- 15 nm fluorescence, respectively) buried 3 mm below the surface of a tissue mimicking phantom are acquired. The results demonstrate that the system can provide complementary information in fluorescence imaging. The design diagram of the proposed system is given to explain the advantage of compactness and flexibility in integrating with other imaging platforms.

  14. Fluorescence microscopy imaging of cells with a plasmonic dish integrally molded

    Science.gov (United States)

    Tawa, Keiko; Sasakawa, Chisato; Fujita, Tsuyoshi; Kiyosue, Kazuyuki; Hosokawa, Chie; Nishii, Junji; Oike, Makoto; Kakinuma, Norihiro

    2016-03-01

    A plastic dish with a wavelength-scale periodic structure at a bottom panel was integrally molded and coated with thin metal films. The integrally molded dish called plasmonic dish was applied to bioimaging under a fluorescence microscope. On the plasmonic substrate, the enhanced electric field based on a grating-coupled surface plasmon resonance (GC-SPR) can provide an enhanced fluorescence. In this study, two kinds of cells, human embryonic kidney (HEK) cells and neuronal cells, were observed in our plasmonic dish. Fluorescence images of HEK cells were above 10 times brighter than those obtained on a conventional glass-bottomed dish. Neuronal cells were successfully cultured for 10 d on the plasmonic dish integrally molded, and in fluorescence images with transmitted light, a higher contrast was obtained than in epifluorescence images. The plasmonic dish integrally molded, as well as that fabricated by the UV nanoimprint method, was also found to be useful for sensitive bioimaging.

  15. Fluorescence Modified Chitosan-Coated Magnetic Nanoparticles for High-Efficient Cellular Imaging

    Directory of Open Access Journals (Sweden)

    Nie Fang

    2009-01-01

    Full Text Available Abstract Labeling of cells with nanoparticles for living detection is of interest to various biomedical applications. In this study, novel fluorescent/magnetic nanoparticles were prepared and used in high-efficient cellular imaging. The nanoparticles coated with the modified chitosan possessed a magnetic oxide core and a covalently attached fluorescent dye. We evaluated the feasibility and efficiency in labeling cancer cells (SMMC-7721 with the nanoparticles. The nanoparticles exhibited a high affinity to cells, which was demonstrated by flow cytometry and magnetic resonance imaging. The results showed that cell-labeling efficiency of the nanoparticles was dependent on the incubation time and nanoparticles’ concentration. The minimum detected number of labeled cells was around 104by using a clinical 1.5-T MRI imager. Fluorescence and transmission electron microscopy instruments were used to monitor the localization patterns of the magnetic nanoparticles in cells. These new magneto-fluorescent nanoagents have demonstrated the potential for future medical use.

  16. CEA-targeted nanoparticles allow specific in vivo fluorescent imaging of colorectal cancer models.

    Science.gov (United States)

    Tiernan, James P; Ingram, Nicola; Marston, Gemma; Perry, Sarah L; Rushworth, Jo V; Coletta, P Louise; Millner, Paul A; Jayne, David G; Hughes, Thomas A

    2015-01-01

    Fluorescent imaging of colorectal tumor cells would improve tumor localization and allow intra-operative staging, facilitating stratification of surgical resections thereby improving patient outcomes. We aimed to develop and test fluorescent nanoparticles capable of allowing this in vivo. Dye-doped silica nanoparticles were synthesized. Anti-CEA (carcinoembryonic antigen) or control IgGs were conjugated to nanoparticles using various chemical strategies. Binding of CEA-targeted or control nanoparticles to colorectal cancer cells was quantified in vitro, and in vivo after systemic-delivery to murine xenografts. CEA-targeted, polyamidoamine dendrimer-conjugated, nanoparticles, but not control nanoparticles, allowed strong tumor-specific imaging. We are the first to demonstrate live, specific, in vivo imaging of colorectal cancer cells using antibody-targeted fluorescent nanoparticles. These nanoparticles have potential to allow intra-operative fluorescent visualization of tumor cells.

  17. Hyperspectral imaging of endogenous fluorescent metabolic molecules to identify pain states in central nervous system tissue

    Science.gov (United States)

    Staikopoulos, Vasiliki; Gosnell, Martin E.; Anwer, Ayad G.; Mustafa, Sanam; Hutchinson, Mark R.; Goldys, Ewa M.

