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Sample records for fluorescence lifetime standards

  1. Expanding the potential of standard flow cytometry by extracting fluorescence lifetimes from cytometric pulse shifts

    National Research Council Canada - National Science Library

    Cao, Ruofan; Naivar, Mark A; Wilder, Mark; Houston, Jessica P

    2014-01-01

    Fluorescence lifetime measurements provide information about the fluorescence relaxation, or intensity decay, of organic fluorophores, fluorescent proteins, and other inorganic molecules that fluoresce...

  2. Stroboscopic fluorescence lifetime imaging.

    Science.gov (United States)

    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.

  3. FLUORESCENCE LIFETIME DISTRIBUTIONS IN PROTEINS

    OpenAIRE

    ALCALA, JR; Gratton, E; PRENDERGAST, FG

    1987-01-01

    The fluorescence lifetime value of tryptophan residues varies by more than a factor of 100 in different proteins and is determined by several factors, which include solvent exposure and interactions with other elements of the protein matrix. Because of the variety of different elements that can alter the lifetime value and the sensitivity to the particular environment of the tryptophan residue, it is likely that non-unique lifetime values result in protein systems. The emission decay of most ...

  4. In vivo fluorescence lifetime tomography

    Science.gov (United States)

    Nothdurft, Ralph E.; Patwardhan, Sachin V.; Akers, Walter; Ye, Yunpeng; Achilefu, Samuel; Culver, Joseph P.

    2009-03-01

    Local molecular and physiological processes can be imaged in vivo through perturbations in the fluorescence lifetime (FLT) of optical imaging agents. In addition to providing functional information, FLT methods can quantify specific molecular events and multiplex diagnostic and prognostic information. We have developed a fluorescence lifetime diffuse optical tomography (DOT) system for in vivo preclinical imaging. Data is captured using a time-resolved intensified charge coupled device (ICCD) system to measure fluorescence excitation and emission in the time domain. Data is then converted to the frequency domain, and we simultaneously reconstruct images of yield and lifetime using an extension to the normalized Born approach. By using differential phase measurements, we demonstrate DOT imaging of short lifetimes (from 350 ps) with high precision (+/-5 ps). Furthermore, this system retains the efficiency, speed, and flexibility of transmission geometry DOT. We demonstrate feasibility of FLT-DOT through a progressive series of experiments. Lifetime range and repeatability are first measured in phantoms. Imaging of subcutaneous implants then verifies the FLT-DOT approach in vivo in the presence of inhomogeneous optical properties. Use in a common research scenario is ultimately demonstrated by imaging accumulation of a targeted near-infrared (NIR) fluorescent-labeled peptide probe (cypate-RGD) in a mouse with a subcutaneous tumor.

  5. Lifetime Resolved Fluorescence Fluctuation Spectroscopy

    Science.gov (United States)

    Guo, Peng; Berland, Keith

    2009-11-01

    Fluorescence correlation spectroscopy (FCS) has been widely used investigate molecular dynamics and interactions in biological systems. FCS typically resolves the component species of a sample either through differences in diffusion coefficient or molecular brightness. Diffusion based assays currently have a major limitation which requires that the diffusion coefficients of component species in a sample must be substantially different in order to be resolved. This criterion is not met in many important cases, such as when molecules of similar molecular weight bind to each other. This limitation can be overcome, and resolution of FCS measurements enhanced, by combining FCS measurements with measurements of fluorescence lifetimes. By using of global analysis on simultaneously acquired FCS and lifetime data we show that we can dramatically enhance resolution in FCS measurements, and accurately resolve the concentration and diffusion coefficients of multiple sample components even when their diffusion coefficients are identical provided there is a difference in the lifetime of the component species. We show examples of this technique using both simulations and experiments. It is expected that this method will be of significance for binding assays studying molecular interactions.

  6. Mean fluorescence lifetime and its error

    Energy Technology Data Exchange (ETDEWEB)

    Fiserova, Eva [Department of Mathematical Analysis and Applications of Mathematics, Faculty of Science, Palacky University in Olomouc, tr. 17. listopadu 12, CZE-77146 Olomouc (Czech Republic); Kubala, Martin, E-mail: mkubala@prfnw.upol.cz [Department of Biophysics, Faculty of Science, Palacky University in Olomouc, tr. 17. listopadu 12, CZE-77146 Olomouc (Czech Republic)

    2012-08-15

    Mean excited-state lifetime is one of the fundamental fluorescence characteristics and enters as an important parameter into numerous calculations characterizing molecular interactions, such as e.g. FRET or fluorescence quenching. Our experiments demonstrated that the intensity-weighted mean fluorescence lifetime is very robust characteristic, in contrast to the amplitude-weighted one, which value is dependent on the data quality and particularly on the used fitting model. For the first time, we also report the procedure for the error estimation for both the intensity- and amplitude-weighted mean fluorescence lifetimes. Furthermore, we present a method for estimation of the mean fluorescence lifetime directly from the fluorescence-decay curve recorded by TCSPC (Time-Correlated Single-Photon Counting) method. For its simplicity and low computational demands, it could be a useful tool in the high-throughput applications, such as FACS, FLIM-FRET or HPLC detectors. - Highlights: Black-Right-Pointing-Pointer Intensity-weighted mean fluorescence lifetime is very robust characteristic. Black-Right-Pointing-Pointer The amplitude-weighted mean lifetime depends on the selection of fitting model. Black-Right-Pointing-Pointer Rigorous procedure for estimation of confidence intervals for mean lifetime. Black-Right-Pointing-Pointer The mean lifetime can be estimated directly from the TCSPC histogram.

  7. Remote UV Fluorescence Lifetime Spectrometer Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The goal of this project is to develop, demonstrate, and deliver to NASA an innovative, portable, and power efficient Remote UV Fluorescence Lifetime Spectrometer...

  8. 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

  9. 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.

  10. 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.

  11. Fluorescence lifetimes: fundamentals and interpretations.

    Science.gov (United States)

    Noomnarm, Ulai; Clegg, Robert M

    2009-01-01

    Fluorescence measurements have been an established mainstay of photosynthesis experiments for many decades. Because in the photosynthesis literature the basics of excited states and their fates are not usually described, we have presented here an easily understandable text for biology students in the style of a chapter in a text book. In this review we give an educational overview of fundamental physical principles of fluorescence, with emphasis on the temporal response of emission. Escape from the excited state of a molecule is a dynamic event, and the fluorescence emission is in direct kinetic competition with several other pathways of de-excitation. It is essentially through a kinetic competition between all the pathways of de-excitation that we gain information about the fluorescent sample on the molecular scale. A simple probability allegory is presented that illustrates the basic ideas that are important for understanding and interpreting most fluorescence experiments. We also briefly point out challenges that confront the experimenter when interpreting time-resolved fluorescence responses.

  12. 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.

  13. Digital analysis and sorting of fluorescence lifetime by flow cytometry.

    Science.gov (United States)

    Houston, Jessica P; Naivar, Mark A; Freyer, James P

    2010-09-01

    Frequency-domain flow cytometry techniques are combined with modifications to the digital signal-processing capabilities of the open reconfigurable cytometric acquisition system (ORCAS) to analyze fluorescence decay lifetimes and control sorting. Real-time fluorescence lifetime analysis is accomplished by rapidly digitizing correlated, radiofrequency (RF)-modulated detector signals, implementing Fourier analysis programming with ORCAS' digital signal processor (DSP) and converting the processed data into standard cytometric list mode data. To systematically test the capabilities of the ORCAS 50 MS/sec analog-to-digital converter (ADC) and our DSP programming, an error analysis was performed using simulated light scatter and fluorescence waveforms (0.5-25 ns simulated lifetime), pulse widths ranging from 2 to 15 micros, and modulation frequencies from 2.5 to 16.667 MHz. The standard deviations of digitally acquired lifetime values ranged from 0.112 to >2 ns, corresponding to errors in actual phase shifts from 0.0142 degrees to 1.6 degrees. The lowest coefficients of variation (digital analysis system to a previous analog phase-sensitive flow cytometer demonstrated similar precision and accuracy on measurements of a range of fluorescent microspheres, unstained cells, and cells stained with three common fluorophores. Sorting based on fluorescence lifetime was accomplished by adding analog outputs to ORCAS and interfacing with a commercial cell sorter with a RF-modulated solid-state laser. Two populations of fluorescent microspheres with overlapping fluorescence intensities but different lifetimes (2 and 7 ns) were separated to approximately 98% purity. Overall, the digital signal acquisition and processing methods we introduce present a simple yet robust approach to phase-sensitive measurements in flow cytometry. The ability to simply and inexpensively implement this system on a commercial flow sorter will allow both better dissemination of this technology and better

  14. Origin of tryptophan fluorescence lifetimes. Part 2: fluorescence lifetimes origin of tryptophan in proteins.

    Science.gov (United States)

    Albani, J R

    2014-01-01

    Fluorescence intensity decays of L-tryptophan in proteins dissolved in pH 7 buffer, in ethanol and in 6 M guanidine pH 7.8 and in lyophilized proteins were measured. In all protein conditions, three lifetimes were obtained along the emission spectrum (310-410 nm). The two shortest lifetimes are in the same range of those obtained for L-Trp in water or in ethanol. Thus, these two lifetimes originate from specific two sub-structures existing in the excited state and are inherent to the tryptophan structure independently of the surrounding environment (amino acids residues, solvent, etc.) In proteins, the third lifetime originates from the interactions that are occurring between tryptophan residues and neighboring amino acids. Populations of these lifetimes are independent of the excitation wavelength and thus originate from pre-defined sub structures existing in the excited state and put into evidence after tryptophan excitation. Fluorescence decay studies of different tripeptides having a tryptophan residue in second position show that the best analysis is obtained with two fluorescence lifetimes. Consequently, this result seems to exclude the possibility that peptide bond induces the third fluorescence lifetimes. Indole dissolved in water and/or in ethanol emits with two fluorescence lifetimes that are completely different from those observed for L-Trp. Absence of the third lifetime in ethanol demonstrates that indole behaves differently when compared to tryptophan. Thus, it seems not adequate to attribute fluorescence lifetime or fluorescence properties of tryptophan to indole ring and to compare tryptophan fluorescence properties in proteins to molecules having close structures such as NATA which fluoresces with one lifetime.

  15. Solid-State Camera System for Fluorescence Lifetime Microscopy

    NARCIS (Netherlands)

    Zhao, Q.

    2014-01-01

    Fluorescence microscopy is a well-established platform for biology and biomedical research (Chapter 2). Based on this platform, fluorescence lifetime imaging microscopy (FLIM) has been developed to measure fluorescence lifetimes, which are independent of fluorophore concentration and excitation inte

  16. 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.

  17. 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

  18. 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.

  19. Modulated CMOS camera for fluorescence lifetime microscopy.

    Science.gov (United States)

    Chen, Hongtao; Holst, Gerhard; Gratton, Enrico

    2015-12-01

    Widefield frequency-domain fluorescence lifetime imaging microscopy (FD-FLIM) is a fast and accurate method to measure the fluorescence lifetime of entire images. However, the complexity and high costs involved in construction of such a system limit the extensive use of this technique. PCO AG recently released the first luminescence lifetime imaging camera based on a high frequency modulated CMOS image sensor, QMFLIM2. Here we tested and provide operational procedures to calibrate the camera and to improve the accuracy using corrections necessary for image analysis. With its flexible input/output options, we are able to use a modulated laser diode or a 20 MHz pulsed white supercontinuum laser as the light source. The output of the camera consists of a stack of modulated images that can be analyzed by the SimFCS software using the phasor approach. The nonuniform system response across the image sensor must be calibrated at the pixel level. This pixel calibration is crucial and needed for every camera settings, e.g. modulation frequency and exposure time. A significant dependency of the modulation signal on the intensity was also observed and hence an additional calibration is needed for each pixel depending on the pixel intensity level. These corrections are important not only for the fundamental frequency, but also for the higher harmonics when using the pulsed supercontinuum laser. With these post data acquisition corrections, the PCO CMOS-FLIM camera can be used for various biomedical applications requiring a large frame and high speed acquisition.

  20. Origin of tryptophan fluorescence lifetimes part 1. Fluorescence lifetimes origin of tryptophan free in solution.

    Science.gov (United States)

    Albani, J R

    2014-01-01

    Fluorescence intensity decays of L-tryptophan free in polar, hydrophobic and mixture of polar-hydrophobic solvents were recorded along the emission spectrum (310-410 nm). Analysis of the data show that emission of tryptophan occurs with two lifetimes in 100% polar and hydrophobic environments. The values of the two lifetimes are not the same in both environments while their populations (pre-exponentials values) are identical. Fluorescence lifetimes and pre-exponentials values do not change with the excitation wavelength and thus are independent of excitation energy. Our results indicate that tryptophan emission occurs from two specific sub-structures existing in the excited state. These sub-structures differ from those present in the ground states and characterize an internal property and/or organization of the tryptophan structure in the excited state. By sub-substructure, we mean here tryptophan backbone and its electronic cloud. In ethanol, three fluorescence lifetimes were measured; two lifetimes are very close to those observed in water (0.4-0.5 ns and 2-4 ns). Presence of a third lifetime for tryptophan in ethanol results from the interaction of both hydrophobic and hydrophilic dipoles or chemical functions of ethanol with the fluorophore.

  1. 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.

  2. 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.

  3. 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

  4. 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.

  5. Flow cytometric fluorescence lifetime analysis of DNA binding fluorochromes

    Energy Technology Data Exchange (ETDEWEB)

    Crissman, Harry A.; Cui, H. H. (H. Helen); Steinkamp, J. A.

    2002-01-01

    Most flow cytometry (FCM) applications monitor fluorescence intensity to quantitate the various cellular parameters; however, the fluorescence emission also contains information relative to the fluorescence lifetime. Recent developments in FCM (Pinsky et al., 1993; Steinkamp & Crissman, 1993; Steinkamp et al., 1993), provide for the measurement of fluorescence lifetime which is also commonly referred to as fluorescence decay, or the time interval in which a fluorochrome remains in the excited state. Many unbound fluorochromes have characteristic lifetime values that are determined by their molecular structure; however, when the probe becomes bound, the lifetime value is influenced by a number of factors that affect the probe interaction with a target molecule. Monitoring the changes in the lifetime of the probe yields information relating to the molecular conformation, the functional state or activity of the molecular target. In addition, the lifetime values can be used as signatures to resolve the emissions of multiple fluorochrome labels with overlapping emission spectra that cannot be resolved by conventional FCM methodology. Such strategies can increase the number of fluorochrome combinations used in a flow cytometer with a single excitation source. Our studies demonstrate various applications of lifetime measurements for the analysis of the binding of different fluorochromes to DNA in single cells. Data presented in this session will show the utility of lifetime measurements for monitoring changes in chromatin structure associated with cell cycle progression, cellular differentiation, or DNA damage, such as induced during apoptosis. Several studies show that dyes with specificity for nucleic acids display different lifetime values when bound to DNA or to dsRNA. The Phase Sensitive Flow Cytometer is a multiparameter instrument, capable of performing lifetime measurements in conjunction with all the conventional FCM measurements. Future modifications of this

  6. 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.

  7. 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.

  8. 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.

  9. 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.

  10. Finding of Optimal Calcium Ion Probes for Fluorescence Lifetime Measurement

    Science.gov (United States)

    Yoshiki, Keisuke; Azuma, Hiroki; Yoshioka, Kazuhiko; Hashimoto, Mamoru; Araki, Tsutomu

    We have investigated the fluorescence lifetime properties of 8 calcium ion probes, calcium-green-1, calcium green-2, calcium green-5N, calcium orange, oregon green 488 BAPTA-6F, fluo-3, fluo-4, and fluo-5N. We found that the decay time of calcium green-5N varied more sensitively with calcium concentration than calcium green-1 which was known to be a highly sensitive probe. We have also found that the center of observable range of calcium concentration by fluorescence lifetime measurement is lower than that by fluorescence intensity measurement.

  11. Fluorescence-lifetime-based sensors for anions

    Science.gov (United States)

    Teichmann, Maria; Draxler, Sonja; Kieslinger, Dietmar; Lippitsch, Max E.

    1997-05-01

    Sensing of anions has been investigated using the fluorescence decaytime as the information carrier. The sensing mechanism is based on the coextraction of an anion and a proton, and the presence of a fluorophore with a rather long fluorescence decaytime inside the membrane to act as a pH indicator. The relevant theory is discussed shortly. As an example a sensor for nitrate is shown, and the influence of ionic additives on the working function has been investigated.

  12. Frequency domain fluorescence lifetime microwell-plate platform for respirometry measurements

    Science.gov (United States)

    Chatni, M. R.; Yale, G.; Van Ryckeghem, A.; Porterfield, D. M.

    2010-04-01

    Traditionally micro-well plate based platforms used in biology utilize fluorescence intensity based methods to measure processes of biological relevance. However, fluorescence intensity measurements suffer from calibration drift due to a variety of factors. Photobleaching and self-quenching of the fluorescent dyes cause the intensity signal to drop over the lifetime of sensor immobilized inside the well. Variation in turbidity of the sample during the course of the measurement affects the measured fluorescence intensity. In comparison, fluorescence lifetime measurements are not significantly affected by these factors because fluorescence lifetime is a physico-chemical property of the fluorescent dye. Reliable and inexpensive frequency domain fluorescence lifetime instrumentation platforms are possible because the greater tolerance for optical alignment, and because they can be performed using inexpensive light sources such as LEDs. In this paper we report the development of a frequency domain fluorescence lifetime well-plate platform utilizing an oxygen sensitive transition-metal ligand complex fluorophore with a lifetime in the microsecond range. The fluorescence lifetime dye is incorporated in a polymer matrix and immobilized on the base of micro-well of a 60 well micro-well plate. Respiration measurements are performed in both aqueous and non-aqueous environment. Respirometry measurements were recorded from single Daphnia magna egg in hard water. Daphnia is an aquatic organism, important in environmental toxicology as a standard bioassay and early warning indicator for water quality monitoring. Also respirometry measurements were recorded from Tribolium castaneum eggs, which are common pests in the processed flour industry. These eggs were subjected to mitochondrial electron transport chain inhibitor such as potassium cyanide (KCN) and its effects on egg respiration were measured in real-time.

  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. 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

  15. Time-domain microfluidic fluorescence lifetime flow cytometry for high-throughput Förster resonance energy transfer screening.

    Science.gov (United States)

    Nedbal, Jakub; Visitkul, Viput; Ortiz-Zapater, Elena; Weitsman, Gregory; Chana, Prabhjoat; Matthews, Daniel R; Ng, Tony; Ameer-Beg, Simon M

    2015-02-01

    Sensing ion or ligand concentrations, physico-chemical conditions, and molecular dimerization or conformation change is possible by assays involving fluorescent lifetime imaging. The inherent low throughput of imaging impedes rigorous statistical data analysis on large cell numbers. We address this limitation by developing a fluorescence lifetime-measuring flow cytometer for fast fluorescence lifetime quantification in living or fixed cell populations. The instrument combines a time-correlated single photon counting epifluorescent microscope with microfluidics cell-handling system. The associated computer software performs burst integrated fluorescence lifetime analysis to assign fluorescence lifetime, intensity, and burst duration to each passing cell. The maximum safe throughput of the instrument reaches 3,000 particles per minute. Living cells expressing spectroscopic rulers of varying peptide lengths were distinguishable by Förster resonant energy transfer measured by donor fluorescence lifetime. An epidermal growth factor (EGF)-stimulation assay demonstrated the technique's capacity to selectively quantify EGF receptor phosphorylation in cells, which was impossible by measuring sensitized emission on a standard flow cytometer. Dual-color fluorescence lifetime detection and cell-specific chemical environment sensing were exemplified using di-4-ANEPPDHQ, a lipophilic environmentally sensitive dye that exhibits changes in its fluorescence lifetime as a function of membrane lipid order. To our knowledge, this instrument opens new applications in flow cytometry which were unavailable due to technological limitations of previously reported fluorescent lifetime flow cytometers. The presented technique is sensitive to lifetimes of most popular fluorophores in the 0.5-5 ns range including fluorescent proteins and is capable of detecting multi-exponential fluorescence lifetime decays. This instrument vastly enhances the throughput of experiments involving fluorescence

  16. 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).

  17. 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.

  18. Measuring and sorting cell populations expressing isospectral fluorescent proteins with different fluorescence lifetimes.

    Directory of Open Access Journals (Sweden)

    Bryan Sands

    Full Text Available Study of signal transduction in live cells benefits from the ability to visualize and quantify light emitted by fluorescent proteins (XFPs fused to different signaling proteins. However, because cell signaling proteins are often present in small numbers, and because the XFPs themselves are poor fluorophores, the amount of emitted light, and the observable signal in these studies, is often small. An XFP's fluorescence lifetime contains additional information about the immediate environment of the fluorophore that can augment the information from its weak light signal. Here, we constructed and expressed in Saccharomyces cerevisiae variants of Teal Fluorescent Protein (TFP and Citrine that were isospectral but had shorter fluorescence lifetimes, ∼ 1.5 ns vs ∼ 3 ns. We modified microscopic and flow cytometric instruments to measure fluorescence lifetimes in live cells. We developed digital hardware and a measure of lifetime called a "pseudophasor" that we could compute quickly enough to permit sorting by lifetime in flow. We used these abilities to sort mixtures of cells expressing TFP and the short-lifetime TFP variant into subpopulations that were respectively 97% and 94% pure. This work demonstrates the feasibility of using information about fluorescence lifetime to help quantify cell signaling in living cells at the high throughput provided by flow cytometry. Moreover, it demonstrates the feasibility of isolating and recovering subpopulations of cells with different XFP lifetimes for subsequent experimentation.

  19. Fluorescence lifetime measurements of boronate derivatives to determine glucose concentration

    Energy Technology Data Exchange (ETDEWEB)

    Gable, J H

    2000-06-01

    A novel investigation into the fluorescence lifetimes of molecules, both established and newly designed, was performed. These molecules are the basis of a continuous, minimally invasive, glucose sensor based on fluorescence lifetime measurements. This sensor, if coupled with an automated insulin delivery device, would effectively create an artificial pancreas allowing for the constant monitoring and control of glucose levels in a person with diabetes. The proposed sensor includes a fluorescent molecule that changes its' fluorescence properties upon binding selectively and reversibly to glucose. One possible sensor molecule is N-methyl-N-(9-methylene anthryl)-2-methylenephenylboronic acid (AB). The fluorescence intensity of AB was shown to change in response to changing glucose concentrations. (James, 1994) James proposed that when glucose binds to AB the fluorescence intensity increases due to an enhancement of the N{yields}B dative bond which prevents photoinduced electron transfer (PET). PET from the amine (N) to the fluorophore (anthracene) quenches the fluorescence. The dative bond between the boron and the amine can prevent PET by involving the lone pair of electrons on the amine in interactions with the boron rather than allowing them to be transferred to the fluorophore. Results of this research show the average fluorescence lifetime of AB also changes with glucose concentration. It is proposed that fluorescence is due to two components: (1) AB with an enhanced N{yields}B interaction, and no PET, and (2) AB with a weak N{yields}B interaction, resulting in fluorescence quenching by PET. Lifetime measurements of AB as a function of both the pH of the solvent and glucose concentration in the solution were made to characterize this two component system and investigate the nature of the N{yields}B bond. Measurements of molecules similar to AB were also performed in order to isolate behavior of specific AB constituents. These molecules are 9

  20. 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

  1. 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.

  2. Developing and Testing a Bayesian Analysis of Fluorescence Lifetime Measurements

    Science.gov (United States)

    Needleman, Daniel J.

    2017-01-01

    FRET measurements can provide dynamic spatial information on length scales smaller than the diffraction limit of light. Several methods exist to measure FRET between fluorophores, including Fluorescence Lifetime Imaging Microscopy (FLIM), which relies on the reduction of fluorescence lifetime when a fluorophore is undergoing FRET. FLIM measurements take the form of histograms of photon arrival times, containing contributions from a mixed population of fluorophores both undergoing and not undergoing FRET, with the measured distribution being a mixture of exponentials of different lifetimes. Here, we present an analysis method based on Bayesian inference that rigorously takes into account several experimental complications. We test the precision and accuracy of our analysis on controlled experimental data and verify that we can faithfully extract model parameters, both in the low-photon and low-fraction regimes. PMID:28060890

  3. 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.

  4. Fluorescence lifetime imaging using a single photon avalanche diode array sensor (Conference Presentation)

    Science.gov (United States)

    Wargocki, Piotr M.; Spence, David J.; Goldys, Ewa M.; Charbon, Edoardo; Bruschini, Claudio E.; Antalović, Ivan Michel; Burri, Samuel

    2017-02-01

    Single photon detectors allows us work with the weakest fluorescence signals. Single photon arrays, combined with ps-controlled gating allow us to create image maps of fluorescence lifetimes, which can be used for in-vivo discrimination of tissue activity. Here we present fluorescence lifetime imaging using the `SwissSPAD' sensor, a 512-by-128-pixel array of gated single photon detectors, fabricated in a standard high-voltage 0.35 μm CMOS process. We present a protocol for spatially resolved lifetime measurements where the lifetime can be retrieved for each pixel. We demonstrate the system by imaging patterns of Fluorescein and Rhodamine B on test slides, as well as measuring mixed samples to retrieve both components of the decay lifetime. The single photon sensitivity of the sensor creates a valuable instrument to perform live cell or live animal (in vivo) measurements of the weak autofluorescent signals, for example distinguishing unlabelled free and bound NADH. Our ultimate goal is to create a real time fluorescence lifetime imaging system, possibly integrated into augmented reality goggles, which could allow immediate discrimination of in vivo tissues.

  5. 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

  6. Fluorescence-lifetime-based sensors using inhomogeneous waveguiding

    Science.gov (United States)

    Draxler, Sonja; Kieslinger, Dietmar; Trznadel, Karolina; Lippitsch, Max E.

    1996-12-01

    Most intrinsic fiberoptic sensors are based on the evanescent-wave scheme, where the evanescent field of modes guided in a fiber reaches out into a chemically sensitive coating. In the commonly used multimode waveguides, the evanescent field contains only a small part of the total energy, however, thus making evanescent-wave sensors rather insensitive. Combining a transparent substrate and a transparent sensing layer of rather similar refractive index into a common waveguiding structure produces an inhomogeneous waveguide, where a large portion of the total energy transverses the sensing layer. This yields much superior sensor performance. The transmission through a waveguide is subject to various disturbing influences. Thus it is advantageous to combine the inhomogeneous waveguiding approach with a measuring scheme that is not prone to those disturbances. Such a scheme is available with fluorescence lifetime-based sensors. The fluorescence lifetime of an indicator incorporated into the sensing layer is changed by the presence of the respective analyte. This lifetime is independent of the transmission through the waveguide. Thus inhomogeneous waveguiding together with fluorescence lifetime measurement paves the way for optical chemical sensors with high analyte sensitivity and immunity to external disturbances.

  7. Investigations on exponential lifetime measurements for fluorescence thermometry

    Science.gov (United States)

    Fernicola, V. C.; Rosso, L.; Galleano, R.; Sun, T.; Zhang, Z. Y.; Grattan, K. T. V.

    2000-07-01

    Lifetime-based methods have been, on the whole, one of the most successful schemes for fiber optic temperature sensing, using fluorescent materials whose response is intensity independent. Several approaches for determining the fluorescence lifetime, and with that the measurand, have been investigated. An experimental comparison of direct and indirect measurement methods, i.e., involving actual signals from representative optical media instead of simply using Monte Carlo simulations, has been carried out. Direct fitting methods, including Marquardt, log-fit and Prony, were used to estimate the fluorescence lifetime of a Cr3+:YAG-based sensor system and the results were compared. An agreement to better than 0.5% between Marquardt and log-fit algorithms and an agreement of about 1.5% between Marquardt and Prony approaches was found. Thus, a temperature reproducibility, of 0.5 and 1.2 °C, respectively, can be obtained with the Cr3+:YAG sensor system. An indirect measurement approach based on a phase-locked (analog-to-digital signal processor) (A-DSP) was also tested. It was found that when the A-DSP output is used to estimate the lifetime, it performs only slightly better than using direct fitting methods. On the contrary, when the whole A-DSP sensor system was directly calibrated against temperature, the measurement accuracy improves by at least a factor of 10.

  8. UV fluorescence lifetime modification by aluminum and magnesium nanoapertures

    Science.gov (United States)

    Wang, Yunshan; Jiao, Xiaojin; Peterson, Eric M.; Harris, Joel M.; Appusamy, Kanagasundar; Guruswamy, Sivaraman; Blair, Steve

    2016-09-01

    Ultra-violet (UV) fluorescence lifetime modification by aluminum (Al) and magnesium (Mg) nanoapertures are reported in this manuscript. Nanoapertures with diameter ranging from 30nm to 90nm are fabricated using focused ion beam (FIB). Largest lifetime reduction are observed for apertures with smallest diameters and undercuts into glass substrate. For Al nanoapertures, largest lifetime reduction is 5.30×, larger than perviously reported 3.50×.1 For Mg nanoapertures, largest lifetime reduction is 6.90×, which is the largest lifetime reduction of UV fluorescence dye reported so far in literature. The dependence of count rate per molecule (CRM) on aperture size and undercut is also investigated, revealing that CRM increases with increasing undercut, however, the CRM is small (less than 2) for the entire range of aperture size and undercut we investigated. FDTD simulation were conducted and in order to favorably compare experimental results with simulated results, it is critical to take into account the exact shape and material properties of the nano aperture. Simulation results revealed the fundamental difference between Al and Mg nano aperture under 266nm illumination-Mg nano aperture presents a waveguide mode in which the maximum field enhancement and Purcell factor is within the nano aperture instead of on the surface which is the case for Al nano aperture.

  9. Photochromicity and fluorescence lifetimes of green fluorescent protein

    OpenAIRE

    1999-01-01

    The green fluorescent protein (GFP) of the bioluminescent jellyfish Aequorea and its mutants have gained widespread usage as an indicator of structure and function within cells. Proton transfer has been implicated in the complex photophysics of the wild-type molecule, exhibiting a protonated A species excited at 400 nm, and two deprotonated excited-state species I* and B* with red-shifted excitation similar to 475 nm. Photochromicity between the protonated and deprotonated species has been re...

  10. Family of fluorescence lifetime sensors for environmental purposes

    Science.gov (United States)

    Draxler, Sonja; Lippitsch, Max E.

    1995-09-01

    A family of indicators has been developed for measuring different analytes, all the indicators being derivatives of the same chemical compound and having identical spectral and lifetime properties. The indicators show an absorption accessible to low-cost light sources, a large Stokes shift, and a long fluorescence decay time. All indicators can be excited at the same excitation wavelength, monitored at the same emission wavelength, and measured within the same time range. This opens the possibility for a compact lifetime-based instrument for water monitoring.

  11. Fluorescent Protein Based FRET Pairs with Improved Dynamic Range for Fluorescence Lifetime Measurements.

    Directory of Open Access Journals (Sweden)

    Bobin George Abraham

    Full Text Available Fluorescence Resonance Energy Transfer (FRET using fluorescent protein variants is widely used to study biochemical processes in living cells. FRET detection by fluorescence lifetime measurements is the most direct and robust method to measure FRET. The traditional cyan-yellow fluorescent protein based FRET pairs are getting replaced by green-red fluorescent protein variants. The green-red pair enables excitation at a longer wavelength which reduces cellular autofluorescence and phototoxicity while monitoring FRET. Despite the advances in FRET based sensors, the low FRET efficiency and dynamic range still complicates their use in cell biology and high throughput screening. In this paper, we utilized the higher lifetime of NowGFP and screened red fluorescent protein variants to develop FRET pairs with high dynamic range and FRET efficiency. The FRET variations were analyzed by proteolytic activity and detected by steady-state and time-resolved measurements. Based on the results, NowGFP-tdTomato and NowGFP-mRuby2 have shown high potentials as FRET pairs with large fluorescence lifetime dynamic range. The in vitro measurements revealed that the NowGFP-tdTomato has the highest Förster radius for any fluorescent protein based FRET pairs yet used in biological studies. The developed FRET pairs will be useful for designing FRET based sensors and studies employing Fluorescence Lifetime Imaging Microscopy (FLIM.

  12. Fluorescence of ceramic color standards.

    Science.gov (United States)

    Koo, Annette; Clare, John F; Nield, Kathryn M; Deadman, Andrew; Usadi, Eric

    2010-04-20

    Fluorescence has been found in color standards available for use in calibration and verification of color measuring instruments. The fluorescence is excited at wavelengths below about 600?nm and emitted above 700?nm, within the response range of silicon photodiodes, but at the edge of the response of most photomultipliers and outside the range commonly scanned in commercial colorimeters. The degree of fluorescence on two of a set of 12 glossy ceramic tiles is enough to introduce significant error when those tiles have been calibrated in one mode of measurement and are used in another. We report the nature of the fluorescence and the implications for color measurement.

  13. 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.

  14. 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

  15. Evaluation of actinic cheilitis using fluorescence lifetime spectroscopy

    Science.gov (United States)

    Saito Nogueira, Marcelo; Cosci, Alessandro; Pratavieira, Sebastião.; Takahama, Ademar; Souza Azevedo, Rebeca; Kurachi, Cristina

    2016-03-01

    Actinic cheilitis is a potentially malignant disorder that mostly affects the vermilion border of the lower lip and can lead to squamous cell carcinoma. Because of its heterogeneous clinical aspect, it is difficult to indicate representative biopsy area. Late diagnosis is a limiting factor of therapeutic possibilities available to treat oral cancer. The diagnosis of actinic cheilitis is mainly based on clinical and histopathological analysis and it is a time consuming procedure to get the results. Information about the organization and chemical composition of the tissues can be obtained using fluorescence lifetime spectroscopy techniques without the need for biopsy. The main targeted fluorophores are NADH (nicotinamide adenine dinucleotide) and FAD (flavin adenine dinucleotide), which have free and bound states, each one with different average lifetimes. The average lifetimes for free and bound NADH and FAD change according to tissue metabolic alterations and allow a quick and non-invasive clinical investigation of injuries and to help clinicians with the early diagnosis of actinic cheilitis. This study aims to evaluate the fluorescence lifetime parameters at the discrimination of three degrees of epithelial dysplasia, the most important predictor of malignant development, described in up to 100% of actinic cheilitis cases.

  16. Effect of refractive index on the fluorescence lifetime of green fluorescent protein.

    Science.gov (United States)

    Tregidgo, Carolyn; Levitt, James A; Suhling, Klaus

    2008-01-01

    The average fluorescence lifetime of the green fluorescent protein (GFP) in solution is a function of the refractive index of its environment. We report that this is also the case for GFP-tagged proteins in cells. Using time-correlated single-photon counting (TCSPC)-based fluorescence lifetime imaging (FLIM) with a confocal scanning microscope, images of GFP-tagged proteins in cells suspended in different refractive index media are obtained. It is found that the average fluorescence lifetime of GFP decreases on addition of glycerol or sucrose to the media in which the fixed cells are suspended. The inverse GFP lifetime is proportional to the refractive index squared. This is the case for GFP-tagged major histocompatibility complex (MHC) proteins with the GFP located inside the cytoplasm, and also for GPI-anchored GFP that is located outside the cell membrane. The implications of these findings are discussed with regard to total internal reflection fluorescence (TIRF) techniques where the change in refractive index is crucial in producing an evanescent wave to excite fluorophores near a glass interface. Our findings show that the GFP fluorescence lifetime is shortened in TIRF microscopy in comparison to confocal microscopy.

  17. pH Dependence of the Fluorescence Lifetime of FAD in Solution and in Cells

    OpenAIRE

    Nobuhiro Ohta; Takakazu Nakabayashi; Masataka Kinjo; Md. Serajul Islam; Masato Honma

    2013-01-01

    We have studied physiological parameters in a living cell using fluorescence lifetime imaging of endogenous chromophores. In this study, pH dependence of the fluorescence lifetime of flavin adenine dinucleotide (FAD), that is a significant cofactor exhibiting autofluorescence, has been investigated in buffer solution and in cells. The fluorescence lifetime of FAD remained unchanged with pH 5 to 9 in solution. However, the fluorescence lifetime in HeLa cells was found to decrease with increasi...

  18. pH dependence of the fluorescence lifetime of FAD in solution and in cells.

    Science.gov (United States)

    Islam, Md Serajul; Honma, Masato; Nakabayashi, Takakazu; Kinjo, Masataka; Ohta, Nobuhiro

    2013-01-18

    We have studied physiological parameters in a living cell using fluorescence lifetime imaging of endogenous chromophores. In this study, pH dependence of the fluorescence lifetime of flavin adenine dinucleotide (FAD), that is a significant cofactor exhibiting autofluorescence, has been investigated in buffer solution and in cells. The fluorescence lifetime of FAD remained unchanged with pH 5 to 9 in solution. However, the fluorescence lifetime in HeLa cells was found to decrease with increasing intracellular pH, suggesting that pH in a single cell can be estimated from the fluorescence lifetime imaging of FAD without adding exogenous fluorescent probes.

  19. A hyperspectral fluorescence lifetime probe for skin cancer diagnosis

    Science.gov (United States)

    De Beule, P. A. A.; Dunsby, C.; Galletly, N. P.; Stamp, G. W.; Chu, A. C.; Anand, U.; Anand, P.; Benham, C. D.; Naylor, A.; French, P. M. W.

    2007-12-01

    The autofluorescence of biological tissue can be exploited for the detection and diagnosis of disease but, to date, its complex nature and relatively weak signal levels have impeded its widespread application in biology and medicine. We present here a portable instrument designed for the in situ simultaneous measurement of autofluorescence emission spectra and temporal decay profiles, permitting the analysis of complex fluorescence signals. This hyperspectral fluorescence lifetime probe utilizes two ultrafast lasers operating at 355 and 440nm that can excite autofluorescence from many different biomolecules present in skin tissue including keratin, collagen, nicotinamide adenine dinucleotide (phosphate), and flavins. The instrument incorporates an optical fiber probe to provide sample illumination and fluorescence collection over a millimeter-sized area. We present a description of the system, including spectral and temporal characterizations, and report the preliminary application of this instrument to a study of recently resected (skin lesions, illustrating its potential for skin cancer detection and diagnosis.

  20. 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.

  1. Bessel beam fluorescence lifetime tomography of live embryos (Conference Presentation)

    Science.gov (United States)

    Xu, Dongli; Peng, Leilei

    2016-03-01

    Optical tomography allows isotropic 3D imaging of embryos. Scanning-laser optical tomography (SLOT) has superior light collecting efficiency than wide-field optical tomography, making it ideal for fluorescence imaging of live embryos. We previously reported an imaging system that combines SLOT with a novel Fourier-multiplexed fluorescence lifetime imaging (FmFLIM) technique named FmFLIM-SLOT. FmFLIM-SLOT performs multiplexed FLIM-FRET readout of multiple FRET sensors in live embryos. Here we report a recent effort on improving the spatial resolution of the FmFLIM-SLOT system in order to image complex biochemical processes in live embryos at the cellular level. Optical tomography has to compromise between resolution and the depth of view. In SLOT, the commonly-used focused Gaussian beam diverges quickly from the focal plane, making it impossible to achieve high resolution imaging in a large volume specimen. We thus introduce Bessel beam laser-scanning tomography, which illuminates the sample with a spatial-light-modulator-generated Bessel beam that has an extended focal depth. The Bessel beam is scanned across the whole specimen. Fluorescence projection images are acquired at equal angular intervals as the sample rotates. Reconstruction artifacts due to annular-rings of the Bessel beam are removed by a modified 3D filtered back projection algorithm. Furthermore, in combination of Fourier-multiplexing fluorescence lifetime imaging (FmFLIM) method, the Bessel FmFLIM-SLOT system is capable of perform 3D lifetime imaging of live embryos at cellular resolution. The system is applied to in-vivo imaging of transgenic Zebrafish embryos. Results prove that Bessel FmFLIM-SLOT is a promising imaging method in development biology research.

  2. Shifts in the fluorescence lifetime of EGFP during bacterial phagocytosis measured by phase-sensitive flow cytometry

    Science.gov (United States)

    Li, Wenyan; Houston, Kevin D.; Houston, Jessica P.

    2017-01-01

    Phase-sensitive flow cytometry (PSFC) is a technique in which fluorescence excited state decay times are measured as fluorescently labeled cells rapidly transit a finely focused, frequency-modulated laser beam. With PSFC the fluorescence lifetime is taken as a cytometric parameter to differentiate intracellular events that are challenging to distinguish with standard flow cytometry. For example PSFC can report changes in protein conformation, expression, interactions, and movement, as well as differences in intracellular microenvironments. This contribution focuses on the latter case by taking PSFC measurements of macrophage cells when inoculated with enhanced green fluorescent protein (EGFP)-expressing E. coli. During progressive internalization of EGFP-E. coli, fluorescence lifetimes were acquired and compared to control groups. It was hypothesized that fluorescence lifetimes would correlate well with phagocytosis because phagosomes become acidified and the average fluorescence lifetime of EGFP is known to be affected by pH. We confirmed that average EGFP lifetimes consistently decreased (3 to 2 ns) with inoculation time. The broad significance of this work is the demonstration of how high-throughput fluorescence lifetime measurements correlate well to changes that are not easily tracked by intensity-only cytometry, which is affected by heterogeneous protein expression, cell-to-cell differences in phagosome formation, and number of bacterium engulfed.

  3. 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.

  4. Fluorescence lifetime multiplexing with nanocrystals and organic labels.

    Science.gov (United States)

    Grabolle, Markus; Kapusta, Peter; Nann, Thomas; Shu, Xu; Ziegler, Jan; Resch-Genger, Ute

    2009-09-15

    The potential of semiconducting nanocrystals or so-called quantum dots (QDs) for lifetime multiplexing has not been investigated yet, despite the increasing use of QDs in (bio)analytical detection, biosensing, and fluorescence imaging and the obvious need for simple and cost-effective tools and strategies for the simultaneous detection of multiple analytes or events. This is most likely related to their multiexponential decay behavior as for multiplex chromophores, typically monoexponential decay kinetics are requested. The fluorescence decay kinetics of various mixtures of a long-lived, multiexponentially decaying CdSe QD and a short-lived organic dye were analyzed, and a model was developed for the quantification of these labels from the measured complex decay kinetics as a first proof-of-concept for the huge potential of these labels for lifetime multiplexing. In a second step, we evaluated the potential of mixtures of two types of QDs, varying in constituent material to realize distinguishable, yet multiexponential decay kinetics and similar absorption and emission spectra. Strategies for lifetime multiplexing with nanocrystalline labels were derived on the basis of these measurements.

  5. Intracellular distribution of fluorescent copper and zinc bis(thiosemicarbazonato) complexes measured with fluorescence lifetime spectroscopy.

    Science.gov (United States)

    Hickey, James L; James, Janine L; Henderson, Clare A; Price, Katherine A; Mot, Alexandra I; Buncic, Gojko; Crouch, Peter J; White, Jonathan M; White, Anthony R; Smith, Trevor A; Donnelly, Paul S

    2015-10-05

    The intracellular distribution of fluorescently labeled copper and zinc bis(thiosemicarbazonato) complexes was investigated in M17 neuroblastoma cells and primary cortical neurons with a view to providing insights into the neuroprotective activity of a copper bis(thiosemicarbazonato) complex known as Cu(II)(atsm). Time-resolved fluorescence measurements allowed the identification of the Cu(II) and Zn(II) complexes as well as the free ligand inside the cells by virtue of the distinct fluorescence lifetime of each species. Confocal fluorescent microscopy of cells treated with the fluorescent copper(II)bis(thiosemicarbazonato) complex revealed significant fluorescence associated with cytoplasmic puncta that were identified to be lysosomes in primary cortical neurons and both lipid droplets and lysosomes in M17 neuroblastoma cells. Fluorescence lifetime imaging microscopy confirmed that the fluorescence signal emanating from the lipid droplets could be attributed to the copper(II) complex but also that some degree of loss of the metal ion led to diffuse cytosolic fluorescence that could be attributed to the metal-free ligand. The accumulation of the copper(II) complex in lipid droplets could be relevant to the neuroprotective activity of Cu(II)(atsm) in models of amyotrophic lateral sclerosis and Parkinson's disease.

  6. 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.

  7. Bloodstain age analysis: toward solid state fluorescent lifetime measurements

    Science.gov (United States)

    Guo, Kevin; Zhegalova, Natalia; Achilefu, Samuel; Berezin, Mikhail Y.

    2013-03-01

    One of the most pressing unsolved challenges in forensic science is the determination of time since deposition (TSD) of bloodstains at crime scenes. Despite a number of high profile cases over the past couple hundred years involving controversy over TSD methods, no reliable quantitative method has been established. We present here an approach that has yet to be explored by forensic scientist: measuring the fluorescence lifetime of solid-state blood. Such a method would allow for on-site measurements of bloodstains utilizing the appropriate device, and would allow for rapid results returned in real-time to investigators.

  8. 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.

  9. Effect of surface modification on semiconductor nanocrystal fluorescence lifetime.

    Science.gov (United States)

    Ruedas-Rama, Maria J; Orte, Angel; Hall, Elizabeth A H; Alvarez-Pez, Jose M; Talavera, Eva M

    2011-04-04

    Semiconductor nanocrystals, namely, quantum dots (QDs), present a set of unique photoluminescence properties, which has led to increased interest in using them as advantageous alternatives to conventional organic dyes. Many applications of QDs involve surface modification to enhance the solubility or biocompatibility of the QDs. One of the least exploited properties of QDs is the very long photoluminescence lifetime that usually has complex kinetics owing to the effect of quantum confinement. Herein, we describe the effect of different surface modifications on the photoluminescence decay kinetics of QDs. The different surface modifications were carefully chosen to provide lipophilic or water-soluble QDs with either positive or negative surface net charges. We also survey the effect on the QD lifetime of several ligands that interact with the QD surface, such as organic chromophores or fluorescent proteins. The results obtained demonstrate that time-resolved fluorescence is a useful tool for QD-based sensing to set the basis for the development of time-resolved-based nanosensors.

  10. 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.

  11. One- and two-photon excited fluorescence lifetimes and anisotropy decays of green fluorescent proteins.

    OpenAIRE

    2000-01-01

    We have used one- (OPE) and two-photon (TPE) excitation with time-correlated single-photon counting techniques to determine time-resolved fluorescence intensity and anisotropy decays of the wild-type Green Fluorescent Protein (GFP) and two red-shifted mutants, S65T-GFP and RSGFP. WT-GFP and S65T-GFP exhibited a predominant approximately 3 ns monoexponential fluorescence decay, whereas for RSGFP the main lifetimes were approximately 1.1 ns (main component) and approximately 3.3 ns. The anisotr...

  12. Using multiphoton fluorescence lifetime imaging to characterize liver damage and fluorescein disposition in liver in vivo

    Science.gov (United States)

    Thorling, Camilla A.; Studier, Hauke; Crawford, Darrell; Roberts, Michael S.

    2016-03-01

    Liver disease is the fifth most common cause of death and unlike many other major causes of mortality, liver disease rates are increasing rather than decreasing. There is no ideal measurement of liver disease and although biopsies are the gold standard, this only allows for a spot examination and cannot follow dynamic processes of the liver. Intravital imaging has the potential to extract detailed information over a larger sampling area continuously. The aim of this project was to investigate whether multiphoton and fluorescence lifetime imaging microscopy could detect early liver damage and to assess whether it could detect changes in metabolism of fluorescein in normal and diseased livers. Four experimental groups were used in this study: 1) control; 2) ischemia reperfusion injury; 3) steatosis and 4) steatosis with ischemia reperfusion injury. Results showed that multiphoton microscopy could visualize morphological changes such as decreased fluorescence of endogenous fluorophores and the presence of lipid droplets, characteristic of steatosis. Fluorescence lifetime imaging microscopy showed increase in NADPH in steatosis with and without ischemia reperfusion injury and could detect changes in metabolism of fluorescein to fluorescein monoglurcuronide, which was impaired in steatosis with ischemia reperfusion injury. These results concluded that the combination of multiphoton microscopy and fluorescence lifetime imaging is a promising method of assessing early stage liver damage and that it can be used to study changes in drug metabolism in the liver as an indication of liver disease and has the potential to replace the traditional static liver biopsy currently used.

  13. Multiphoton fluorescence spectra and lifetimes of biliverdins and their protein-associated complex

    Science.gov (United States)

    Huang, Chin-Jie; Wu, Cheng-Ham; Liu, Tzu-Ming

    2012-03-01

    To investigate whether endogenous biliverdins can serve as a fluorescence metabolic marker in cancer diagnosis, we measured their multiphoton fluorescence spectra and lifetimes with femtosecond Cr:forsterite laser. Excited at 1230nm, the two-photon fluorescence of biliverdins peaks around 670nm. The corresponding lifetime (catabolism in human cells or tissues.

  14. 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

  15. 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...

  16. Differentiating quiescent cancer cell populations in heterogeneous samples with fluorescence lifetime imaging

    Science.gov (United States)

    Heaster, Tiffany M.; Walsh, Alex J.; Skala, Melissa C.

    2016-03-01

    Measurement of relative fluorescence intensities of NAD(P)H and FAD with fluorescence lifetime imaging (FLIM) allows metabolic characterization of cancerous populations and correlation to treatment response. However, quiescent populations of cancer cells introduce heterogeneity to the tumor and exhibit resistance to standard therapies, requiring a better understanding of this influence on treatment outcome. Significant differences were observed between proliferating and quiescent cell populations upon comparison of respective redox ratios (pFAD lifetimes (p<0.05) across monolayers and in mixed samples. These results demonstrate that metabolic activity may function as a marker for separation and characterization of proliferating and quiescent cancer cells within mixed samples, contributing to comprehensive investigation of heterogeneity-dependent drug resistance.

  17. 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.

  18. 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.

  19. Fluorescence lifetime spectroscopy for breast cancer margins assessment

    Science.gov (United States)

    Gorpas, Dimitris; Fatakdawala, Hussain; Zhang, Yanhong; Bold, Richard; Marcu, Laura

    2015-03-01

    During breast conserving surgery (BCS), which is the preferred approach to treat most early stage breast cancers, the surgeon attempts to excise the tumor volume, surrounded by thin margin of normal tissue. The intra-operative assessment of cancerous areas is a challenging procedure, with the surgeon usually relying on visual or tactile guidance. This study evaluates whether time-resolved fluorescence spectroscopy (TRFS) presents the potential to address this problem. Point TRFS measurements were obtained from 19 fresh tissue slices (7 patients) and parameters that characterize the transient signals were quantified via constrained least squares deconvolution scheme. Fibrotic tissue (FT, n=69), adipose tissue (AT, n=76), and invasive ductal carcinoma (IDC, n=27) were identified in histology and univariate statistical analysis, followed by multi-comparison test, was applied to the corresponding lifetime data. Significant differentiation between the three tissue types exists at 390 nm and 500 nm bands. The average lifetime is 3.23+/-0.74 ns for AT, 4.21+/-0.83 ns for FT and 4.71+/-0.35 ns (ptissue in real-time and assess tumor margins.

  20. Photon efficiency optimization in time-correlated single photon counting technique for fluorescence lifetime imaging systems.

    Science.gov (United States)

    Turgeman, Lior; Fixler, Dror

    2013-06-01

    In time-correlated single photon counting (TCSPC) systems, the maximum signal throughput is limited by the occurrence of pile-up and other effects. In many biological applications that exhibit high levels of fluorescence intensity (FI), pile-up-related distortions yield serious distortions in the fluorescence lifetime (FLT) calculation as well as significant decrease in the signal-to-noise ratio (SNR). Recent developments that allow the use of high-repetition-rate light sources (in the range of 50-100 MHz) in fluorescence lifetime imaging (FLIM) experiments enable minimization of pile-up-related distortions. However, modern TCSPC configurations that use high-repetition-rate excitation sources for FLIM suffer from dead-time-related distortions that cause unpredictable distortions of the FI signal. In this study, the loss of SNR is described by F- value as it is typically done in FLIM systems. This F-value describes the relation of the relative standard deviation in the estimated FLT to the relative standard deviation in FI measurements. Optimization of the F-value allows minimization of signal distortion, as well as shortening of the acquisition time for certain samples. We applied this method for Fluorescein, Rhodamine B, and Erythrosine fluorescent solutions that have different FLT values (4 ns, 1.67 ns, and 140 ps, respectively).

  1. 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.

  2. Note: Rapid measurement of fluorescence lifetimes using SiPM detection and waveform sampling

    Science.gov (United States)

    Tsai, H.-M.; Souris, J. S.; Kim, H.-J.; Cheng, S.-H.; Chen, L.; Lo, L.-W.; Chen, C.-T.; Kao, C.-M.

    2017-09-01

    In fluorescence spectroscopy and imaging, fluorescence lifetime measurement—assessing the average time fluorophores spend in their excited state before returning to their ground state—offers a number of advantages over quantifying fluorescence intensities that include resistance to photo-bleaching and independence from fluorophore concentration, excitation intensity, and measurement methodology. Despite growing interest, fluorescence lifetime techniques frequently mandate relatively complex instrumentation, slow data acquisition rates, and significant data analyses. In this work, we demonstrate the feasibility of measuring fluorescence lifetimes using off-the-shelf analog silicon photomultipliers and switched-capacitor array waveform sampling techniques, with precision matching that of much larger and more elaborate commercial instruments.

  3. 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

  4. 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

  5. Early-photon guided reconstruction method for time-domain fluorescence lifetime tomography

    Institute of Scientific and Technical Information of China (English)

    Lin Zhang; Chuangjian Cai; Yanlu Lv; Jianwen Luo

    2016-01-01

    A reconstruction method guided by early-photon fluorescence yield tomography is proposed for time-domain fluorescence lifetime tomography (FLT) in this study.The method employs the early-arriving photons to reconstruct a fluorescence yield map,which is utilized as a priori information to reconstruct the FLT via all the photons along the temporal-point spread functions.Phantom experiments demonstrate that,compared with the method using all the photons for reconstruction of fluorescence yield and lifetime maps,the proposed method can achieve higher spatial resolution and reduced crosstalk between different targets without sacrificing the quantification accuracy of lifetime and contrast between heterogeneous targets.

  6. 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.

  7. Spectrally resolved fluorescence lifetime imaging of Nile red for measurements of intracellular polarity

    Science.gov (United States)

    Levitt, James A.; Chung, Pei-Hua; Suhling, Klaus

    2015-09-01

    Spectrally resolved confocal microscopy and fluorescence lifetime imaging have been used to measure the polarity of lipid-rich regions in living HeLa cells stained with Nile red. The emission peak from the solvatochromic dye in lipid droplets is at a shorter wavelength than other, more polar, stained internal membranes, and this is indicative of a low polarity environment. We estimate that the dielectric constant, ɛ, is around 5 in lipid droplets and 25<ɛ<40 in other lipid-rich regions. Our spectrally resolved fluorescence lifetime imaging microscopy (FLIM) data show that intracellular Nile red exhibits complex, multiexponential fluorescence decays due to emission from a short lifetime locally excited state and a longer lifetime intramolecular charge transfer state. We measure an increase in the average fluorescence lifetime of the dye with increasing emission wavelength, as shown using phasor plots of the FLIM data. We also show using these phasor plots that the shortest lifetime decay components arise from lipid droplets. Thus, fluorescence lifetime is a viable contrast parameter for distinguishing lipid droplets from other stained lipid-rich regions. Finally, we discuss the FLIM of Nile red as a method for simultaneously mapping both polarity and relative viscosity based on fluorescence lifetime measurements.

  8. 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.

  9. A Single-Photon Avalanche Diode Array for Fluorescence Lifetime Imaging Microscopy.

    Science.gov (United States)

    Schwartz, David Eric; Charbon, Edoardo; Shepard, Kenneth L

    2008-11-21

    We describe the design, characterization, and demonstration of a fully integrated single-photon avalanche diode (SPAD) imager for use in time-resolved fluorescence imaging. The imager consists of a 64-by-64 array of active SPAD pixels and an on-chip time-to-digital converter (TDC) based on a delay-locked loop (DLL) and calibrated interpolators. The imager can perform both standard time-correlated single-photon counting (TCSPC) and an alternative gated-window detection useful for avoiding pulse pile-up when measuring bright signal levels. To illustrate the use of the imager, we present measurements of the decay lifetimes of fluorescent dyes of several types with a timing resolution of 350 ps.

  10. Recommendations for fluorescence instrument qualification: the new ASTM Standard Guide.

    Science.gov (United States)

    DeRose, Paul C; Resch-Genger, Ute

    2010-03-01

    Aimed at improving quality assurance and quantitation for modern fluorescence techniques, ASTM International (ASTM) is about to release a Standard Guide for Fluorescence, reviewed here. The guide's main focus is on steady state fluorometry, for which available standards and instrument characterization procedures are discussed along with their purpose, suitability, and general instructions for use. These include the most relevant instrument properties needing qualification, such as linearity and spectral responsivity of the detection system, spectral irradiance reaching the sample, wavelength accuracy, sensitivity or limit of detection for an analyte, and day-to-day performance verification. With proper consideration of method-inherent requirements and limitations, many of these procedures and standards can be adapted to other fluorescence techniques. In addition, procedures for the determination of other relevant fluorometric quantities including fluorescence quantum yields and fluorescence lifetimes are briefly introduced. The guide is a clear and concise reference geared for users of fluorescence instrumentation at all levels of experience and is intended to aid in the ongoing standardization of fluorescence measurements.

  11. Fluorescence lifetime-based biosensing of zinc: Origin of the broad dynamic range.

    Science.gov (United States)

    Thompson, R B; Patchan, M W

    1995-06-01

    Fluorescence lifetime-based chemical sensors have recently been described for applications in medicine, environmental monitoring, and bioprocess control. These sensors transduce the level of the analyte as a change in the apparent fluorescence lifetime of an indicator phase. We have previously developed a wavelength-ratiometric fluorescence biosensor for zinc based on binding of zinc and dansylamide to apo-carbonic anhydrase which exhibited high sensitivity and selectivity. We demonstrate that the apo-carbonic anhydrase/dansylamide indicator system is very well suited for lifetime-based sensing, with a subnanomolar detection limit and greater than 1000-fold dynamic range. The theoretical basis for the wide dynamic range is discussed.

  12. 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.

  13. 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.

  14. Fluorescence lifetime measurements of intrinsically unstructured proteins: application to α-synuclein.

    Science.gov (United States)

    Schreurs, Sarah; Kluba, Malgorzata; Meuvis, Jessika; Engelborghs, Yves

    2012-01-01

    Lifetimes of fluorescent states and their fluorescence intensities are strictly coupled and very sensitive to the environment of the fluorophores. The advantage of measuring lifetimes, next to intensities, comes from the fact that it can reveal heterogeneity and dynamic properties of this environment. In this way lifetime analysis can be used to characterize static and dynamic conformational properties and heterogeneity of fluorescent groups in different areas of a protein and as a function of time for an evolving protein. The phenomena that determine the lifetime of a label are its intrinsic properties, dynamic quenching by neighboring groups, exposure to the solvent, as well as Förster resonance energy transfer (FRET) between different groups. The basic principles of these fluorescence phenomena can be found extensively described in the excellent book of Lakowicz (Principles of fluorescence spectroscopy, 3rd edn. Springer, New York, 2006). The fluorescent groups involved are either natural amino acid side chains like tryptophan (Trp) or tyrosine (Tyr), or fluorescent labels covalently engineered into the protein. Even a single fluorescent group can show indications of heterogeneity in the local environment. If several natural fluorescent groups are present, the properties of the individual groups can be separated using site-directed mutagenesis, and additivity of their contributions can be analyzed (Engelborghs, Spectrochim Acta A Mol Biomol Spectrosc 57(11):2255-2270, 2001). If no fluorescent group is naturally present, site-directed mutagenesis can be used to introduce either a fluorescent amino acid or a cysteine allowing chemical labeling.

  15. Fluorescence optimisation and lifetime studies of fingerprints treated with magnetic powders.

    Science.gov (United States)

    Seah, L K; Dinish, U S; Phang, W F; Chao, Z X; Murukeshan, V M

    2005-09-10

    Fluorescence study plays a significant role in fingerprint detection when conventional chemical enhancement methods fail. The basic properties of fluorescence emission such as colour, intensity and lifetime could be well exploited in the detection of latent fingerprints under steady state and in dynamic methods. This paper describes a systematic study of fluorescence emission intensity from fingerprint samples treated with different magnetic powders. Understanding of suitable excitation wavelength required for getting maximum fluorescence emission intensity could be beneficial when selecting the appropriate fluorescent powders for the fingerprint detection. Lifetime study of fingerprints treated with various magnetic powders was also carried out. The importance of lifetime study is well explained through the time-resolved (TR) imaging of fingerprints with nanosecond resolution. Results from the TR imaging study revealed an improvement in the fingerprint image contrast. This is significant when the print is deposited on fluorescing background and its emission wavelength is close to that of treated fingerprint.

  16. Multiphoton microscopy, fluorescence lifetime imaging and optical spectroscopy for the diagnosis of neoplasia

    Science.gov (United States)

    Skala, Melissa Caroline

    2007-12-01

    Cancer morbidity and mortality is greatly reduced when the disease is diagnosed and treated early in its development. Tissue biopsies are the gold standard for cancer diagnosis, and an accurate diagnosis requires a biopsy from the malignant portion of an organ. Light, guided through a fiber optic probe, could be used to inspect regions of interest and provide real-time feedback to determine the optimal tissue site for biopsy. This approach could increase the diagnostic accuracy of current biopsy procedures. The studies in this thesis have characterized changes in tissue optical signals with carcinogenesis, increasing our understanding of the sensitivity of optical techniques for cancer detection. All in vivo studies were conducted on the dimethylbenz[alpha]anthracene treated hamster cheek pouch model of epithelial carcinogenesis. Multiphoton microscopy studies in the near infrared wavelength region quantified changes in tissue morphology and fluorescence with carcinogenesis in vivo. Statistically significant morphological changes with precancer included increased epithelial thickness, loss of stratification in the epithelium, and increased nuclear diameter. Fluorescence changes included a statistically significant decrease in the epithelial fluorescence intensity per voxel at 780 nm excitation, a decrease in the fluorescence lifetime of protein-bound nicotinamide adenine dinucleotide (NADH, an electron donor in oxidative phosphorylation), and an increase in the fluorescence lifetime of protein-bound flavin adenine dinucleotide (FAD, an electron acceptor in oxidative phosphorylation) with precancer. The redox ratio (fluorescence intensity of FAD/NADH, a measure of the cellular oxidation-reduction state) did not significantly change with precancer. Cell culture experiments (MCF10A cells) indicated that the decrease in protein-bound NADH with precancer could be due to increased levels of glycolysis. Point measurements of diffuse reflectance and fluorescence spectra in

  17. Molecular Fluorescence Lifetime Fluctuations: On the Possible Role of Conformational Effects

    NARCIS (Netherlands)

    Vallée, R.A.L.; Vancso, G.J.; Hulst, van N.F.; Calbert, J.-P.; Cornil, J.; Brédas, J.L.

    2003-01-01

    The radiative lifetime of single 1,1'-dioctadecyl-3,3,3',3'- etramethylindodicarbocyanine molecules, embedded in a polymer thin film, has been characterized. At room temperature the chemically identical molecules exhibit strong fluctuations in their fluorescence lifetime. The possible conformational

  18. Fluorescence lifetime plate reader: resolution and precision meet high-throughput.

    Science.gov (United States)

    Petersen, Karl J; Peterson, Kurt C; Muretta, Joseph M; Higgins, Sutton E; Gillispie, Gregory D; Thomas, David D

    2014-11-01

    We describe a nanosecond time-resolved fluorescence spectrometer that acquires fluorescence decay waveforms from each well of a 384-well microplate in 3 min with signal-to-noise exceeding 400 using direct waveform recording. The instrument combines high-energy pulsed laser sources (5-10 kHz repetition rate) with a photomultiplier and high-speed digitizer (1 GHz) to record a fluorescence decay waveform after each pulse. Waveforms acquired from rhodamine or 5-((2-aminoethyl)amino) naphthalene-1-sulfonic acid dyes in a 384-well plate gave lifetime measurements 5- to 25-fold more precise than the simultaneous intensity measurements. Lifetimes as short as 0.04 ns were acquired by interleaving with an effective sample rate of 5 GHz. Lifetime measurements resolved mixtures of single-exponential dyes with better than 1% accuracy. The fluorescence lifetime plate reader enables multiple-well fluorescence lifetime measurements with an acquisition time of 0.5 s per well, suitable for high-throughput fluorescence lifetime screening applications.

  19. Determination of biological activity from fluorescence-lifetime measurements in Saccharomyces cerevisiae

    Science.gov (United States)

    Rudek, F.; Baselt, T.; Lempe, B.; Taudt, C.; Hartmann, P.

    2015-03-01

    The importance of fluorescence lifetime measurement as an optical analysis tool is growing. Many applications already exist in order to determine the fluorescence lifetime, but the majority of these require the addition of fluorescence-active substances to enable measurements. Every usage of such foreign materials has an associated risk. This paper investigates the use of auto-fluorescing substances in Saccharomyces cerevisiae (Baker's yeast) as a risk free alternative to fluorescence-active substance enabled measurements. The experimental setup uses a nitrogen laser with a pulse length of 350 ps and a wavelength of 337 nm. The excited sample emits light due to fluorescence of NADH/NADPH and collagen. A fast photodiode collects the light at the output of an appropriate high-pass edge-filter at 400 nm. Fluorescence lifetimes can be determined from the decay of the measurement signals, which in turn characterizes the individual materials and their surrounding environment. Information about the quantity of the fluorescence active substances can also be measured based on the received signal intensity. The correlation between the fluorescence lifetime and the metabolic state of Saccharomyces cerevisiae was investigated and is presented here.

  20. 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.

  1. DBD dyes as fluorescence lifetime probes to study conformational changes in proteins.

    Science.gov (United States)

    Wawrzinek, Robert; Ziomkowska, Joanna; Heuveling, Johanna; Mertens, Monique; Herrmann, Andreas; Schneider, Erwin; Wessig, Pablo

    2013-12-16

    Previously, [1,3]dioxolo[4,5-f][1,3]benzodioxole (DBD)-based fluorophores used as highly sensitive fluorescence lifetime probes reporting on their microenvironmental polarity have been described. Now, a new generation of DBD dyes has been developed. Although they are still sensitive to polarity, in contrast to the former DBD dyes, they have extraordinary spectroscopic properties even in aqueous surroundings. They are characterized by long fluorescence lifetimes (10-20 ns), large Stokes shifts (≈100 nm), high photostabilities, and high quantum yields (>0.56). Here, the spectroscopic properties and synthesis of functionalized derivatives for labeling biological targets are described. Furthermore, thio-reactive maleimido derivatives of both DBD generations show strong intramolecular fluorescence quenching. This mechanism has been investigated and is found to undergo a photoelectron transfer (PET) process. After reaction with a thiol group, this fluorescence quenching is prevented, indicating successful bonding. Being sensitive to their environmental polarity, these compounds have been used as powerful fluorescence lifetime probes for the investigation of conformational changes in the maltose ATP-binding cassette transporter through fluorescence lifetime spectroscopy. The differing tendencies of the fluorescence lifetime change for both DBD dye generations promote their combination as a powerful toolkit for studying microenvironments in proteins.

  2. Nanoantenna array-induced fluorescence enhancement and reduced lifetimes

    DEFF Research Database (Denmark)

    Bakker, R. M.; Drachev, V. P.; Liu, Z.;

    2008-01-01

    Enhanced fluorescence is observed from dye molecules interacting with optical nanoantenna arrays. Elliptical gold dimers form individual nanoantennae with tunable plasmon resonances depending upon the geometry of the two particles and the size of the gap between them. A fluorescent dye, Rhodamine...

  3. Time-domain measurement of fluorescence lifetime variation with pH

    Science.gov (United States)

    Ryder, Alan G.; Power, Sarah; Glynn, Thomas J.; Morrison, John J.

    2001-07-01

    Advances in the design and miniaturization of the lasers and electronics required for Time Correlated Single Photon Counting (TCSPC) measurement of fluorescence lifetime have simplified the use of the time domain method. We have assembled a compact portable system that is capable of measuring lifetimes down to approximately 200 ps (with deconvolution) and that can operate at a range of excitation and emission wavelengths. The excitation sources are pulsed LEDs and laser diodes with a maximum pulse rate of 40 MHz and are easily interchanged. Furthermore, the development of violet and blue GaN LEDs and laser diodes is expanding the range of fluorophores available for fluorescence lifetime measurement of ion concentrations. pH sensitive fluorophores have a wide range of biological and clinical applications. The use of fluorescence lifetime rather than intensity to measure pH has a number of advantages including the reduction of effects due to the photobleaching, scattering, and intensity variations in the excitation source. Using our compact TCSPC instrumentation we have measured the dependence of fluorescence lifetimes on pH for a range of dyes in phosphate buffer over the physiologically important range of 6.0 to 8.0. Most dyes exhibit only a small variation in lifetime (pH range; however, acridine exhibits a large variation in lifetime and hence shows promise as a pH indicator.

  4. Assessing the photoaging process at sun exposed and non-exposed skin using fluorescence lifetime spectroscopy

    Science.gov (United States)

    Saito Nogueira, Marcelo; Kurachi, Cristina

    2016-03-01

    Photoaging is the skin premature aging due to exposure to ultraviolet light, which damage the collagen, elastin and can induce alterations on the skin cells DNA, and, then, it may evolve to precancerous lesions, which are widely investigated by fluorescence spectroscopy and lifetime. The fluorescence spectra and fluorescence lifetime analysis has been presented as a technique of great potential for biological tissue characterization at optical diagnostics. The main targeted fluorophores are NADH (nicotinamide adenine dinucleotide) and FAD (flavin adenine dinucleotide), which have free and bound states, each one with different average lifetimes. The average lifetimes for free and bound NADH and FAD change according to tissue metabolic alterations and may contribute to a non-invasive clinical investigation of injuries such as skin lesions. These lesions and the possible areas where they may develop can be interrogated using fluorescence lifetime spectroscopy taking into account the variability of skin phototypes and the changes related to melanin, collagen and elastin, endogenous fluorophores which have emissions that spectrally overlap to the NADH and FAD emission. The objective of this study is to assess the variation on fluorescence lifetimes of normal skin at sun exposed and non-exposed areas and associate this variation to the photoaging process.

  5. Ruby fluorescence lifetime measurements for temperature determinations at high (p, T)

    Science.gov (United States)

    Bauer, Johannes D.; Bayarjargal, Lkhamsuren; Winkler, Björn

    2012-06-01

    The lifetime of the ruby R1 fluorescence line was measured as a function of pressure (up to about 20 GPa) and temperature (550 K) in an externally heated diamond anvil cell (DAC). At constant temperatures, the lifetime is increasing linearly with increasing pressure. The slope of the pressure dependence is constant up to a temperature of 450 K and it is decreasing at higher temperatures. At constant pressure, the lifetime is exponentially decreasing with increasing temperature. The (p, T)-dependence can be parametrized by the combination of a linear and an exponential function. This allows an accurate p, T-determination by the combination of fluorescence spectroscopy using Sm2+-doped strontium tetraborate and lifetime measurements of ruby, as the energy of the Sm2+ fluorescence is nearly temperature-independent.

  6. Quantitative mapping of aqueous microfluidic temperature with sub-degree resolution using fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Graham, Emmelyn M; Iwai, Kaoru; Uchiyama, Seiichi; de Silva, A Prasanna; Magennis, Steven W; Jones, Anita C

    2010-05-21

    The use of a water-soluble, thermo-responsive polymer as a highly sensitive fluorescence-lifetime probe of microfluidic temperature is demonstrated. The fluorescence lifetime of poly(N-isopropylacrylamide) labelled with a benzofurazan fluorophore is shown to have a steep dependence on temperature around the polymer phase transition and the photophysical origin of this response is established. The use of this unusual fluorescent probe in conjunction with fluorescence lifetime imaging microscopy (FLIM) enables the spatial variation of temperature in a microfluidic device to be mapped, on the micron scale, with a resolution of less than 0.1 degrees C. This represents an increase in temperature resolution of an order of magnitude over that achieved previously by FLIM of temperature-sensitive dyes.

  7. The use of DAPI fluorescence lifetime imaging for investigating chromatin condensation in human chromosomes.

    Science.gov (United States)

    Estandarte, Ana Katrina; Botchway, Stanley; Lynch, Christophe; Yusuf, Mohammed; Robinson, Ian

    2016-08-16

    Chromatin undergoes dramatic condensation and decondensation as cells transition between the different phases of the cell cycle. The organization of chromatin in chromosomes is still one of the key challenges in structural biology. Fluorescence lifetime imaging (FLIM), a technique which utilizes a fluorophore's fluorescence lifetime to probe changes in its environment, was used to investigate variations in chromatin compaction in fixed human chromosomes. Fixed human metaphase and interphase chromosomes were labeled with the DNA minor groove binder, DAPI, followed by measurement and imaging of the fluorescence lifetime using multiphoton excitation. DAPI lifetime variations in metaphase chromosome spreads allowed mapping of the differentially compacted regions of chromatin along the length of the chromosomes. The heteromorphic regions of chromosomes 1, 9, 15, 16, and Y, which consist of highly condensed constitutive heterochromatin, showed statistically significant shorter DAPI lifetime values than the rest of the chromosomes. Differences in the DAPI lifetimes for the heteromorphic regions suggest differences in the structures of these regions. DAPI lifetime variations across interphase nuclei showed variation in chromatin compaction in interphase and the formation of chromosome territories. The successful probing of differences in chromatin compaction suggests that FLIM has enormous potential for application in structural and diagnostic studies.

  8. Improved Fluorescent Protein Contrast and Discrimination by Optically Controlling Dark State Lifetimes.

    Science.gov (United States)

    Chen, Yen-Cheng; Dickson, Robert M

    2017-02-16

    Modulation and optical control of photoswitchable fluorescent protein (PS-FP) dark state lifetimes drastically improves sensitivity and selectivity in fluorescence imaging. The dark state population of PS-FPs generates an out-of-phase fluorescence component relative to the sinusoidally modulated 488 nm laser excitation. Because this apparent phase advanced emission results from slow recovery to the fluorescent manifold, we hasten recovery and, therefore, modulation frequency by varying coillumination intensity at 405 nm. As 405 nm illumination regenerates the fluorescent ground state more rapidly than via thermal recovery, we experimentally demonstrate that secondary illumination can control PS-FPs dark state lifetime to act as an additional dimension for discriminating spatially and spectrally overlapping emitters. This experimental combination of out of phase imaging after optical modulation (OPIOM) and synchronously amplified fluorescence image recovery (SAFIRe) optically controls the fluorescent protein dark state lifetimes for improved time resolution, with the resulting modulation-based selective signal recovery being quantitatively modeled. The combined experimental results and quantitative numerical simulations further demonstrate the potential of SAFIRe-OPIOM for wide-field biological imaging with improved speed, sensitivity, and optical resolution over other modulation-based fluorescence microscopies.

  9. Fluorescence lifetime imaging of the oxygen distribution in the skin

    Science.gov (United States)

    Kieslinger, Dietmar; Draxler, Sonja; Puzon, Janusz; Lippitsch, Max E.

    1997-05-01

    An instrument has been designed and implemented capable of mapping oxygen distribution in skin tissue over an area of several square centimeters with a spatial resolution of better than 1 mm and with a resolution in oxygen partial pressure of better than 5 torr. The measurement scheme is optical and is based on luminescence lifetime. It is non- invasive and avoids any patient contact with electrical parts. The instrument should be a valuable supplement to other clinical methods for monitoring microcirculation and peripheral oxygen supply.

  10. Use of multiphoton tomography and fluorescence lifetime imaging to investigate skin pigmentation in vivo

    Science.gov (United States)

    Dancik, Yuri; Favre, Amandine; Loy, Chong Jin; Zvyagin, Andrei V.; Roberts, Michael S.

    2013-02-01

    There is a growing body of literature showing the usefulness of multiphoton tomography (MPT) and fluorescence lifetime imaging for in situ characterization of skin constituents and the ensuing development of noninvasive diagnostic tools against skin diseases. Melanin and pigmentation-associated skin cancers constitute some of the major applications. We show that MPT and fluorescence lifetime imaging can be used to measure changes in cutaneous melanin concentration and that these can be related to the visible skin color. Melanin in the skin of African, Indian, Caucasian, and Asian volunteers is detected on the basis of its emission wavelength and fluorescence lifetimes in solution and in a melanocyte-keratinocyte cell culture. Fluorescence intensity is used to characterize the melanin content and distribution as a function of skin type and depth into the skin (stratum granulosum and stratum basale). The measured fluorescence intensities in given skin types agree with melanin amounts reported by others using biopsies. Our results suggest that spatial distribution of melanin in skin can be studied using MPT and fluorescence lifetime imaging, but further studies are needed to ascertain that the method can resolve melanin amount in smaller depth intervals.

  11. 3D printed miniaturized spectral system for tissue fluorescence lifetime measurements

    Science.gov (United States)

    Zou, Luwei; Mahmoud, Mohamad; Fahs, Mehdi; Liu, Rui; Lo, Joe F.

    2016-04-01

    Various types of collagens, e.g. type I and III, represent the main load-bearing components in biological tissues. Their composition changes during processes like wound healing and fibrosis. Collagens exhibit autofluorescence when excited by ultra-violet light, distinguishable by their unique fluorescent lifetimes across a range of emission wavelengths. Therefore, we designed a miniaturized spectral-lifetime detection system for collagens as a non-invasive probe for monitoring tissue in wound healing and scarring applications. A sine modulated LED illumination was applied to enable frequency domain (FD) fluorescence lifetime measurements under different wavelengths bands, separated via a series of longpass dichroics at 387nm, 409nm and 435nm. To achieve the minute scale of optomechanics, we employed a stereolithography based 3D printer with types were measured, where the separate spectral bands enhanced the differentiation of their lifetimes.

  12. High-speed confocal fluorescence lifetime imaging microscopy by analog mean-delay method

    Science.gov (United States)

    Won, Youngjae; Kim, Donguk; Yang, Wenzhong; Kim, Dug Y.

    2010-02-01

    We have demonstrated the high-speed confocal fluorescence lifetime imaging microscopy (FLIM) by analog mean-delay (AMD) method. The AMD method is a new signal processing technique for calculation of fluorescence lifetime and it is very suitable for the high-speed confocal FLIM with good accuracy and photon economy. We achieved the acquisition speed of 7.7 frames per second for confocal FLIM imaging. Here, the highest photon detection rate for one pixel was larger than 125 MHz and averaged photon detection rate was more than 62.5 MHz. Based on our system, we successfully obtained a sequence of confocal fluorescence lifetime images of RBL-2H3 cell labeled with Fluo-3/AM and excited by 4αPDD (TRPV channel agonist) within one second.

  13. Exciton-polaron quenching in organic thin-film transistors studied by fluorescence lifetime imaging microscopy

    DEFF Research Database (Denmark)

    Jensen, Per Baunegaard With; Leißner, Till; Osadnik, Andreas

    Organic semiconductors show great potential in electronic and optical applications. However, a major challenge is the degradation of the semiconductor materials that cause a reduction in device performance. Here, we present our investigations of Organic Thin Film Transistors (OTFT) based...... that correlates with the local charge density indicates a pronounced exciton quenching by the injected charges. Subsequent FLIM measurements on previously biased OTFT devices show a general decrease in fluorescence lifetime suggesting degradation of the organic semiconductor. This is correlated with the results...... on the material 5,5-bis(naphthyl)-2,20-bithiophene (NaT2). These types of OTFT have previously been shown to have light emitting properties. Fluorescence Lifetime Imaging Microscopy (FLIM) has been used to investigate the exciton-polaron quenching in biased OTFTs. A clear reduction in fluorescence lifetime...

  14. Temperature-dependent fluorescence lifetime of a fluorescent polymeric thermometer, poly(N-isopropylacrylamide), labeled by polarity and hydrogen bonding sensitive 4-sulfamoyl-7-aminobenzofurazan.

    Science.gov (United States)

    Gota, Chie; Uchiyama, Seiichi; Yoshihara, Toshitada; Tobita, Seiji; Ohwada, Tomohiko

    2008-03-13

    Fluorescent molecular thermometers showing temperature-dependent fluorescence lifetimes enable thermal mapping of small spaces such as a microchannel and a living cell. We report the temperature-dependent fluorescence lifetimes of poly(NIPAM-co-DBD-AA), which is a random copolymer of N-isopropylacrylamide (NIPAM) and an environment-sensitive fluorescent monomer (DBD-AA) containing a 4-sulfamoyl-7-aminobenzofurazan structure. The average fluorescence lifetime of poly(NIPAM-co-DBD-AA) in aqueous solution increased from 4.22 to 14.1 ns with increasing temperature from 30 to 35 degrees C. This drastic change in fluorescence lifetime (27% increase per 1 degrees C) is the sharpest ever reported. Concentration independency, one of the advantages of fluorescence lifetime measurements, was seen in average fluorescence lifetime (13.7 +/- 0.18 ns) of poly(NIPAM-co-DBD-AA) at 33 degrees C over a wide concentration range (0.005-1 w/v%). With increasing temperature, polyNIPAM units in poly(NIPAM-co-DBD-AA) change their structure from an extended form to a globular form, providing apolar and aprotic environments to the fluorescent DBD-AA units. Consequently, the environment-sensitive DBD-AA units translate the local environmental changes into the extension of the fluorescence lifetime. This role of the DBD-AA units was revealed by a study of solvent effects on fluorescence lifetime of a model environment-sensitive fluorophore.

  15. 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.

  16. Fluorescence lifetime measurements of native and glycated human serum albumin and bovine serum albumin

    Science.gov (United States)

    Joshi, Narahari V.; Joshi, Virgina O. d.; Contreras, Silvia; Gil, Herminia; Medina, Honorio; Siemiarczuk, Aleksander

    1999-05-01

    Nonenzymatic glycation, also known as Maillard reaction, plays an important role in the secondary complications of the diabetic pathology and aging, therefore, human serum albumin (HSA) and bovine serum albumin (BSA) were glycated by a conventional method in our laboratory using glucose as the glycating agent. Fluorescence lifetime measurements were carried out with a laser strobe fluorometer equipped with a nitrogen/dye laser and a frequency doubler as a pulsed excitation source. The samples were excited at 295 nm and the emission spectra were recorded at 345 nm. The obtained decay curves were tried for double and triple exponential functions. It has been found that the shorter lifetime increases for glycated proteins as compared with that of the native ones. For example, in the case of glycated BSA the lifetime increased from 1.36 ns to 2.30 ns. Similarly, for HSA, the lifetime increases from 1.58 ns to 2.26 ns. Meanwhile, the longer lifetime changed very slightly for both proteins (from 6.52 ns to 6.72 ns). The increase in the lifetime can be associated with the environmental effect; originated from the attachment of glucose to some lysine residues. A good example is Trp 214 which is in the cage of Lys 225, Lys 212, Lys 233, Lys 205, Lys 500, Lys 199 and Lys 195. If fluorescence lifetime technique is calibrated and properly used it could be employed for assessing glycation of proteins.

  17. Spectrally resolved fluorescence lifetime imaging to investigate cell metabolism in malignant and nonmalignant oral mucosa cells.