    2016-12-01

    Fluorescence-based bio-imaging methods have been extensively used to identify molecular changes occurring in biological samples in various pathological adaptations. Auto-fluorescence generated by endogenous fluorescent molecules within these samples can interfere with signal to background noise making positive antibody based fluorescent staining difficult to resolve. Hyperspectral imaging uses spectral and spatial imaging information for target detection and classification, and can be used to resolve changes in endogenous fluorescent molecules such as flavins, bound and free NADH and retinoids that are involved in cell metabolism. Hyperspectral auto-fluorescence imaging of spinal cord slices was used in this study to detect metabolic differences within pain processing regions of non-pain versus sciatic chronic constriction injury (CCI) animals, an established animal model of peripheral neuropathy. By using an endogenous source of contrast, subtle metabolic variations were detected between tissue samples, making it possible to distinguish between animals from non-injured and injured groups. Tissue maps of native fluorophores, flavins, bound and free NADH and retinoids unveiled subtle metabolic signatures and helped uncover significant tissue regions with compromised mitochondrial function. Taken together, our results demonstrate that hyperspectral imaging provides a new non-invasive method to investigate central changes of peripheral neuropathic injury and other neurodegenerative disease models, and paves the way for novel cellular characterisation in health, disease and during treatment, with proper account of intrinsic cellular heterogeneity.

  18. Evaluation of Mobile Phone Performance for Near-Infrared Fluorescence Imaging.

    Science.gov (United States)

    Ghassemi, Pejhman; Wang, Bohan; Wang, Jianting; Wang, Quanzeng; Chen, Yu; Pfefer, T Joshua

    2016-08-19

    We have investigated the potential for contrast-enhanced near-infrared fluorescence imaging of tissue on a mobile phone platform. CCD- and phone-based cameras were used to image molded and 3Dprinted tissue phantoms, and an ex vivo animal model. Quantitative and qualitative evaluations of image quality demonstrate the viability of this approach and elucidate variations in performance due to wavelength, pixel color and image processing.

  19. The diversity of (68)Ga-based imaging agents.

    Science.gov (United States)

    Velikyan, Irina

    2013-01-01

    Development of new radiopharmaceuticals and their availability are crucial factors influencing the expansion of clinical nuclear medicine. The number of new (68)Ga-based imaging agents for positron emission tomography (PET) is increasing greatly. (68)Ga has been used for labeling of a broad range of molecules (small organic molecules, peptides, proteins, and oligonucleotides) as well as particles, thus demonstrating its potential to become a PET analog of the legendary generator-produced gamma-emitting (99m)Tc but with added value of higher sensitivity and resolution as well as quantitation and dynamic scanning. Further, the availability of technology for GMP-compliant automated tracer production can facilitate the introduction of new radiopharmaceuticals and enable standardized, harmonized multicenter studies to be conducted for regulatory approval. This chapter presents some examples of tracers for targeted, pretargeted, and nontargeted imaging with emphasis on the potential of (68)Ga to facilitate clinically practical PET development and to promote the PET technique worldwide for earlier and better diagnostics, and personalized medicine with the ultimate objective of improved therapeutic outcome.

  20. Zinc Phthalocyanine Labelled Polyethylene Glycol: Preparation, Characterization, Interaction with Bovine Serum Albumin and Near Infrared Fluorescence Imaging in Vivo

    Directory of Open Access Journals (Sweden)

    Tianjun Liu

    2012-05-01

    Full Text Available Zinc phthalocyanine labelled polyethylene glycol was prepared to track and monitor the in vivo fate of polyethylene glycol. The chemical structures were characterized by nuclear magnetic resonance and infrared spectroscopy. Their light stability and fluorescence quantum yield were evaluated by UV-Visible and fluorescence spectroscopy methods. The interaction of zinc phthalocyanine labelled polyethylene glycol with bovine serum albumin was evaluated by fluorescence titration and isothermal titration calorimetry methods. Optical imaging in vivo, organ aggregation as well as distribution of fluorescence experiments for tracking polyethylene glycol were performed with zinc phthalocyanine labelled polyethylene glycol as fluorescent agent. Results show that zinc phthalocyanine labelled polyethylene glycol has good optical stability and high emission ability in the near infrared region. Imaging results demonstrate that zinc phthalocyanine labelled polyethylene glycol can track and monitor the in vivo process by near infrared fluorescence imaging, which implies its potential in biomaterials evaluation in vivo by a real-time noninvasive method.

  1. Quantitative method to assess caries via fluorescence imaging from the perspective of autofluorescence spectral analysis

    Science.gov (United States)

    Chen, Q. G.; Zhu, H. H.; Xu, Y.; Lin, B.; Chen, H.

    2015-08-01

    A quantitative method to discriminate caries lesions for a fluorescence imaging system is proposed in this paper. The autofluorescence spectral investigation of 39 teeth samples classified by the International Caries Detection and Assessment System levels was performed at 405 nm excitation. The major differences in the different caries lesions focused on the relative spectral intensity range of 565-750 nm. The spectral parameter, defined as the ratio of wavebands at 565-750 nm to the whole spectral range, was calculated. The image component ratio R/(G + B) of color components was statistically computed by considering the spectral parameters (e.g. autofluorescence, optical filter, and spectral sensitivity) in our fluorescence color imaging system. Results showed that the spectral parameter and image component ratio presented a linear relation. Therefore, the image component ratio was graded as 1.62 to quantitatively classify sound, early decay, established decay, and severe decay tissues, respectively. Finally, the fluorescence images of caries were experimentally obtained, and the corresponding image component ratio distribution was compared with the classification result. A method to determine the numerical grades of caries using a fluorescence imaging system was proposed. This method can be applied to similar imaging systems.