    Science.gov (United States)

    Rück, Angelika; Hauser, Carmen; Mosch, Simone; Kalinina, Sviatlana

    2014-09-01

    Fluorescence-guided diagnosis of tumor tissue is in many cases insufficient, because false positive results interfere with the outcome. Improvement through observation of cell metabolism might offer the solution, but needs a detailed understanding of the origin of autofluorescence. With respect to this, spectrally resolved multiphoton fluorescence lifetime imaging was investigated to analyze cell metabolism in metabolic phenotypes of malignant and nonmalignant oral mucosa cells. The time-resolved fluorescence characteristics of NADH were measured in cells of different origins. The fluorescence lifetime of bound and free NADH was calculated from biexponential fitting of the fluorescence intensity decay within different spectral regions. The mean lifetime was increased from nonmalignant oral mucosa cells to different squamous carcinoma cells, where the most aggressive cells showed the longest lifetime. In correlation with reports in the literature, the total amount of NADH seemed to be less for the carcinoma cells and the ratio of free/bound NADH was decreased from nonmalignant to squamous carcinoma cells. Moreover for squamous carcinoma cells a high concentration of bound NADH was found in cytoplasmic organelles (mainly mitochondria). This all together indicates that oxidative phosphorylation and a high redox potential play an important role in the energy metabolism of these cells.

  18. Spectrally resolved fluorescence lifetime imaging to investigate cell metabolism in malignant and nonmalignant oral mucosa cells

    Science.gov (United States)

    Rück, Angelika; Hauser, Carmen; Mosch, Simone; Kalinina, Sviatlana

    2014-09-01

    Fluorescence-guided diagnosis of tumor tissue is in many cases insufficient, because false positive results interfere with the outcome. Improvement through observation of cell metabolism might offer the solution, but needs a detailed understanding of the origin of autofluorescence. With respect to this, spectrally resolved multiphoton fluorescence lifetime imaging was investigated to analyze cell metabolism in metabolic phenotypes of malignant and nonmalignant oral mucosa cells. The time-resolved fluorescence characteristics of NADH were measured in cells of different origins. The fluorescence lifetime of bound and free NADH was calculated from biexponential fitting of the fluorescence intensity decay within different spectral regions. The mean lifetime was increased from nonmalignant oral mucosa cells to different squamous carcinoma cells, where the most aggressive cells showed the longest lifetime. In correlation with reports in the literature, the total amount of NADH seemed to be less for the carcinoma cells and the ratio of free/bound NADH was decreased from nonmalignant to squamous carcinoma cells. Moreover for squamous carcinoma cells a high concentration of bound NADH was found in cytoplasmic organelles (mainly mitochondria). This all together indicates that oxidative phosphorylation and a high redox potential play an important role in the energy metabolism of these cells.

  19. Multiphoton fluorescence lifetime imaging of metabolic status in mesenchymal stem cell during adipogenic differentiation

    Science.gov (United States)

    Meleshina, A. V.; Dudenkova, V. V.; Shirmanova, M. V.; Bystrova, A. S.; Zagaynova, E. V.

    2016-03-01

    Non-invasive imaging of cell metabolism is a valuable approach to assess the efficacy of stem cell therapy and understand the tissue development. In this study we analyzed metabolic trajectory of the mesenchymal stem cells (MCSs) during differentiation into adipocytes by measuring fluorescence lifetimes of free and bound forms of the reduced nicotinamide adenine dinucleotide (NAD(P)H) and flavine adenine dinucleotide (FAD). Undifferentiated MSCs and MSCs on the 5, 12, 19, 26 days of differentiation were imaged on a Zeiss 710 microscope with fluorescence lifetime imaging (FLIM) system B&H (Germany). Fluorescence of NAD(P)H and FAD was excited at 750 nm and 900 nm, respectively, by a femtosecond Ti:sapphire laser and detected in a range 455-500 nm and 500-550 nm, correspondingly. We observed the changes in the NAD(P)H and FAD fluorescence lifetimes and their relative contributions in the differentiated adipocytes compare to undifferentiated MSCs. Increase of fluorescence lifetimes of the free and bound forms of NAD(P)H and the contribution of protein-bound NAD(P)H was registered, that can be associated with a metabolic switch from glycolysis to oxidative phosphorylation and/or synthesis of lipids in adipogenically differentiated MSCs. We also found that the contribution of protein-bound FAD decreased during differentiation. After carrying out appropriate biochemical measurements, the observed changes in cellular metabolism can potentially serve to monitor stem cell differentiation by FLIM.

  20. 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.

  1. [The analysis of sinusoidal modulated method used for measuring fluorescence lifetime].

    Science.gov (United States)

    Feng, Ying; Huang, Shi-hua

    2007-12-01

    This paper has built a system with a sinusoidal modulated LED as the excitation source. Such exciter was used upon the sample Eu2 L'3 x nH2O (L' = C4H4O4). Both the excitation light and the 5Do-7F2 emission of Eu3+ ion were measured. Fluorescence lifetime, which approximate to 0.680 ms, can then be obtained from the measured excitation and fluorescence waveforms by non-linear least square curve fitting based on the principle of phase-shift measurement of fluorescence lifetime. Data processing methods considering respectively the high order harmonics in the modulation and multi-exponential decay of the fluorescence were discussed. A method of utilizing Fourier series expandedness to amendatory the result was put forward. Accordingly, the applicability for phase-shift method was expanded as well as a more exact result was acquired.

  2. The Gray Institute 'open' high-content, fluorescence lifetime microscopes.

    Science.gov (United States)

    Barber, P R; Tullis, I D C; Pierce, G P; Newman, R G; Prentice, J; Rowley, M I; Matthews, D R; Ameer-Beg, S M; Vojnovic, B

    2013-08-01

    We describe a microscopy design methodology and details of microscopes built to this 'open' design approach. These demonstrate the first implementation of time-domain fluorescence microscopy in a flexible automated platform with the ability to ease the transition of this and other advanced microscopy techniques from development to use in routine biology applications. This approach allows easy expansion and modification of the platform capabilities, as it moves away from the use of a commercial, monolithic, microscope body to small, commercial off-the-shelf and custom made modular components. Drawings and diagrams of our microscopes have been made available under an open license for noncommercial use at http://users.ox.ac.uk/~atdgroup. Several automated high-content fluorescence microscope implementations have been constructed with this design framework and optimized for specific applications with multiwell plates and tissue microarrays. In particular, three platforms incorporate time-domain FLIM via time-correlated single photon counting in an automated fashion. We also present data from experiments performed on these platforms highlighting their automated wide-field and laser scanning capabilities designed for high-content microscopy. Devices using these designs also form radiation-beam 'end-stations' at Oxford and Surrey Universities, showing the versatility and extendibility of this approach. © 2013 The Authors. Journal of Microscopy published by John Wiley & Sons Ltd on behalf of Royal Microscopical Society.

  3. Real-time fluorescence lifetime actuation for cell sorting using a CMOS SPAD silicon photomultiplier.

    Science.gov (United States)

    Rocca, Francescopaolo Mattioli Della; Nedbal, Jakub; Tyndall, David; Krstajić, Nikola; Li, David Day-Uei; Ameer-Beg, Simon M; Henderson, Robert K

    2016-02-15

    Time-correlated single photon counting (TCSPC) is a fundamental fluorescence lifetime measurement technique offering high signal to noise ratio (SNR). However, its requirement for complex software algorithms for histogram processing restricts throughput in flow cytometers and prevents on-the-fly sorting of cells. We present a single-point digital silicon photomultiplier (SiPM) detector accomplishing real-time fluorescence lifetime-activated actuation targeting cell sorting applications in flow cytometry. The sensor also achieves burst-integrated fluorescence lifetime (BIFL) detection by TCSPC. The SiPM is a single-chip complementary metal-oxide-semiconductor (CMOS) sensor employing a 32×32 single-photon avalanche diode (SPAD) array and eight pairs of time-interleaved time to digital converters (TI-TDCs) with a 50 ps minimum timing resolution. The sensor's pile-up resistant embedded center of mass method (CMM) processor accomplishes low-latency measurement and thresholding of fluorescence lifetime. A digital control signal is generated with a 16.6 μs latency for cell sorter actuation allowing a maximum cell throughput of 60,000 cells per second and an error rate of 0.6%.

  4. Visualizing heterogeneity of photosynthetic properties of plant leaves with two-photon fluorescence lifetime imaging microscopy

    NARCIS (Netherlands)

    Iermak, Ievgeniia; Vink, Jochem; Bader, Arjen N.; Wientjes, Emilie; Amerongen, van Herbert

    2016-01-01

    Two-photon fluorescence lifetime imaging microscopy (FLIM) was used to analyse the distribution and properties of Photosystem I (PSI) and Photosystem II (PSII) in palisade and spongy chloroplasts of leaves from the C3 plant Arabidopsis thaliana and the C4 plant Miscanthus x giganteus. This was ac

  5. Fluorescence lifetime biosensing with DNA microarrays and a CMOS-SPAD imager

    NARCIS (Netherlands)

    Giraud, G.; Schulze, H.; Li, D.U.; Bachmann, T.T.; Crain, J.; Tyndall, D.; Richardson, J.; Walker, R.; Stoppa, D.; Charbon, E.; Henderson, R.; Arlt, J.

    2010-01-01

    Fluorescence lifetime of dye molecules is a sensitive reporter on local microenvironment which is generally independent of fluorophores concentration and can be used as a means of discrimination between molecules with spectrally overlapping emission. It is therefore a potentially powerful multiplexe

  6. The Use of Chlorophyll Fluorescence Lifetime to Assess Phytoplankton Physiology within a River-Dominated Environment

    Science.gov (United States)

    Hall, Callie M.; Miller, Richard L.; Redalje, Donald G.; Fernandez, Salvador M.

    2002-01-01

    Chlorophyll a fluorescence lifetime was measured for phytoplankton populations inhabiting the three physical zones surrounding the Mississippi River's terminus in the Gulf of Mexico. Observations of river discharge volume, nitrate + nitrite, silicate, phosphate, PAR (Photosynthetically Active Radiation) diffuse attenuation within the water column, salinity, temperature, SPM, and chl a concentration were used to characterize the distribution of chl fluorescence lifetime within a given region within restricted periods of time. 33 stations extending from the Mississippi River plume to the shelf break of the Louisiana coast were surveyed for analysis of chlorophyll fluorescence lifetime during two cruises conducted March 31 - April 6, 2000, and October 24 - November 1, 2000. At each station, two to three depths were chosen for fluorescence lifetime measurement to represent the vertical characteristics of the water column. Where possible, samples were taken from just below the surface and from just above and below the pycnocline. All samples collected were within the 1% light level of the water column (the euphotic zone). Upon collection, samples were transferred to amber Nalgene bottles and left in the dark for at least 15 minutes to reduce the effects of non-photochemical quenching and to insure that photosynthetic reaction centers were open. Before measurements within the phase fluorometer were begun, the instrument was allowed to warm up for no less than one hour.

  7. Fluorescence lifetime of emitters with broad homogeneous linewidths modified in opal photonic crystals

    DEFF Research Database (Denmark)

    Nikolaev, Ivan S.; Lodahl, Peter; Vos, Willem L.

    2008-01-01

    We have investigated the dynamics of spontaneous emission from dye molecules embedded in opal photonic crystals. Fluorescence lifetimes of Rhodamine 6G (R6G) dye were measured as a function of both optical frequency and crystal lattice parameter of the polystyrene opals. Due to the broad homogene...

  8. Fluorescence lifetime imaging of lipids during 3T3-L1 cell differentiation

    Science.gov (United States)

    Song, Young Sik; Won, Young Jae; Lee, Sang-Hak; Kim, Dug Young

    2014-03-01

    Obesity is becoming a big health problem in these days. Since increased body weight is due to increased number and size of the triglyceride-storing adipocytes, many researchers are working on differentiation conditions and processes of adipocytes. Adipocytes also work as regulators of whole-body energy homeostasis by secreting several proteins that regulate processes as diverse as haemostasis, blood pressure, immune function, angiogenesis and energy balance. 3T3-L1 cells are widely used cell line for studying adipogenesis because it can differentiate into an adipocyte-like phenotype under appropriate conditions. In this paper, we propose an effective fluorescence lifetime imaging technique which can easily distinguish lipids in membrane and those in lipid droplets. Nile red dyes are attached to lipids in 3T3-L1 cells. Fluorescence lifetime images were taken for 2 week during differentiation procedure of 3T3-L1 cells into adipocytes. We used 488 nm pulsed laser with 5MHz repetition rate and emission wavelength is 520 nm of Nile Red fluorescent dye. Results clearly show that the lifetime of Nile red in lipid droplets are smaller than those in cell membrane. Our results suggest that fluorescence lifetime imaging can be a very powerful tool to monitor lipid droplet formation in adipocytes from 3T3-L1 cells.

  9. Towards sensitive, high-throughput, biomolecular assays based on fluorescence lifetime

    Science.gov (United States)

    Ioanna Skilitsi, Anastasia; Turko, Timothé; Cianfarani, Damien; Barre, Sophie; Uhring, Wilfried; Hassiepen, Ulrich; Léonard, Jérémie

    2017-09-01

    Time-resolved fluorescence detection for robust sensing of biomolecular interactions is developed by implementing time-correlated single photon counting in high-throughput conditions. Droplet microfluidics is used as a promising platform for the very fast handling of low-volume samples. We illustrate the potential of this very sensitive and cost-effective technology in the context of an enzymatic activity assay based on fluorescently-labeled biomolecules. Fluorescence lifetime detection by time-correlated single photon counting is shown to enable reliable discrimination between positive and negative control samples at a throughput as high as several hundred samples per second.

  10. Evidence for covalent binding of epicocconone with proteins from synchronous fluorescence spectra and fluorescence lifetimes

    Indian Academy of Sciences (India)

    Debashis Panda; Anindya Datta

    2007-03-01

    Synchronous fluorescence and time-resolved fluorescence spectroscopic studies that reveal the interaction of epicocconone with human serum albumin is significantly different from its interaction with surfactant assemblies. This observation, along with steady-state fluorescence data, indicates groundstate interaction between the fluorophore epicocconone and the protein. Similarity in fluorescence properties with the adduct of the fluorophore with -butylamine indicates that bonding occurs at the Nterminus of the protein.

  11. Lifetime

    Institute of Scientific and Technical Information of China (English)

    姚祎

    2004-01-01

    @@ 继ESPN刊出同名杂志之后,2003年赫斯特公司(Hearst Corp.)和迪斯尼(Walt Disney Co.)的合作促成了一本新杂志的诞生:(Lifetime),其目标读者是成百万收看同名有线电视网节目的妇女们.

  12. Online multispectral fluorescence lifetime values estimation and overlay onto tissue white-light video frames

    Science.gov (United States)

    Gorpas, Dimitris; Ma, Dinglong; Bec, Julien; Yankelevich, Diego R.; Marcu, Laura

    2016-03-01

    Fluorescence lifetime imaging has been shown to be a robust technique for biochemical and functional characterization of tissues and to present great potential for intraoperative tissue diagnosis and guidance of surgical procedures. We report a technique for real-time mapping of fluorescence parameters (i.e. lifetime values) onto the location from where the fluorescence measurements were taken. This is achieved by merging a 450 nm aiming beam generated by a diode laser with the excitation light in a single delivery/collection fiber and by continuously imaging the region of interest with a color CMOS camera. The interrogated locations are then extracted from the acquired frames via color-based segmentation of the aiming beam. Assuming a Gaussian profile of the imaged aiming beam, the segmentation results are fitted to ellipses that are dynamically scaled at the full width of three automatically estimated thresholds (50%, 75%, 90%) of the Gaussian distribution's maximum value. This enables the dynamic augmentation of the white-light video frames with the corresponding fluorescence decay parameters. A fluorescence phantom and fresh tissue samples were used to evaluate this method with motorized and hand-held scanning measurements. At 640x512 pixels resolution the area of interest augmented with fluorescence decay parameters can be imaged at an average 34 frames per second. The developed method has the potential to become a valuable tool for real-time display of optical spectroscopy data during continuous scanning applications that subsequently can be used for tissue characterization and diagnosis.

  13. Real-Time Visualization of Tissue Surface Biochemical Features Derived From Fluorescence Lifetime Measurements.

    Science.gov (United States)

    Gorpas, Dimitris; Ma, Dinglong; Bec, Julien; Yankelevich, Diego R; Marcu, Laura

    2016-08-01

    Fiber based fluorescence lifetime imaging has shown great potential for intraoperative diagnosis and guidance of surgical procedures. Here we describe a novel method addressing a significant challenge for the practical implementation of this technique, i.e., the real-time display of the quantified biochemical or functional tissue properties superimposed on the interrogated area. Specifically, an aiming beam (450 nm) generated by a continuous-wave laser beam was merged with the pulsed fluorescence excitation light in a single delivery/collection fiber and then imaged and segmented using a color-based algorithm. We demonstrate that this approach enables continuous delineation of the interrogated location and dynamic augmentation of the acquired frames with the corresponding fluorescence decay parameters. The method was evaluated on a fluorescence phantom and fresh tissue samples. Current results demonstrate that 34 frames per second can be achieved for augmenting videos of 640 × 512 pixels resolution. Also we show that the spatial resolution of the fluorescence lifetime map depends on the tissue optical properties, the scanning speed, and the frame rate. The dice similarity coefficient between the fluorescence phantom and the reconstructed maps was estimated to be as high as 93%. The reported method could become a valuable tool for augmenting the surgeon's field of view with diagnostic information derived from the analysis of fluorescence lifetime data in real-time using handheld, automated, or endoscopic scanning systems. Current method provides also a means for maintaining the tissue light exposure within safety limits. This study provides a framework for using an aiming beam with other point spectroscopy applications.

  14. A chloride ion nanosensor for time-resolved fluorimetry and fluorescence lifetime imaging.

    Science.gov (United States)

    Ruedas-Rama, Maria J; Orte, Angel; Hall, Elizabeth A H; Alvarez-Pez, Jose M; Talavera, Eva M

    2012-03-21

    In this work, the first CdSe/ZnS quantum dot (QD) photoluminescence lifetime based chloride ion nanosensor is reported. The acridinium dication lucigenin was self-assembled on the surface of negatively charged mercaptopropionic acid capped QDs to achieve QD-lucigenin conjugates. Upon attachment, a drastic decrease of the photoluminescence lifetime of both QD nanoparticles and lucigenin is observed by virtue of a charge transfer mechanism. Since lucigenin is a chloride-sensitive indicator dye, the photoluminescence decay of QD-lucigenin conjugates changes by adding chloride ion. The photoluminescence lifetime of the QDs in the conjugate increases after reacting with Cl(-), but also shows a concomitant decrease in the lucigenin lifetime immobilized on the surface. The photoluminescence lifetime of QD-lucigenin nanosensors shows a linear response in the Cl(-) concentration range between 0.5 and 50 mM. Moreover, the ratio τ(ave)(QD)/τ(ave)(luc) can be used as an analytical signal since the lifetime ratio presents a linear response in the same Cl(-) concentration range. The system also shows good selectivity towards most of the main anions and molecules that can be found in biological fluids. These nanosensors have been satisfactorily applied for Cl(-) determination in simulated intracellular media with high sensitivity and high selectivity. Finally, we demonstrate the potential application of the proposed nanosensor in confocal fluorescence lifetime imaging (FLIM). These results show the promising application of the QD-lucigenin nanosensors in FLIM, particularly for intracellular sensing, with the invaluable advantages of the time-resolved fluorescence techniques.

  15. Fluorescence lifetime imaging of endogenous molecules in live mouse cancer models (Conference Presentation)

    Science.gov (United States)

    Svindrych, Zdenek; Wang, Tianxiong; Hu, Song; Periasamy, Ammasi

    2017-02-01

    NADH and FAD are important endogenous fluorescent coenzymes participating in key enzymatic reactions of cellular metabolism. While fluorescence intensities of NADH and FAD have been used to determine the redox state of cells and tissues, this simple approach breaks down in the case of deep-tissue intravital imaging due to depth- and wavelength-dependent light absorption and scattering. To circumvent this limitation, our research focuses on fluorescence lifetimes of two-photon excited NADH and FAD emission to study the metabolic state of live tissues. In our custom-built scanning microscope we combine tunable femtosecond Ti:sapphire laser (operating at 740 nm for NADH excitation and 890 nm for FAD excitation), two GaAsP hybrid detectors for registering individual fluorescence photons and two Becker and Hickl time correlator boards for high precision lifetime measurements. Together with our rigorous FLIM analysis approach (including image segmentation, multi-exponential decay fitting and detailed statistical analysis) we are able to detect metabolic changes in cancer xenografts (human pancreatic cancer MPanc96 cells injected subcutaneously into the ear of an immunodeficient nude mouse), relative to surrounding healthy tissue. Advantageously, with the same instrumentation we can also take high-resolution and high-contrast images of second harmonic signal (SHG) originating from collagen fibers of both the healthy skin and the growing tumor. The combination of metabolic measurements (NADH and FAD lifetime) and morphological information (collagen SHG) allows us to follow the tumor growth in live mouse model and the changes in tumor microenvironment.

  16. Dynamic noninvasive monitoring of renal function in vivo by fluorescence lifetime imaging

    Science.gov (United States)

    Goiffon, Reece J.; Akers, Walter J.; Berezin, Mikhail Y.; Lee, Hyeran; Achilefu, Samuel

    2009-03-01

    Kidneys normally filter the blood of excess salts and metabolic products, such as urea, while retaining plasma proteins. In diseases such as multiple myeloma and diabetes mellitus, the renal function is compromised and protein escapes into the urine. In this study, we present the use of fluorescence lifetime imaging (FLI) to image excess serum protein in urine (proteinuria). The near-infrared fluorescent dye LS-288 has distinct lifetimes when bound to protein versus free in solution, providing contrast between the protein-rich viscera and the mostly protein-free bladder. FLI with LS-288 in mice revealed that fluorescence lifetime (FLT) differences in the bladder relative to surrounding tissues was due to the fractional contributions of the bound and unbound dye molecules. The FLT of LS-288 decreased in the case of proteinuria while fluorescence intensity was unchanged. The results show that FLI can be useful for the dynamic imaging of protein-losing nephropathy due to diabetes mellitus and other renal diseases and suggest the potential use of the FLI to distinguish tumors from fluid-filled cysts in the body.

  17. Protein-protein interaction analysis in single microfluidic droplets using FRET and fluorescence lifetime detection.

    Science.gov (United States)

    Benz, Christian; Retzbach, Heiko; Nagl, Stefan; Belder, Detlev

    2013-07-21

    Herein, we demonstrate the feasibility of a protein-protein interaction analysis and reaction progress monitoring in microfluidic droplets using FRET and microscopic fluorescence lifetime measurements. The fabrication of microdroplet chips using soft- and photolithographic techniques is demonstrated and the resulting chips reliably generate microdroplets of 630 pL and 6.71 nL at frequencies of 7.9 and 0.75 Hz, respectively. They were used for detection of protein-protein interactions in microdroplets using a model system of Alexa Fluor 488 labelled biotinylated BSA, Alexa Fluor 594 labelled streptavidin and unlabelled chicken egg white avidin. These microchips could be used for quantitative detection of avidin and streptavidin in microdroplets in direct and competitive assay formats with nanomolar detection limits, corresponding to attomole protein amounts. Four droplets were found to be sufficient for analytical determination. Fluorescence intensity ratio and fluorescence lifetime measurements were performed and compared for microdroplet FRET determination. A competitive on-chip binding assay for determination of unlabelled avidin using fluorescence lifetime detection could be performed within 135 s only.

  18. Two-photon excited fluorescence lifetime imaging and spectroscopy of melanins in vitro and in vivo

    Science.gov (United States)

    Krasieva, Tatiana B.; Stringari, Chiara; Liu, Feng; Sun, Chung-Ho; Kong, Yu; Balu, Mihaela; Meyskens, Frank L.; Gratton, Enrico; Tromberg, Bruce J.

    2013-03-01

    Changes in the amounts of cellular eumelanin and pheomelanin have been associated with carcinogenesis. The goal of this work is to develop methods based on two-photon-excited-fluorescence (TPEF) for measuring relative concentrations of these compounds. We acquire TPEF emission spectra (λex=1000 nm) of melanin in vitro from melanoma cells, hair specimens, and in vivo from healthy volunteers. We find that the pheomelanin emission peaks at approximately 615 to 625 nm and eumelanin exhibits a broad maximum at 640 to 680 nm. Based on these data we define an optical melanin index (OMI) as the ratio of fluorescence intensities at 645 and 615 nm. The measured OMI for the MNT-1 melanoma cell line is 1.6±0.22 while the Mc1R gene knockdown lines MNT-46 and MNT-62 show substantially greater pheomelanin production (OMI=0.5±0.05 and 0.17±0.03, respectively). The measured values are in good agreement with chemistry-based melanin extraction methods. In order to better separate melanin fluorescence from other intrinsic fluorophores, we perform fluorescence lifetime imaging microscopy of in vitro specimens. The relative concentrations of keratin, eumelanin, and pheomelanin components are resolved using a phasor approach for analyzing lifetime data. Our results suggest that a noninvasive TPEF index based on spectra and lifetime could potentially be used for rapid melanin ratio characterization both in vitro and in vivo.

  19. Fluorescence lifetime spectroscopy in multiple-scattering environments: an application to biotechnology

    Science.gov (United States)

    Cerussi, Albert E.; Gratton, Enrico; Fantini, Sergio

    1999-07-01

    Over the past few years, there has been significant research activity devoted to the application of fluorescence spectroscopy to strongly scattering media, where photons propagate diffusely. Much of this activity focused on fluorescence as a source of contrast enhancement in optical tomography. Our efforts have emphasized the quantitative recovery of fluorescence parameters for spectroscopy. Using a frequency-domain diffusion-based model, we have successfully recovered the lifetime, the absolute quantum yield, the fluorophore concentration, and the emission spectrum of the fluorophore, as well as the absorption and the reduced scattering coefficients at the emission wavelength of the medium in different measurements. In this contribution, we present a sensitive monitor of the binding between ethidium bromide and bovine cells in fresh milk. The spectroscopic contrast was the approximately tenfold increase in the ethidium bromide lifetime upon binding to DNA. The measurement clearly demonstrated that we could quantitatively measure the density of cells in the milk, which is an application vital to the tremendous economic burden of bovine subclinical mastitis detection. Furthermore, we may in principle use the spirit of this technique as a quantitative monitor of the binding of fluorescent drugs inside tissues. This is a first step towards lifetime spectroscopy in tissues.

  20. In vivo fluorescence lifetime imaging monitors binding of specific probes to cancer biomarkers.

    Directory of Open Access Journals (Sweden)

    Yasaman Ardeshirpour

    Full Text Available One of the most important factors in choosing a treatment strategy for cancer is characterization of biomarkers in cancer cells. Particularly, recent advances in Monoclonal Antibodies (MAB as primary-specific drugs targeting tumor receptors show that their efficacy depends strongly on characterization of tumor biomarkers. Assessment of their status in individual patients would facilitate selection of an optimal treatment strategy, and the continuous monitoring of those biomarkers and their binding process to the therapy would provide a means for early evaluation of the efficacy of therapeutic intervention. In this study we have demonstrated for the first time in live animals that the fluorescence lifetime can be used to detect the binding of targeted optical probes to the extracellular receptors on tumor cells in vivo. The rationale was that fluorescence lifetime of a specific probe is sensitive to local environment and/or affinity to other molecules. We attached Near-InfraRed (NIR fluorescent probes to Human Epidermal Growth Factor 2 (HER2/neu-specific Affibody molecules and used our time-resolved optical system to compare the fluorescence lifetime of the optical probes that were bound and unbound to tumor cells in live mice. Our results show that the fluorescence lifetime changes in our model system delineate HER2 receptor bound from the unbound probe in vivo. Thus, this method is useful as a specific marker of the receptor binding process, which can open a new paradigm in the "image and treat" concept, especially for early evaluation of the efficacy of the therapy.

  1. Efficacy of photodynamic therapy against larvae of Aedes aegypti: confocal microscopy and fluorescence-lifetime imaging

    Science.gov (United States)

    de Souza, L. M.; Pratavieira, S.; Inada, N. M.; Kurachi, C.; Corbi, J.; Guimarães, F. E. G.; Bagnato, V. S.

    2014-03-01

    Recently a few demonstration on the use of Photodynamic Reaction as possibility to eliminate larvae that transmit diseases for men has been successfully demonstrated. This promising tool cannot be vastly used due to many problems, including the lake of investigation concerning the mechanisms of larvae killing as well as security concerning the use of photosensitizers in open environment. In this study, we investigate some of the mechanisms in which porphyrin (Photogem) is incorporated on the Aedes aegypti larvae previously to illumination and killing. Larvae at second instar were exposed to the photosensitizer and after 30 minutes imaged by a confocal fluorescence microscope. It was observed the presence of photosensitizer in the gut and at the digestive tract of the larva. Fluorescence-Lifetime Imaging showed greater photosensitizer concentration in the intestinal wall of the samples, which produces a strong decrease of the Photogem fluorescence lifetime. For Photodynamic Therapy exposition to different light doses and concentrations of porphyrin were employed. Three different light sources (LED, Fluorescent lamp, Sun light) also were tested. Sun light and fluorescent lamp shows close to 100% of mortality after 24 hrs. of illumination. These results indicate the potential use of photodynamic effect against the LARVAE of Aedes aegypti.

  2. High-Speed Fluorescence Microscopy: Lifetime Imaging in the Biomedical Sciences

    Science.gov (United States)

    Periasamy, Ammasi; Wang, Xue F.; Wodnick, Pawel; Gordon, Gerald W.; Kwon, Seongwook; Diliberto, Pamela A.; Herman, Brian

    1995-02-01

    The ability to observe the behavior of living cells and tissues provides unparalleled access to information regarding the organization and dynamics of complex cellular structures. While great strides have been made over the past 30 to 40 years in the design and application of a variety of novel optical microscopic techniques, until recently, it has not been possible to image biological phenomena that occur over very short time periods (nanosecond to millisecond) or over short distances (10 to 1000 [Angstrom capital A, ring]). However, the recent combination of (1) very rapidly gated and sensitive image intensifiers and (2) the ability to deliver fluorescence excitation energy to intact living biological specimens in a pulsed or sinusoidally modulated fashion has allowed such measurements to become a reality through the imaging of the lifetimes of fluorescent molecules. This capability has resulted in the ability to observe the dynamic organization and interaction of cellular components on a spatial and temporal scale previously not possible using other microscopic techniques. This paper discusses the implementation of a fluorescence lifetime imaging microscope (FLIM) and provides a review of some of the applications of such an instrument. These include measurements of receptor topography and subunit interactions using fluorescence resonance energy transfer (FRET), fluorescence anisotropy of phospholipids in cell membranes, cytosolic free calcium (Ca2+)i and the detection of human papillomavirus (HPV) infection in clinical cervicovaginal smears.

  3. Fluorescence lifetime measurement with confocal endomicroscopy for direct analysis of tissue biochemistry in vivo

    Directory of Open Access Journals (Sweden)

    Youngjae Won

    2016-08-01

    Full Text Available Confocal endomicroscopy is a powerful tool for in vivo real-time imaging at cellular resolution inside a living body without tissue resection. Microscopic fluorescence lifetime measurement can provide information about localized biochemical conditions such as pH and the concentrations of oxygen and calcium. We hypothesized that combining these techniques could assist accurate cancer discrimination by providing both biochemical and morphological information. We designed a dual-mode experimental setup for confocal endomicroscopic imaging and fluorescence lifetime measurement and applied it to a mouse xenograft model of activated human pancreatic cancer generated by subcutaneous injection of AsPC-1 tumor cells. Using this method with pH-sensitive sodium fluorescein injection, we demonstrated discrimination between normal and cancerous tissues in a living mouse. With further development, this method may be useful for clinical cancer detection.

  4. Use of Fluorescence Lifetime Imaging Microscopy (FLIM) as a Timer of Cell Cycle S Phase.

    Science.gov (United States)

    Okkelman, Irina A; Dmitriev, Ruslan I; Foley, Tara; Papkovsky, Dmitri B

    2016-01-01

    Incorporation of thymidine analogues in replicating DNA, coupled with antibody and fluorophore staining, allows analysis of cell proliferation, but is currently limited to monolayer cultures, fixed cells and end-point assays. We describe a simple microscopy imaging method for live real-time analysis of cell proliferation, S phase progression over several division cycles, effects of anti-proliferative drugs and other applications. It is based on the prominent (~ 1.7-fold) quenching of fluorescence lifetime of a common cell-permeable nuclear stain, Hoechst 33342 upon the incorporation of 5-bromo-2'-deoxyuridine (BrdU) in genomic DNA and detection by fluorescence lifetime imaging microscopy (FLIM). We show that quantitative and accurate FLIM technique allows high-content, multi-parametric dynamic analyses, far superior to the intensity-based imaging. We demonstrate its uses with monolayer cell cultures, complex 3D tissue models of tumor cell spheroids and intestinal organoids, and in physiological study with metformin treatment.

  5. Hardware-friendly bi-exponential fluorescence lifetime imaging algorithms and phasor approaches

    Science.gov (United States)

    Li, David; Chen, Yu

    2015-07-01

    A newly developed hardware-friendly non-iterative fluorescence lifetime imaging (FLIM) analysis method was verified in an FPGA chip. Its performances were also demonstrated on two-photon FLIM images of gold nanorods (GNRs)-Cy5 labelled Hela cells. The results obtained by the proposed method can be presented in a polor plot to be compared to the widely used phasor (Phasor) approach. Combining our method with Phasor will be very useful in FLIM analysis.

  6. Three-dimensional printed miniaturized spectral system for collagen fluorescence lifetime measurements

    Science.gov (United States)

    Zou, Luwei; Koslakiewicz, Ronald; Mahmoud, Mohamad; Fahs, Mehdi; Liu, Rui; Lo, Joe Fujiou

    2016-07-01

    Various types of collagens, e.g., type I and III, represent the main load-bearing components in biological tissues. Their composition changes during processes such as wound healing and fibrosis. When excited by ultraviolet light, collagens exhibit autofluorescence distinguishable by their unique fluorescent lifetimes across a range of emission wavelengths. Here, we designed a miniaturized spectral-lifetime detection system as a noninvasive probe for monitoring tissue collagen compositions. A sine-modulated LED illumination was applied to enable frequency domain fluorescence lifetime measurements under three wavelength bands, separated via a series of longpass dichroics at 387, 409, and 435 nm. We employed a lithography-based three-dimensional (3-D) printer with modeling to simulate the effect of thermal (from LED) and mechanical (from handling) strain on the optical system. The geometry was further optimized with ray tracing to form the final 3-D printed structure. Using this device, the phase shift and demodulation of collagen types were measured, where the separate spectral bands enhanced the differentiation of their lifetimes. This system represents a low cost, handheld probe for clinical tissue monitoring applications.

  7. A comparative study of metabolic state of stem cells during osteogenic and adipogenic differentiations via fluorescence lifetime imaging microscopy

    Science.gov (United States)

    Chakraborty, Sandeep; Ou, Meng-Hsin; Kuo, Jean-Cheng; Chiou, Arthur

    2016-10-01

    Cellular metabolic state can serve as a biomarker to indicate the differentiation potential of stem cells into other specialized cell lineages. In this study, two-photon fluorescence lifetime imaging microscopy (2P-FLIM) was applied to determine the fluorescence lifetime and the amounts of the auto-fluorescent metabolic co-factor reduced nicotinamide adenine dinucleotide (NADH) to elucidate the cellular metabolism of human mesenchymal stem cells (hMSCs) in osteogenic and adipogenic differentiation processes. 2P-FLIM provides the free to protein-bound NADH ratio which can serve as the indicator of cellular metabolic state. We measured NADH fluorescence lifetime at 0, 7, and 14 days after hMSCs were induced for either osteogenesis or adipogenesis. In both cases, the average fluorescence lifetime increased significantly at day 14 (P stem cells into other specialized cell lineages.

  8. Diagnosis of basal cell carcinoma by two photon excited fluorescence combined with lifetime imaging

    Science.gov (United States)

    Fan, Shunping; Peng, Xiao; Liu, Lixin; Liu, Shaoxiong; Lu, Yuan; Qu, Junle

    2014-02-01

    Basal cell carcinoma (BCC) is the most common type of human skin cancer. The traditional diagnostic procedure of BCC is histological examination with haematoxylin and eosin staining of the tissue biopsy. In order to reduce complexity of the diagnosis procedure, a number of noninvasive optical methods have been applied in skin examination, for example, multiphoton tomography (MPT) and fluorescence lifetime imaging microscopy (FLIM). In this study, we explored two-photon optical tomography of human skin specimens using two-photon excited autofluorescence imaging and FLIM. There are a number of naturally endogenous fluorophores in skin sample, such as keratin, melanin, collagen, elastin, flavin and porphyrin. Confocal microscopy was used to obtain structures of the sample. Properties of epidermic and cancer cells were characterized by fluorescence emission spectra, as well as fluorescence lifetime imaging. Our results show that two-photon autofluorescence lifetime imaging can provide accurate optical biopsies with subcellular resolution and is potentially a quantitative optical diagnostic method in skin cancer diagnosis.

  9. Temperature and bath gas composition dependence of effective fluorescence lifetimes of toluene excited at 266 nm

    Science.gov (United States)

    Faust, S.; Dreier, T.; Schulz, C.

    2011-05-01

    Time-resolved fluorescence spectra of gas-phase toluene upon picosecond excitation at 266 nm were investigated as a function of temperature (296-1074 K) and bath gas composition (varying amounts of N 2, O 2, and CO 2) at 1 bar total pressure with a temporal resolution of 50 ps. In the investigated temperature range the effective fluorescence lifetime drops with increasing temperature from 46 ± 3 ns to 0.05 ± 0.01 ns in N 2 and CO 2. In the presence of O 2 at constant temperature the lifetimes also decrease significantly (e.g., from 46 ± 3 ns without O 2 to 0.63 ± 0.05 ns in air at room temperature), whereas lifetimes are independent on the CO 2 concentration. The implications of the results for the existing phenomenological model of predicting temporally integrated fluorescence intensities in toluene [W. Koban, J.D. Koch, R.K. Hanson, C. Schulz, Appl. Phys. B 80 (2005) 777] are discussed.

  10. 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.

  11. Silica nanodisks as platforms for fluorescence lifetime-based sensing of pH

    Indian Academy of Sciences (India)

    Subhasree Banerjee; Anjali Dhir; Tuseeta Banerjee; Avinash Kumar Singh; Anindya Datta

    2011-11-01

    Core-shell conjugates of silica nanodisks and fluorescent dyes have been prepared. Rhodamine B, the reference, has been attached to the core, by surface functionalization of the pristine SNDs. Then, a layer of silica has been deposited on the composite nanodisks. Finally, the surface has been functionalized with fluorescein in one case and protoporphyrin IX in the other. These dyes exhibit pH-dependent fluorescence properties. The nanoconjugates are found to sense the pH of the medium, through systematic variation of the fluorescence intensity ratios of the reporter dye at the surface and the reference dye at the core. Moreover, the fluorescence lifetimes and corresponding amplitudes of the reporter dyes have been found to be reliable parameters for assessing the pH of the medium, even though the variation in lifetimes of fluorescein is rather small. In case of protoporphyrin, however, this variation is significantly large. Besides, the change in amplitudes is prominent in acidic as well as alkaline solutions. The temporal parameters can thus be used to ascertain the pH of the medium, when used in conjunction with each other.

  12. Fluorescence intensity and lifetime-based cyanide sensitive probes for physiological safeguard

    Energy Technology Data Exchange (ETDEWEB)

    Badugu, Ramachandram [Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, Medical Biotechnology Center, University of Maryland School of Medicine, 725 West Lombard Street, Baltimore, MD 21201 (United States); Lakowicz, Joseph R. [Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, Medical Biotechnology Center, University of Maryland School of Medicine, 725 West Lombard Street, Baltimore, MD 21201 (United States)]. E-mail: lakowicz@cfs.umbi.umd.edu; Geddes, Chris D. [Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, Medical Biotechnology Center, University of Maryland School of Medicine, 725 West Lombard Street, Baltimore, MD 21201 (United States) and Institute of Fluorescence and Center for Fluorescence Spectroscopy, Medical Biotechnology Center, University of Maryland Biotechnology Institute, 725 West Lombard Street, Baltimore, MD 21201 (United States)]. E-mail: chris@cfs.umbi.umd.edu

    2004-09-20

    We characterize six new fluorescent probes that show both intensity and lifetime changes in the presence of free uncomplexed aqueous cyanide, allowing for fluorescence based cyanide sensing up to physiological safeguard levels, i.e. <30 {mu}M. One of the probes, m-BMQBA, shows a {approx}15-fold reduction in intensity and a {approx}10% change in mean lifetime at this level. The response of the new probes is based on their ability to bind the cyanide anion through a boronic acid functional group, changing from the neutral form of the boronic acid group R-B(OH){sub 2} to the anionic R-B{sup -}(CN){sub 3} form, a new cyanide binding mechanism which we have recently reported. The presence of an electron deficient quaternary heterocyclic nitrogen nucleus, and the electron rich cyanide bound form, provides for the intensity changes observed. We have determined the disassociation constants of the probes to be in the range {approx}15-84 {mu}M{sup 3}. In addition we have synthesized control compounds which do not contain the boronic acid moiety, allowing for a rationale of the cyanide responses between the probe isomers to be made. The lifetime of the cyanide bound probes are significantly shorter than the free R-B(OH){sub 2} probe forms, providing for the opportunity of lifetime based cyanide sensing up to physiologically lethal levels. Finally, while fluorescent probes containing the boronic acid moiety have earned a well-deserved reputation for monosaccharide sensing, we show that strong bases such as CN{sup -} and OH{sup -} preferentially bind as compared to glucose, enabling the potential use of these probes for cyanide safeguard and determination in physiological fluids, especially given that physiologies do not experience any notable changes in pH.