  2. Targeted contrast agents--an adjunct to whole-body imaging: current concepts.

    Science.gov (United States)

    Foran, Paul; Bolster, Ferdia; Crosbie, Ian; MacMahon, Peter; O'Kennedy, Richard; Eustace, Stephen J

    2010-03-01

    This article reviews the potential use of a combination of whole-body imaging and targeted contrast agents in improving diagnostics, with a particular focus on oncology imaging. It looks at the rationale for nanoparticles and their development as targeted contrast agents. It subsequently describes many of the advances made thus far in developing tissue-specific contrast agents capable of targeting tumors that combined with whole-body imaging may enable superior cancer detection and characterization.

  3. Efficient synthesis of highly fluorescent nitrogen-doped carbon dots for cell imaging using unripe fruit extract of Prunus mume

    Energy Technology Data Exchange (ETDEWEB)

    Atchudan, Raji; Edison, Thomas Nesakumar Jebakumar Immanuel [School of Chemical Engineering, Yeungnam University, Gyeongsan 38541 (Korea, Republic of); Sethuraman, Mathur Gopalakrishnan, E-mail: mgsethu@gmail.com [Department of Chemistry, Gandhigram Rural Institute-Deemed University, Gandhigram 624 302, Tamilnadu (India); Lee, Yong Rok, E-mail: yrlee@yu.ac.kr [School of Chemical Engineering, Yeungnam University, Gyeongsan 38541 (Korea, Republic of)

    2016-10-30

    Graphical abstract: The green synthesis of highly fluorescent N-CDs was achieved using the extract of unripe P. mume fruit as a carbon precursor by a one-pot simple hydrothermal-carbonization method. The resulting N-CDs were used as a staining agent for the fluorescence imaging of MDA-MB-231 cells. Display Omitted - Highlights: • The green synthesis of highly fluorescent N-CDs using the extract of unripe P. mume. • The N-CDs were synthesized by one-pot hydrothermal-carbonization method. • This method of synthesis is a simple, cost effective and eco-friendly route. • N-CDs will be a good alternative for fluorescent dyes and SQDs for bio-applications. - Abstract: Highly fluorescent nitrogen-doped carbon dots (N-CDs) were synthesized using the extract of unripe Prunus mume (P. mume) fruit by a simple one step hydrothermal-carbonization method. The N-CDs were synthesized at different pH ranges, 2.3, 5, 7, and 9. The pH of the P. mume extract was adjusted using an aqueous ammonia solution (25%). The optical properties of N-CDs were examined by UV–vis and fluorescence spectroscopy. The N-CDs synthesized at pH 9 emitted high fluorescence intensity compared to other obtained N-CDs. The N-CDs synthesized at pH 9 was further characterized by high resolution transmission electron microscopy (HR-TEM), X-ray diffraction (XRD), Raman spectroscopy, X-ray photoelectron spectroscopy (XPS), and Fourier transform-infra red (FT-IR) spectroscopy. HR-TEM showed that the average size of the synthesized N-CDs was approximately 9 nm and the interlayer distance was 0.21 nm, which was validated by XRD. The graphitic nature of the synthesized N-CDs were confirmed by Raman spectroscopy. XPS and FT-IR spectroscopy confirmed the doping of the nitrogen moiety over the synthesized CDs. The synthesized nitrogen doped CDs (N-CDs) were low toxicity and were used as a staining probe for fluorescence cell imaging.

  4. Cell-permeable Ln(III) chelate-functionalized InP quantum dots as multimodal imaging agents.

    Science.gov (United States)

    Stasiuk, Graeme J; Tamang, Sudarsan; Imbert, Daniel; Poillot, Cathy; Giardiello, Marco; Tisseyre, Céline; Barbier, Emmanuel L; Fries, Pascal Henry; de Waard, Michel; Reiss, Peter; Mazzanti, Marinella

    2011-10-25

    Quantum dots (QDs) are ideal scaffolds for the development of multimodal imaging agents, but their application in clinical diagnostics is limited by the toxicity of classical CdSe QDs. A new bimodal MRI/optical nanosized contrast agent with high gadolinium payload has been prepared through direct covalent attachment of up to 80 Gd(III) chelates on fluorescent nontoxic InP/ZnS QDs. It shows a high relaxivity of 900 mM(-1) s(-1) (13 mM(-1 )s(-1) per Gd ion) at 35 MHz (0.81 T) and 298 K, while the bright luminescence of the QDs is preserved. Eu(III) and Tb(III) chelates were also successfully grafted to the InP/ZnS QDs. The absence of energy transfer between the QD and lanthanide emitting centers results in a multicolor system. Using this convenient direct grafting strategy additional targeting ligands can be included on the QD. Here a cell-penetrating peptide has been co-grafted in a one-pot reaction to afford a cell-permeable multimodal multimeric MRI contrast agent that reports cellular localization by fluorescence and provides high relaxivity and increased tissue retention with respect to commercial contrast agents.