  13. Time-resolved microspectrofluorometry and fluorescence lifetime imaging of photosensitizers using picosecond pulsed diode lasers in laser scanning microscopes.

    Science.gov (United States)

    Kress, Matthias; Meier, Thomas; Steiner, Rudolf; Dolp, Frank; Erdmann, Rainer; Ortmann, Uwe; Rück, Angelika

    2003-01-01

    This work describes the time-resolved fluorescence characteristics of two different photosensitizers in single cells, in detail mTHPC and 5-ALA induced PPIX, which are currently clinically used in photodynamic therapy. The fluorescence lifetime of the drugs was determined in the cells from time-gated spectra as well as single photon counting, using a picosecond pulsed diode laser for fluorescence excitation. The diode laser, which emits pulses at 398 nm with 70 ps full width at half maximum duration, was coupled to a confocal laser scanning microscope. For time-resolved spectroscopy a setup consisting of a Czerny Turner spectrometer and a MCP-gated and -intensified CCD camera was used. Time-gated spectra within the cells were acquired by placing the laser beam in "spot scan" mode. In addition, a time-correlated single photon counting module was used to determine the fluorescence lifetime from single spots and to record lifetime images. The fluorescence lifetime of mTHPC decreased from 7.5 to 5.5 ns during incubation from 1 to 6 h. This decrease was probably attributed to enhanced formation of aggregates during incubation. Fluorescence lifetime imaging showed that longer lifetimes were correlated with accumulation in the cytoplasm in the neighborhood of the cell nucleus, whereas shorter lifetimes were found in the outer cytoplasm. For cells that were incubated with 5-ALA, a fluorescence lifetime of 7.4 ns was found for PPIX; a shorter lifetime at 3.6 ns was probably attributed to photoproducts and aggregates of PPIX. In contrast from fluorescence intensity images alone, different fluorescence species could not be distinguished. However, in the lifetime image a structured fluorescence distribution in the cytoplasm was correlated with the longer lifetime and probably coincides with mitochondria. In conclusion, picosecond diode lasers coupled to a laser scanning microscope equipped with appropriate detection units allows time-resolved spectroscopy and lifetime imaging

  14. Relationship between the Fluorescence Lifetime of Chlorophyll 'a' and Primary Productivity within the Mississippi River Plume and Adjacent Shelf Region

    Science.gov (United States)

    Hall, Callie; Miller, Richard L.; Fernandez, Salvador M.; McKee, Brent A.

    2000-01-01

    In situ measurements of chlorophyll fluorescence intensity have been widely used to estimate phytoplankton biomass. However, because the fluorescence quantum yield of chlorophyll a in vivo can be highly variable, measurements of chlorophyll fluorescence intensity cannot be directly correlated with phytoplankton biomass and do not provide information on the physiological state of the phytoplankton under study. Conversely, lifetime-based measurements of chlorophyll fluorescence provide a framework in which photosynthetic rates of phytoplankton can be analyzed according to phytoplankton physiology. Along with the measurement of primary production and ambient nutrient concentrations within the Mississippi River plume in the northern Gulf of Mexico, phytoplankton fluorescence lifetimes were measured using a Fluorescence Lifetime Phytoplankton Analyzer (developed under a NASA Small Business Innovative Research contract to Ciencia, Inc.). Variability of fluorescence lifetimes within the plume can be used as a background from which to interpret variations in the maximum quantum yield of photochemistry. The extent to which nutrient and effluent loading in this dynamic coastal area affect the photosynthetic performance of phytoplankton will be presented as a function of phytoplankton fluorescence lifetimes.

  15. Relationship between the Fluorescence Lifetime of Chlorophyll 'a' and Primary Productivity within the Mississippi River Plume and Adjacent Shelf Region

    Science.gov (United States)

    Hall, Callie; Miller, Richard L.; Fernandez, Salvador M.; McKee, Brent A.

    2000-01-01

    In situ measurements of chlorophyll fluorescence intensity have been widely used to estimate phytoplankton biomass. However, because the fluorescence quantum yield of chlorophyll a in vivo can be highly variable, measurements of chlorophyll fluorescence intensity cannot be directly correlated with phytoplankton biomass and do not provide information on the physiological state of the phytoplankton under study. Conversely, lifetime-based measurements of chlorophyll fluorescence provide a framework in which photosynthetic rates of phytoplankton can be analyzed according to phytoplankton physiology. Along with the measurement of primary production and ambient nutrient concentrations within the Mississippi River plume in the northern Gulf of Mexico, phytoplankton fluorescence lifetimes were measured using a Fluorescence Lifetime Phytoplankton Analyzer (developed under a NASA Small Business Innovative Research contract to Ciencia, Inc.). Variability of fluorescence lifetimes within the plume can be used as a background from which to interpret variations in the maximum quantum yield of photochemistry. The extent to which nutrient and effluent loading in this dynamic coastal area affect the photosynthetic performance of phytoplankton will be presented as a function of phytoplankton fluorescence lifetimes.

  16. Advances in frequency-domain fluorometry, gigahertz instrumentation, time-dependent photomigration, and fluorescence lifetime imaging

    Science.gov (United States)

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

    1992-02-01

    During the past seven years, there have been remarkable advances in the frequency-domain method for measurement of time-resolved emission or light scattering. In this presentation we describe the recent extension of the frequency range to 10 GHz using a specially designed microchannel plate PMT. Experimental data will be shown for measurement of picosecond rotational diffusion and for sub-picosecond resolution of time delays. The resolution of ps to ns timescale processes is not obtained at the expense of sensitivity or is it shown by measurements on the intrinsic tryptophan emission from hemoglobin. We also describe a time- resolved reflectance imaging experiment on a scattering medium containing an absorbing object. Time-resolved imaging of the back-scattered light is realized by means of a RF-phase- sensitive camera, synchronized to the laser pulses. By processing the stored images, a final image can be created, the contrast of which is based only on time differences of the back- scattered photons. This image reveals the presence and position of the absorber within the scattering medium. And finally, we describe a new methodology, fluorescence lifetime imaging (FLIM), in which the contrast depends on the fluorescence lifetime at each point in a two-dimensional image, and not 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. FLIM has numerous potential applications in cell biology and imaging.

  17. Compressive hyperspectral time-resolved wide-field fluorescence lifetime imaging

    Science.gov (United States)

    Pian, Qi; Yao, Ruoyang; Sinsuebphon, Nattawut; Intes, Xavier

    2017-07-01

    Spectrally resolved fluorescence lifetime imaging and spatial multiplexing have offered information content and collection-efficiency boosts in microscopy, but efficient implementations for macroscopic applications are still lacking. An imaging platform based on time-resolved structured light and hyperspectral single-pixel detection has been developed to perform quantitative macroscopic fluorescence lifetime imaging (MFLI) over a large field of view (FOV) and multiple spectral bands simultaneously. The system makes use of three digital micromirror device (DMD)-based spatial light modulators (SLMs) to generate spatial optical bases and reconstruct N by N images over 16 spectral channels with a time-resolved capability (∼40 ps temporal resolution) using fewer than N2 optical measurements. We demonstrate the potential of this new imaging platform by quantitatively imaging near-infrared (NIR) Förster resonance energy transfer (FRET) both in vitro and in vivo. The technique is well suited for quantitative hyperspectral lifetime imaging with a high sensitivity and paves the way for many important biomedical applications.

  18. Differentiation of ocular fundus fluorophores by fluorescence lifetime imaging using multiple excitation and emission wavelengths

    Science.gov (United States)

    Hammer, M.; Schweitzer, D.; Schenke, S.; Becker, W.; Bergmann, A.

    2006-10-01

    Ocular fundus autofluorescence imaging has been introduced into clinical diagnostics recently. It is in use for the observation of the age pigment lipofuscin, a precursor of age - related macular degeneration (AMD). But other fluorophores may be of interest too: The redox pair FAD - FADH II provides information on the retinal energy metabolism, advanced glycation end products (AGE) indicate protein glycation associated with pathologic processes in diabetes as well as AMD, and alterations in the fluorescence of collagen and elastin in connective tissue give us the opportunity to observe fibrosis by fluorescence imaging. This, however, needs techniques able to differentiate particular fluorophores despite limited permissible ocular exposure as well as excitation wavelength (limited by the transmission of the human ocular lens to >400 nm). We present an ophthalmic laser scanning system (SLO), equipped with picosecond laser diodes (FWHM 100 ps, 446 nm or 468 nm respectively) and time correlated single photon counting (TCSPC) in two emission bands (500 - 560 nm and 560 - 700 nm). The decays were fitted by a bi-exponential model. Fluorescence spectra were measured by a fluorescence spectrometer fluorolog. Upon excitation at 446 nm, the fluorescence of AGE, FAD, and lipofuscin were found to peak at 503 nm, 525 nm, and 600 nm respectively. Accordingly, the statistical distribution of the fluorescence decay times was found to depend on the different excitation wavelengths and emission bands used. The use of multiple excitation and emission wavelengths in conjunction with fluorescence lifetime imaging allows us to discriminate between intrinsic fluorophores of the ocular fundus. Taken together with our knowledge on the anatomical structure of the fundus, these findings suggest an association of the short, middle and long fluorescence decay time to the retinal pigment epithelium, the retina, and connective tissue respectively.

  19. Fluorescence lifetime technique for surgical imaging, guidance and augmented reality (Conference Presentation)

    Science.gov (United States)

    Marcu, Laura

    2017-02-01

    The surgeon's limited ability to accurately delineate the tumor margin during surgical interventions is one key challenge in clinical management of cancer. New methods for guiding tumor resection decisions are needed. Numerous studies have shown that tissue autofluorescence properties have the potential to asses biochemical features associates with distinct pathologies in tissue and to distinguish various cancers from normal tissues. However, despite these promising reports, autofluorescence techniques were sparsely adopted in clinical settings. Moreover, when adopted they were primarily used for pre-operative diagnosis rather than guiding interventions. To address this need, we have researched and engineered instrumentation that utilizes label-free fluorescence lifetime contrast to characterize tissue biochemical features in vivo in patients and methodologies conducive to real-time (few seconds) diagnosis of tissue pathologies during surgical procedures. This presentation overviews clinically-compatible multispectral fluorescence lifetime imaging techniques developed in our laboratory and their ability to operate as stand-alone tools, integrated in a biopsy needle and in conjunction with the da Vinci surgical robot. We present pre-clinical and clinical studies in patients that demonstrate the potential of these techniques for intraoperative assessment of brain tumors and head and neck cancer. Current results demonstrate that intrinsic fluorescence signals can provide useful contrast for delineation distinct types of tissues including tumors intraoperatively. Challenges and solutions in the clinical implementation of these techniques are discussed.

  20. Studying Biological Tissue with Fluorescence Lifetime Imaging: Microscopy, Endoscopy, and Complex Decay Profiles

    Science.gov (United States)

    Siegel, Jan; Elson, Daniel S.; Webb, Stephen E. D.; Lee, K. C. Benny; Vlandas, Alexis; Gambaruto, Giovanni L.; Léveque-Fort, Sandrine; Lever, M. John; Tadrous, Paul J.; Stamp, Gordon W. H.; Wallace, Andrew L.; Sandison, Ann; Watson, Tim F.; Alvarez, Fernando; French, Paul M. W.

    2003-06-01

    We have applied fluorescence lifetime imaging (FLIM) to the autofluorescence of different kinds of biological tissue in vitro , including animal tissue sections and knee joints as well as human teeth, obtaining two-dimensional maps with functional contrast. We find that fluorescence decay profiles of biological tissue are well described by the stretched exponential function (StrEF), which can represent the complex nature of tissue. The StrEF yields a continuous distribution of fluorescence lifetimes, which can be extracted with an inverse Laplace transformation, and additional information is provided by the width of the distribution. Our experimental results from FLIM microscopy in combination with the StrEF analysis indicate that this technique is ready for clinical deployment, including portability that is through the use of a compact picosecond diode laser as the excitation source. The results obtained with our FLIM endoscope successfully demonstrated the viability of this modality, though they need further optimization. We expect a custom-designed endoscope with optimized illumination and detection efficiencies to provide significantly improved performance.

  1. Nanoscale fluorescence lifetime imaging of an optical antenna with a single diamond NV center.

    Science.gov (United States)

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

    2013-08-14

    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.

  2. Determining a fluorophore's transition dipole moment from fluorescence lifetime measurements in solvents of varying refractive index.

    Science.gov (United States)

    Chung, Pei-Hua; Tregidgo, Carolyn; Suhling, Klaus

    2016-11-11

    The transition dipole moment of organic dyes PM546 and rhodamine 123 is calculated from fluorescence lifetime measurements in solutions of different refractive index. A model proposed by Toptygin et al (2002 J. Phys. Chem. B 106 3724-34) provides a relationship between the radiative rate constant and refractive index of the solvent, and allows the electronic transition dipole moments to be found: it is (7.1  ±  1.1) D for PM546 which matches that found in the literature, and (8.1  ±  0.1) D for rhodamine 123. Toptygin's model goes further in predicting the shape of the fluorescent dye and here we predict the shape of PM546 and rhodamine 123 to be ellipsoidal.

  3. Spectral and lifetime fluorescence imaging microscopies: new modalities of multiphoton microscopy applied to tissue or cell engineering.

    Science.gov (United States)

    Dumas, D; Gaborit, N; Grossin, L; Riquelme, B; Gigant-Huselstein, C; De Isla, N; Gillet, P; Netter, P; Stoltz, J F

    2004-01-01

    Spectral and multiphoton imaging is the preferred approach for non-invasive study allowing deeper penetration to image molecular processes in living cells. But currently available fluorescence microscopic techniques based on fluorescence intensity, such as confocal or multiphoton excitation, cannot provide detailed quantitative information about the dynamic of complex cellular structure (molecular interaction). Due to the variation of the probe concentration, photostability, cross-talking, its effects cannot be distinguished in simple intensity images. Therefore, Time Resolved fluorescence image is required to investigate molecular interactions in biological systems. Fluorescence lifetimes are generally absolute, sensitive to environment, independent of the concentration of the probe and allow the use of probes with overlapping spectra but that not have the same fluorescence lifetime. In this work, we present the possibilities that are opened up by Fluorescence Lifetime Imaging Microscopy, firstly to collect images based on fluorescence lifetime contrast of GFP variants used as a reporter of gene expression in chondrocytes and secondly, to measure molecular proximity in erythrocyte (glycophorin/membrane) by Fluorescence Resonance Energy Transfer (FLIM-FRET).

  4. Center for Fluorescence Spectroscopy: advanced studies of fluorescence dynamics, lifetime imaging, clinical sensing, two-photon excitation, and light quenching

    Science.gov (United States)

    Lakowicz, Joseph R.; Malak, Henryk M.; Gryczynski, Ignacy; Szmacinski, Henryk; Kusba, Jozef; Akkaya, Engin; Terpetschnig, Ewald A.; Johnson, Michael L.

    1994-08-01

    The Center for Fluorescence Spectroscopy (CFS) is a multi-user facility providing state of the art time-resolved fluorescence instrumentation and software for scientists, whose research can be enhanced by such experimental data. The CFS is a national center, supported by the National Center for Research Resources Division of the National Institutes of Health, and in part by the National Science Foundation. Both time-domain (TD) and frequency- domain (FD) measurements (10 MHz to 10 Ghz) are available, with a wide range of excitation and emission wavelengths (UV to NIR). The data can be used to recover distances and site-to-site diffusion in protein, interactions between macromolecules, accessibility of fluorophores to quenchers, and the dynamic properties of proteins, membranes and nucleic acids. Current software provides for analysis of multi-exponential intensity and anisotropy decays, lifetime distribution, distance distributions for independent observation of fluorescence donors and acceptors, transient effects in collisional quenching, phase-modulation spectra and time-resolved emission spectra. Most programs provide for global analysis of multiple data sets obtained under similar experimental conditions. Data can be analyzed on-site by connection with the CFS computers through the internet. During six years of operation we have established scientific collaborations with over 30 academic and industrial groups in the United States. These collaborations have resulted in 63 scientific papers.

  5. New insights in the interpretation of tryptophan fluorescence : origin of the fluorescence lifetime and characterization of a new fluorescence parameter in proteins: the emission to excitation ratio.

    Science.gov (United States)

    Albani, J R

    2007-07-01

    Origin of tryptophan fluorescence is still up to these days a quiz which is not completely solved. Fluorescence emission properties of tryptophan within proteins are in general considered as the result of fluorophore interaction within its environment. For example, a low fluorescence quantum yield is supposed to be the consequence of an important fluorophore-environment interaction. However, are we sure that the fluorophore has been excited upon light absorption? What if fluorophore excitation did not occur as the result of internal conformation specific to the fluorophore environment? Are we sure that all absorbed energy is used for the excitation process? Fluorescence lifetimes of Trp residues are considered to originate from rotamers or conformers resulting from the rotation of the indole ring within the peptide bonds. However, how can we explain the fact that in most of the proteins, the two lifetimes 0.5 and 3 ns, attributed to the conformers, are also observed for free tryptophan in solution? The present work, performed on free tryptophan and tyrosine in solution and on different proteins, shows that absorption and excitation spectra overlap but their intensities at the different excitation wavelengths are not necessarily equal. Also, we found that fluorescence emission intensities recorded at different excitation wavelengths depend on the intensities at these excitation wavelengths and not on the optical densities. Thus, excitation is not equal to absorption. In our interpretation of the data, we consider that absorbed photons are not necessary used only for the excitation, part of them are used to reorganize fluorophore molecules in a new state (excited structure) and another part is used for the excitation process. A new parameter that characterizes the ratio of the number of emitted photons over the real number of photons used to excite the fluorophore can be defined. We call this parameter, the emission to excitation ratio. Since our results were

  6. Measurement of pH micro-heterogeneity in natural cheese matrices by fluorescence lifetime imaging

    Directory of Open Access Journals (Sweden)

    Zuzana eBurdikova

    2015-03-01

    Full Text Available Cheese, a product of microbial fermentation may be defined as a protein matrix entrapping fat, moisture, minerals and solutes as well as dispersed bacterial colonies. The growth and physiology of bacterial cells in these colonies may be influenced by the microenvironment around the colony, or alternatively the cells within the colony may modify the microenvironment (e.g. pH, redox potential due to their metabolic activity. While cheese pH may be measured at macro level there remains a significant knowledge gap relating to the degree of micro-heterogeneity of pH within the cheese matrix and its relationship with microbial, enzymatic and physiochemical parameters and ultimately with cheese quality, consistency and ripening patterns. The pH of cheese samples was monitored both at macroscopic scale and at microscopic scale, using a non-destructive microscopic technique employing C-SNARF-4 and Oregon Green 488 fluorescent probes. The objectives of this work were to evaluate the suitability of these dyes for microscale pH measurements in natural cheese matrices and to enhance the sensitivity and extend the useful pH range of these probes using fluorescence lifetime imaging (FLIM. In particular, fluorescence lifetime of Oregon Green 488 proved to be sensitive probe to map pH micro heterogeneity within cheese matrices. Good agreement was observed between macroscopic scale pH measurement by FLIM and by traditional pH methods, but in addition considerable localized microheterogeneity in pH was evident within the curd matrix with pH range between 4.0 and 5.5. This technique provides significant potential to further investigate the relationship between cheese matrix physico-chemistry and bacterial metabolism during cheese manufacture and ripening.

  7. Measurement of pH micro-heterogeneity in natural cheese matrices by fluorescence lifetime imaging.

    Science.gov (United States)

    Burdikova, Zuzana; Svindrych, Zdenek; Pala, Jan; Hickey, Cian D; Wilkinson, Martin G; Panek, Jiri; Auty, Mark A E; Periasamy, Ammasi; Sheehan, Jeremiah J

    2015-01-01

    Cheese, a product of microbial fermentation may be defined as a protein matrix entrapping fat, moisture, minerals and solutes as well as dispersed bacterial colonies. The growth and physiology of bacterial cells in these colonies may be influenced by the microenvironment around the colony, or alternatively the cells within the colony may modify the microenvironment (e.g., pH, redox potential) due to their metabolic activity. While cheese pH may be measured at macro level there remains a significant knowledge gap relating to the degree of micro-heterogeneity of pH within the cheese matrix and its relationship with microbial, enzymatic and physiochemical parameters and ultimately with cheese quality, consistency and ripening patterns. The pH of cheese samples was monitored both at macroscopic scale and at microscopic scale, using a non-destructive microscopic technique employing C-SNARF-4 and Oregon Green 488 fluorescent probes. The objectives of this work were to evaluate the suitability of these dyes for microscale pH measurements in natural cheese matrices and to enhance the sensitivity and extend the useful pH range of these probes using fluorescence lifetime imaging (FLIM). In particular, fluorescence lifetime of Oregon Green 488 proved to be sensitive probe to map pH micro heterogeneity within cheese matrices. Good agreement was observed between macroscopic scale pH measurement by FLIM and by traditional pH methods, but in addition considerable localized microheterogeneity in pH was evident within the curd matrix with pH range between 4.0 and 5.5. This technique provides significant potential to further investigate the relationship between cheese matrix physico-chemistry and bacterial metabolism during cheese manufacture and ripening.

  8. Fluorescence lifetimes of tryptophan: structural origin and relation with So --> 1Lb and So --> 1La transitions.

    Science.gov (United States)

    Albani, Jihad René

    2009-11-01

    We measured fluorescence lifetimes of L-Tryptophan dissolved in de-ionized water and in ethanol in the absence and the presence of high progesterone concentrations. The hormone absorbs between 220 and 280 with a peak around 250 nm, while its absorption is equal to zero beyond 280 nm. Tryptophan excitation spectrum recorded in presence of progesterone shows that the S(o) --> 1L(a) transition is completely abolished while the S(o) --> 1L(b) transition is not affected. Emission of L-tryptophan in water occurs with two fluorescence lifetimes, 0.40 and 2.8 ns. In ethanol, three fluorescence lifetimes equal to around 0.2, 1.8 and 4.8 ns were observed. Addition of progesterone to the medium does not affect any of the fluorescence lifetimes indicating clearly that both transitions could induce tryptophan excitation and that recorded fluorescence lifetimes could be assigned to sub-structures generated in the excited state.

  9. Visualizing heterogeneity of photosynthetic properties of plant leaves with two-photon fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Iermak, Ievgeniia; Vink, Jochem; Bader, Arjen N; Wientjes, Emilie; van Amerongen, Herbert

    2016-09-01

    Two-photon fluorescence lifetime imaging microscopy (FLIM) was used to analyse the distribution and properties of Photosystem I (PSI) and Photosystem II (PSII) in palisade and spongy chloroplasts of leaves from the C3 plant Arabidopsis thaliana and the C4 plant Miscanthus x giganteus. This was achieved by separating the time-resolved fluorescence of PSI and PSII in the leaf. It is found that the PSII antenna size is larger on the abaxial side of A. thaliana leaves, presumably because chloroplasts in the spongy mesophyll are "shaded" by the palisade cells. The number of chlorophylls in PSI on the adaxial side of the A. thaliana leaf is slightly higher. The C4 plant M. x giganteus contains both mesophyll and bundle sheath cells, which have a different PSI/PSII ratio. It is shown that the time-resolved fluorescence of bundle sheath and mesophyll cells can be analysed separately. The relative number of chlorophylls, which belong to PSI (as compared to PSII) in the bundle sheath cells is at least 2.5 times higher than in mesophyll cells. FLIM is thus demonstrated to be a useful technique to study the PSI/PSII ratio and PSII antenna size in well-defined regions of plant leaves without having to isolate pigment-protein complexes.

  10. A CTRW-based model of time-resolved fluorescence lifetime imaging in a turbid medium.

    Science.gov (United States)

    Chernomordik, Victor; Gandjbakhche, Amir H; Hassan, Moinuddin; Pajevic, Sinisa; Weiss, George H

    2010-12-01

    We develop an analytic model of time-resolved fluorescent imaging of photons migrating through a semi-infinite turbid medium bounded by an infinite plane in the presence of a single stationary point fluorophore embedded in the medium. In contrast to earlier models of fluorescent imaging in which photon motion is assumed to be some form of continuous diffusion process, the present analysis is based on a continuous-time random walk (CTRW) on a simple cubic lattice, the object being to estimate the position and lifetime of the fluorophore. Such information can provide information related to local variations in pH and temperature with potential medical significance. Aspects of the theory were tested using time-resolved measurements of the fluorescence from small inclusions inside tissue-like phantoms. The experimental results were found to be in good agreement with theoretical predictions provided that the fluorophore was not located too close to the planar boundary, a common problem in many diffusive systems.

  11. Steam-sterilizable, fluorescence lifetime-based sensing film for dissolved carbon dioxide.

    Science.gov (United States)

    Chang, Q; Randers-Eichhorn, L; Lakowicz, J R; Rao, G

    1998-01-01

    An autoclavable sensing film was developed for monitoring dissolved CO2. The sensing film, based on fluorescence resonance energy transfer (FRET), consisted of a fluorescent donor, an acceptor, and a quaternary ammonium hydroxide, which were doped in a two-component silicone film. As no aqueous solution was used in the sensing film matrix, the sensing film was unaffected by osmotic pressure. Fluorescence lifetime was selected as the sensing parameter, and measured in frequency domain using phase fluorometry. Upon exposure to 20% CO2-saturated water, a 43 degrees increase in phase angle was observed at 100 MHz. The process was fully reversible when the sensing film was exposed to nitrogen-saturated water. The estimated response and recovery times for 90% signal change were 1 min (for a step change from 0 to 6.7% CO2-saturated water) and 1.5 min (for a step change from 6.7 to 3.3% CO2-saturated water). When used for on-line monitoring of dissolved CO2 produced by a culture of Escherichia coli, the sensing film showed a similar trend to that obtained from off-line measurements using a wet chemistry analyzer.

  12. FPGA-based multi-channel fluorescence lifetime analysis of Fourier multiplexed frequency-sweeping lifetime imaging.

    Science.gov (United States)

    Zhao, Ming; Li, Yu; Peng, Leilei

    2014-09-22

    We report a fast non-iterative lifetime data analysis method for the Fourier multiplexed frequency-sweeping confocal FLIM (Fm-FLIM) system [Opt. Express 22, 10221 (2014)]. The new method, named R-method, allows fast multi-channel lifetime image analysis in the system's FPGA data processing board. Experimental tests proved that the performance of the R-method is equivalent to that of single-exponential iterative fitting, and its sensitivity is well suited for time-lapse FLIM-FRET imaging of live cells, for example cyclic adenosine monophosphate (cAMP) level imaging with GFP-Epac-mCherry sensors. With the R-method and its FPGA implementation, multi-channel lifetime images can now be generated in real time on the multi-channel frequency-sweeping FLIM system, and live readout of FRET sensors can be performed during time-lapse imaging.

  13. Carrier Lifetimes in Fluorescent 6H-SiC for LEDs Application

    DEFF Research Database (Denmark)

    Grivickas, Vytautas; Gulbinas, Karolis; Jokubavičius, Valdas

    Recently it was shown a new approach based on all-semiconductor material technology which is composed with a near ultra-violet GaN LED excitation source and fluorescent silicon carbide (f-6H-SiC) substrate which generates a visible broad spectral light by N and B dopants and an efficient donor...... to acceptor pair recombination [1,2]. This combination can achieve higher electric-light conversion efficiency and high color rendering in comparison with today’s used blue GaN LED based and phosphors. The devices are promising candidates for general lightning applications and may obtain stability....../reproducibility, and potentially low cost in high performance LEDs. However, there are still many problems to obtain best optimization for f-6H-SiC material since neither carrier transport, nor the carrier recombination is known in such co-doped carbides. From the existing data of carrier lifetimes in the SiC materials...

  14. Resolution of heterogeneous fluorescence emission signals and decay lifetime measurement on fluorochrome-labeled cells by phase-sensitive FCM

    Energy Technology Data Exchange (ETDEWEB)

    Steinkamp, J.A.; Crissman, H.A.

    1993-02-01

    A phase-sensitive flow cytometer has been developed to resolve signals from heterogeneous fluorescence emission spectra and quantify fluorescence decay times on cells labeled with fluorescent dyes. This instrument combines flow cytometry (FCM) and fluorescence spectroscopy measurement principles to provide unique capabilities for making phase-resolved measurements on single cells in flow, while preserving conventional FCM measurement capabilities. Stained cells are analyzed as they pass through an intensity-modulated (sinusoid) laser excitation beam. Fluorescence is measured orthogonally using a s barrier filter to block scattered laser excitation light, and a photomultiplier tube detector output signals, which are shifted in phase from a reference signal and amplitude demodulated, are processed by phase-sensitive detection electronics to resolve signals from heterogeneous emissions and quantify decay lifetimes directly. The output signals are displayed as frequency distribution histograms and bivariate diagrams using a computer-based data acquisition system. Results have demonstrated signal phase shift, amplitude demodulation, and average measurement of fluorescence lifetimes on stained cells; a detection limit threshold of 300 to 500 fluorescein isothiocyanate (FITC); fluorescence measurement precision of 1.3% on alignment fluorospheres and 3.4% on propidium iodide (PI)-stained cells; the resolution of PI and FITC signals from cells stainedin combination with PI and FITC, based on differences in their decay lifetimes; and the ability to measure single decay nines by the two-phase, phase comparator, method.

  15. Resolution of heterogeneous fluorescence emission signals and decay lifetime measurement on fluorochrome-labeled cells by phase-sensitive FCM

    Energy Technology Data Exchange (ETDEWEB)

    Steinkamp, J.A.; Crissman, H.A.

    1993-01-01

    A phase-sensitive flow cytometer has been developed to resolve signals from heterogeneous fluorescence emission spectra and quantify fluorescence decay times on cells labeled with fluorescent dyes. This instrument combines flow cytometry (FCM) and fluorescence spectroscopy measurement principles to provide unique capabilities for making phase-resolved measurements on single cells in flow, while preserving conventional FCM measurement capabilities. Stained cells are analyzed as they pass through an intensity-modulated (sinusoid) laser excitation beam. Fluorescence is measured orthogonally using a s barrier filter to block scattered laser excitation light, and a photomultiplier tube detector output signals, which are shifted in phase from a reference signal and amplitude demodulated, are processed by phase-sensitive detection electronics to resolve signals from heterogeneous emissions and quantify decay lifetimes directly. The output signals are displayed as frequency distribution histograms and bivariate diagrams using a computer-based data acquisition system. Results have demonstrated signal phase shift, amplitude demodulation, and average measurement of fluorescence lifetimes on stained cells; a detection limit threshold of 300 to 500 fluorescein isothiocyanate (FITC); fluorescence measurement precision of 1.3% on alignment fluorospheres and 3.4% on propidium iodide (PI)-stained cells; the resolution of PI and FITC signals from cells stainedin combination with PI and FITC, based on differences in their decay lifetimes; and the ability to measure single decay nines by the two-phase, phase comparator, method.

  16. Fluorescence lifetime and acrylamide quenching studies of the interactions between troponin subunits.

    Science.gov (United States)

    Leavis, P C; Gowell, E; Tao, T

    1984-08-28

    Fluorescence lifetime and acrylamide quenching studies were carried out to characterize the interactions between the subunits of troponin under various conditions of metal ion binding. Troponin C was labeled at Cys-98 with N-(iodoacetyl)-N'-(5-sulfo-1-naphthyl)ethylenediamine. In the presence of Ca2+, the fluorescence decay of labeled troponin C (TnC*) was monoexponential, lifetime tau = 15.5 ns and quenching rate constant kq = 2.97 X 10(8) M-1 s-1. In the absence of Ca2+, the decay was resolvable into a major component with tau = 11.9 ns and a minor component with tau = 20.5 ns, with corresponding values of kq = 4.80 X 10(8) and 0.66 X 10(8) M-1 s-1, respectively. Upon the binding of either troponin I (TnI) or troponin T (TnT) in the presence of Ca2+, tau increased to approximately 18 ns, and kq decreased to approximately 0.8 X 10(8) M-1 s-1. For the Ca2+ form of the TnC*-TnI-TnT ternary complex, values of tau = 17.6 ns and kq = 1.73 X 10(8) M-1 s-1 were obtained. These values did not vary significantly when Ca2+ was removed, or when Mg2+ replaced Ca2+. These findings were interpreted as follows: the region around Cys-98 of TnC* adopts a looser conformation upon the removal of Ca2+ from the high-affinity sites. Both TnI and TnT bind to TnC* in the region containing Cys-98. The probe is shielded from the solvent to a greater extent in the binary complexes than in the ternary complex.(ABSTRACT TRUNCATED AT 250 WORDS)

  17. In vivo wound healing diagnosis with second harmonic and fluorescence lifetime imaging

    Science.gov (United States)

    Deka, Gitanjal; Wu, Wei-Wen; Kao, Fu-Jen

    2013-06-01

    Skin wounds heal when a series of cell lineages are triggered, followed by collagen deposition, to reconstruct damaged tissues. This study evaluates the regeneration of collagen and change in cellular metabolic rate in vivo during wound healing in rats, with second harmonic generation (SHG) and fluorescence lifetime imaging microscopy respectively. The metabolic rate of cells is reflected through the lifetime of the autofluorescence from the co-enzyme protein, reduced nicotinamide adenine dinucleotide, due to its change in the relative concentration of bound and free forms. A higher than normal cellular metabolic rate is observed during the first week of healing, which decreases gradually after eight days of wound formation. SHG signal intensity change indicates the net degradation of collagen during the inflammatory phase, and net regeneration begins on day five. Eventually, the quantity of collagen increases gradually to form a scar tissue as the final product. Importantly, this work demonstrates the feasibility of an in vivo imaging approach for a normal wound on rat skin, which has the potential to supplement the noninvasive clinical diagnosis of wounds.

  18. Analysis of energy metabolism of HeLa cancer cells in vitro and in vivo using fluorescence lifetime microscopy

    Science.gov (United States)

    Lukina, Maria; Shirmanova, Marina; Dudenkova, Varvara; Druzhkova, Irina; Shumilova, Anastasia; Zagaynova, Elena

    2016-04-01

    The aim of the present work was to study energy metabolism in human cervical carcinoma (HeLa) cells in vitro and in vivo using two-photon FLIM. Cellular metabolism was examined by monitoring of the fluorescence lifetimes of free and protein-bound forms of NAD(P)H and FAD and their relative contributions. Two-photon fluorescence and second harmonic generation microscopy as well as standard histopathology with hematoxylin and eosin were used to characterize tissue structure. Cellular metabolism was analyzed in cancer cells co-cultured with human fibroblasts and in tumor xenografts transplanted to nude mice. In the HeLa-huFB co-culture we observed a metabolic shift from OXPHOS toward glycolysis in cancer cells, and from glycolysis to OXPHOS in fibroblasts, starting from Day 2 of co-culturing. In the tumor tissue we detected metabolic heterogeneity with more glycolytic metabolism of cancer cells in the stroma-rich zones. The results of the study are of a great importance for understanding metabolic behavior of tumors and for development of anticancer drugs targeted to metabolic pathways.

  19. Combined nonlinear laser imaging (two-photon excitation fluorescence, second and third-harmonic generation, and fluorescence lifetime imaging microscopies) in ovarian tumors

    Science.gov (United States)

    Adur, J.; Pelegati, V. B.; de Thomaz, A. A.; Bottcher-Luiz, F.; Andrade, L. A. L. A.; Almeida, D. B.; Carvalho, H. F.; Cesar, C. L.

    2012-03-01

    We applied Two-photon Excited Fluorescence (TPEF), Second/Third Harmonic Generation (SHG and THG) and Fluorescence Lifetime Imaging (FLIM) Non Linear Optics (NLO) Laser-Scanning Microscopy within the same imaging platform to evaluate their use as a diagnostic tool in ovarian tumors. We assess of applicability of this multimodal approach to perform a pathological evaluation of serous and mucinous tumors in human samples. The combination of TPEF-SHG-THG imaging provided complementary information about the interface epithelium/stromal, such as the transformation of epithelium surface (THG) and the overall fibrillar tissue architecture (SHG). The fact that H&E staining is the standard method used in clinical pathology and that the stored samples are usually fixed makes it important a re-evaluation of these samples with NLO microscopy to compare new results with a library of already existing samples. FLIM, however, depends on the chemical environment around the fluorophors that was completely changed after fixation; therefore it only makes sense in unstained samples. Our FLIM results in unstained samples demonstrate that it is possible to discriminate healthy epithelia from serous or mucinous epithelia. Qualitative and quantitative analysis of the different imaging modalities used showed that multimodal nonlinear microscopy has the potential to differentiate between cancerous and healthy ovarian tissue.

  20. Persistent luminescence nanoprobe for biosensing and lifetime imaging of cell apoptosis via time-resolved fluorescence resonance energy transfer.

    Science.gov (United States)

    Zhang, Lei; Lei, Jianping; Liu, Jintong; Ma, Fengjiao; Ju, Huangxian

    2015-10-01

    Time-resolved fluorescence technique can reduce the short-lived background luminescence and auto-fluorescence interference from cells and tissues by exerting the delay time between pulsed excitation light and signal acquisition. Here, we prepared persistent luminescence nanoparticles (PLNPs) to design a universal time-resolved fluorescence resonance energy transfer (TR-FRET) platform for biosensing, lifetime imaging of cell apoptosis and in situ lifetime quantification of intracellular caspase-3. Three kinds of PLNPs-based nanoprobes are assembled by covalently binding dye-labeled peptides or DNA to carboxyl-functionalized PLNPs for the efficient detection of caspase-3, microRNA and protein. The peptides-functionalized nanoprobe is also employed for fluorescence lifetime imaging to monitor cell apoptosis, which shows a dependence of cellular fluorescence lifetime on caspase-3 activity and thus leads to an in situ quantification method. This work provides a proof-of-concept for PLNPs-based TR-FRET analysis and demonstrates its potential in exploring dynamical information of life process. Copyright © 2015 Elsevier Ltd. All rights reserved.

  1. Fluorescence Characteristics and Lifetime Images of Photosensitizers of Talaporfin Sodium and Sodium Pheophorbide a in Normal and Cancer Cells

    Directory of Open Access Journals (Sweden)

    Kamlesh Awasthi

    2015-05-01

    Full Text Available Fluorescence spectra and fluorescence lifetime images of talaporfin sodium and sodium-pheophorbide a, which can be regarded as photosensitizers for photodynamic therapy, were measured in normal and cancer cells. The reduction of the fluorescence intensity by photoirradiation was observed for both photosensitizers in both cells, but the quenching rate was much faster in cancer cells than in normal cells. These results are explained in terms of the excessive generation of reactive oxygen species via photoexcitation of these photosensitizers in cancer cells. The fluorescence lifetimes of both photosensitizers in cancer cells are different from those in normal cells, which originates from the different intracellular environments around the photosensitizers between normal and cancer cells.

  2. Two-photon excitation with pico-second fluorescence lifetime imaging to detect nuclear association of flavanols

    Energy Technology Data Exchange (ETDEWEB)

    Mueller-Harvey, Irene, E-mail: i.mueller-harvey@reading.ac.uk [Chemistry and Biochemistry Laboratory, Food Production and Quality Research Division, School of Agriculture, Policy and Development, University of Reading, P O Box 236, Reading RG6 6AT (United Kingdom); Feucht, Walter, E-mail: walter.feucht@gmail.com [Department of Plant Sciences, Technical University of Munich (TUM), Wissenschaftszentrum Weihenstephan (WZW), D-85354 Freising (Germany); Polster, Juergen, E-mail: j.polster@wzw.tum.de [Department of Physical Biochemistry, Technical University of Munich (TUM), Wissenschaftszentrum Weihenstephan (WZW), D-85354 Freising (Germany); Trnkova, Lucie, E-mail: lucie.trnkova@uhk.cz [University of Hradec Kralove, Faculty of Science, Department of Chemistry, Rokitanskeho 62, 50003 Hradec Kralove (Czech Republic); Burgos, Pierre, E-mail: pierre.burgos@stfc.ac.uk [Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell-Oxford, Didcot, Oxfordshire, OX11 0QX (United Kingdom); Parker, Anthony W., E-mail: tony.parker@stfc.ac.uk [Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell-Oxford, Didcot, Oxfordshire, OX11 0QX (United Kingdom); Botchway, Stanley W., E-mail: stan.botchway@stfc.ac.uk [Central Laser Facility, Research Complex at Harwell, Science and Technology Facilities Council, Rutherford Appleton Laboratory, Harwell-Oxford, Didcot, Oxfordshire, OX11 0QX (United Kingdom)

    2012-03-16

    Highlights: Black-Right-Pointing-Pointer This fluorescence lifetime imaging microscopy (FLIM) technique for flavanols overcomes autofluorescence interference in cells. Black-Right-Pointing-Pointer Plant flavanols differed in their lifetimes. Black-Right-Pointing-Pointer Dissolved and bound flavanols revealed contrasting lifetime changes. Black-Right-Pointing-Pointer This technique will allow studying of flavanol trafficking in live cells. - Abstract: Two-photon excitation enabled for the first time the observation and measurement of excited state fluorescence lifetimes from three flavanols in solution, which were {approx}1.0 ns for catechin and epicatechin, but <45 ps for epigallocatechin gallate (EGCG). The shorter lifetime for EGCG is in line with a lower fluorescence quantum yield of 0.003 compared to catechin (0.015) and epicatechin (0.018). In vivo experiments with onion cells demonstrated that tryptophan and quercetin, which tend to be major contributors of background fluorescence in plant cells, have sufficiently low cross sections for two-photon excitation at 630 nm and therefore do not interfere with detection of externally added or endogenous flavanols in Allium cepa or Taxus baccata cells. Applying two-photon excitation to flavanols enabled 3-D fluorescence lifetime imaging microscopy and showed that added EGCG penetrated the whole nucleus of onion cells. Interestingly, EGCG and catechin showed different lifetime behaviour when bound to the nucleus: EGCG lifetime increased from <45 to 200 ps, whilst catechin lifetime decreased from 1.0 ns to 500 ps. Semi-quantitative measurements revealed that the relative ratios of EGCG concentrations in nucleoli associated vesicles: nucleus: cytoplasm were ca. 100:10:1. Solution experiments with catechin, epicatechin and histone proteins provided preliminary evidence, via the appearance of a second lifetime ({tau}{sub 2} = 1.9-3.1 ns), that both flavanols may be interacting with histone proteins. We conclude that there

  3. Two-photon excitation with pico-second fluorescence lifetime imaging to detect nuclear association of flavanols.

    Science.gov (United States)

    Mueller-Harvey, Irene; Feucht, Walter; Polster, Juergen; Trnková, Lucie; Burgos, Pierre; Parker, Anthony W; Botchway, Stanley W

    2012-03-16

    Two-photon excitation enabled for the first time the observation and measurement of excited state fluorescence lifetimes from three flavanols in solution, which were ~1.0 ns for catechin and epicatechin, but <45 ps for epigallocatechin gallate (EGCG). The shorter lifetime for EGCG is in line with a lower fluorescence quantum yield of 0.003 compared to catechin (0.015) and epicatechin (0.018). In vivo experiments with onion cells demonstrated that tryptophan and quercetin, which tend to be major contributors of background fluorescence in plant cells, have sufficiently low cross sections for two-photon excitation at 630 nm and therefore do not interfere with detection of externally added or endogenous flavanols in Allium cepa or Taxus baccata cells. Applying two-photon excitation to flavanols enabled 3-D fluorescence lifetime imaging microscopy and showed that added EGCG penetrated the whole nucleus of onion cells. Interestingly, EGCG and catechin showed different lifetime behaviour when bound to the nucleus: EGCG lifetime increased from <45 to 200 ps, whilst catechin lifetime decreased from 1.0 ns to 500 ps. Semi-quantitative measurements revealed that the relative ratios of EGCG concentrations in nucleoli associated vesicles: nucleus: cytoplasm were ca. 100:10:1. Solution experiments with catechin, epicatechin and histone proteins provided preliminary evidence, via the appearance of a second lifetime (τ(2)=1.9-3.1 ns), that both flavanols may be interacting with histone proteins. We conclude that there is significant nuclear absorption of flavanols. This advanced imaging using two-photon excitation and biophysical techniques described here will prove valuable for probing the intracellular trafficking and functions of flavanols, such as EGCG, which is the major flavanol of green tea.

  4. Open Source High Content Analysis Utilizing Automated Fluorescence Lifetime Imaging Microscopy.

    Science.gov (United States)

    Görlitz, Frederik; Kelly, Douglas J; Warren, Sean C; Alibhai, Dominic; West, Lucien; Kumar, Sunil; Alexandrov, Yuriy; Munro, Ian; Garcia, Edwin; McGinty, James; Talbot, Clifford; Serwa, Remigiusz A; Thinon, Emmanuelle; da Paola, Vincenzo; Murray, Edward J; Stuhmeier, Frank; Neil, Mark A A; Tate, Edward W; Dunsby, Christopher; French, Paul M W

    2017-01-18

    We present an open source high content analysis instrument utilizing automated fluorescence lifetime imaging (FLIM) for assaying protein interactions using Förster resonance energy transfer (FRET) based readouts of fixed or live cells in multiwell plates. This provides a means to screen for cell signaling processes read out using intramolecular FRET biosensors or intermolecular FRET of protein interactions such as oligomerization or heterodimerization, which can be used to identify binding partners. We describe here the functionality of this automated multiwell plate FLIM instrumentation and present exemplar data from our studies of HIV Gag protein oligomerization and a time course of a FRET biosensor in live cells. A detailed description of the practical implementation is then provided with reference to a list of hardware components and a description of the open source data acquisition software written in µManager. The application of FLIMfit, an open source MATLAB-based client for the OMERO platform, to analyze arrays of multiwell plate FLIM data is also presented. The protocols for imaging fixed and live cells are outlined and a demonstration of an automated multiwell plate FLIM experiment using cells expressing fluorescent protein-based FRET constructs is presented. This is complemented by a walk-through of the data analysis for this specific FLIM FRET data set.

  5. Direct Vpr-Vpr Interaction in Cells monitored by two Photon Fluorescence Correlation Spectroscopy and Fluorescence Lifetime Imaging

    Directory of Open Access Journals (Sweden)

    Mély Yves

    2008-09-01

    Full Text Available Abstract Background The human immunodeficiency virus type 1 (HIV-1 encodes several regulatory proteins, notably Vpr which influences the survival of the infected cells by causing a G2/M arrest and apoptosis. Such an important role of Vpr in HIV-1 disease progression has fuelled a large number of studies, from its 3D structure to the characterization of specific cellular partners. However, no direct imaging and quantification of Vpr-Vpr interaction in living cells has yet been reported. To address this issue, eGFP- and mCherry proteins were tagged by Vpr, expressed in HeLa cells and their interaction was studied by two photon fluorescence lifetime imaging microscopy and fluorescence correlation spectroscopy. Results Results show that Vpr forms homo-oligomers at or close to the nuclear envelope. Moreover, Vpr dimers and trimers were found in the cytoplasm and in the nucleus. Point mutations in the three α helices of Vpr drastically impaired Vpr oligomerization and localization at the nuclear envelope while point mutations outside the helical regions had no effect. Theoretical structures of Vpr mutants reveal that mutations within the α-helices could perturb the leucine zipper like motifs. The ΔQ44 mutation has the most drastic effect since it likely disrupts the second helix. Finally, all Vpr point mutants caused cell apoptosis suggesting that Vpr-mediated apoptosis functions independently from Vpr oligomerization. Conclusion We report that Vpr oligomerization in HeLa cells relies on the hydrophobic core formed by the three α helices. This oligomerization is required for Vpr localization at the nuclear envelope but not for Vpr-mediated apoptosis.

  6. Measurement of radiative lifetime in atomic samarium using simultaneous detection of laser-induced fluorescence and photoionization signals

    Indian Academy of Sciences (India)

    A C Sahoo; M L Shah; P K Mandal; A K Pulhani; G P Gupta; Vas Dev; B M Suri

    2014-02-01

    In this paper, we report the investigations of lifetime measurement of odd-parity energy level 19009.52 cm-1 of Sm I using simultaneous detection of laser-induced fluorescence and laserinduced photoionization signals employing pump–probe technique. To the best of our knowledge, this is for the first time that the results obtained using laser-induced fluorescence and photoionization techniques have been compared with each other. The obtained results match well with those reported in the literature.

  7. Support vector machine based classification and mapping of atherosclerotic plaques using fluorescence lifetime imaging (Conference Presentation)

    Science.gov (United States)

    Fatakdawala, Hussain; Gorpas, Dimitris S.; Bec, Julien; Ma, Dinglong M.; Yankelevich, Diego R.; Bishop, John W.; Marcu, Laura

    2016-02-01

    The progression of atherosclerosis in coronary vessels involves distinct pathological changes in the vessel wall. These changes manifest in the formation of a variety of plaque sub-types. The ability to detect and distinguish these plaques, especially thin-cap fibroatheromas (TCFA) may be relevant for guiding percutaneous coronary intervention as well as investigating new therapeutics. In this work we demonstrate the ability of fluorescence lifetime imaging (FLIm) derived parameters (lifetime values from sub-bands 390/40 nm, 452/45 nm and 542/50 nm respectively) for generating classification maps for identifying eight different atherosclerotic plaque sub-types in ex vivo human coronary vessels. The classification was performed using a support vector machine based classifier that was built from data gathered from sixteen coronary vessels in a previous study. This classifier was validated in the current study using an independent set of FLIm data acquired from four additional coronary vessels with a new rotational FLIm system. Classification maps were compared to co-registered histological data. Results show that the classification maps allow identification of the eight different plaque sub-types despite the fact that new data was gathered with a different FLIm system. Regions with diffuse intimal thickening (n=10), fibrotic tissue (n=2) and thick-cap fibroatheroma (n=1) were correctly identified on the classification map. The ability to identify different plaque types using FLIm data alone may serve as a powerful clinical and research tool for studying atherosclerosis in animal models as well as in humans.

  8. Actin cytoskeleton-dependent Rab GTPase-regulated angiotensin type I receptor lysosomal degradation studied by fluorescence lifetime imaging microscopy

    Science.gov (United States)

    Li, Hewang; Yu, Peiying; Sun, Yuansheng; Felder, Robin A.; Periasamy, Ammasi; Jose, Pedro A.

    2010-09-01

    The dynamic regulation of the cellular trafficking of human angiotensin (Ang) type 1 receptor (AT1R) is not well understood. Therefore, we investigated the cellular trafficking of AT1R-enhanced green fluorescent protein (EGFP) (AT1R-EGFP) heterologously expressed in HEK293 cells by determining the change in donor lifetime (AT1R-EGFP) in the presence or absence of acceptor(s) using fluorescence lifetime imaging-fluorescence resonance energy transfer (FRET) microscopy. The average lifetime of AT1R-EGFP in our donor-alone samples was ~2.33 ns. The basal state lifetime was shortened slightly in the presence of Rab5 (2.01+/-0.10 ns) or Rab7 (2.11+/-0.11 ns) labeled with Alexa 555, as the acceptor fluorophore. A 5-min Ang II treatment markedly shortened the lifetime of AT1R-EGFP in the presence of Rab5-Alexa 555 (1.78+/-0.31 ns) but was affected minimally in the presence of Rab7-Alexa 555 (2.09+/-0.37 ns). A 30-min Ang II treatment further decreased the AT1R-EGFP lifetime in the presence of both Rab5- and Rab7-Alexa 555. Latrunculin A but not nocodazole pretreatment blocked the ability of Ang II to shorten the AT1R-EGFP lifetime. The occurrence of FRET between AT1R-EGFP (donor) and LAMP1-Alexa 555 (acceptor) with Ang II stimulation was impaired by photobleaching the acceptor. These studies demonstrate that Ang II-induced AT1R lysosomal degradation through its association with LAMP1 is regulated by Rab5/7 via mechanisms that are dependent on intact actin cytoskeletons.

  9. FLIMX: A Software Package to Determine and Analyze the Fluorescence Lifetime in Time-Resolved Fluorescence Data from the Human Eye

    Science.gov (United States)

    Klemm, Matthias; Schweitzer, Dietrich; Peters, Sven; Sauer, Lydia; Hammer, Martin; Haueisen, Jens

    2015-01-01

    Fluorescence lifetime imaging ophthalmoscopy (FLIO) is a new technique for measuring the in vivo autofluorescence intensity decays generated by endogenous fluorophores in the ocular fundus. Here, we present a software package called FLIM eXplorer (FLIMX) for analyzing FLIO data. Specifically, we introduce a new adaptive binning approach as an optimal tradeoff between the spatial resolution and the number of photons required per pixel. We also expand existing decay models (multi-exponential, stretched exponential, spectral global analysis, incomplete decay) to account for the layered structure of the eye and present a method to correct for the influence of the crystalline lens fluorescence on the retina fluorescence. Subsequently, the Holm-Bonferroni method is applied to FLIO measurements to allow for group comparisons between patients and controls on the basis of fluorescence lifetime parameters. The performance of the new approaches was evaluated in five experiments. Specifically, we evaluated static and adaptive binning in a diabetes mellitus patient, we compared the different decay models in a healthy volunteer and performed a group comparison between diabetes patients and controls. An overview of the visualization capabilities and a comparison of static and adaptive binning is shown for a patient with macular hole. FLIMX’s applicability to fluorescence lifetime imaging microscopy is shown in the ganglion cell layer of a porcine retina sample, obtained by a laser scanning microscope using two-photon excitation. PMID:26192624

  10. A dual-modality optical coherence tomography and fluorescence lifetime imaging microscopy system for simultaneous morphological and biochemical tissue characterization.

    Science.gov (United States)

    Park, Jesung; Jo, Javier A; Shrestha, Sebina; Pande, Paritosh; Wan, Qiujie; Applegate, Brian E

    2010-07-16

    Most pathological conditions elicit changes in the tissue optical response that may be interrogated by one or more optical imaging modalities. Any single modality typically only furnishes an incomplete picture of the tissue optical response, hence an approach that integrates complementary optical imaging modalities is needed for a more comprehensive non-destructive and minimally-invasive tissue characterization. We have developed a dual-modality system, incorporating optical coherence tomography (OCT) and fluorescence lifetime imaging microscopy (FLIM), that is capable of simultaneously characterizing the 3-D tissue morphology and its biochemical composition. The Fourier domain OCT subsystem, at an 830 nm center wavelength, provided high-resolution morphological volumetric tissue images with an axial and lateral resolution of 7.3 and 13.4 µm, respectively. The multispectral FLIM subsystem, based on a direct pulse-recording approach (upon 355 nm laser excitation), provided two-dimensional superficial maps of the tissue autofluorescence intensity and lifetime at three customizable emission bands with 100 µm lateral resolution. Both subsystems share the same excitation/illumination optical path and are simultaneously raster scanned on the sample to generate coregistered OCT volumes and FLIM images. The developed OCT/FLIM system was capable of a maximum A-line rate of 59 KHz for OCT and a pixel rate of up to 30 KHz for FLIM. The dual-modality system was validated with standard fluorophore solutions and subsequently applied to the characterization of two biological tissue types: postmortem human coronary atherosclerotic plaques, and in vivo normal and cancerous hamster cheek pouch epithelial tissue.

  11. Confirmation of temperature independence in the fluorescence lifetime of the 3P 0 → 3F 2 transition in praseodymium-doped fluoride glass

    Science.gov (United States)

    Nguyen, Thinh B.; Vella, Vince; Baxter, Greg W.; Collins, Stephen F.; Newman, Peter J.; MacFarlane, Douglas R.

    2006-05-01

    The dependence of the fluorescence lifetime from the 3P0 → 3F2 transition in praseodymium-doped fluoride glass as a function of dopant concentration and temperature was investigated. It was found that the fluorescence lifetime at the concentration of 7000 ppm was constant with temperature, confirming the prediction of temperature independence in the lifetime for this transition in Pr3+-doped ZBLAN glass.

  12. Cytosolic NADH-NAD+ Redox Visualized in Brain Slices by Two-Photon Fluorescence Lifetime Biosensor Imaging

    Science.gov (United States)

    Mongeon, Rebecca; Venkatachalam, Veena

    2016-01-01

    Abstract Aim: Cytosolic NADH-NAD+ redox state is central to cellular metabolism and a valuable indicator of glucose and lactate metabolism in living cells. Here we sought to quantitatively determine NADH-NAD+ redox in live cells and brain tissue using a fluorescence lifetime imaging of the genetically-encoded single-fluorophore biosensor Peredox. Results: We show that Peredox exhibits a substantial change in its fluorescence lifetime over its sensing range of NADH-NAD+ ratio. This allows changes in cytosolic NADH redox to be visualized in living cells using a two-photon scanning microscope with fluorescence lifetime imaging capabilities (2p-FLIM), using time-correlated single photon counting. Innovation: Because the lifetime readout is absolutely calibrated (in nanoseconds) and is independent of sensor concentration, we demonstrate that quantitative assessment of NADH redox is possible using a single fluorophore biosensor. Conclusion: Imaging of the sensor in mouse hippocampal brain slices reveals that astrocytes are typically much more reduced (with higher NADH:NAD+ ratio) than neurons under basal conditions, consistent with the hypothesis that astrocytes are more glycolytic than neurons. Antioxid. Redox Signal. 25, 553–563. PMID:26857245

  13. Improved maximum entropy method for the analysis of fluorescence spectroscopy data: evaluating zero-time shift and assessing its effect on the determination of fluorescence lifetimes.

    Science.gov (United States)

    Esposito, Rosario; Mensitieri, Giuseppe; de Nicola, Sergio

    2015-12-21

    A new algorithm based on the Maximum Entropy Method (MEM) is proposed for recovering both the lifetime distribution and the zero-time shift from time-resolved fluorescence decay intensities. The developed algorithm allows the analysis of complex time decays through an iterative scheme based on entropy maximization and the Brent method to determine the minimum of the reduced chi-squared value as a function of the zero-time shift. The accuracy of this algorithm has been assessed through comparisons with simulated fluorescence decays both of multi-exponential and broad lifetime distributions for different values of the zero-time shift. The method is capable of recovering the zero-time shift with an accuracy greater than 0.2% over a time range of 2000 ps. The center and the width of the lifetime distributions are retrieved with relative discrepancies that are lower than 0.1% and 1% for the multi-exponential and continuous lifetime distributions, respectively. The MEM algorithm is experimentally validated by applying the method to fluorescence measurements of the time decays of the flavin adenine dinucleotide (FAD).

  14. Alterations in cerebral metabolism observed in living rodents using fluorescence lifetime microscopy of intrinsic NADH (Conference Presentation)

    Science.gov (United States)

    Yaseen, Mohammad A.; Sakadžić, Sava; Sutin, Jason; Wu, Weicheng; Fu, Buyin; Boas, David A.

    2017-02-01

    Monitoring cerebral energy metabolism at a cellular level is essential to improve our understanding of healthy brain function and its pathological alterations. In this study, we resolve specific alterations in cerebral metabolism utilizing minimally-invasive 2-Photon fluorescence lifetime imaging (2P-FLIM) measurements of reduced nicotinamide adenine dinucleotide (NADH) fluorescence, collected in vivo from anesthetized rats and mice. Time-resolved lifetime measurements enables distinction of different components contributing to NADH autofluorescence. These components reportedly represent different enzyme-bound formulations of NADH. Our observations from this study confirm the hypothesis that NADH FLIM can identify specific alterations in cerebral metabolism. Using time-correlated single photon counting (TCSPC) equipment and a custom-built multimodal imaging system, 2-photon fluorescence lifetime imaging (FLIM) was performed in cerebral tissue with high spatial and temporal resolution. Multi-exponential fits for NADH fluorescence lifetimes indicate 4 distinct components, or 'species.' We observed distinct variations in the relative proportions of these components before and after pharmacological-induced impairments to several reactions involved in anaerobic glycolysis and aerobic oxidative metabolism. Classification models developed with experimental data correctly predict the metabolic impairments associated with bicuculline-induced focal seizures in separate experiments. Compared to traditional intensity-based NADH measurements, lifetime imaging of NADH is less susceptible to the adverse effects of overlying blood vessels. Evaluating NADH measurements will ultimately lead to a deeper understanding of cerebral energetics and its pathology-related alterations. Such knowledge will likely aid development of therapeutic strategies for neurodegenerative diseases such as Alzheimer's Disease, Parkinson's disease, and stroke.

  15. Monitoring of labeled antisense oligonucleotides within living cells by using a multifrequency phase/modulation approach for fluorescence lifetime measurements

    Science.gov (United States)

    Kocisova, E.; Sureau, F.; Praus, P.; Rosenberg, I.; Stepanek, J.; Turpin, P.-Y.

    2003-06-01

    A multifrequency phase/modulation method has been developed for our UV confocal laser microspectrofluorimeter (modulation frequency 1-200 MHz) for fluorescence lifetime measurements. This technique enables excited state lifetimes of mixed fluorescent components to be resolved and the fluorescence spectral contribution of each species to be determined without using any model spectra. This approach is very efficient for analyzing intracellular multicomponent fluorescence signals. Our effort is focused on the elucidation of the intracellular behavior of synthetic modified oligonucleotides - potential drugs for antisense and/or antigene strategies of curing viral and malignant diseases. A novel type single stranded dT 15 oligomer analogue containing isopolar, non-isosteric, phosphonate-based internucleotide linkages (3'-O-P-CH 2-O-5'), labeled with tetramethylrhodamine dye at the 3'-end, has been utilized. This method, along with fluorescence micro-imaging, was used to monitor uptake, distribution and stability of our modified oligonucleotide inside living cells. Binding to Escort™ vector leads to an homogeneous intracellular distribution of fluorescent labeled oligonucleotide, including nucleus staining, while point distribution only is achieved for its free form.

  16. Low-pressure effective fluorescence lifetimes and photo-physical rate constants of one- and two-ring aromatics

    Science.gov (United States)

    Benzler, Thorsten; Faust, Stephan; Dreier, Thomas; Schulz, Christof

    2015-12-01

    One- and two-ring aromatics such as toluene and naphthalene are frequently used molecular tracer species in laser-induced fluorescence (LIF) imaging diagnostics. Quantifying LIF signal intensities requires knowledge of the photo-physical processes that determine the fluorescence quantum yield. Collision-induced and intramolecular energy transfer processes in the excited electronic state closely interact under practical conditions. They can be separated through experiments at variable low pressures. Effective fluorescence lifetimes of gaseous toluene, 1,2,4-trimethylbenzene, anisole, naphthalene, and 1-methylnaphthalene diluted in CO2 were measured after picosecond laser excitation at 266 nm and time-resolved detection of fluorescence intensities. Measurements in an optically accessible externally heated cell between 296 and 475 K and 0.010-1 bar showed that effective fluorescence lifetimes generally decrease with temperature, while the influence of the bath-gas pressure depends on the respective target species and temperature. The results provide non-radiative and fluorescence rate constants and experimentally validate the effect of photo-induced cooling.

  17. Evaluation of the oxidative stress of psoriatic fibroblasts based on spectral two-photon fluorescence lifetime imaging

    Science.gov (United States)

    Kapsokalyvas, Dimitrios; Barygina, Victoria; Cicchi, Riccardo; Fiorillo, Claudia; Pavone, Francesco S.

    2013-02-01

    Psoriasis is an autoimmune disease of the skin characterized by hyperkeratosis, hyperproliferation of the epidermis, inflammatory cell accumulation and increased dilatation of dermal papillary blood vessels. Metabolic activity is increased in the epidermis and the dermis. Oxidative stress is high mainly due to reactive oxygen species (ROS) originating from the skin environment and cellular metabolism. We employed a custom multiphoton microscope coupled with a FLIM setup to image primary culture fibroblast cells from perilesional and lesional psoriatic skin in-vitro. Twophoton excited fluorescence images revealed the morphological differences between healthy and psoriatic fibroblasts. Based on the spectral analysis of the NADH and FAD components the oxidative stress was assessed and found to be higher in psoriatic cells. Furthermore the fluorescence lifetime properties were investigated with a TCSPC FLIM module. Mean fluorescence lifetime was found to be longer in psoriatic lesional cells. Analysis of the fast (τ1) and slow (τ2) decay lifetimes revealed a decrease of the ratio of the contribution of the fast (α1) parameter to the contribution of the slow (α2) parameter. The fluorescence in the examined part of the spectrum is attributed mainly to NADH. The decrease of the ratio (α1)/ (α2) is believed to correlate strongly with the anti-oxidant properties of NADH which can lead to the variation of its population in high ROS environment. This methodology could serve as an index of the oxidative status in cells and furthermore could be used to probe the oxidative stress of tissues in-vivo.

  18. Label-free fluorescence lifetime and second harmonic generation imaging microscopy improves quantification of experimental renal fibrosis.

    Science.gov (United States)

    Ranjit, Suman; Dobrinskikh, Evgenia; Montford, John; Dvornikov, Alexander; Lehman, Allison; Orlicky, David J; Nemenoff, Raphael; Gratton, Enrico; Levi, Moshe; Furgeson, Seth

    2016-11-01

    All forms of progressive renal diseases develop a final pathway of tubulointerstitial fibrosis and glomerulosclerosis. Renal fibrosis is usually quantified using histological staining, a process that is time-consuming and pathologist dependent. Here we develop a fast and operator-independent method to measure fibrosis utilizing the murine unilateral ureteral obstruction model which manifests a time-dependent fibrotic increase in obstructed kidneys while the contralateral kidneys are used as controls. After ureteral obstruction, kidneys were analyzed at 7, 14, and 21 days. Fibrosis was quantified using fluorescence lifetime imaging (FLIM) and second harmonic generation (SHG) in a Deep Imaging via Enhanced photon Recovery deep tissue imaging microscope. This microscope was developed for deep tissue along with second and third harmonic generation imaging and has extraordinary sensitivity toward harmonic generation. SHG data suggest the presence of more fibrillar collagen in the obstructed kidneys. The combination of short-wavelength FLIM and SHG analysis results in a robust assessment procedure independent of observer interpretation and let us create criteria to quantify the extent of fibrosis directly from the image. Thus, the FLIM-SHG technique shows remarkable improvement in quantification of renal fibrosis compared to standard histological techniques. Copyright © 2016 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

  19. Parallelized TCSPC for dynamic intravital fluorescence lifetime imaging: quantifying neuronal dysfunction in neuroinflammation.

    Directory of Open Access Journals (Sweden)

    Jan Leo Rinnenthal

    Full Text Available Two-photon laser-scanning microscopy has revolutionized our view on vital processes by revealing motility and interaction patterns of various cell subsets in hardly accessible organs (e.g. brain in living animals. However, current technology is still insufficient to elucidate the mechanisms of organ dysfunction as a prerequisite for developing new therapeutic strategies, since it renders only sparse information about the molecular basis of cellular response within tissues in health and disease. In the context of imaging, Förster resonant energy transfer (FRET is one of the most adequate tools to probe molecular mechanisms of cell function. As a calibration-free technique, fluorescence lifetime imaging (FLIM is superior for quantifying FRET in vivo. Currently, its main limitation is the acquisition speed in the context of deep-tissue 3D and 4D imaging. Here we present a parallelized time-correlated single-photon counting point detector (p-TCSPC (i for dynamic single-beam scanning FLIM of large 3D areas on the range of hundreds of milliseconds relevant in the context of immune-induced pathologies as well as (ii for ultrafast 2D FLIM in the range of tens of milliseconds, a scale relevant for cell physiology. We demonstrate its power in dynamic deep-tissue intravital imaging, as compared to multi-beam scanning time-gated FLIM suitable for fast data acquisition and compared to highly sensitive single-channel TCSPC adequate to detect low fluorescence signals. Using p-TCSPC, 256×256 pixel FLIM maps (300×300 µm(2 are acquired within 468 ms while 131×131 pixel FLIM maps (75×75 µm(2 can be acquired every 82 ms in 115 µm depth in the spinal cord of CerTN L15 mice. The CerTN L15 mice express a FRET-based Ca-biosensor in certain neuronal subsets. Our new technology allows us to perform time-lapse 3D intravital FLIM (4D FLIM in the brain stem of CerTN L15 mice affected by experimental autoimmune encephalomyelitis and, thereby, to truly quantify

  20. Investigation of the Co-Dependence of Morphology and Fluorescence Lifetime in a Metal-Organic Framework.

    Science.gov (United States)

    Schrimpf, Waldemar; Ossato, Giulia; Hirschle, Patrick; Wuttke, Stefan; Lamb, Don C

    2016-07-01

    Porous materials, due to their large surface-to-volume ratio, are important for a broad range of applications and are the subject of intense research. Most studies investigate the bulk properties of these materials, which are not sensitive to the effect of heterogeneities within the sample. Herein, a new strategy based on correlative fluorescence lifetime imaging and scanning electron microscopy is presented that allows the detection and localization of those heterogeneities, and connects them to morphological and structural features of the material. By applying this method to a dye-modified metal-organic framework (MOF), two independent fluorescence quenching mechanisms in the MOF scaffold are identified and quantified. The first mechanism is based on quenching via amino groups, while the second mechanism is influenced by morphology. Furthermore, a similar correlation between the inherent luminescence lifetime and the morphology of the unmodified MOF structure is demonstrated.

  1. CMOS image sensor with lateral electric field modulation pixels for fluorescence lifetime imaging with sub-nanosecond time response

    Science.gov (United States)

    Li, Zhuo; Seo, Min-Woong; Kagawa, Keiichiro; Yasutomi, Keita; Kawahito, Shoji

    2016-04-01

    This paper presents the design and implementation of a time-resolved CMOS image sensor with a high-speed lateral electric field modulation (LEFM) gating structure for time domain fluorescence lifetime measurement. Time-windowed signal charge can be transferred from a pinned photodiode (PPD) to a pinned storage diode (PSD) by turning on a pair of transfer gates, which are situated beside the channel. Unwanted signal charge can be drained from the PPD to the drain by turning on another pair of gates. The pixel array contains 512 (V) × 310 (H) pixels with 5.6 × 5.6 µm2 pixel size. The imager chip was fabricated using 0.11 µm CMOS image sensor process technology. The prototype sensor has a time response of 150 ps at 374 nm. The fill factor of the pixels is 5.6%. The usefulness of the prototype sensor is demonstrated for fluorescence lifetime imaging through simulation and measurement results.

  2. From morphology to clinical pathophysiology: multiphoton fluorescence lifetime imaging at patients' bedside

    Science.gov (United States)

    Mess, Christian; Zens, Katharina; Gorzelanny, Christian; Metze, Dieter; Luger, Thomas A.; König, Karsten; Schneider, Stefan W.; Huck, Volker

    2017-02-01

    Application of multiphoton microscopy in the field of biomedical research and advanced diagnostics promises unique insights into the pathophysiology of skin diseases. By means of multiphoton excitation, endogenous biomolecules like NADH, collagen or elastin show autofluorescence or second harmonic generation. Thus, these molecules provide information about the subcellular morphology, epidermal architecture and physiological condition of the skin. To gain a deeper understanding of the linkage between cellular structure and physiological processes, non-invasive multiphotonbased intravital tomography (MPT) and fluorescence lifetime imaging (FLIM) were combined within the scopes of inflammatory skin, chronic wounds and drug delivery in clinical application. The optical biopsies generated via MPT were morphologically analyzed and aligned with classical skin histology. Because of its subcellular resolution, MPT provided evidence of a redistribution of mitochondria in keratinocytes, indicating an altered cellular metabolism. Independent morphometric algorithms reliably showed a perinuclear accumulation in lesional skin in contrast to an even distribution in healthy skin. Confirmatively, MPT-FLIM showed an obvious metabolic shift in lesions. Moreover, detection of the onset and progression of inflammatory processes could be achieved. The feasibility of primary in vivo tracking of applied therapeutic agents further broadened our scope: We examined the permeation and subsequent distribution of agents directly visualized in patientś skin in short-term repetitive measurements. Furthermore, we performed MPT-FLIM follow-up investigations in the long-term course of therapy. Therefore, clinical MPT-FLIM application offers new insights into the pathophysiology and the individual therapeutic course of skin diseases, facilitating a better understanding of the processes of inflammation and wound healing.

  3. Fluorescence lifetime FRET imaging of receptor-ligand complexes in tumor cells in vitro and in vivo

    Science.gov (United States)

    Rudkouskaya, Alena; Sinsuebphon, Nattawut; Intes, Xavier; Mazurkiewicz, Joseph E.; Barroso, Margarida

    2017-02-01

    To guide the development of targeted therapies with improved efficacy and accelerated clinical acceptance, novel imaging methodologies need to be established. Toward this goal, fluorescence lifetime Förster resonance energy transfer (FLIM-FRET) imaging assays capitalize on the ability of antibodies or protein ligands to bind dimerized membrane bound receptors to measure their target engagement levels in cancer cells. Conventional FLIM FRET microscopy has been widely applied at visible wavelengths to detect protein-protein interactions in vitro. However, operation at these wavelengths restricts imaging quality and ability to quantitate lifetime changes in in vivo small animal optical imaging due to high auto-fluorescence and light scattering. Here, we have analyzed the uptake of iron-bound transferrin (Tf) probes into human breast cancer cells using FLIM-FRET microscopy in the visible and near-infrared (NIR) range. The development of NIR FLIM FRET microscopy allows for the use of quantitative lifetime-based molecular assays to measure drug-target engagement levels at multiple scales: from in vitro microscopy to in vivo small animal optical imaging (macroscopy). This novel approach can be extended to other receptors, currently targeted in oncology. Hence, lifetime-based molecular imaging can find numerous applications in drug delivery and targeted therapy assessment and optimization.

  4. Room-temperature fluorescence lifetime of pseudoisocyanine (PIC) J excitons with various aggregate morphologies in relation to microcavity polariton formation.

    Science.gov (United States)

    Obara, Yuki; Saitoh, Keita; Oda, Masaru; Tani, Toshiro

    2012-01-01

    The results of room-temperature fluorescence lifetime measurements are reported for the excitation of J aggregates (Js) of pseudoisocyanine chloride (PIC-Cl) prepared in potassium polyvinyl sulfate (PVS) polymer thin films, their aqueous solutions, and NaCl aqueous solutions. Variations of the microscopic morphologies of the aggregates were investigated. The results show that fluorescence decay features correlated to the morphology change. The observed fluorescence lifetime and quantum efficiency of PIC J aggregates (PIC-Js) in a NaCl aqueous solution were 310 ps and 28%, respectively. The lifetime of the fibril-shaped macroaggregates prepared in PVS thin films was below the instrumental time resolution of 5 ps, and the efficiency decreased to below 3%. The results indicate that PIC-Js prepared with PVS polymers have an increased nonradiative contribution to the excitation deactivation process. In particular, macro-Js with isolated fibril-shaped structures revealed nonradiative pathway(s) that are closely associated to the specific packaging morphology of the constituent meso-Js. The possibility of a destructive effect on the formation of cavity-polaritons is also discussed.

  5. Validation of a regression model for standardizing lifetime racing performances of thoroughbreds.

    Science.gov (United States)

    Martin, G S; Strand, E; Kearney, M T

    1997-06-01

    To determine the relationship between prediction errors of a regression model of racing finish times and earnings or finish position; the relationship between standardized finish times, determined by use of this model, and earnings or finish position; and whether this model was valid when applied to data for horses that underwent surgical treatment. Survey. Records of 6,700 healthy Thoroughbreds racing in Louisiana and of 31 Thoroughbreds with idiopathic left laryngeal hemiplegia that underwent surgical treatment. Predicted and standardized finish times were calculated by use of the regression model for healthy horses, and the relationships between prediction error (actual--predicted finish time) and standardized finish times, and earnings and finish position, were examined. Then, the regression model was applied to data for horses with hemiplegia to determine whether the model was valid when used to calculate predicted and standardized finish times for lifetime performance data. Prediction error and standardized finish times were negatively correlated with earnings and positively correlated with finish position and, thus, appeared to be reliable measures of racing performance. The regression model was found to be valid when applied to lifetime performance records of horses with laryngeal hemiplegia. Prediction error and standardized finish times are measures of racing performance that can be used to compare performances among Thoroughbred racehorses across a variety of circumstances that would otherwise confound comparison.

  6. Study on the effect of deposition rate and concentration of Eu on the fluorescent lifetime of CsI: Tl thin film

    Science.gov (United States)

    Xie, Yijun; Guo, Lina; Liu, Shuang; Wang, Qianfeng; Zhang, Shangjian; Liu, Yong; Zhong, Zhiyong

    2017-06-01

    Although there are many new scintillators being developed recently, CsI: Tl is still very efficient among them. The fluorescent lifetime is a very important parameter of CsI: Tl thin film and two series of experiments have been conducted to learn about it. Our experiments, however, have demonstrated that the deposition rate and the codoping of Eu2+ will significantly influence its fluorescent lifetime. In order to increase the efficiency of the imaging system, we intend to obtain a higher fluorescent lifetime for CsI: Tl thin film by controlling these two conditions.

  7. A corrected likelihood approach for the nonlinear transformation model with application to fluorescence lifetime measurements using exponential mixtures.

    Science.gov (United States)

    Rebafka, Tabea; Roueff, François; Souloumiac, Antoine

    2010-01-01

    A fast and efficient estimation method is proposed that compensates the distortion in nonlinear transformation models. A likelihood-based estimator is developed that can be computed by an EM-type algorithm. The consistency of the estimator is shown and its limit distribution is provided. The new estimator is particularly well suited for fluorescence lifetime measurements, where only the shortest arrival time of a random number of emitted fluorescence photons can be detected and where arrival times are often modeled by a mixture of exponential distributions. The method is evaluated on real and synthetic data. Compared to currently used methods in fluorescence, the new estimator should allow a reduction of the acquisition time of an order of magnitude.

  8. Lifetime growth in wild meerkats: incorporating life history and environmental factors into a standard growth model.

    Science.gov (United States)

    English, Sinéad; Bateman, Andrew W; Clutton-Brock, Tim H

    2012-05-01

    Lifetime records of changes in individual size or mass in wild animals are scarce and, as such, few studies have attempted to model variation in these traits across the lifespan or to assess the factors that affect them. However, quantifying lifetime growth is essential for understanding trade-offs between growth and other life history parameters, such as reproductive performance or survival. Here, we used model selection based on information theory to measure changes in body mass over the lifespan of wild meerkats, and compared the relative fits of several standard growth models (monomolecular, von Bertalanffy, Gompertz, logistic and Richards). We found that meerkats exhibit monomolecular growth, with the best model incorporating separate growth rates before and after nutritional independence, as well as effects of season and total rainfall in the previous nine months. Our study demonstrates how simple growth curves may be improved by considering life history and environmental factors, which may be particularly relevant when quantifying growth patterns in wild populations.

  9. Absorption, steady-state fluorescence, fluorescence lifetime, and 2D self-assembly properties of engineered fluorescent S-layer fusion proteins of Geobacillus stearothermophilus NRS 2004/3a.

    Science.gov (United States)

    Kainz, Birgit; Steiner, Kerstin; Möller, Marco; Pum, Dietmar; Schäffer, Christina; Sleytr, Uwe B; Toca-Herrera, José L

    2010-01-11

    S-layer fusion protein technology was used to design four different fluorescent fusion proteins with three different GFP mutants and the red fluorescent protein mRFP1. Their absorption spectra, steady-state fluorescence, and fluorescence lifetime were investigated as a function of pH. It was found that fluorescence intensities and lifetime of the GFP mutant S-layer fusion proteins decreased about 50% between pH 6 and pH 5. The spectral properties of the red S-layer fusion protein were minimally affected by pH variations. These results were compared with His-tagged reference fluorescent proteins, demonstrating that the S-layer protein did not change the general spectral properties of the whole fusion protein. In addition, the pK(a) values of the fluorescent S-layer fusion proteins were calculated. Finally, it was shown that the S-layer fusion proteins were able to self-assemble forming 2D nanostructures of oblique p2 symmetry with lattice parameters of about a = 11 nm, b = 14 nm, and gamma = 80 degrees . The fluorescence tag did not hinder the natural self-assembly process of the S-layer protein. The combination of the fluorescence properties and the self-assembly ability of the engineered fusion proteins make them a promising tool to generate biomimetic surfaces for future applications in nanobiotechnology at a wide range of pH.

  10. Constraining the lifetime and opening angle of quasars using fluorescent Ly a emission: the case of Q0420-388

    CERN Document Server

    Borisova, Elena; Cantalupo, Sebastiano; Prochaska, J Xavier; Rakic, Olivera; Worseck, Gabor

    2015-01-01

    A toy model is developed to understand how the spatial distribution of fluorescent emitters in the vicinity of bright quasars could be affected by the geometry of the quasar bi-conical radiation field and by the quasar lifetime. We then compare the predictions of this model to a sample of high equivalent width Ly a emitters (EW0 > 100 A) that were identified in a deep narrow-band 36x36 arcmin2 image centered on the luminous quasar Q0420-388. These are identified to the edge of the field and show some evidence of an azimuthal asymmetry on the sky of the type expected if the quasar is radiating in a bipolar cone. If these sources are being fluorescently illuminated by the quasar, then the two most distant sources require a lifetime of at least 15 Myr for an opening angle of 60 degrees or more, increasing to more than 40 Myr if the opening angle is reduced to a minimum 30 degrees. The overall distribution of all of the sources across the field gives best fit lifetimes in the range 20 < t < 50 Myr for openi...

  11. DNA origami-based standards for quantitative fluorescence microscopy.

    Science.gov (United States)

    Schmied, Jürgen J; Raab, Mario; Forthmann, Carsten; Pibiri, Enrico; Wünsch, Bettina; Dammeyer, Thorben; Tinnefeld, Philip

    2014-01-01

    Validating and testing a fluorescence microscope or a microscopy method requires defined samples that can be used as standards. DNA origami is a new tool that provides a framework to place defined numbers of small molecules such as fluorescent dyes or proteins in a programmed geometry with nanometer precision. The flexibility and versatility in the design of DNA origami microscopy standards makes them ideally suited for the broad variety of emerging super-resolution microscopy methods. As DNA origami structures are durable and portable, they can become a universally available specimen to check the everyday functionality of a microscope. The standards are immobilized on a glass slide, and they can be imaged without further preparation and can be stored for up to 6 months. We describe a detailed protocol for the design, production and use of DNA origami microscopy standards, and we introduce a DNA origami rectangle, bundles and a nanopillar as fluorescent nanoscopic rulers. The protocol provides procedures for the design and realization of fluorescent marks on DNA origami structures, their production and purification, quality control, handling, immobilization, measurement and data analysis. The procedure can be completed in 1-2 d.

  12. Fluorescence lifetime and UV-Vis spectroscopy to evaluate the interactions between quercetin and its yeast microcapsule.

    Science.gov (United States)

    Pham-Hoang, Bao-Ngoc; Winckler, Pascale; Waché, Yves

    2017-09-09

    Quercetin is a fragile bioactive compound. Several works have tried to preserve it by encapsulation but the form of encapsulation (mono- or supra-molecular structure, tautomeric form), though important for stability and bioavailability, remains unknown. The present work aims at developing a fluorescence lifetime technique to evaluate the structure of quercetin during encapsulation in a vector capsule that has already proven efficiency, yeast cells. Molecular stabilization was observed during a four-month storage period. The time-correlated single-photon counting (TCSPC) technique was used to evaluate the interaction between quercetin molecules and the yeast capsule. The various tautomeric forms, as identified by UV-Vis spectroscopy, resulted in various lifetimes in TCSPC, although they varied also with the buffer environment. Quercetin in buffer exhibited a three-to-four longer long time after 24 h (changing from 6-7 to 18-23 ns), suggesting an aggregation of molecules. In yeast microcapsules, the long-time population exhibited a longer lifetime (around 27 ns) from the beginning and concerned about 20% of molecules compared to dispersed quercetin. This shows that lifetime analysis can show the monomolecular instability of quercetin in buffer and the presence of interactions between quercetin molecules and their microcapsules. This article is protected by copyright. All rights reserved.