  5. Benzothiadiazole Derivatives as Fluorescence Imaging Probes: Beyond Classical Scaffolds.

    Science.gov (United States)

    Neto, Brenno A D; Carvalho, Pedro H P R; Correa, Jose R

    2015-06-16

    This Account describes the origins, features, importance, and trends of the use of fluorescent small-molecule 2,1,3-benzothiadiazole (BTD) derivatives as a new class of bioprobes applied to bioimaging analyses of several (live and fixed) cell types. BTDs have been successfully used as probes for a plethora of biological analyses for only a few years, and the impressive responses obtained by using this important class of heterocycle are fostering the development of new fluorescent BTDs and expanding the biological applications of such derivatives. The first use of a fluorescent small-molecule BTD derivative as a selective cellular probe dates back to 2010, and since then impressive advances have been described by us and others. The well-known limitations of classical scaffolds urged the development of new classes of bioprobes. Although great developments have been achieved by using classical scaffolds such as coumarins, BODIPYs, fluoresceins, rhodamines, cyanines, and phenoxazines, there is still much to be done, and BTDs aim to succeed where these dyes have shown their limitations. Important organelles and cell components such as nuclear DNA, mitochondria, lipid droplets, and others have already been successfully labeled by fluorescent small-molecule BTD derivatives. New technological systems that use BTDs as the fluorophores for bioimaging experiments have been described in recent scientific literature. The successful application of BTDs as selective bioprobes has led some groups to explore their potential for use in studying membrane pores or tumor cells under hypoxic conditions. Finally, BTDs have also been used as fluorescent tags to investigate the action mechanism of some antitumor compounds. The attractive photophysical data typically observed for π-extended BTD derivatives is fostering interest in the use of this new class of bioprobes. Large Stokes shifts, large molar extinction coefficients, high quantum yields, high stability when stored in solution or

  6. Fluorescence endoscopic imaging study of anastomotic recurrence of Crohn's disease after right ileocolonic resection

    Science.gov (United States)

    Mordon, Serge R.; Maunoury, Vincent; Klein, Olivier; Colombel, Jean-Frederic

    1995-12-01

    Crohn's disease is an inflammatory bowel disease of unknown etiology. Vasculitis is hypothesized but it was never demonstrated in vivo. This study aimed to evaluate the vascular mucosa perfusion using fluorescence imaging in 13 patients who had previously undergone eileocolonic resection and who agreed to participate in a prospective endoscopic study of anastomotic recurrence. This anastomotic recurrence rate is known to be high (73% after 1 year follow-up) and is characterized by ulcerations. The fluorescence study was started with an I.V. bolus injection of sodium fluorescein. The pre-anastomotic mucosa was endoscopically examined with blue light that stimulates fluorescein fluorescence. Fluorescence emission was recorded with an ultra-high-sensitivity camera connected to the endoscope via an interference filter (520 - 560 nm). A uniform fluorescence was observed a few seconds after the injection and lasted for 15 min in healthy subjects. In case of recurrence, the centers of the ulcerations displayed a very low fluorescence indicating localized ischemia. In contrast, the rims of the ulcers revealed brighter fluorescent images than those of normal mucosa. The anastomotic ulcerations of Crohn's disease recurrence exhibit a high fluorescence intensity at their margins indicating an increased mucosal blood flow and/or enhanced transcapillary diffusion. These findings support the hypothesis of a primary vasculitis in Crohn's disease.

  7. Cyanine-loaded lipid nanoparticles for improved in vivo fluorescence imaging

    Science.gov (United States)

    Texier, Isabelle; Goutayer, Mathieu; da Silva, Anabela; Guyon, Laurent; Djaker, Nadia; Josserand, Véronique; Neumann, Emmanuelle; Bibette, Jérôme; Vinet, Françoise

    2009-09-01

    Fluorescence is a very promising radioactive-free technique for functional imaging in small animals and, in the future, in humans. However, most commercial near-infrared dyes display poor optical properties, such as low fluorescence quantum yields and short fluorescence lifetimes. In this paper, we explore whether the encapsulation of infrared cyanine dyes within the core of lipid nanoparticles (LNPs) could improve their optical properties. Lipophilic dialkylcarbocyanines DiD and DiR are loaded very efficiently in 30-35-nm-diam lipid droplets stabilized in water by