  13. A Rotational BODIPY Nucleotide: An Environment-Sensitive Fluorescence-Lifetime Probe for DNA Interactions and Applications in Live-Cell Microscopy.

    Science.gov (United States)

    Dziuba, Dmytro; Jurkiewicz, Piotr; Cebecauer, Marek; Hof, Martin; Hocek, Michal

    2016-01-04

    Fluorescent probes for detecting the physical properties of cellular structures have become valuable tools in life sciences. The fluorescence lifetime of molecular rotors can be used to report on variations in local molecular packing or viscosity. We used a nucleoside linked to a meso-substituted BODIPY fluorescent molecular rotor (dC(bdp)) to sense changes in DNA microenvironment both in vitro and in living cells. DNA incorporating dC(bdp) can respond to interactions with DNA-binding proteins and lipids by changes in the fluorescence lifetimes in the range 0.5-2.2 ns. We can directly visualize changes in the local environment of exogenous DNA during transfection of living cells. Relatively long fluorescence lifetimes and extensive contrast for detecting changes in the microenvironment together with good photostability and versatility for DNA synthesis make this probe suitable for analysis of DNA-associated processes, cellular structures, and also DNA-based nanomaterials.

  14. Two-photon-excited fluorescence (TPEF) and fluorescence lifetime imaging (FLIM) with sub-nanosecond pulses and a high analog bandwidth signal detection

    Science.gov (United States)

    Eibl, Matthias; Karpf, Sebastian; Hakert, Hubertus; Weng, Daniel; Huber, Robert

    2017-02-01

    Two-photon excited fluorescence (TPEF) microscopy and fluorescence lifetime imaging (FLIM) are powerful imaging techniques in bio-molecular science. The need for elaborate light sources for TPEF and speed limitations for FLIM, however, hinder an even wider application. We present a way to overcome this limitations by combining a robust and inexpensive fiber laser for nonlinear excitation with a fast analog digitization method for rapid FLIM imaging. The applied sub nanosecond pulsed laser source is synchronized to a high analog bandwidth signal detection for single shot TPEF- and single shot FLIM imaging. The actively modulated pulses at 1064nm from the fiber laser are adjustable from 50ps to 5ns with kW of peak power. At a typically applied pulse lengths and repetition rates, the duty cycle is comparable to typically used femtosecond pulses and thus the peak power is also comparable at same cw-power. Hence, both types of excitation should yield the same number of fluorescence photons per time on average when used for TPEF imaging. However, in the 100ps configuration, a thousand times more fluorescence photons are generated per pulse. In this paper, we now show that the higher number of fluorescence photons per pulse combined with a high analog bandwidth detection makes it possible to not only use a single pulse per pixel for TPEF imaging but also to resolve the exponential time decay for FLIM. To evaluate the performance of our system, we acquired FLIM images of a Convallaria sample with pixel rates of 1 MHz where the lifetime information is directly measured with a fast real time digitizer. With the presented results, we show that longer pulses in the many-10ps to nanosecond regime can be readily applied for TPEF imaging and enable new imaging modalities like single pulse FLIM.

  15. New solutions for standardization, monitoring and quality management of fluorescence-based imaging systems (Conference Presentation)

    Science.gov (United States)

    Royon, Arnaud; Papon, Gautier

    2016-03-01

    Fluorescence microscopes have become ubiquitous in life sciences laboratories, including those focused on pharmaceuticals, diagnosis, and forensics. For the past few years, the need for both performance guarantees and quantifiable results has driven development in this area. However, the lack of appropriate standards and reference materials makes it difficult or impossible to compare the results of two fluorescence microscopes, or to measure performance fluctuations of one microscope over time. Therefore, the operation of fluorescence microscopes is not monitored as often as their use warrants - an issue that is recognized by both systems manufacturers and national metrology institutes. We have developed a new process that enables the etching of long-term stable fluorescent patterns with sub-micrometer sizes in three dimensions inside glass. In this paper, we present, based on this new process, a fluorescent multi-dimensional ruler and a dedicated software that are suitable for monitoring and quality management of fluorescence-based imaging systems (wide-field, confocal, multiphoton, high content machines). In addition to fluorescence, the same patterns exhibit bright- and dark-field contrast, DIC, and phase contrast, which make them also relevant to monitor these types of microscopes. Non-exhaustively, this new solution enables the measurement of: The stage repositioning accuracy; The illumination and detection homogeneities; The field flatness; The detectors' characteristics; The lateral and axial spatial resolutions; The spectral response (spectrum, intensity and lifetime) of the system. Thanks to the stability of the patterns, microscope performance assessment can be carried out as well in a daily basis as in the long term.

  16. Determining a fluorophore’s transition dipole moment from fluorescence lifetime measurements in solvents of varying refractive index

    Science.gov (United States)

    Chung, Pei-Hua; Tregidgo, Carolyn; Suhling, Klaus

    2016-12-01

    The transition dipole moment of organic dyes PM546 and rhodamine 123 is calculated from fluorescence lifetime measurements in solutions of different refractive index. A model proposed by Toptygin et al (2002 J. Phys. Chem. B 106 3724-34) provides a relationship between the radiative rate constant and refractive index of the solvent, and allows the electronic transition dipole moments to be found: it is (7.1  ±  1.1) D for PM546 which matches that found in the literature, and (8.1  ±  0.1) D for rhodamine 123. Toptygin’s model goes further in predicting the shape of the fluorescent dye and here we predict the shape of PM546 and rhodamine 123 to be ellipsoidal.

  17. Signal peptide peptidase (SPP dimer formation as assessed by fluorescence lifetime imaging microscopy (FLIM in intact cells

    Directory of Open Access Journals (Sweden)

    Nyborg Andrew C

    2006-11-01

    Full Text Available Abstract Background Signal peptide peptidase (SPP is an intramembrane cleaving protease identified by its cleavage of several type II membrane signal peptides. Conservation of intramembrane active site residues demonstrates that SPP, SPP family members, and presenilins (PSs make up a family of intramembrane cleaving proteases. Because SPP appears to function without additional protein cofactors, the study of SPP may provide structural insights into the mechanism of intramembrane proteolysis by this biomedically important family of proteins. Previous studies have shown that SPP isolated from cells appears to be a homodimer, but some evidence exists that in vitro SPP may be active as a monomer. We have conducted additional experiments to determine if SPP exists as a monomer or dimer in vivo. Results Fluorescence lifetime imaging microscopy (FLIM can be is used to determine intra- or intermolecular interactions by fluorescently labeling epitopes on one or two different molecules. If the donor and acceptor fluorophores are less than 10 nm apart, the donor fluorophore lifetime shortens proportionally to the distance between the fluorophores. In this study, we used two types of fluorescence energy transfer (FRET pairs; cyan fluorescent protein (CFP with yellow fluorescent protein (YFP or Alexa 488 with Cy3 to differentially label the NH2- or COOH-termini of SPP molecules. A cell based SPP activity assay was used to show that all tagged SPP proteins are proteolytically active. Using FLIM we were able to show that the donor fluorophore lifetime of the CFP tagged SPP construct in living cells significantly decreases when either a NH2- or COOH-terminally YFP tagged SPP construct is co-transfected, indicating close proximity between two different SPP molecules. These data were then confirmed in cell lines stably co-expressing V5- and FLAG-tagged SPP constructs. Conclusion Our FLIM data strongly suggest dimer formation between two separate SPP proteins

  18. Asante Calcium Green and Asante Calcium Red--novel calcium indicators for two-photon fluorescence lifetime imaging.

    Science.gov (United States)

    Jahn, Karolina; Hille, Carsten

    2014-01-01

    For a comprehensive understanding of cellular processes and potential dysfunctions therein, an analysis of the ubiquitous intracellular second messenger calcium is of particular interest. This study examined the suitability of the novel Ca2+-sensitive fluorescent dyes Asante Calcium Red (ACR) and Asante Calcium Green (ACG) for two-photon (2P)-excited time-resolved fluorescence measurements. Both dyes displayed sufficient 2P fluorescence excitation in a range of 720-900 nm. In vitro, ACR and ACG exhibited a biexponential fluorescence decay behavior and the two decay time components in the ns-range could be attributed to the Ca(2+)-free and Ca(2+)-bound dye species. The amplitude-weighted average fluorescence decay time changed in a Ca(2+)-dependent way, unraveling in vitro dissociation constants K(D) of 114 nM and 15 nM for ACR and ACG, respectively. In the presence of bovine serum albumin, the absorption and steady-state fluorescence behavior of ACR was altered and its biexponential fluorescence decay showed about 5-times longer decay time components indicating dye-protein interactions. Since no ester derivative of ACG was commercially available, only ACR was evaluated for 2P-excited fluorescence lifetime imaging microscopy (2P-FLIM) in living cells of American cockroach salivary glands. In living cells, ACR also exhibited a biexponential fluorescence decay with clearly resolvable short (0.56 ns) and long (2.44 ns) decay time components attributable to the Ca(2+)-free and Ca(2+)-bound ACR species. From the amplitude-weighted average fluorescence decay times, an in situ K(D) of 180 nM was determined. Thus, quantitative [Ca(2+)]i recordings were realized, unraveling a reversible dopamine-induced [Ca(2+)]i elevation from 21 nM to 590 nM in salivary duct cells. It was concluded that ACR is a promising new Ca(2+) indicator dye for 2P-FLIM recordings applicable in diverse biological systems.

  19. Asante Calcium Green and Asante Calcium Red--novel calcium indicators for two-photon fluorescence lifetime imaging.

    Directory of Open Access Journals (Sweden)

    Karolina Jahn

    Full Text Available For a comprehensive understanding of cellular processes and potential dysfunctions therein, an analysis of the ubiquitous intracellular second messenger calcium is of particular interest. This study examined the suitability of the novel Ca2+-sensitive fluorescent dyes Asante Calcium Red (ACR and Asante Calcium Green (ACG for two-photon (2P-excited time-resolved fluorescence measurements. Both dyes displayed sufficient 2P fluorescence excitation in a range of 720-900 nm. In vitro, ACR and ACG exhibited a biexponential fluorescence decay behavior and the two decay time components in the ns-range could be attributed to the Ca(2+-free and Ca(2+-bound dye species. The amplitude-weighted average fluorescence decay time changed in a Ca(2+-dependent way, unraveling in vitro dissociation constants K(D of 114 nM and 15 nM for ACR and ACG, respectively. In the presence of bovine serum albumin, the absorption and steady-state fluorescence behavior of ACR was altered and its biexponential fluorescence decay showed about 5-times longer decay time components indicating dye-protein interactions. Since no ester derivative of ACG was commercially available, only ACR was evaluated for 2P-excited fluorescence lifetime imaging microscopy (2P-FLIM in living cells of American cockroach salivary glands. In living cells, ACR also exhibited a biexponential fluorescence decay with clearly resolvable short (0.56 ns and long (2.44 ns decay time components attributable to the Ca(2+-free and Ca(2+-bound ACR species. From the amplitude-weighted average fluorescence decay times, an in situ K(D of 180 nM was determined. Thus, quantitative [Ca(2+]i recordings were realized, unraveling a reversible dopamine-induced [Ca(2+]i elevation from 21 nM to 590 nM in salivary duct cells. It was concluded that ACR is a promising new Ca(2+ indicator dye for 2P-FLIM recordings applicable in diverse biological systems.

  20. Incorporating a piperidinyl group in the fluorophore extends the fluorescence lifetime of click-derived cyclam-naphthalimide conjugates.

    Science.gov (United States)

    Yu, Mingfeng; Ast, Sandra; Yu, Qun; Lo, Anthony T S; Flehr, Roman; Todd, Matthew H; Rutledge, Peter J

    2014-01-01

    Ligands incorporating a tetraazamacrocycle receptor, a 'click'-derived triazole and a 1,8-naphthalimide fluorophore have proven utility as probes for metal ions. Three new cyclam-based molecular probes are reported, in which a piperidinyl group has been introduced at the 4-position of the naphthalimide fluorophore. These compounds have been synthesized using the copper(I)-catalyzed azide-alkyne Huisgen cycloaddition and their photophysical properties studied in detail. The alkylamino group induces the expected red-shift in absorption and emission spectra relative to the simple naphthalimide derivatives and gives rise to extended fluorescence lifetimes in aqueous buffer. The photophysical properties of these systems are shown to be highly solvent-dependent. Screening the fluorescence responses of the new conjugates to a wide variety of metal ions reveals significant and selective fluorescence quenching in the presence of copper(II), yet no fluorescence enhancement with zinc(II) as observed previously for the simple naphthalimide derivatives. Reasons for this different behaviour are proposed. Cytotoxicity testing shows that these new cyclam-triazole-dye conjugates display little or no toxicity against either DLD-1 colon carcinoma cells or MDA-MB-231 breast carcinoma cells, suggesting a potential role for these and related systems in biological sensing applications.

  1. Incorporating a piperidinyl group in the fluorophore extends the fluorescence lifetime of click-derived cyclam-naphthalimide conjugates.

    Directory of Open Access Journals (Sweden)

    Mingfeng Yu

    Full Text Available Ligands incorporating a tetraazamacrocycle receptor, a 'click'-derived triazole and a 1,8-naphthalimide fluorophore have proven utility as probes for metal ions. Three new cyclam-based molecular probes are reported, in which a piperidinyl group has been introduced at the 4-position of the naphthalimide fluorophore. These compounds have been synthesized using the copper(I-catalyzed azide-alkyne Huisgen cycloaddition and their photophysical properties studied in detail. The alkylamino group induces the expected red-shift in absorption and emission spectra relative to the simple naphthalimide derivatives and gives rise to extended fluorescence lifetimes in aqueous buffer. The photophysical properties of these systems are shown to be highly solvent-dependent. Screening the fluorescence responses of the new conjugates to a wide variety of metal ions reveals significant and selective fluorescence quenching in the presence of copper(II, yet no fluorescence enhancement with zinc(II as observed previously for the simple naphthalimide derivatives. Reasons for this different behaviour are proposed. Cytotoxicity testing shows that these new cyclam-triazole-dye conjugates display little or no toxicity against either DLD-1 colon carcinoma cells or MDA-MB-231 breast carcinoma cells, suggesting a potential role for these and related systems in biological sensing applications.

  2. Study of thioflavin-T immobilized in porous silicon and the effect of different organic vapors on the fluorescence lifetime.

    Science.gov (United States)

    Hutter, Tanya; Amdursky, Nadav; Gepshtein, Rinat; Elliott, Stephen R; Huppert, Dan

    2011-06-21

    Steady-state and time-resolved emission techniques have been employed to study the fluorescence properties of thioflavin-T (ThT) adsorbed on oxidized porous silicon (PSi) surfaces, with an average pore size of ∼10 nm. We found that the average fluorescence decay time of ThT, when it is adsorbed on the PSi surface, is rather long, τ(av) = 1.3 ns. We attribute this relatively long emission lifetime to the effect of the immobilization of ThT on the PSi surface, which inhibit the rotation of the aniline with respect to the benzothiazole moieties of ThT. We also measured the fluorescence properties of ThT in PSi samples in equilibrium with vapors of several liquids, such as methanol, acetonitrile, and water. We found that the fluorescence intensity drops by a factor of 10, and the average decay time, measured by a time-correlated single-photon counting technique, decreases by a factor of 3. We explain these results in terms of liquid condensation of the vapors in the PSi pores, which leads to partial dissolution of the ThT molecules in the liquid pools.

  3. Novel aspects of fluorescence lifetime for molecules positioned close to metal surfaces

    Science.gov (United States)

    Aussenegg, F. R.; Leitner, A.; Lippitsch, M. E.; Reinisch, H.; Riegler, M.

    1987-10-01

    On metal surfaces with submicroscopic corrugations, surface-enhanced optical processes can be observed. Results obtained by picosecond time-resolved fluorescence spectroscopy for dye molecules in the proximity (0-50 nm) of silver islands films are reported. It is demonstrated how the rather complex dependence of the integral fluorescence intensity on the distance dye-islands, can be resolved in the contributions of different mechanisms by analysing the fluorescence decay curves at various distances. It turns out, that the enhancement of absorption influences only the peak fluorescence intensity without changing the decay time, while the enhancement of emission and dissipative losses reduces the decay time. Thus time-resolved spectroscopy opens the possibility to test theoretical concepts on surface enhancement and provides basic data for tailoring molecule-metal structures with well-defined surface-enhancement properties.

  4. Lifetime Fluorescence and Raman Imaging for Detection of Wound Failure and Heterotopic Ossification

    Science.gov (United States)

    2015-12-01

    5–8]. HO also arises in spinal injuries, severe burns and in the surgical beds resulting from orthopedic surgery complications [1,6,9,10]. The...P. V. Butte, B. K. Pikul, A. Hever, W. H. Yong, K. L. Black, and L. Marcu, "Diagnosis of meningioma by time-resolved fluorescence spectroscopy

  5. A low cost fluorescence lifetime measurement system based on SPAD detectors and FPGA processing

    Science.gov (United States)

    Franch, N.; Alonso, O.; Canals, J.; Vilà, A.; Dieguez, A.

    2017-02-01

    This work presents a low cost fluorescence life time measurement system, aimed at carrying out fast diagnostic tests through label detection in a portable system so it can be used in a medical consultation, within a short time span. The system uses Time Correlated Single Photon Counting (TCSPC), measuring the arrival time of individual photons and building a histogram of those times, showing the fluorescence decay of the label which is characteristic of each fluorescent substance. The system is implemented using a Xilinx FPGA which controls the experiment and includes a Time to Digital Converter (TDC) to perform measurements with a resolution in the order of tenths of picoseconds. Also included are a laser diode and the driving electronics to generate short pulses as well as a HV-CMOS implemented Single Photon Avalanche Diode (SPAD) as a high gain sensor. The system is entirely configurable so it can easily be adapted to the target label molecule and measurement needs. The histogram is constructed within the FPGA and can then be read as convenient. Various performance parameters are also shown, as well as experimental measurements of a quantum dot fluorescence decay as a proof of concept.

  6. A fusion-spliced near-field optical fiber probe using photonic crystal fiber for nanoscale thermometry based on fluorescence-lifetime measurement of quantum dots.

    Science.gov (United States)

    Fujii, Takuro; Taguchi, Yoshihiro; Saiki, Toshiharu; Nagasaka, Yuji

    2011-01-01

    We have developed a novel nanoscale temperature-measurement method using fluorescence in the near-field called fluorescence near-field optics thermal nanoscopy (Fluor-NOTN). Fluor-NOTN enables the temperature distributions of nanoscale materials to be measured in vivo/in situ. The proposed method measures temperature by detecting the temperature dependent fluorescence lifetimes of Cd/Se quantum dots (QDs). For a high-sensitivity temperature measurement, the auto-fluorescence generated from a fiber probe should be reduced. In order to decrease the noise, we have fabricated a novel near-field optical-fiber probe by fusion-splicing a photonic crystal fiber (PCF) and a conventional single-mode fiber (SMF). The validity of the novel fiber probe was assessed experimentally by evaluating the auto-fluorescence spectra of the PCF. Due to the decrease of auto-fluorescence, a six- to ten-fold increase of S/N in the near-field fluorescence lifetime detection was achieved with the newly fabricated fusion-spliced near-field optical fiber probe. Additionally, the near-field fluorescence lifetime of the quantum dots was successfully measured by the fabricated fusion-spliced near-field optical fiber probe at room temperature, and was estimated to be 10.0 ns.

  7. A Fusion-Spliced Near-Field Optical Fiber Probe Using Photonic Crystal Fiber for Nanoscale Thermometry Based on Fluorescence-Lifetime Measurement of Quantum Dots

    Directory of Open Access Journals (Sweden)

    Toshiharu Saiki

    2011-08-01

    Full Text Available We have developed a novel nanoscale temperature-measurement method using fluorescence in the near-field called Fluorescence Near-field Optics Thermal Nanoscopy (Fluor-NOTN. Fluor-NOTN enables the temperature distributions of nanoscale materials to be measured in vivo/in situ. The proposed method measures temperature by detecting the temperature dependent fluorescence lifetimes of Cd/Se Quantum Dots (QDs. For a high-sensitivity temperature measurement, the auto-fluorescence generated from a fiber probe should be reduced. In order to decrease the noise, we have fabricated a novel near-field optical-fiber probe by fusion-splicing a photonic crystal fiber (PCF and a conventional single-mode fiber (SMF. The validity of the novel fiber probe was assessed experimentally by evaluating the auto-fluorescence spectra of the PCF. Due to the decrease of auto-fluorescence, a six- to ten-fold increase of S/N in the near-field fluorescence lifetime detection was achieved with the newly fabricated fusion-spliced near-field optical fiber probe. Additionally, the near-field fluorescence lifetime of the quantum dots was successfully measured by the fabricated fusion-spliced near-field optical fiber probe at room temperature, and was estimated to be 10.0 ns.

  8. Nuclear uptake of ultrasmall gold-doxorubicin conjugates imaged by fluorescence lifetime imaging microscopy (FLIM) and electron microscopy

    Science.gov (United States)

    Zhang, Xuan; Shastry, Sathvik; Bradforth, Stephen E.; Nadeau, Jay L.

    2014-11-01

    Fluorescence lifetime imaging microscopy (FLIM) has been used to image free and encapsulated doxorubicin (Dox) uptake into cells, since interaction of Dox with DNA leads to a characteristic lifetime change. However, none of the reported Dox conjugates were able to enter cell nuclei. In this work, we use FLIM to show nuclear uptake of 2.7 nm mean diameter Au nanoparticles conjugated to Dox. The pattern of labelling differed substantially from what was seen with free Dox, with slower nuclear entry and stronger cytoplasmic labelling at all time points. As the cells died, the pattern of labelling changed further as intracellular structures disintegrated, consistent with association of Au-Dox to membranes. The patterns of Au distribution and intracellular structure changes were confirmed using electron microscopy, and indicate different mechanisms of cytotoxicity with stable Au-Dox conjugates compared to Dox alone. Such conjugates are promising tools for overcoming resistance in Dox-resistant cancers.Fluorescence lifetime imaging microscopy (FLIM) has been used to image free and encapsulated doxorubicin (Dox) uptake into cells, since interaction of Dox with DNA leads to a characteristic lifetime change. However, none of the reported Dox conjugates were able to enter cell nuclei. In this work, we use FLIM to show nuclear uptake of 2.7 nm mean diameter Au nanoparticles conjugated to Dox. The pattern of labelling differed substantially from what was seen with free Dox, with slower nuclear entry and stronger cytoplasmic labelling at all time points. As the cells died, the pattern of labelling changed further as intracellular structures disintegrated, consistent with association of Au-Dox to membranes. The patterns of Au distribution and intracellular structure changes were confirmed using electron microscopy, and indicate different mechanisms of cytotoxicity with stable Au-Dox conjugates compared to Dox alone. Such conjugates are promising tools for overcoming resistance in

  9. The Gray Institute ‘open’ high-content, fluorescence lifetime microscopes

    OpenAIRE

    Barber, PR; TULLIS, IDC; PIERCE, GP; Newman, RG; PRENTICE, J; Rowley, MI; Matthews, DR; AMEER-BEG, SM; Vojnovic, B

    2013-01-01

    Summary We describe a microscopy design methodology and details of microscopes built to this ‘open’ design approach. These demonstrate the first implementation of time-domain fluorescence microscopy in a flexible automated platform with the ability to ease the transition of this and other advanced microscopy techniques from development to use in routine biology applications. This approach allows easy expansion and modification of the platform capabilities, as it moves away from the use of a c...

  10. In vivo and in vitro investigations of retinal fluorophores in age-related macular degeneration by fluorescence lifetime imaging

    Science.gov (United States)

    Hammer, M.; Quick, S.; Klemm, M.; Schenke, S.; Mata, N.; Eitner, A.; Schweitzer, D.

    2009-02-01

    Ocular fundus autofluorescence imaging has been introduced into clinical diagnostics recently for the observation of the age pigment lipofuscin, a precursor of age-related macular degeneration (AMD). However, a deeper understanding of the generation of single compounds contributing to the lipofuscin as well as of the role of other fluorophores such as FAD, glycated proteins, and collagen needs their discrimination by fluorescence lifetime imaging (FLIM). FLIM at the ocular fundus is performed using a scanning laser ophthalmoscope equipped with a picosecond laser source (448nm or 468nm respectively, 100ps, 80 MHz repetition rate) and dual wavelength (490-560nm and 560-7600nm) time-correlated single photon counting. A three-exponential fit of the fluorescence decay revealed associations of decay times to anatomical structures. Disease-related features are identified from alterations in decay times and-amplitudes. The in-vivo investigations in patients were paralleled by experiments in an organ culture of the porcine ocular fundus. Photo-oxidative stress was induced by exposure to blue light (467nm, 0.41 mW/mm2). Subsequent analysis (fluorescence microscopy, HPLC, LC-MS) indicated the accumulation of the pyridinium bis-retinoid A2E and its oxidation products as well as oxidized phospholipids. These compounds contribute to the tissue auto-fluorescence and may play a key role in the pathogenesis of AMD. Thus, FLIM observation at the ocular fundus in vivo enhances our knowledge on the etiology of AMD and may become a diagnostic tool.

  11. Development of a Multi-modal Tissue Diagnostic System Combining High Frequency Ultrasound and Photoacoustic Imaging with Lifetime Fluorescence Spectroscopy

    Science.gov (United States)

    Sun, Yang; Stephens, Douglas N.; Park, Jesung; Sun, Yinghua; Marcu, Laura; Cannata, Jonathan M.; Shung, K. Kirk

    2010-01-01

    We report the development and validate a multi-modal tissue diagnostic technology, which combines three complementary techniques into one system including ultrasound backscatter microscopy (UBM), photoacoustic imaging (PAI), and time-resolved laser-induced fluorescence spectroscopy (TR-LIFS). UBM enables the reconstruction of the tissue microanatomy. PAI maps the optical absorption heterogeneity of the tissue associated with structure information and has the potential to provide functional imaging of the tissue. Examination of the UBM and PAI images allows for localization of regions of interest for TR-LIFS evaluation of the tissue composition. The hybrid probe consists of a single element ring transducer with concentric fiber optics for multi-modal data acquisition. Validation and characterization of the multi-modal system and ultrasonic, photoacoustic, and spectroscopic data coregistration were conducted in a physical phantom with properties of ultrasound scattering, optical absorption, and fluorescence. The UBM system with the 41 MHz ring transducer can reach the axial and lateral resolution of 30 and 65 μm, respectively. The PAI system with 532 nm excitation light from a Nd:YAG laser shows great contrast for the distribution of optical absorbers. The TR-LIFS system records the fluorescence decay with the time resolution of ~300 ps and a high sensitivity of nM concentration range. Biological phantom constructed with different types of tissues (tendon and fat) was used to demonstrate the complementary information provided by the three modalities. Fluorescence spectra and lifetimes were compared to differentiate chemical composition of tissues at the regions of interest determined by the coregistered high resolution UBM and PAI image. Current results demonstrate that the fusion of these techniques enables sequentially detection of functional, morphological, and compositional features of biological tissue, suggesting potential applications in diagnosis of tumors

  12. Single Cell Assay for Molecular Diagnostics and Medicine: Monitoring Intracellular Concentrations of Macromolecules by Two-photon Fluorescence Lifetime Imaging.

    Science.gov (United States)

    Pliss, Artem; Peng, Xiao; Liu, Lixin; Kuzmin, Andrey; Wang, Yan; Qu, Junle; Li, Yuee; Prasad, Paras N

    2015-01-01

    Molecular organization of a cell is dynamically transformed along the course of cellular physiological processes, pathologic developments or derived from interactions with drugs. The capability to measure and monitor concentrations of macromolecules in a single cell would greatly enhance studies of cellular processes in heterogeneous populations. In this communication, we introduce and experimentally validate a bio-analytical single-cell assay, wherein the overall concentration of macromolecules is estimated in specific subcellular domains, such as structure-function compartments of the cell nucleus as well as in nucleoplasm. We describe quantitative mapping of local biomolecular concentrations, either intrinsic relating to the functional and physiological state of a cell, or altered by a therapeutic drug action, using two-photon excited fluorescence lifetime imaging (FLIM). The proposed assay utilizes a correlation between the fluorescence lifetime of fluorophore and the refractive index of its microenvironment varying due to changes in the concentrations of macromolecules, mainly proteins. Two-photon excitation in Near-Infra Red biological transparency window reduced the photo-toxicity in live cells, as compared with a conventional single-photon approach. Using this new assay, we estimated average concentrations of proteins in the compartments of nuclear speckles and in the nucleoplasm at ~150 mg/ml, and in the nucleolus at ~284 mg/ml. Furthermore, we show a profound influence of pharmaceutical inhibitors of RNA synthesis on intracellular protein density. The approach proposed here will significantly advance theranostics, and studies of drug-cell interactions at the single-cell level, aiding development of personal molecular medicine.

  13. Label-free separation of human embryonic stem cells and their differentiating progenies by phasor fluorescence lifetime microscopy

    Science.gov (United States)

    Stringari, Chiara; Sierra, Robert; Donovan, Peter J.; Gratton, Enrico

    2012-04-01

    We develop a label-free optical technique to image and discriminate undifferentiated human embryonic stem cells (hESCs) from their differentiating progenies in vitro. Using intrinsic cellular fluorophores, we perform fluorescence lifetime microscopy (FLIM) and phasor analysis to obtain hESC metabolic signatures. We identify two optical biomarkers to define the differentiation status of hESCs: Nicotinamide adenine dinucleotide (NADH) and lipid droplet-associated granules (LDAGs). These granules have a unique lifetime signature and could be formed by the interaction of reactive oxygen species and unsaturated metabolic precursor that are known to be abundant in hESC. Changes in the relative concentrations of these two intrinsic biomarkers allow for the discrimination of undifferentiated hESCs from differentiating hESCs. During early hESC differentiation we show that NADH concentrations increase, while the concentration of LDAGs decrease. These results are in agreement with a decrease in oxidative phosphorylation rate. Single-cell phasor FLIM signatures reveal an increased heterogeneity in the metabolic states of differentiating H9 and H1 hESC colonies. This technique is a promising noninvasive tool to monitor hESC metabolism during differentiation, which can have applications in high throughput analysis, drug screening, functional metabolomics and induced pluripotent stem cell generation.

  14. Fluorescence lifetime imaging of DAPI-stained nuclei as a novel diagnostic tool for the detection and classification of B-cell chronic lymphocytic leukemia.

    Science.gov (United States)

    Yahav, Gilad; Hirshberg, Abraham; Salomon, Ophira; Amariglio, Ninette; Trakhtenbrot, Luba; Fixler, Dror

    2016-07-01

    B-cell chronic lymphocytic leukaemia (B-CLL) and B-cell precursor acute lymphoblastic leukaemia (B-ALL) are the most common type of leukaemia in adults and children, respectively. Today, fluorescence in situ hybridization (FISH) is the standard for detecting chromosomal aberrations that reflect adverse and favorable outcome. This study revealed a new, simple, and fast diagnostic tool to detect pathological cells by measuring and imaging the fluorescence lifetime (FLT) using FLT imaging microscopy (FLIM) of the peripheral blood (PB) cells of B-CLL samples that were labeled with the DNA binder, DAPI. The FLT of DAPI in healthy individuals was found to be 2.66 ± 0.12 ns. In contrast, PB cells of B-CLL and BM cells of B-ALL patients were characterized by a specific group distribution of the FLT values. The FLT of DAPI was divided into four subgroups, relative to 2.66 ns: short+, normal, prolonged, and prolonged+. These alterations could be related to different chromatin arrangements of B-CLL and B-ALL interphase nuclei. Notably, extremely long FLT of nuclear DAPI correlate with the presence of extra chromosome 12, while moderate increases compared to normal characterize the deletion of p53. Such correlations potentially enable a FLT-based rapid automatic diagnosis and classification of B-CLL even when the frequency of genetic and chromosomal abnormalities is low. © 2016 International Society for Advancement of Cytometry.

  15. Lifetime Autler-Townes Splitting of Dressed Multi-order Fluorescence in Pr3+:YSO

    Directory of Open Access Journals (Sweden)

    Imran Ali

    2016-07-01

    Full Text Available For first time, we study primary and secondary Autler-Townes (AT splitting of multi-order fluorescence (FL in time domain. The AT-splitting of multi-order FL signals are controlled by changing power, detuning, and polarization of single and double dressing in a heteronuclear-like molecule system of Pr+3:YSO. The primary and secondary AT-splitting is caused by double cascaded dressing in time domain. The AT-splitting of multi-order FL in time domain is more sensitive than that of in spectral domain. Such results have potential applications in quantum communication and optical information processing on photonic chip.

  16. Lifetime Autler-Townes Splitting of Dressed Multi-order Fluorescence in Pr3+:YSO

    OpenAIRE

    Imran Ali; Changbiao Li; Abdulkhaleq Hasan; Garuma Abdisa; Zongchen Liu; Feng Ma; Yanpeng Zhang

    2016-01-01

    For first time, we study primary and secondary Autler-Townes (AT) splitting of multi-order fluorescence (FL) in time domain. The AT-splitting of multi-order FL signals are controlled by changing power, detuning, and polarization of single and double dressing in a heteronuclear-like molecule system of Pr+3:YSO. The primary and secondary AT-splitting is caused by double cascaded dressing in time domain. The AT-splitting of multi-order FL in time domain is more sensitive than that of in spectral...

  17. The Gray Institute ‘open’ high-content, fluorescence lifetime microscopes

    Science.gov (United States)

    BARBER, PR; TULLIS, IDC; PIERCE, GP; NEWMAN, RG; PRENTICE, J; ROWLEY, MI; MATTHEWS, DR; AMEER-BEG, SM; VOJNOVIC, B

    2013-01-01

    Summary We describe a microscopy design methodology and details of microscopes built to this ‘open’ design approach. These demonstrate the first implementation of time-domain fluorescence microscopy in a flexible automated platform with the ability to ease the transition of this and other advanced microscopy techniques from development to use in routine biology applications. This approach allows easy expansion and modification of the platform capabilities, as it moves away from the use of a commercial, monolithic, microscope body to small, commercial off-the-shelf and custom made modular components. Drawings and diagrams of our microscopes have been made available under an open license for noncommercial use at http://users.ox.ac.uk/~atdgroup. Several automated high-content fluorescence microscope implementations have been constructed with this design framework and optimized for specific applications with multiwell plates and tissue microarrays. In particular, three platforms incorporate time-domain FLIM via time-correlated single photon counting in an automated fashion. We also present data from experiments performed on these platforms highlighting their automated wide-field and laser scanning capabilities designed for high-content microscopy. Devices using these designs also form radiation-beam ‘end-stations’ at Oxford and Surrey Universities, showing the versatility and extendibility of this approach. PMID:23772985

  18. Multimodal optical setup for nonlinear and fluorescence lifetime imaging microscopies: improvement on a commercial confocal inverted microscope

    Science.gov (United States)

    Pelegati, V. B.; Adur, J.; de Thomaz, A. A.; Almeida, D. B.; Baratti, M. O.; Carvalho, H. F.; Cesar, C. L.

    2012-03-01

    In this work we proposed and built a multimodal optical setup that extends a commercially available confocal microscope (Olympus FV300) to include nonlinear optical (NLO) microscopy and fluorescence lifetime imaging microscopy (FLIM). The NLO microscopies included two-photon fluorescence (TPFE), Second Harmonic Generation (SHG) and Third Harmonic Generation (THG). The whole system, including FLIM, used only one laser source composed of an 80 MHz femtosecond laser. The commercial Ti:sapphire lasers can be tuned up to 690-1040 nm bringing the THG signal to the 350 nm region where most microscope optics do not work. However, the third harmonic is only generated at the sample, meaning that we only have to take care of the collection optics. To do that we used a remote photomultiplier to acquire the THG signal at the 310-350 nm wavelength window. After performing the tests to guarantee that we are observing actually SHG/THG signals we than used this system to acquire multimodal images of several biological samples, from epithelial cancer to vegetables. The ability to see the collagen network together with the cell nuclei proved to be important for cancer tissues diagnosis. Moreover, FLIM provides information about the cell metabolism, also very important for cancer cell processes.

  19. High-efficiency integrated readout circuit for single photon avalanche diode arrays in fluorescence lifetime imaging.

    Science.gov (United States)

    Acconcia, G; Cominelli, A; Rech, I; Ghioni, M

    2016-11-01

    In recent years, lifetime measurements by means of the Time Correlated Single Photon Counting (TCSPC) technique have led to a significant breakthrough in medical and biological fields. Unfortunately, the many advantages of TCSPC-based approaches come along with the major drawback of a relatively long acquisition time. The exploitation of multiple channels in parallel could in principle mitigate this issue, and at the same time it opens the way to a multi-parameter analysis of the optical signals, e.g., as a function of wavelength or spatial coordinates. The TCSPC multichannel solutions proposed so far, though, suffer from a tradeoff between number of channels and performance, and the overall measurement speed has not been increased according to the number of channels, thus reducing the advantages of having a multichannel system. In this paper, we present a novel readout architecture for bi-dimensional, high-density Single Photon Avalanche Diode (SPAD) arrays, specifically designed to maximize the throughput of the whole system and able to guarantee an efficient use of resources. The core of the system is a routing logic that can provide a dynamic connection between a large number of SPAD detectors and a much lower number of high-performance acquisition channels. A key feature of our smart router is its ability to guarantee high efficiency under any operating condition.

  20. Fluorescence lifetime imaging ophthalmoscopy in type 2 diabetic patients who have no signs of diabetic retinopathy

    Science.gov (United States)

    Schweitzer, Dietrich; Deutsch, Lydia; Klemm, Matthias; Jentsch, Susanne; Hammer, Martin; Peters, Sven; Haueisen, Jens; Müller, Ulrich A.; Dawczynski, Jens

    2015-06-01

    The time-resolved autofluorescence of the eye is used for the detection of metabolic alteration in diabetic patients who have no signs of diabetic retinopathy. One eye from 37 phakic and 11 pseudophakic patients with type 2 diabetes, and one eye from 25 phakic and 23 pseudophakic healthy subjects were included in the study. After a three-exponential fit of the decay of autofluorescence, histograms of lifetimes τi, amplitudes αi, and relative contributions Qi were statistically compared between corresponding groups in two spectral channels (490diabetic patients and age-matched controls (p450 ps, and the shift of τ3 from ˜3000 to 3700 ps in ch1 of diabetic patients when compared with healthy subjects indicate an increased production of free flavin adenine dinucleotide, accumulation of advanced glycation end products (AGE), and, probably, a change from free to protein-bound reduced nicotinamide adenine dinucleotide at the fundus. AGE also accumulated in the crystalline lens.

  1. High-efficiency integrated readout circuit for single photon avalanche diode arrays in fluorescence lifetime imaging

    Science.gov (United States)

    Acconcia, G.; Cominelli, A.; Rech, I.; Ghioni, M.

    2016-11-01

    In recent years, lifetime measurements by means of the Time Correlated Single Photon Counting (TCSPC) technique have led to a significant breakthrough in medical and biological fields. Unfortunately, the many advantages of TCSPC-based approaches come along with the major drawback of a relatively long acquisition time. The exploitation of multiple channels in parallel could in principle mitigate this issue, and at the same time it opens the way to a multi-parameter analysis of the optical signals, e.g., as a function of wavelength or spatial coordinates. The TCSPC multichannel solutions proposed so far, though, suffer from a tradeoff between number of channels and performance, and the overall measurement speed has not been increased according to the number of channels, thus reducing the advantages of having a multichannel system. In this paper, we present a novel readout architecture for bi-dimensional, high-density Single Photon Avalanche Diode (SPAD) arrays, specifically designed to maximize the throughput of the whole system and able to guarantee an efficient use of resources. The core of the system is a routing logic that can provide a dynamic connection between a large number of SPAD detectors and a much lower number of high-performance acquisition channels. A key feature of our smart router is its ability to guarantee high efficiency under any operating condition.

  2. Technical Testing of Deep-UV Solid-State Sources for Fluorescence Lifetime Measurements in the Frequency Domain

    Science.gov (United States)

    2007-02-01

    circuits. Depending on the level of fluorescence signal, either standard color -glass optical filters or more expensive custom-designed interference...acids (derivatives of tyrosine and tryptophan), coenzymes NADH and riboflavin as well as dipicolinic acid (DPA) were used for the analysis of the...Roth GmbH, Karlsruhe, Germany), N- acetyl-L-tryptophanamide (NATA), ovalbumin, collagen and elastin (Sigma-Aldrich, St. Louis, MO), riboflavin (Reanal

  3. Luminescence lifetime standards for the nanosecond to microsecond range and oxygen quenching of ruthenium(II) complexes.

    Science.gov (United States)

    Morris, Kaleem J; Roach, Michael S; Xu, Wenying; Demas, J N; DeGraff, B A

    2007-12-15

    A rapid and reproducible method for determining the temperature dependence of luminescence lifetimes has been developed. With the use of this method, a set of standards for the excited-state lifetime oxygen quenching of several ruthenium(II) transition metal complexes was established. With the use of three solvents of different viscosities and two metal complexes with widely different lifetimes, an overlapping range of ca. 100 ns to 6 micros was obtained. The decays are pure single exponentials, which means that they can be used reliably with both phase and pulsed lifetime instruments. For a pure single-exponential decay, a properly operating phase shift instrument will give the same lifetime as a time domain instrument. With the use of a thermal deactivation model and a three-parameter temperature-dependent oxygen quenching constant, the lifetime temperature-dependent data was well fit by a simple six-parameter equation that covers the temperature range of 10-50 degrees C and oxygen pressures from 0 to 1 atm of oxygen with excellent precision (ca. <1%). This permits both laboratory and field calibration of instruments.

  4. Study of plant fluorescence prop erties based on laser-induced chlorophyll fluorescence lifetime imaging technology%基于激光诱导叶绿素荧光寿命成像技术的植物荧光特性研究∗

    Institute of Scientific and Technical Information of China (English)

    万文博; 华灯鑫; 乐静; 闫哲; 周春艳

    2015-01-01

    Plant fluorescence is a susceptible signal in plant fluorescence remote sensing detection. In order to solve this problem, a technique for plant chlorophyll fluorescence lifetime imaging is presented to evaluate living status for plant growth and environmental monitoring. A concave lens is used to expand laser beam at a wavelength of 355 nm, and the living plant is exposed in this laser light source to excite chlorophyll fluorescence. And the chlorophyll fluorescence signals are detected by an intensification charge coupled device. Time resolved measurement method is used in this article, so that every time the same fluorescence signals can be excited by the same laser pulse. Meanwhile, the delay time needed for triggering intensification charge coupled device should be changed consecutively, and the whole discrete fluorescence signal can be obtained. The discrete fluorescence signals from the particular location points of the plant are fitted. An improved method of forward iterative deconvolution is used to retrieve the corresponding fluorescence lifetime, and the high-precision fluorescence lifetime can be obtained. Furthermore, the fluorescence lifetime values at all the location points are retrieved to obtain the distribution map of chlorophyll fluorescence lifetime. This method can give the chlorophyll fluorescence image efficiently. The distribution map of fluorescence lifetime can more effectively reflect the plant chlorophyll concentration than the fluorescence intensity image does. The physical property of chlorophyll fluorescence lifetime from living plants has been studied preliminarily, indicating that the plant physiological status is related to its fluorescence lifetime to a certain extent; and the chlorophyll fluorescence lifetime and plant environment have a subtle and complex correlation. In the future, the relationship between chlorophyll fluorescence lifetime and plant environment will be expected to study with the cooperation of biophysicist.

  5. Fluorescence lifetimes of tyrosine residues in cytochrome c'' as local probes to study protein unfolding.

    Science.gov (United States)

    Noronha, Melinda; Santos, Raquel; Paci, Emanuele; Santos, Helena; Maçanita, António L

    2009-04-01

    Time-resolved fluorescence spectroscopy was used to show that multiple tyrosine residues of a protein can serve as localized probes of structural changes during thermal unfolding. Cytochrome c'' from Methylophilus methylotrophus, which has four tyrosine residues, was chosen as a model protein. The procedure involved, first, the assignment of the experimental decay times to the tyrosine residues, followed by the interpretation of the changes in the decay times and pre-exponential coefficients with temperature. We found that the fluorescence decays of cytochrome c'' are double-exponential from 23 to 80 degrees C, with decay times much shorter than those of the parent compound N-acetyl-tyrosinamide; this quenching was ascribed to dipole-dipole energy transfer from the tyrosine residues to the heme. The tyrosine-heme distances (R) and theoretical decay times, tau(comp), were estimated for each tyrosine residue. The analysis of the simulated decay generated with tau(comp), showed that a double-exponential fit is sufficient to describe the four decay times with two pre-exponential coefficients close to values observed from the experimental decay. Therefore, the decay times at 23 degrees C could be assigned to the individual tyrosine residues as tau(1) to Tyr-10 and Tyr-23 (at 20.3 A) and tau(2) to Tyr-12 and Tyr-115 (at 12-14 A). On the basis of this assignment and MD simulations, the temperature dependence of the decay times and pre-exponential coefficients suggest that upon unfolding, Tyr-12 is displaced from the heme, with loss of the structure of alpha-helix I. Moreover, Tyr-115 remains close to the heme and the structure in this region of the protein is not altered significantly. Altogether the data support the view that the protein core, comprising the heme and the four alpha-helices II to V, is clearly more stable than the remaining region that includes alpha-helix I and the loop between residues 19-27.

  6. Preparation and properties of Nd{sup 3+}:SrAlF{sub 5} nanocrystals embedded fluorophosphate transparent glass-ceramic with long fluorescence lifetime

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Ruilin; Wang, Jinlong; Zhang, Liaolin; Liu, Chunxiao; Wei, Wei [Nanjing University of Posts and Telecommunications, School of Optoelectronic Engineering, Nanjing (China)

    2016-07-15

    Nd{sup 3+}:SrAlF{sub 5} nanocrystals embedded fluorophosphate glass-ceramics were prepared by the melt quenching and subsequent thermal treatment method. The formation of SrAlF{sub 5} nanocrystals in the glass was confirmed by X-ray diffraction and scanning electron microscope. The fluorescence intensity and lifetime of the glass-ceramics increased with the increase of size of nanocrystals. Importantly, by controlling growth of nanocrystals, an obvious enhancement of lifetime (725 μs) emerged in the glass-ceramics heat-treated at 510 C and the transmittance can reach to 72.2 % at 1049 nm. The enhanced fluorescence intensity and lifetime were ascribed to the comfortable local environment to the Nd{sup 3+} ion and scattering of the nanoparticle embedded into the glass matrix. (orig.)

  7. Real-time fluorescence lifetime imaging system with a 32 × 32 0.13μm CMOS low dark-count single-photon avalanche diode array

    NARCIS (Netherlands)

    Li, D.U.; Arlt, J.; Richardson, J.; Walker, R.; Buts, A.; Stoppa, D.; Charbon, E.; Henderson, R.

    2010-01-01

    A compact real-time fluorescence lifetime imaging microscopy (FLIM) system based on an array of low dark count 0.13μm CMOS singlephoton avalanche diodes (SPADs) is demonstrated. Fast background-insensitive fluorescence lifetime determination is achieved by use of a recently proposed algorithm called

  8. The modifier effects of chymotrypsin and trypsin enzymes on fluorescence lifetime distribution of "N-(1-pyrenyl)maleimide-bovine serum albumin" complex

    Science.gov (United States)

    Özyiğit, İbrahim Ethem; Karakuş, Emine; Pekcan, Önder

    2016-02-01

    Chymotrypsin and trypsin are the well known proteolytic enzymes, both of which are synthesized in the pancreas as their precursors - the inactive forms; chymotrypsinogen and trypsinogen - and then are released into the duodenum to cut proteins into smaller peptides. In this paper, the effects of activities of chymotrypsin and trypsin enzymes on fluorescence lifetime distributions of the substrat bovine serum albumin (BSA) modified with N-(1-pyrenyl)maleimide (PM) were examined. In the labeling study of BSA with PM, it is aimed to attach PM to the single free thiol (Cys34) and to all the free amine groups in accessible positions in order to produce excimers of pyrene planes of the possible highest amount to form the lifetime distributions in the widest range, that may show specifically distinguishing changes resulting from the activities of the proteases. The time resolved spectrofluorometer was used to monitor fluorescence decays, which were analyzed by using the exponential series method (ESM) to obtain the changes of lifetime distributions. After the exposure of the synthesized substrat PM-BSA to the enzymes, the fluorescence lifetime distributions exhibited different structures which were attributed to the different activities of the proteases.

  9. Fluorescence lifetime imaging microscopy analysis of defects in multi-tube physical vapor transport grown Cd{sub 1-x}Zn{sub x}Te

    Energy Technology Data Exchange (ETDEWEB)

    Schneider, Andreas; Veale, Matthew C.; Wilson, Matthew D.; Seller, Paul; Botchway, Stanley W. [Science and Technology Facility Council, Rutherford Appleton Laboratory, Detector Development Group and Central Laser Facility, Harwell Oxford, Didcot, OX11 0QX (United Kingdom); Bell, Steven J. [Faculty of Engineering and Physical Sciences, University of Surrey, Guildford, Surrey, GU2 7XH (United Kingdom); Duarte, Diana D. [Science and Technology Facility Council, Rutherford Appleton Laboratory, Detector Development Group and Central Laser Facility, Harwell Oxford, Didcot, OX11 0QX (United Kingdom); Faculty of Engineering and Physical Sciences, University of Surrey, Guildford, Surrey, GU2 7XH (United Kingdom); Choubey, Ashutosh; Halliday, Douglas [Department of Physics, Durham University, Rochester Building, South Road, Durham, DH1 3LE (United Kingdom)

    2014-09-15

    Cadmium zinc telluride (CZT) is the material of choice for high-energy room-temperature X-ray and γ-ray detectors. However, the performance of pixelated detectors is greatly influenced by the quality of CZT. Crystal defects and impurities are one source of shallow and deep level traps for charge carriers. Fluorescence lifetime of the recombination of optically excited charges may indicate the presence and type of defects and impurities in CZT. Fluorescence lifetime imaging microscopy (FLIM) is used to examine the excited-state lifetime in CZT fabricated by different growth methods and conditions. The FLIM set-up analyzes luminescence emitted from the sample following photo excitation. Samples were optically excited above band gap with a pulsed laser (590 nm) for raster scanning a 220 x 165 μm{sup 2} sample area. In-situ room-temperature photoluminescence (PL) and FLIM were recorded simultaneously. In order to analyze the FLIM data, two dominant charge carrier decay processes (τ{sub 1}, τ{sub 2}) were identified. The luminescence signal decays with a rapid lifetime of τ{sub 1} ∼ 50-200 ps, and a large variety of long-lifetime components τ{sub 2} were found in the range of 225-900 ps. CZT grown by multi-tube physical vapor transport (MTPVT) showed extremely long-lived recombination decay times up to 3.5 ns in the vicinity of the interface at growth start. Further away from this interface, the recombination lifetime was in the typical range of fast transitions similar to those found in detector-grade CZT fabricated by travelling heater method. Crystalline material quality strongly influences FLIM lifetime. Time-resolved transients of MTPVT-grown CZT compared with industry-leading detector grade CZT (dots: measured data; lines: fitted exponential decay curves). (copyright 2014 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

  10. Synthesis and Characterisation of Photo-Cross-Linkable Liquid Crystalline Poly(n-[n′-flurobenzoylstyryloxy]alkylmethacrylates and Their Fluorescence Lifetime Properties

    Directory of Open Access Journals (Sweden)

    G. Kumar

    2013-01-01

    Full Text Available This paper reports a study on photo-cross-linkable polymer containing pendant chalcone moiety exhibiting liquid crystalline as well as fluorescence lifetime properties in detail. The photoresponsive polymers were prepared, and their structure has been characterized by 1H-NMR, 13C-NMR, and UV-Visible spectroscopy. The photo-cross-linking behavior of polymers has been studied by UV-Visible and fluorescence spectroscopy. UV spectral studies revealed that the polymers follow 2π+2π cyclo addition reactions when they undergo photo-cross-linking under the influence of UV-light. Number and weight average molecular weight of the polymers were determined by Gel Permeation Chromatography (GPC and polydispersity index value near to 1.5. The thermal and thermooxidative stability of the polymers were determined by Thermogravimetric Analysis (TGA. Thermal transitions were studied by DSC, and presence of mesophases was identified at 147 and 126∘C by hot stage polarized light optical microscopy (HPOM. Fluorescence lifetime measurements using the time-correlated single photon counting (TCSPC method reveal that the average lifetime values decrease from 5.94 ns to 5.32 ns on UV-irradiation were discussed in detail.

  11. Photophysics of cyanine dyes adsorbed onto surfaces. Sub-nanosecond fluorescence lifetime measurements of 3,3'-diethyloxadicarbocyanine iodide and photoisomer

    Energy Technology Data Exchange (ETDEWEB)

    Vieira Ferreira, L.; Oliveira, A.; Henbest, K. [and others

    1998-01-01

    This report describes the experiment entitled 'Photophysics of Cyanine dyes on Surfaces'; carried out at the Central Laser Facility (CLF) from the 6th to the 20th January 1997. The experiment, funded by the Framework IV Large-Scale Facilities Access Scheme, was proposed by Prof. L.F. Vieira Ferreira, Centro de Quimica-Fisica Molecular, Complexo 1, IST, 1096 Lisboa Codex, Portugal, and carried out by visiting researchers from the Institute. They were supported by researchers from the Central Laser Facility, Rutherford Appleton Laboratory. Experimental results: The photo physics of 3,3'-Diethyloxadicarbocyanine iodide (DODCI) adsorbed onto swollen microcrystalline cellulose was investigated. Two fluorescence emissions band have been observed and assigned. One was due to singlet excited momers and a second new emission, seen at high laser fluences, was due to the formation of a photoisomer. The DODCI stays entrapped between the polymer chains and nonradiative pathways for deactivation are reduced, the lifetimes of the excited states were measured using time resolved fluorescence lifetimes techniques. The fluorescence lifetimes of the excited states are longer lived in a swollen cellulose matrix. The photoisomer emission especially lives an order of magnitude longer than in homogeneous media.

  12. The Atmospheric Muon Lifetime, with the Lead Absorption Potential for Muons and References to the Standard Model of Particle Physics

    Science.gov (United States)

    Barazandeh, Cioli; Gutarra-Leon, Angel; Majewski, Walerian

    2017-01-01

    Muon is one of twelve fundamental particles and has the longest free-particle lifetime. It decays into three leptons through an exchange of weak vector bosons W +/W-. Muons are present in atmospheric secondary cosmic rays and reach the sea level. By detecting the time delay between arrival of muons and appearance of decay electrons in a scintillation detector, we will measure muon's lifetime at rest. From the lifetime we can find the ratio gw /MW of the weak coupling constant gw (a weak analog of the electric charge) to mass of the W-boson MW. Vacuum expectation value v of the Higgs field, which determines masses Standard Model (SM) particles, can be calculated as v =2MWc2/gw =(τmμc2/6π3\\hcirc)1/4mμc2 regarding muon mass mμ and muon lifetime τ only. Using the experimental value for MWc2 = 80.4 GeV, we will find weak coupling constant gw. With the SM relation e =gwsin θ√ hcε0 and experimental value of the Z0-photon weak mixing angle θ = 29o we use our muon lifetime to find the elementary electric charge e value. In this experiment we will also determine the sea level fluxes of low-energy (<160 MeV) and high-energy cosmic muons, then will shield the detector with varying thicknesses of lead plates and from the new values of fluxes find the energy-dependent muon stopping power in lead.

  13. 76 FR 52892 - Energy Conservation Program: Energy Conservation Standards for Fluorescent Lamp Ballasts

    Science.gov (United States)

    2011-08-24

    ... Part 430 RIN 1904-AB50 Energy Conservation Program: Energy Conservation Standards for Fluorescent Lamp... fluorescent lamp ballasts (ballasts) pursuant to the Energy Policy and Conservation Act of 1975 (EPCA). During... The EPCA establishes energy conservation standards for certain ballasts and requires that DOE...

  14. 基于同步扫描相机的荧光寿命测量系统研究%A fluorescence lifetime spectrometer based on a synchroscan streak camera

    Institute of Scientific and Technical Information of China (English)

    邵永红; 李恒; 王岩; 屈军乐; 牛憨笨

    2009-01-01

    建立一台基于同步扫描相机的双光子激发荧光寿命显微测量系统,同步扫描相机的重复工作频率为76 MHz,利用钛宝石飞秒激光器作为光源,通过可调延时器和标准具对扫描相机的时间分辨率、扫描速度以及非线性等进行标定.该系统的时间分辨率为9 ps,非线性小于4%,量程为2.8 ns.测量了荧光染料Rose Bengal(RsB)的荧光衰减曲线,通过最小二乘法对荧光的衰减曲线进行拟合,得到RsB的荧光寿命为763 ps,与标准荧光染料对比一致.%A two-photon excitation fluorescence lifetime spectrometer based on a synchroscan streak camera was presented. The spectrometer was calibrated using a Titanium: Sapphire femtosecond laser in conjunction with a custom-made synchronized streak circuit, tunable delay generator and a set of etalons. Experimental results show that the temporal resolution of this spectrometer is 9 ps, nonlinearity is less than 4% , and the sweep range is 2. 8 ns. Fluorescence decay profile of fluorescent dye Rose Bengal ( RsB) is recorded with the spectrometer. Nonlinear least square fitting of the decay profile gives that the lifetime of RsB is 763 ps, consistent with standard fluorescence dyes.

  15. Novel Battery Thermal Management System for Greater Lifetime Ratifying Current Quality and Safety Standard

    DEFF Research Database (Denmark)

    Khan, Mohammad Rezwan; Andreasen, Søren Juhl; Kær, Søren Knudsen

    2014-01-01

    Temperature excursions and non-uniformity of the temperature inside the battery systems are the main concern and drawback for any attempt to scale-up battery cells to the larger sizes as required for high power applications. The applications may include electric generating stations, substations......, vehicles, telecommunications installations, large industrial and commercial installations, large uninterruptible power supply (UPS) installations and renewable energy plant installations etc. The capacity of the battery pack increases as the operating temperature is raised for a battery pack however...... this come with the very high expense of accelerated capacity fade i.e. ageing. Subsequently the lifetime of the battery system is reduced. Moreover poor performance (limited capacity availability) is observed at low operating temperature. In addition, excessive or uneven temperature rise in a system or pack...

  16. Study of intracellular delivery of doxorubicin from poly(lactide-co-glycolide) nanoparticles by means of fluorescence lifetime imaging and confocal Raman microscopy.

    Science.gov (United States)

    Romero, Gabriela; Qiu, Yuan; Murray, Richard A; Moya, Sergio E

    2013-02-01

    The intracellular delivery of Doxorubicin (Dox) from poly(lactide-co-glycolide) (PLGA) nanoparticles stabilised with bovine serum albumin, in HepG2 cells, is studied via flow cytometry, fluorescence lifetime imaging microscopy (FLIM), confocal Raman microscopy (CRM) and cell viability studies. Flow cytometry shows that the initial uptake of PLGA and Dox follow the same kinetics. However, following 8 h of incubation, the fluorescence intensity and cellular uptake of Dox decreases, while in the case of PLGA both parameters remain constant. FLIM shows the presence of a single-lifetime species, with a lifetime of 1.15 ns when measured inside the cells. Cell viability decreases by approximately 20% when incubated for 24 h with PLGA loaded with Dox, with a particle concentration of 100 µg · mL(-1). At the single-cell level, CRM shows changes in the bands from DNA and proteins in the cell nucleus when incubated with PLGA loaded with Dox. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Standardization and quality assurance in fluorescence measurements I state-of-the art and future challenges

    CERN Document Server

    Resch-Genger, Ute

    2008-01-01

    The validation and standardization of fluorescence methods is still in its infancy as compared to other prominent analytical and bioanalytical methods. Appropriate quality assurance standards are however a prerequisite for applications in highly regulated fields such as medical diagnostics, drug development, or food analysis. For the first time, a team of recognized international experts has documented the present status of quality assurance in fluorescence measurements, and outlines concepts for establishing standards in this field. This first of two volumes covers basic aspects and various techniques such as steady-state and time-resolved fluorometry, polarization techniques, and fluorescent chemical sensors

  18. Size-effects on energy relaxation and excited-species desorption in krypton clusters: Fluorescence lifetime measurements with 10 eV laser excitation

    Science.gov (United States)

    Kanaev, A. V.; Museur, L.; Castex, M. C.

    1997-09-01

    Fluorescence lifetime measurements of KrN clusters (N¯=2-2000) have been carried out using intense 10 eV laser excitation near 3P2 metastable atomic energy level. Two principal groups of electronically excited dimers Kr2* have been found in desorption: dimers, loosely bound near the (3P2+1S0) dissociation limit, ejected from cooled clusters and dimers undergoing vibrational relaxation from hot clusters. The desorption is principally terminated when N¯⩾50 at./cluster. The relaxation kinetics seems to converge to the properties of a solid state for 102⩽N¯⩽103 at./cluster. A variation of the Kr2*(1u/0u-) radiative lifetime, from 264 ns (in gas phase) to 440 ns (N¯=102), has been found. An equilibrium cluster temperature of 57 K has been calculated from this τ(N) dependence.

  19. Relation between proteins tertiary structure, tryptophan fluorescence lifetimes and tryptophan S(o)→(1)L(b) and S(o)→(1)L(a) transitions. Studies on α1-acid glycoprotein and β-lactoglobulin.

    Science.gov (United States)

    Albani, Jihad René

    2011-05-01

    We measured fluorescence lifetimes and fluorescence spectra (excitation and emission) of tryptophan residues of α(1)-acid glycoprotein (three Trp residues) and β-lactoglobulin (two Trp residues) in absence and presence of 450 μM progesterone. Progesterone binds only to α(1)-acid glycoprotein. In absence of progesterone, each of the two proteins displays three fluorescence lifetimes. Addition of progesterone induces a partial inhibition of the S(o) → (1)L(a) transition without affecting fluorescence lifetimes. The same experiments performed in presence of denatured proteins in 6 M guanidine show that addition of progesterone inhibits partially the S(o) → (1)L(a) transition and its peak is 15 nm shifted to the red compared to that obtained for native proteins. However, the S(o) → (1)L(b) transition position peak is not affected by protein denaturation. Thus, the tertiary structure of the protein plays an important role by modulating the tryptophan electronic transitions. Fluorescence emission decay recorded in absence and presence of progesterone yields three fluorescence lifetimes whether proteins are denatured or not. Thus, protein tertiary structure is not responsible for the presence of three fluorescence lifetimes. These characterize tryptophan substructures reached at the excited states and which population (pre-exponential values) depend on the tryptophan residues interaction with their microenvironment(s) and thus on the global conformation of the protein.

  20. 基于荧光寿命机理的光纤温度传感器研究%Fiber Temperature Sensor Based on Fluorescent Lifetime

    Institute of Scientific and Technical Information of China (English)

    江小峰; 李亚东; 李欣; 夏添艺; 林春

    2015-01-01

    为了实现恶劣环境下温度的测量,设计了一种基于荧光寿命机理的光纤温度传感系统.温度传感系统选用415 nm LED作为光源,以稀土荧光材料Y2 O2 S:Eu作为温度敏感材料,通过探测放大器和信号采集模块测量了敏感材料的荧光寿命,并由荧光寿命与温度的单调关系最终实现了温度的测量.采用油浴加热的方法进行温度实验,实验结果表明,温度传感系统在25~80℃实现了温度的测量,分辨率为0.5℃.%In order to achieve the measurement of temperature in harsh environments, a fiber temperature sensor system based on the fluorescent lifetime was designed. 415 nm LED was selected as the light source and a rare earth material Y2 O2 S:Eu was selected as the sensitive material. Finally, the fluorescence lifetime of sensitive material was measured by a probe amplifier and signal acquisition module and temperature measurements were realized due to the monotonic relationship between the fluorescence lifetime and temperature. Temperature experiments were carried out using an oil bath heating method, and the experimental results showed that: the temperature measurement can be realized in the range of 25 ~80 ℃ and the temperature sensor has an average resolution of 0 . 5 ℃.

  1. Two-photon spectral fluorescence lifetime and second-harmonic generation imaging of the porcine cornea with a 12-femtosecond laser microscope

    Science.gov (United States)

    Batista, Ana; Breunig, Hans Georg; Uchugonova, Aisada; Morgado, António Miguel; König, Karsten

    2016-03-01

    Five dimensional microscopy with a 12-fs laser scanning microscope based on spectrally resolved two-photon autofluorescence lifetime and second-harmonic generation (SHG) imaging was used to characterize all layers of the porcine cornea. This setup allowed the simultaneous excitation of both metabolic cofactors, NAD(P)H and flavins, and their discrimination based on their spectral emission properties and fluorescence decay characteristics. Furthermore, the architecture of the stromal collagen fibrils was assessed by SHG imaging in both forward and backward directions. Information on the metabolic state and the tissue architecture of the porcine cornea were obtained with subcellular resolution, and high temporal and spectral resolutions.

  2. Assessing solvent effects on the singlet excited state lifetime of uracil derivatives: A femtosecond fluorescence upconversion study in alcohols and D{sub 2}O

    Energy Technology Data Exchange (ETDEWEB)

    Gustavsson, Thomas [Laboratoire Francis Perrin, CEA/DSM/DRECAM/SPAM - CNRS URA 2453, CEA/Saclay, F-91191 Gif-sur-Yvette (France)], E-mail: thomas.gustavsson@cea.fr; Banyasz, Akos [Laboratoire Francis Perrin, CEA/DSM/DRECAM/SPAM - CNRS URA 2453, CEA/Saclay, F-91191 Gif-sur-Yvette (France); Sarkar, Nilmoni [Department of Chemistry, Indian Institute of Technology, Kharagpur 721 302, WB (India); Markovitsi, Dimitra [Laboratoire Francis Perrin, CEA/DSM/DRECAM/SPAM - CNRS URA 2453, CEA/Saclay, F-91191 Gif-sur-Yvette (France); Improta, Roberto [Dipartimento di Chimica, Universita Federico II, Complesso Universitario Monte S. Angelo, Via Cintia, I-80126 Napoli (Italy); Istituto Biostrutture e Bioimmagini/CNR, V. Mezzocannone 6 - 80134 Napoli (Italy)

    2008-06-23

    The excited state lifetimes of uracil, thymine and 5-fluorouracil have been measured using femtosecond UV fluorescence upconversion in various protic and aprotic polar solvents. The fastest decays are observed in acetonitrile and the slowest in aqueous solution while those observed in alcohols are intermediate. No direct correlation with macroscopic solvent parameters such as polarity or viscosity is found, but hydrogen bonding is one key factor affecting the fluorescence decay. It is proposed that the solvent modulates the relative energy of two close-lying electronically excited states, the bright {pi}{pi}* and the dark n{pi}* states. This relative energy gap controls the non-radiative relaxation of the {pi}{pi}* state through a conical intersection close to the Franck-Condon region competing with the ultrafast internal conversion to the ground state. In addition, an inverse isotope effect is observed in D{sub 2}O where the decays are faster than in H{sub 2}O.

  3. Frequency-domain fluorescence lifetime imaging system (pco.flim) based on a in-pixel dual tap control CMOS image sensor

    Science.gov (United States)

    Franke, Robert; Holst, Gerhard A.

    2015-03-01

    The luminescence lifetime as a beneficial analytical parameter is known for many years and is well described by a large variety of publications. Many instruments including 2D measuring systems with cameras have been developed and applied in the past years. However, since the current instrumentation to perform either time- or frequency-domain lifetime measurements is rather complex, new developments in CMOS image sensor technology have achieved to create new image sensors, which can efficiently be integrated into easier-to-handle luminescence lifetime measuring systems. The principle of these modulatable CMOS image sensors, while initially being designed for distance measurements, shows a clear analogy to frequency-domain FLIM measurements, which was proven by researchers [1, 2]. Based on this principle a new CMOS image sensor has been developed, integrated into a camera system and has been investigated within a research project. The image sensor has a resolution of 1024 × 1024 pixels with a 5.6 μm pitch and can be modulated up to 50 MHz. First measurements show an effective dynamic range of larger than 1:1024 (corresponding to 10 bit dynamic). The maximum frame rate is in the range of 90 frames/s in dual-tap mode, resulting in an effective lifetime image frame rate for realistic measurements of approximately 22 frames/s. The camera system pco.flim, featuring that image sensor, generates all required modulation signals from 5 kHz to 50 MHz (sinusoidal and rectangular). It performs advanced pixel correction to generate linear and high-quality images, while the basic lifetime image processing is done in the computer. The modulation frequency can be freely adjusted within the specified range. The characteristics of the camera systems are presented, and first results are discussed using different representations of the data like for example the phasor approach [3], which has been established to provide a more global view to pixelwise fluorescence lifetime data and

  4. Interaction of poxvirus intracellular mature virion proteins with the TPR domain of kinesin light chain in live infected cells revealed by two-photon-induced fluorescence resonance energy transfer fluorescence lifetime imaging microscopy.

    Science.gov (United States)

    Jeshtadi, Ananya; Burgos, Pierre; Stubbs, Christopher D; Parker, Anthony W; King, Linda A; Skinner, Michael A; Botchway, Stanley W

    2010-12-01

    Using two-photon-induced fluorescence lifetime imaging microscopy, we corroborate an interaction (previously demonstrated by yeast two-hybrid domain analysis) of full-length vaccinia virus (VACV; an orthopoxvirus) A36 protein with the cellular microtubule motor protein kinesin. Quenching of enhanced green fluorescent protein (EGFP), fused to the C terminus of VACV A36, by monomeric red fluorescent protein (mDsRed), fused to the tetratricopeptide repeat (TPR) domain of kinesin, was observed in live chicken embryo fibroblasts infected with either modified vaccinia virus Ankara (MVA) or wild-type fowlpox virus (FWPV; an avipoxvirus), and the excited-state fluorescence lifetime of EGFP was reduced from 2.5 ± 0.1 ns to 2.1 ± 0.1 ns due to resonance energy transfer to mDsRed. FWPV does not encode an equivalent of intracellular enveloped virion surface protein A36, yet it is likely that this virus too must interact with kinesin to facilitate intracellular virion transport. To investigate possible interactions between innate FWPV proteins and kinesin, recombinant FWPVs expressing EGFP fused to the N termini of FWPV structural proteins Fpv140, Fpv168, Fpv191, and Fpv198 (equivalent to VACV H3, A4, p4c, and A34, respectively) were generated. EGFP fusions of intracellular mature virion (IMV) surface protein Fpv140 and type II membrane protein Fpv198 were quenched by mDsRed-TPR in recombinant FWPV-infected cells, indicating that these virion proteins are found within 10 nm of mDsRed-TPR. In contrast, and as expected, EGFP fusions of the IMV core protein Fpv168 did not show any quenching. Interestingly, the p4c-like protein Fpv191, which demonstrates late association with preassembled IMV, also did not show any quenching.

  5. Formation of Gel-like Nanodomains in Cholesterol-Containing Sphingomyelin or Phosphatidylcholine Binary Membrane As Examined by Fluorescence Lifetimes and (2)H NMR Spectra.

    Science.gov (United States)

    Yasuda, Tomokazu; Matsumori, Nobuaki; Tsuchikawa, Hiroshi; Lönnfors, Max; Nyholm, Thomas K M; Slotte, J Peter; Murata, Michio

    2015-12-29

    In this study, we measured the time-resolved fluorescence of trans-parinaric acid (tPA), steady-state fluorescence anisotropy of diphenylhexatriene (DPH), and (2)H NMR of 10,10-d2-stearoyl lipids in stearoyl sphingomyelin with cholesterol (SSM/Chol) and l-palmitoyl-2-stearoyl-sn-glycero-3-phosphocholine with Chol (PSPC/Chol) binary membranes. The results suggest that the membrane order obtained from the fluorescence experiments shows a similar temperature dependency as those of the (2)H NMR data. More importantly, the time-resolved fluorescence data implied the presence of at least two types of domains, cholesterol-poor gel-like domains (CPGLD) and cholesterol-enriched liquid-ordered (Lo) domains. These domains appear on a nano-to-micro second time scale for both SSM-Chol and PSPC-Chol membranes. The relative size of the gel-like domain was also estimated from the temperature-dependent lifetime measurements and (2)H NMR spectral changes. The results imply that the size of the gel-like domains is very small, probably on the nanometer scale, and smaller in SSM-Chol membrane than those in PSPC-Chol bilayers, which could account for the higher thermal stability of SM-Chol membranes. The present study demonstrates that gel-like nanodomains occur in SM-Chol binary membrane even with Chol content of over 33 mol %, which has been thought to consist exclusively of Lo phase, implying that not only Lo domains but also gel-like nanodomains are important for formation of lipid-ordered phase in SM-Chol and PC-Chol membranes.

  6. Ideal Molecular Design of Blue Thermally Activated Delayed Fluorescent Emitter for High Efficiency, Small Singlet-Triplet Energy Splitting, Low Efficiency Roll-Off, and Long Lifetime.

    Science.gov (United States)

    Lee, Dong Ryun; Choi, Jeong Min; Lee, Chil Won; Lee, Jun Yeob

    2016-09-07

    Highly efficient thermally activated delayed fluorescent (TADF) emitters, 5-(2-(4,6-diphenyl-1,3,5-triazin-2-yl)phenyl)-5H-benzofuro[3,2-c]carbazole (oBFCzTrz), 5-(3-(4,6-diphenyl-1,3,5-triazin-2-yl)phenyl)-5H-benzofuro[3,2-c]carbazole (mBFCzTrz), and 5-(4-(4,6-diphenyl-1,3,5-triazin-2-yl)phenyl)-5H-benzofuro[3,2-c]carbazole (pBFCzTrz), were synthesized to study the effects of ortho-, meta-, and para- linkages between donor and acceptor moieties. oBFCzTrz having ortho- linked donor and acceptor moieties showed smaller singlet-triplet energy gap, shorter excited state lifetime, and higher photoluminescence quantum yield than mBFCzTrz and pBFCzTrz which are interconnected by meta- and para- positions. The TADF device using oBFCzTrz as a blue emitter exhibited high external quantum efficiency over 20%, little efficiency roll-off, and long device lifetime.

  7. Exploiting p-Type Delayed Fluorescence in Hybrid White OLEDs: Breaking the Trade-off between High Device Efficiency and Long Lifetime.

    Science.gov (United States)

    Zhang, Dongdong; Zhang, Deqiang; Duan, Lian

    2016-09-01

    Despite that the majority of practical organic light-emitting diodes (OLEDs) still rely on blue fluorophors with low triplet (T1) for creating blue light, hybrid white OLEDs based on low T1 blue fluorophors are still much lagged behind in power efficiency. Here, "ideal" hybrid WOLEDs with recorded efficiency as well as low roll-off, good color-stability and long lifetime were realized by utilizing the bipolar mixed materials as the host of green phosphor as well as the spacer to reduce T1 trap, while blue fluorophors with p-type delayed fluorescence to recycle the trapped T1. An electron transport material with both high electron mobility and good exciton confinement ability was used to boost the TTA efficiency. Hybrid WOLEDs with maximum current efficiency, external quantum efficiency and power efficiency of 49.6 cd/A, 19.1%, and 49.3 lm/W, respectively, together with a high color rendering index of 80 and a half lifetime of over 7000 h at an initial luminescence of 1000 cd/m(2) were realized, manifesting the high potential of the strategy.

  8. Cost-effectiveness of dronedarone and standard of care compared with standard of care alone: US results of an ATHENA lifetime model

    Directory of Open Access Journals (Sweden)

    Reynolds MR

    2013-01-01

    Full Text Available Matthew R Reynolds,1 Jonas Nilsson,2 Örjan Åkerborg,2 Mehul Jhaveri,3 Peter Lindgren2,41Beth Israel Deaconess Medical Center, VA Boston Healthcare System, Boston, MA, USA; 2OptumInsight, Stockholm, Sweden; 3sanofi-aventis Inc, Bridgewater, NJ, USA; 4Institute of Environmental Medicine, Karolinska Institute, Stockholm, SwedenBackground: The first antiarrhythmic drug to demonstrate a reduced rate of cardiovascular hospitalization in atrial fibrillation/flutter (AF/AFL patients was dronedarone in a placebo-controlled, double-blind, parallel arm Trial to assess the efficacy of dronedarone 400 mg bid for the prevention of cardiovascular Hospitalization or death from any cause in patiENts with Atrial fibrillation/atrial flutter (ATHENA trial. The potential cost-effectiveness of dronedarone in this patient population has not been reported in a US context. This study assesses the cost-effectiveness of dronedarone from a US health care payers’ perspective.Methods and results: ATHENA patient data were applied to a patient-level health state transition model. Probabilities of health state transitions were derived from ATHENA and published data. Associated costs used in the model (2010 values were obtained from published sources when trial data were not available. The base-case model assumed that patients were treated with dronedarone for the duration of ATHENA (mean 21 months and were followed over a lifetime. Cost-effectiveness, from the payers' perspective, was determined using a Monte Carlo microsimulation (1 million fictitious patients. Dronedarone plus standard care provided 0.13 life years gained (LYG, and 0.11 quality-adjusted life years (QALYs, over standard care alone; cost/QALY was $19,520 and cost/LYG was $16,930. Compared to lower risk patients, patients at higher risk of stroke (Congestive heart failure, history of Hypertension, Age ≥ 75 years, Diabetes mellitus, and past history of Stroke or transient ischemic attack (CHADS2 scores 3

  9. Motor neuron disease mortality and lifetime petrol lead exposure: Evidence from national age-specific and state-level age-standardized death rates in Australia.

    Science.gov (United States)

    Zahran, Sammy; Laidlaw, Mark A S; Rowe, Dominic B; Ball, Andrew S; Mielke, Howard W

    2017-02-01

    The age standardized death rate from motor neuron disease (MND) for persons 40-84 years of age in the Australian States of New South Wales, Victoria, and Queensland increased dramatically from 1958 to 2013. Nationally, age-specific MND death rates also increased over this time period, but the rate of the rise varied considerably by age-group. The historic use of lead (Pb) additives in Australian petrol is a candidate explanation for these trends in MND mortality (International Classification of Disease (ICD)-10 G12.2). Leveraging temporal and spatial variation in petrol lead exposure risk resulting from the slow rise and rapid phase-out of lead as a constituent in gasoline in Australia, we analyze relationships between (1) national age-specific MND death rates in Australia and age-specific lifetime petrol lead exposure, (2) annual between-age dispersions in age-specific MND death rates and age-specific lifetime petrol lead exposure; and (3) state-level age-standardized MND death rates as a function of age-weighted lifetime petrol lead exposure. Other things held equal, we find that a one percent increase in lifetime petrol lead exposure increases the MND death rate by about one-third of one percent in both national age-specific and state-level age-standardized models of MND mortality. Lending support to the supposition that lead exposure is a driver of MND mortality risk, we find that the annual between-age group standard deviation in age-specific MND death rates is strongly correlated with the between-age standard deviation in age-specific lifetime petrol lead exposure. Legacy petrol lead emissions are associated with age-specific MND death rates as well as state-level age-standardized MND death rates in Australia. Results indicate that we are approaching peak lead exposure-attributable MND mortality. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. Fluorescence Lifetime Imaging of Physiological Free Cu(II) Levels in Live Cells with a Cu(II)-Selective Carbonic Anhydrase-Based Biosensor

    Science.gov (United States)

    McCranor, Bryan J.; Szmacinski, Henryk; Zeng, Hui Hui; Stoddard, A.K.; Hurst, Tamiika; Fierke, Carol A.; Lakowicz, J.R.

    2014-01-01

    Copper is a required trace element that plays key roles in a number of human enzymes, such that copper deficiency or genetic defects in copper transport lead to serious or fatal disease. Rae, et al., had famously predicted that free copper ion levels in the cell cytoplasm were extremely low, typically too low to be observable. We recently developed a variant of human apocarbonic anhydrase II for sensing metal ions that exhibits 25-fold better selectivity for Cu(II) over Zn(II) than the wild type protein, enabling us to accurately measure Cu(II) in the presence of ordinary cellular (picomolar) concentrations of free zinc. We inserted a fluorescent labeled Cu(II)-specific variant of human apocarbonic anhydrase into PC-12 cells and found that the levels are indeed extremely low (in the femtomolar range). We imaged the free Cu(II) levels in living cells by means of frequency-domain fluorescence lifetime microscopy. Implications of this finding are discussed. PMID:24671220

  11. 76 FR 20089 - Energy Conservation Program: Energy Conservation Standards for Fluorescent Lamp Ballasts

    Science.gov (United States)

    2011-04-11

    ... April 11, 2011 Part II Department of Energy 10 CFR Part 430 Energy Conservation Program: Energy... Conservation Program: Energy Conservation Standards for Fluorescent Lamp Ballasts AGENCY: Office of Energy... public meeting. SUMMARY: The Energy Policy and Conservation Act (EPCA) prescribes energy......

  12. 76 FR 70547 - Energy Conservation Program: Energy Conservation Standards for Fluorescent Lamp Ballasts

    Science.gov (United States)

    2011-11-14

    ... November 14, 2011 Part II Department of Energy 10 CFR Part 430 Energy Conservation Program: Energy...-AB50 Energy Conservation Program: Energy Conservation Standards for Fluorescent Lamp Ballasts AGENCY... Energy Policy and Conservation Act of 1975 (EPCA), as amended, prescribes energy conservation...

  13. Reliable measurement of E. coli single cell fluorescence distribution using a standard microscope set-up.

    Science.gov (United States)

    Cortesi, Marilisa; Bandiera, Lucia; Pasini, Alice; Bevilacqua, Alessandro; Gherardi, Alessandro; Furini, Simone; Giordano, Emanuele

    2017-01-01

    Quantifying gene expression at single cell level is fundamental for the complete characterization of synthetic gene circuits, due to the significant impact of noise and inter-cellular variability on the system's functionality. Commercial set-ups that allow the acquisition of fluorescent signal at single cell level (flow cytometers or quantitative microscopes) are expensive apparatuses that are hardly affordable by small laboratories. A protocol that makes a standard optical microscope able to acquire quantitative, single cell, fluorescent data from a bacterial population transformed with synthetic gene circuitry is presented. Single cell fluorescence values, acquired with a microscope set-up and processed with custom-made software, are compared with results that were obtained with a flow cytometer in a bacterial population transformed with the same gene circuitry. The high correlation between data from the two experimental set-ups, with a correlation coefficient computed over the tested dynamic range > 0.99, proves that a standard optical microscope- when coupled with appropriate software for image processing- might be used for quantitative single-cell fluorescence measurements. The calibration of the set-up, together with its validation, is described. The experimental protocol described in this paper makes quantitative measurement of single cell fluorescence accessible to laboratories equipped with standard optical microscope set-ups. Our method allows for an affordable measurement/quantification of intercellular variability, whose better understanding of this phenomenon will improve our comprehension of cellular behaviors and the design of synthetic gene circuits. All the required software is freely available to the synthetic biology community (MUSIQ Microscope flUorescence SIngle cell Quantification).

  14. Fluorescent dye labeled DNA size standards for molecular mass detection in visible/infrared range

    Directory of Open Access Journals (Sweden)

    Sreelakshmi Yellamaraju

    2011-01-01

    Full Text Available Abstract Background Targeting Induced Local Lesions in Genomes (TILLING is a high throughput reverse genetics tool which detects mismatches (single point mutations or small indels in large number of individuals of mutagenized populations. Currently, TILLING is intensively used for genomics assisted molecular breeding of several crop plants for desired traits. Most commonly used platform for mutation detection is Li-COR DNA Analyzer, where PCR amplified products treated with single strand mismatch specific nuclease are resolved on denaturing gels. The molecular size of any cut product can be easily estimated by comparing with IR dye labeled markers of known sizes. Similar fluorescent dye labeled size markers are also used for several genotyping experiments. Currently, commercially available size standards are expensive and are restricted up to only 700 bp which renders estimation of products of sizes greater than 700 bases inaccurate. Findings A simple protocol was developed for labeling 5' end of multiple DNA size markers with fluorescent dyes. This method involves cloning a pool of different size markers of DNA in a plasmid vector. PCR amplification of plasmid using IR dye labeled universal primers generates 5' fluorescent labeled products of various sizes. The size of products constituting the ladder can be customized as per the need. The generated size markers can be used without any further purification and were found to be stable up to one year at -20°C. Conclusions A simple method was developed for generating fluorescent dye labeled size standards. This method can be customized to generate different size standards as per experimental needs. The protocol described can also be adapted for developing labeled size standards for detection on platforms other than Li-COR i.e. other than infra red range of the spectrum.

  15. Development of suitable plastic standards for X-ray fluorescence analysis

    Science.gov (United States)

    Mans, Christian; Hanning, Stephanie; Simons, Christoph; Wegner, Anne; Janβen, Anton; Kreyenschmidt, Martin

    2007-02-01

    For the adoption of the EU directive "Restriction on use of certain Hazardous Substances" and "Waste Electrical and Electronic Equipment" using X-ray fluorescence analysis suitable standard materials are required. Plastic standards based on acrylonitrile-butadiene-styrene terpolymer, containing the regulated elements Br, Cd, Cr, Hg and Pb were developed and produced as granulates and solid bodies. The calibration materials were not generated as a dilution from one master batch but rather the element concentrations were distributed over nine independent calibration samples. This was necessary to enable inter-elemental corrections and empirical constant mass absorption coefficients. The produced standard materials are characterized by a homogenous element distribution, which is more than sufficient for X-ray fluorescence analysis. Concentrations for all elements except for Br could be determined by Inductively Coupled Plasma Atomic Emission Spectroscopy after microwave assisted digestion. The concentration of Br was determined by use of Neutron Activation Analysis at Hahn-Meitner-Institute in Berlin, Germany. The correlation of the X-ray fluorescence analysis measurements with the values determined using Inductively Coupled Plasma Atomic Emission Spectroscopy and Neutron Activation Analysis showed a very good linearity.

  16. A light diet for a giant appetite: An assessment of China's proposed fluorescent lamp standard

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Jiang

    2002-04-11

    Lighting has been one of the fastest growing electric end-uses in China over the last twenty years, with an average annual growth rate of 14%. Fluorescent lighting provides a significant portion of China's lighting need. In 1998, China produced 680 million fluorescent lamps, of which 420 million were linear fluorescent lamps of various diameters (T8 to T12). There are substantial variations both in energy efficiency and lighting performance among locally produced fluorescent lamps. Such variations present a perfect opportunity for policy intervention through efficiency standards to promote the adoption of more efficient fluorescent lamps in China. This paper analyzes China's proposed minimum efficiency standard for fluorescent lamps and presents an assessment of its likely impacts on China's lighting energy consumption and GHG emissions.

  17. Near-infrared fluorescence image quality test methods for standardized performance evaluation

    Science.gov (United States)

    Kanniyappan, Udayakumar; Wang, Bohan; Yang, Charles; Ghassemi, Pejhman; Wang, Quanzeng; Chen, Yu; Pfefer, Joshua

    2017-03-01

    Near-infrared fluorescence (NIRF) imaging has gained much attention as a clinical method for enhancing visualization of cancers, perfusion and biological structures in surgical applications where a fluorescent dye is monitored by an imaging system. In order to address the emerging need for standardization of this innovative technology, it is necessary to develop and validate test methods suitable for objective, quantitative assessment of device performance. Towards this goal, we develop target-based test methods and investigate best practices for key NIRF imaging system performance characteristics including spatial resolution, depth of field and sensitivity. Characterization of fluorescence properties was performed by generating excitation-emission matrix properties of indocyanine green and quantum dots in biological solutions and matrix materials. A turbid, fluorophore-doped target was used, along with a resolution target for assessing image sharpness. Multi-well plates filled with either liquid or solid targets were generated to explore best practices for evaluating detection sensitivity. Overall, our results demonstrate the utility of objective, quantitative, target-based testing approaches as well as the need to consider a wide range of factors in establishing standardized approaches for NIRF imaging system performance.

  18. Bias in the absorption coefficient determination of a fluorescent dye, standard reference material 1932 fluorescein solution

    Energy Technology Data Exchange (ETDEWEB)

    DeRose, Paul C. [Analytical Chemistry Division, National Institute of Standards and Technology, 100 Bureau Dr., Gaithersburg, MD 20899-8394 (United States)]. E-mail: paul.derose@nist.gov; Kramer, Gary W. [Analytical Chemistry Division, National Institute of Standards and Technology, 100 Bureau Dr., Gaithersburg, MD 20899-8394 (United States)

    2005-06-15

    The absorption coefficient of standard reference material[registered] (SRM[registered]) 1932, fluorescein in a borate buffer solution (pH=9.5) has been determined at {lambda}=488.0, 490.0, 490.5 and 491.0 nm using the US national reference UV/visible spectrophotometer. The purity of the fluorescein was determined to be 97.6% as part of the certification of SRM 1932. The solution measured was prepared gravimetrically by diluting SRM 1932 with additional borate buffer. The value of the absorption coefficient was corrected for bias due to fluorescence that reaches the detector and for dye purity. Bias due to fluorescence was found to be on the order of -1% for both monochromatic and polychromatic (e.g., diode-array based) spectrophotometers.

  19. Section E9 of the American College of Medical Genetics technical standards and guidelines: fluorescence in situ hybridization.

    Science.gov (United States)

    Mascarello, James T; Hirsch, Betsy; Kearney, Hutton M; Ketterling, Rhett P; Olson, Susan B; Quigley, Denise I; Rao, Kathleen W; Tepperberg, James H; Tsuchiya, Karen D; Wiktor, Anne E

    2011-07-01

    This updated Section E9 has been incorporated into and supersedes the previous Section E9 in Section E: Clinical Cytogenetics of the 2008 Edition (Revised 02/2007) American College of Medical Genetics Standards and Guidelines for Clinical Genetics Laboratories. This section deals specifically with the standards and guidelines applicable to fluorescence in situ hybridization analysis.

  20. Quantitative micro x-ray fluorescence analyses without reference standard material; Referenzprobenfreie quantitative Mikro-Roentgenfluoreszenzanalyse

    Energy Technology Data Exchange (ETDEWEB)

    Wolff, Timo

    2009-07-15

    X-ray fluorescence analysis (XRF) is a standard method for non-destructive investigations. Due to the development of polycapillary optics and SDDdetectors requiring no cooling with liquid nitrogen, XRF becomes a suitable method for a large number of applications, e. g. for the analysis of objects in arts and archaeology. Spectrometers developed for those purposes allow investigations outside of laboratories und provide excitation areas with diameters of 10-70 {mu}m. In most applications, quantification of XRF data is realized by the usage of standard reference materials. Due to absorption processes in the samples the accuracy of the results depends strongly on the similarity of the sample and the reference standard. In cases where no suitable references are available, quantification can be done based on the ''fundamental parameter (fp) method''. This quantification procedure is based on a set of equations describing the fluorescence production and detection mathematical. The cross sections for the interaction of x-rays with matter can be taken from different databases. During an iteration process the element concentrations can be determined. Quantitative XRF based on fundamental parameters requires an accurate knowledge of the excitation spectrum. In case of a conventional setup this spectrum is given by the X-ray tube spectrum and can be calculated. The use of polycapillary optics in micro-XRF spectrometers changes the spectral distribution of the excitation radiation. For this reason it is necessary to access the transmission function of the used optic. The aim of this work is to find a procedure to describe this function for routine quantification based on fundamental parameters. Most of the measurements have been carried out using a commercial spectrometer developed for applications in arts and archaeology. On the one hand the parameters of the lens, used in the spectrometer, have been investigated by different experimental characterization

  1. New Homogeneous Standards by Atomic Layer Deposition for Synchrotron X-ray Fluorescence and Absorption Spectroscopies.

    Energy Technology Data Exchange (ETDEWEB)

    Butterworth, A.L.; Becker, N.; Gainsforth, Z.; Lanzirotti, A.; Newville, M.; Proslier, T.; Stodolna, J.; Sutton, S.; Tyliszczak, T.; Westphal, A.J.; Zasadzinski, J. (UCB)

    2012-03-13

    Quantification of synchrotron XRF analyses is typically done through comparisons with measurements on the NIST SRM 1832/1833 thin film standards. Unfortunately, these standards are inhomogeneous on small scales at the tens of percent level. We are synthesizing new homogeneous multilayer standards using the Atomic Layer Deposition technique and characterizing them using multiple analytical methods, including ellipsometry, Rutherford Back Scattering at Evans Analytical, Synchrotron X-ray Fluorescence (SXRF) at Advanced Photon Source (APS) Beamline 13-ID, Synchrotron X-ray Absorption Spectroscopy (XAS) at Advanced Light Source (ALS) Beamlines 11.0.2 and 5.3.2.1 and by electron microscopy techniques. Our motivation for developing much-needed cross-calibration of synchrotron techniques is borne from coordinated analyses of particles captured in the aerogel of the NASA Stardust Interstellar Dust Collector (SIDC). The Stardust Interstellar Dust Preliminary Examination (ISPE) team have characterized three sub-nanogram, {approx}1{micro}m-sized fragments considered as candidates to be the first contemporary interstellar dust ever collected, based on their chemistries and trajectories. The candidates were analyzed in small wedges of aerogel in which they were extracted from the larger collector, using high sensitivity, high spatial resolution >3 keV synchrotron x-ray fluorescence spectroscopy (SXRF) and <2 keV synchrotron x-ray transmission microscopy (STXM) during Stardust ISPE. The ISPE synchrotron techniques have complementary capabilities. Hard X-ray SXRF is sensitive to sub-fg mass of elements Z {ge} 20 (calcium) and has a spatial resolution as low as 90nm. X-ray Diffraction data were collected simultaneously with SXRF data. Soft X-ray STXM at ALS beamline 11.0.2 can detect fg-mass of most elements, including cosmochemically important oxygen, magnesium, aluminum and silicon, which are invisible to SXRF in this application. ALS beamline 11.0.2 has spatial resolution

  2. QSO Lifetimes

    CERN Document Server

    Martini, P

    2003-01-01

    The QSO lifetime t_Q is one of the most fundamental quantities for understanding black hole and QSO evolution, yet it remains uncertain by several orders of magnitude. If t_Q is long, then only a small fraction of galaxies went through a luminous QSO phase. In contrast, a short lifetime would require most galaxies today to have undergone a QSO phase in their youth. The current best estimates or constraints on t_Q from black hole demographics and the radiative properties of QSOs vary from at least 10^6 to 10^8 years. This broad range still allows both possibilities: that QSOs were either a rare or a common stage of galaxy evolution. These constraints also do not rule out the possibility that QSO activity is episodic, with individual active periods much shorter than the total active lifetime. In the next few years a variety of additional observational constraints on the lifetimes of QSOs will become available, including clustering measurements and the proximity effect. These new constraints can potentially dete...

  3. Standard coupling unification in SO(10), hybrid seesaw neutrino mass and leptogenesis, dark matter, and proton lifetime predictions

    Science.gov (United States)

    Parida, M. K.; Nayak, Bidyut Prava; Satpathy, Rajesh; Awasthi, Ram Lal

    2017-04-01

    We discuss gauge coupling unification of SU(3) C × SU(2) L × U(1) Y descending directly from non-supersymmetric SO(10) while providing solutions to the three out-standing problems of the standard model: neutrino masses, dark matter, and the baryon asymmetry of the universe. Conservation of matter parity as gauged discrete symmetry for the stability and identification of dark matter in the model calls for high-scale spontaneous symmetry breaking through 126 H Higgs representation. This naturally leads to the hybrid seesaw formula for neutrino masses mediated by heavy scalar triplet and right-handed neutrinos. Being quadratic in the Majorana coupling, the seesaw formula predicts two distinct patterns of right-handed neutrino masses, one hierarchical and another not so hierarchical (or compact), when fitted with the neutrino oscillation data. Predictions of the baryon asymmetry via leptogenesis are investigated through the decays of both the patterns of RH ν masses. A complete flavor analysis has been carried out to compute CP-asymmetries including washouts and solutions to Boltzmann equations have been utilised to predict the baryon asymmetry. The additional contribution to vertex correction mediated by the heavy left-handed triplet scalar is noted to contribute as dominantly as other Feynman diagrams. We have found successful predictions of the baryon asymmetry for both the patterns of right-handed neutrino masses. The SU(2) L triplet fermionic dark matter at the TeV scale carrying even matter parity is naturally embedded into the non-standard fermionic representation 45 F of SO(10). In addition to the triplet scalar and the triplet fermion, the model needs a nonstandard color octet fermion of mass ˜ 5 × 107 GeV to achieve precision gauge coupling unification at the GUT mass scale M U 0 = 1015.56 GeV. Threshold corrections due to superheavy components of 126H and other representations are estimated and found to be substantial. It is noted that the proton life

  4. Comparative study of the fatty acid binding process of a new FABP from Cherax quadricarinatus by fluorescence intensity, lifetime and anisotropy.

    Directory of Open Access Journals (Sweden)

    Jiayao Li

    Full Text Available Fatty acid-binding proteins (FABPs are small cytosolic proteins, largely distributed in invertebrates and vertebrates, which accomplish uptake and intracellular transport of hydrophobic ligands such as fatty acids. Although long chain fatty acids play multiple crucial roles in cellular functions (structural, energy metabolism, regulation of gene expression, the precise functions of FABPs, especially those of invertebrate species, remain elusive. Here, we have identified and characterized a novel FABP family member, Cq-FABP, from the hepatopancreas of red claw crayfish Cherax quadricarinatus. We report the characterization of fatty acid-binding affinity of Cq-FABP by four different competitive fluorescence-based assays. In the two first approaches, the fluorescent probe 8-Anilino-1-naphthalenesulfonate (ANS, a binder of internal cavities of protein, was used either by directly monitoring its fluorescence emission or by monitoring the fluorescence resonance energy transfer occurring between the single tryptophan residue of Cq-FABP and ANS. The third and the fourth approaches were based on the measurement of the fluorescence emission intensity of the naturally fluorescent cis-parinaric acid probe or the steady-state fluorescence anisotropy measurements of a fluorescently labeled fatty acid (BODIPY-C16, respectively. The four methodologies displayed consistent equilibrium constants for a given fatty acid but were not equivalent in terms of analysis. Indeed, the two first methods were complicated by the existence of non specific binding modes of ANS while BODIPY-C16 and cis-parinaric acid specifically targeted the fatty acid binding site. We found a relationship between the affinity and the length of the carbon chain, with the highest affinity obtained for the shortest fatty acid, suggesting that steric effects primarily influence the interaction of fatty acids in the binding cavity of Cq-FABP. Moreover, our results show that the binding affinities

  5. Direct solid surface fluorescence spectroscopy of standard chemicals and humic acid in ternary system.

    Science.gov (United States)

    Mounier, S; Nicolodelli, G; Redon, R; Milori, D M B P

    2017-04-15

    The front face fluorescence spectroscopy is often used to quantify chemicals in well-known matrices as it is a rapid and powerful technique, with no sample preparation. However it was not used to investigate extracted organic matter like humic substances. This work aims to fully investigate for the first time front face fluorescence spectroscopy response of a ternary system including boric acid, tryptophan and humic substances, and two binaries system containing quinine sulfate or humic substance in boric acid. Pure chemicals, boric acid, tryptophan, quinine sulfate and humic acid were mixed together in solid pellet at different contents from 0 to 100% in mass. The measurement of excitation emission matrix of fluorescence (3D fluorescence) and laser induced fluorescence were then done in the front face mode. Fluorescence matrices were decomposed using the CP/PARAFAC tools after scattering treatments. Results show that for 3D fluorescence there is no specific component for tryptophan and quinine sulfate, and that humic substances lead to a strong extinction effect for mixture containing quinine sulfate. Laser induced fluorescence gives a very good but non-specific related response for both quinine sulfate and tryptophan. No humic substances fluorescence response was found, but extinction effect is observed as for 3D fluorescence. This effect is stronger for quinine sulfate than for tryptophan. These responses were modeled using a simple absorbance versus emission model.

  6. Direct solid surface fluorescence spectroscopy of standard chemicals and humic acid in ternary system

    Science.gov (United States)

    Mounier, S.; Nicolodelli, G.; Redon, R.; Milori, D. M. B. P.

    2017-04-01

    The front face fluorescence spectroscopy is often used to quantify chemicals in well-known matrices as it is a rapid and powerful technique, with no sample preparation. However it was not used to investigate extracted organic matter like humic substances. This work aims to fully investigate for the first time front face fluorescence spectroscopy response of a ternary system including boric acid, tryptophan and humic substances, and two binaries system containing quinine sulfate or humic substance in boric acid. Pure chemicals, boric acid, tryptophan, quinine sulfate and humic acid were mixed together in solid pellet at different contents from 0 to 100% in mass. The measurement of excitation emission matrix of fluorescence (3D fluorescence) and laser induced fluorescence were then done in the front face mode. Fluorescence matrices were decomposed using the CP/PARAFAC tools after scattering treatments. Results show that for 3D fluorescence there is no specific component for tryptophan and quinine sulfate, and that humic substances lead to a strong extinction effect for mixture containing quinine sulfate. Laser induced fluorescence gives a very good but non-specific related response for both quinine sulfate and tryptophan. No humic substances fluorescence response was found, but extinction effect is observed as for 3D fluorescence. This effect is stronger for quinine sulfate than for tryptophan. These responses were modeled using a simple absorbance versus emission model.

  7. Dual Lifetimes for Complexes between Glutathione-S-transferase (hGSTA1-1) and Product-like Ligands Detected by Single-Molecule Fluorescence Imaging.

    Science.gov (United States)

    Pettersson, John R; Lanni, Frederick; Rule, Gordon S

    2017-08-08

    Single-molecule fluorescence techniques were used to characterize the binding of products and inhibitors to human glutathione S-transferase A1-1 (hGSTA1-1). The identification of at least two different bound states for the wild-type enzyme suggests that there are at least two conformations of the protein, consistent with the model that ligand binding promotes closure of the carboxy-terminal helix over the active site. Ligand induced changes in ensemble fluorescence energy transfer support this proposed structural change. The more predominant state in the ensemble of single molecules shows a significantly faster off-rate, suggesting that the carboxy-terminal helix is delocalized in this state, permitting faster exit of the bound ligand. A point mutation (I219A), which is known to interfere with the association of the carboxy-terminal helix with the enzyme, shows increased rates of interconversion between the open and closed state. Kinematic traces of fluorescence from single molecules show that a single molecule readily samples a number of different conformations, each with a characteristic off-rate.

  8. Interactions between epinastine and human serum albumin: Investigation by fluorescence, UV-vis, FT-IR, CD, lifetime measurement and molecular docking

    Science.gov (United States)

    Ariga, Girish G.; Naik, Praveen N.; Chimatadar, Shivamurti A.; Nandibewoor, Sharanappa T.

    2017-06-01

    The fluorescence quenching of human serum albumin (HSA) by epinastine hydrochloride (EPN) at pH 7.4 buffer was studied using absorption, fluorescence quenching, time-resolved, circular-dichroism, synchronous and molecular docking studies have been employed in the system. The fluorescence quenching study revealed that the static quenching mechanism was involved in the interaction of EPN with human serum albumin. The value number of binding sites, n, is close to unity, EPN-HSA, indicated the presence of a single class of binding site for the drug in protein. The binding constant value of EPN_HSA was observed to be 2.72 × 104 M-1 at 298 K. The spectral results attest that the binding of EPN-HSA induced conformational changes in the HSA. The metal ions viz., Ca2+, Co2+, Cu2+, Ni2+ and Zn2+ were found to influence the binding of the EPN to HSA. Based on the Forster's theory of non-radiation energy transfer, the binding average distance, r, between the donor (HSA) and acceptor (EPN) was found to be 4.33 nm. The circular dichroism data revealed that the presence of EPN decreased the α-helix content of serum albumin, which indicated conformation changes in HSA upon interaction with EPN.

  9. Steady-State Fluorescence and Lifetime Emission Study of pH-Sensitive Probes Based on i-motif Forming Oligonucleotides Single and Double Labeled with Pyrene

    Directory of Open Access Journals (Sweden)

    Anna Dembska

    2015-09-01

    Full Text Available Cytosine-rich nucleic acids undergo pH-stimulated structural transitions leading to formation of an i-motif architecture at an acidic pH. Thus, i-motifs are good foundation for designing simple pH-sensitive fluorescent probes. We report here steady-state and time-resolved fluorescence studies of pyrene-labeled probes based on RET sequence: C4GC4GC4GC4TA (RET21, AC4GC4GC4GC4TA (RET21A and C4GC4GC4GC4T (RET20. Comparative studies with single- and double-labeled i-motif probes were carried out. For each probe, we have measured fluorescence spectra and decays for emission wavelength of 390 nm over a wide range of pH (from 4.0 to 8.0. Effect of the oligonucleotide sequence and the number of pyrene labels on the spectral characteristics of probes were discussed.

  10. Upgrading the GSI beamline microscope with a confocal fluorescence lifetime scanner to monitor charged particle induced chromatin decondensation in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Abdollahi, Elham; Taucher-Scholz, Gisela [Department of Biophysics, GSI Helmholtz Center for Heavy Ion Research, Planckstrasse 1, 64291 Darmstadt (Germany); Durante, Marco [Department of Biophysics, GSI Helmholtz Center for Heavy Ion Research, Planckstrasse 1, 64291 Darmstadt (Germany); Institute for Condensed Matter Physics, Darmstadt University of Technology, 64289 Darmstadt (Germany); Jakob, Burkhard, E-mail: B.Jakob@gsi.de [Department of Biophysics, GSI Helmholtz Center for Heavy Ion Research, Planckstrasse 1, 64291 Darmstadt (Germany)

    2015-12-15

    We report the upgrade of the GSI beamline microscope coupled to the linear accelerator UNILAC by a confocal FLIM scanner utilizing time correlated single photon counting technique (TCSPC). The system can now be used to address the radiation induced chromatin decondensation in more detail and with higher sensitivity compared to intensity based methods. This decondensation of heterochromatic areas is one of the early DNA damage responses observed after charged particle irradiation and might facilitate the further processing of the induced lesions. We describe here the establishment of different DNA dyes as chromatin compaction probes usable for quantification of the DNA condensation status in living cells utilizing lifetime imaging. In addition, we find an evidence of heterochromatic chromatin decondensation in ion irradiated murine chromocenters detected after subsequent fixation using FLIM measurements.

  11. Standard practice for fluorescent liquid penetrant testing using the hydrophilic Post-Emulsification process

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2010-01-01

    1.1 This practice covers procedures for fluorescent penetrant examination utilizing the hydrophilic post-emulsification process. It is a nondestructive testing method for detecting discontinuities that are open to the surface such as cracks, seams, laps, cold shuts, laminations, isolated porosity, through leaks, or lack of fusion and is applicable to in-process, final, and maintenance examination. It can be effectively used in the examination of nonporous, metallic materials, both ferrous and nonferrous, and of nonmetallic materials such as glazed or fully densified ceramics and certain nonporous plastics and glass. 1.2 This practice also provides a reference: 1.2.1 By which a fluorescent penetrant examination hydrophilic post-emulsification process recommended or required by individual organizations can be reviewed to ascertain their applicability and completeness. 1.2.2 For use in the preparation of process specifications dealing with the fluorescent penetrant examination of materials and parts using the hy...

  12. Standard practice for fluorescent liquid penetrant testing using the Solvent-Removable process

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2010-01-01

    1.1 This practice covers procedures for fluorescent penetrant examination utilizing the solvent-removable process. It is a nondestructive testing method for detecting discontinuities that are open to the surface, such as cracks, seams, laps, cold shuts, laminations, isolated porosity, through leaks, or lack of fusion and is applicable to in-process, final, and maintenance examination. It can be effectively used in the examination of nonporous, metallic materials, both ferrous and nonferrous, and of nonmetallic materials such as glazed or fully densified ceramics and certain nonporous plastics and glass. 1.2 This practice also provides a reference: 1.2.1 By which a fluorescent penetrant examination solvent-removable process recommended or required by individual organizations can be reviewed to ascertain its applicability and completeness. 1.2.2 For use in the preparation of process specifications dealing with the fluorescent solvent-removable liquid penetrant examination of materials and parts. Agreement by th...

  13. Standard practice for fluorescent liquid penetrant testing using the Water-Washable process

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2010-01-01

    1.1 This practice covers procedures for water-washable fluorescent penetrant examination of materials. It is a nondestructive testing method for detecting discontinuities that are open to the surface such as cracks, seams, laps, cold shuts, laminations, isolated porosity, through leaks, or lack of porosity and is applicable to in-process, final, and maintenance examination. It can be effectively used in the examination of nonporous, metallic materials, both ferrous and nonferrous, and of nonmetallic materials such as glazed or fully densified ceramics and certain nonporous plastics and glass. 1.2 This practice also provides a reference: 1.2.1 By which a fluorescent penetrant examination method using the water-washable process recommended or required by individual organizations can be reviewed to ascertain its applicability and completeness. 1.2.2 For use in the preparation of process specifications dealing with the water-washable fluorescent penetrant examination of materials and parts. Agreement by the purch...

  14. A fluorescence lifetime-based binding assay for acetylpolyamine amidohydrolases from Pseudomonas aeruginosa using a [1,3]dioxolo[4,5-f][1,3]benzodioxole (DBD) ligand probe.

    Science.gov (United States)

    Meyners, Christian; Wawrzinek, Robert; Krämer, Andreas; Hinz, Steffen; Wessig, Pablo; Meyer-Almes, Franz-Josef

    2014-08-01

    High-throughput assays for drug screening applications have to fulfill particular specifications. Besides the capability to identify even compounds with low potency, one of the major issues is to minimize the number of false-positive hits in a screening campaign in order to reduce the logistic effort for the subsequent cherry picking and confirmation procedure. In this respect, fluorescence lifetime (FLT) appears as an ideal readout parameter that is supposed to be robust against autofluorescent and light-absorbing compounds, the most common source of systematic false positives. The extraordinary fluorescence features of the recently discovered [1,3]dioxolo[4,5-f][1,3] benzodioxole dyes were exploited to develop an FLT-based binding assay with exceptionally robust readout. The assay setup was comprehensively validated and shown to comply not only with all requirements for a powerful high-throughput screening assay but also to be suitable to determine accurate binding constants for inhibitors against enzymes of the histone deacetylase family. Using the described binding assay, the first inhibitors against three members of this enzyme family from Pseudomonas aeruginosa were identified. The compounds were characterized in terms of potency and selectivity profile. The novel ligand probe should also be applicable to other homologues of the histone deacetylase family that are inhibited by N-hydroxy-N'-phenyloctandiamide.

  15. Optical-sectioning microscopy of protoporphyrin IX fluorescence in human gliomas: standardization and quantitative comparison with histology

    Science.gov (United States)

    Wei, Linpeng; Chen, Ye; Yin, Chengbo; Borwege, Sabine; Sanai, Nader; Liu, Jonathan T. C.

    2017-04-01

    Systemic delivery of 5-aminolevulinic acid leads to enhanced fluorescence image contrast in many tumors due to the increased accumulation of protoporphyrin IX (PpIX), a fluorescent porphyrin that is associated with tumor burden and proliferation. The value of PpIX-guided resection of malignant gliomas has been demonstrated in prospective randomized clinical studies in which a twofold greater extent of resection and improved progression-free survival have been observed. In low-grade gliomas and at the diffuse infiltrative margins of all gliomas, PpIX fluorescence is often too weak to be detected with current low-resolution surgical microscopes that are used in operating rooms. However, it has been demonstrated that high-resolution optical-sectioning microscopes are capable of detecting the sparse and punctate accumulations of PpIX that are undetectable via conventional low-power surgical fluorescence microscopes. To standardize the performance of high-resolution optical-sectioning devices for future clinical use, we have developed an imaging phantom and methods to ensure that the imaging of PpIX-expressing brain tissues can be performed reproducibly. Ex vivo imaging studies with a dual-axis confocal microscope demonstrate that these methods enable the acquisition of images from unsectioned human brain tissues that quantitatively and consistently correlate with images of histologically processed tissue sections.

  16. In-vitro and in-vivo detection of p53 by fluorescence lifetime on a hybrid FITC-gold nanosensor

    Science.gov (United States)

    Sironi, L.; Freddi, S.; D'Alfonso, L.; Collini, M.; Gorletta, T.; Soddu, S.; Chirico, G.

    2010-02-01

    P53 is a tumor suppressor used as marker for early cancer diagnosis and prognosis. We have studied constructs based on gold nanoparticles (NPs) decorated with specific anti-p53 antibodies and with a fluoresceine derivative, FITC. The interaction of gold surface plasmons with fluorophores bound within few nanometers from the surface, likely induces changes in the fluorophore excited state lifetime. Indeed we found previously that this parameter follows linearly the p53 concentration in solutions (in vitro conditions) up to 200-400 pM, depending on the size of the NP, with a 5 pM uncertainty. We have evaluated here the nanosensor specificity for p53 by testing it in-vitro against bovine serum albumine, beta-lactolglobulin and lysozyme. Moreover, the titration of total cell extracts from p53+/+ or p53-/- cells with the p53antibody decorated gold NPs, indicates that this construct can also be used to detect the presence of p53 in total cell extracts and it will be therefore a valuable tool also for in vivo screening.

  17. Study on fluorescence absolute quantum yield and lifetime of europium complexes by doping yttrium%掺杂钇的铕稀土配合物的荧光绝对量子产率和寿命的研究

    Institute of Scientific and Technical Information of China (English)

    费邦忠; 陶栋梁; 张宏; 崔玉民; 张坤; 王永忠; 杨森林; 鲁仕梅

    2016-01-01

    A series of Co-luminescence EuxY1-x(TTA)3phen were synthesized in anhydrous ethanol by using Eu3+and Y3+as central ions and 2-Thenoyltrifluoroacetone (TTA) and 1,10-phenanthroline (phen) as ligands. IR spectra of the ligand TTA and EuxY1-x(TTA)3phen were determined. The absolute fluorescence quantum yields and average fluorescence lifetimes of europium complexes undergo great change after the europium complexes are doped Y into. With the Y content increasing, the absolute quantum yields of EuxY1- x(TTA)3phen first increase and then decrease, and the average fluorescence lifetimes of EuxY1- x (TTA)3phen become shorter in a wave-like pattern. These results indicate that Y-doped results in intramolecular microstructure change of EuxY1-x(TTA)3phen, which results in change of intramolecular energy transfer system of EuxY1-x(TTA)3phen.%在无水乙醇中,利用Eu3+和Y3+作为中心离子,α-噻吩甲酰三氟丙酮(TTA)和1,10-邻菲啰啉(phen)作为配体制备了一系列共发光稀土配合物EuxY1-x(TTA)3phen,并对TTA和EuxY1-x(TTA)3phen进行了红外表征。掺杂钇的铕配合物与没掺杂钇相比,荧光绝对量子产率和平均寿命都发生了很大变化,随着钇含量的增大,EuxY1-x(TTA)3phen的荧光绝对量子产率先增大,然后减小,而平均寿命则以波动方式逐渐减小,说明钇的掺杂改变了EuxY1-x(TTA)3phen的分子微观结构,从而改变了EuxY1-x(TTA)3phen的能量传递方式。

  18. Family of lifetime sensors for medical purposes

    Science.gov (United States)

    Lippitsch, Max E.; Draxler, Sonja

    1995-05-01

    A family of indicators has been developed for fluorescence lifetime-based measurement of oxygen, pH, carbon dioxide, and potassium, all the indicators being derivatives of the same chemical compound and having identical spectral and lifetime properties. The indicators show an absorption accessible to low- cast light sources, a large Stokes shift, and long fluorescence decay time. all indicators can be excited at the same excitation wavelength, monitored at the same emission wavelength, and measured within the same time range. This opens the possibility of building a compact lifetime-based instrument to simultaneously measure blood gases and cations.

  19. The development of a single molecule fluorescence standard and its application in estimating the stoichiometry of the nuclear pore complex.

    Science.gov (United States)

    Tie, Hieng Chiong; Madugula, Viswanadh; Lu, Lei

    2016-09-30

    We report here an image-based method to quantify the stoichiometry of diffraction-limited sub-cellular protein complexes in vivo under spinning disk confocal microscopy. A GFP single molecule fluorescence standard was first established by immobilizing His-tagged GFP molecules onto the glass surface via nickel nitrilotriacetic acid functionalized polyethylene glycol. When endogenous nucleoporins were knocked down and replaced by the exogenously expressed and knockdown-resistant GFP-nucleoporins, the stoichiometry of the nucleoporin was estimated by the ratio of its fluorescence intensity to that of the GFP single molecules. Our measured stoichiometry of Nup35, Nup93, Nup133 and Nup88 is 23, 18, 14 and 9 and there are possibly16 copies of Nup107-160 complex per nuclear pore complex.

  20. Standards for Analysis of Ce, La, Pb, Rb, Se, Sr, Y, AND Zr in Rock Samples Using Laser-induced Breakdown Spectroscopy and X-ray Fluorescence

    Science.gov (United States)

    Mackie, Jason; Dyar, M. Darby; Ytsma, Caroline; Lepore, Kate; Fassett, Caleb I.; Hanlon, Avery; Wagoner, Carlie; Treiman, Allan

    2017-01-01

    Analytical geochemistry has long depended on the availability of robust suites of rock standards with well-characterized compositions. Standard rock powders for wet chemistry and x-ray fluorescence were initially characterized and supplied to the community by the U.S. Geological Survey, which continues to distribute a few dozen standards. Many other rock standards have subsequently been developed by organizations such as the Centre de Recherches Pétrographiques et Géochimiques (CRPG) and Brammer Standard Company, Inc.

  1. Standard test method for uranium analysis in natural and waste water by X-ray fluorescence

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2004-01-01

    1.1 This test method applies for the determination of trace uranium content in waste water. It covers concentrations of U between 0.05 mg/L and 2 mg/L. 1.2 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard. 1.3 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

  2. Standard test method for determination of low concentrations of uranium in oils and organic liquids by X-ray fluorescence

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2011-01-01

    1.1 This test method covers the steps necessary for the preparation and analysis by X-ray fluorescence (XRF) of oils and organic solutions containing uranium. Two different preparation techniques are described. 1.2 The procedure is valid for those solutions containing 20 to 2000 μg uranium per mL as presented to the spectrometer for the solution technique and 200 to 50 000 μg uranium per g for the pellet technique. 1.3 This test method requires the use of an appropriate internal standard. Care must be taken to ascertain that samples analyzed by this test method do not contain the internal standard or that this contamination, whenever present, has been corrected for mathematically. Such corrections are not addressed in this procedure. Care must be taken that the internal standard and sample medium are compatible; that is, samples must be miscible with tri-n-butyl phosphate (TBP) and must not remove the internal standard from solution. Alternatively, a scatter line may be used as the internal standard. 1....

  3. Development of Standard Samples for on-board Calibration of a New Planetary X-Ray Fluorescence Spectrometer

    Science.gov (United States)

    Dreißigacker, Anne; Köhler, Eberhard; Fabel, Oliver; van Gasselt, Stephan

    2014-05-01

    At the Planetary Sciences and Remote Sensing research group at Freie Universität Berlin an SCD-based X-Ray Fluorescence Spectrometer is being developed to be employed on planetary orbiters to conduct direct, passive energy-dispersive x-ray fluorescence measurements of planetary surfaces through measuring the emitted X-Ray fluorescence induced by solar x-rays and high energy particles. Because the Sun is a highly variable radiation source, the intensity of solar X-Ray radiation has to be monitored constantly to allow for comparison and signal calibration of X-Ray radiation from lunar surface materials. Measurements are obtained by indirectly monitoring incident solar x-rays emitted from a calibration sample. This has the additional advantage of minimizing the risk of detector overload and damage during extreme solar events such as high-energy solar flares and particle storms as only the sample targets receive the higher radiation load directly (while the monitor is never directly pointing towards the Sun). Quantitative data are being obtained and can be subsequently analysed through synchronous measurement of fluorescence of the Moon's surface by the XRF-S main instrument and the emitted x-ray fluorescence of calibration samples by the XRF-S-ISM (Indirect Solar Monitor). We are currently developing requirements for 3 sample tiles for onboard correction and calibration of XRF-S, each with an area of 3-9 cm2 and a maximum weight of 45 g. This includes development of design concepts, determination of techniques for sample manufacturing, manufacturing and testing of prototypes and statistical analysis of measurement characteristics and quantification of error sources for the advanced prototypes and final samples. Apart from using natural rock samples as calibration sample, we are currently investigating techniques for sample manufacturing including laser sintering of rock-glass on metals, SiO2-stabilized mineral-powders, or artificial volcanic glass. High precision

  4. Steady-state and time-resolved fluorometry of fluorescent pollutants and heavy metal complexes

    Science.gov (United States)

    Resch, Ute; Rurack, Knut

    1997-05-01

    Time-resolved laser-induced fluorescence spectroscopy is one of the most sensitive optical methods which is well suited for on-line in situ analysis. Here, three examples for the steady- state and time-resolved fluorescence analysis of environmentally important analytes, the fluorescent monoaromatic hydrocarbons benzene, toluene, and xylene as well as non fluorescent heavy metal ions forming a fluorescent complex with a cation coordinating fluorescence probe, are presented and the potential of both methods is discussed. For BTX, various mixtures of the spectrally similar compounds B, T, and X showing different fluorescence lifetimes were studied with both methods. As an example for fluorometric metal ion analysis, the fluorescence probe BP(OH)2 (2,2'-bipyridyl- 3,3'-diol) was employed for the determination of d10 metal ions in water and the newly developed fluorescence probe APTA for the detection of Cu(II). Cation complexation of BP(OH2 yields spectrally very similar complexes which differ in their fluorescence lifetimes. Complexation of APTA to Cu(II) leads to small spectral changes and a strong increase in fluorescence quantum yield and lifetime. For the analytes studied, a comparison of the detection limits, standard deviations, and linear dynamic range of both methods clearly demonstrates the analytical potential of time-resolved fluorometry.

  5. Real time optical Biopsy: Time-resolved Fluorescence Spectroscopy instrumentation and validation

    Science.gov (United States)

    Kittle, David S.; Vasefi, Fartash; Patil, Chirag G.; Mamelak, Adam; Black, Keith L.; Butte, Pramod V.

    2016-12-01

    The Time-resolved fluorescence spectroscopy (TR-FS) has the potential to differentiate tumor and normal tissue in real time during surgical excision. In this manuscript, we describe the design of a novel TR-FS device, along with preliminary data on detection accuracy for fluorophores in a mixture. The instrument is capable of near real-time fluorescence lifetime acquisition in multiple spectral bands and analysis. It is also able to recover fluorescence lifetime with sub-20ps accuracy as validated with individual organic fluorescence dyes and dye mixtures yielding lifetime values for standard fluorescence dyes that closely match with published data. We also show that TR-FS is able to quantify the relative concentration of fluorescence dyes in a mixture by the unmixing of lifetime decays. We show that the TR-FS prototype is able to identify in near-real time the concentrations of dyes in a complex mixture based on previously trained data. As a result, we demonstrate that in complex mixtures of fluorophores, the relative concentration information is encoded in the fluorescence lifetime across multiple spectral bands. We show for the first time the temporal and spectral measurements of a mixture of fluorochromes and the ability to differentiate relative concentrations of each fluorochrome mixture in real time.

  6. Standard test methods for chemical analysis of ceramic whiteware materials using wavelength dispersive X-Ray fluorescence spectrometry

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2004-01-01

    1.1 These test methods cover the determination of ten major elements (SiO2, Al2O3, Fe2O3, MgO, CaO, Na2O, K2O, TiO2, P2O5, MnO, and LOI in ceramic whitewares clays and minerals using wavelength dispersive X-ray fluorescence spectrometry (WDXRF). The sample is first ignited, then fused with lithium tetraborate and the resultant glass disc is introduced into a wavelength dispersive X-ray spectrometer. The disc is irradiated with X-rays from an X-ray tube. X-ray photons emitted by the elements in the samples are counted and concentrations determined using previously prepared calibration standards. (1) In addition to 10 major elements, the method provides a gravimetric loss-on-ignition. Note 1—Much of the text of this test method is derived directly from Major element analysis by wavelength dispersive X-ray fluorescence spectrometry, included in Ref (1). 1.2 Interferences, with analysis by WDXRF, may result from mineralogical or other structural effects, line overlaps, and matrix effects. The structure of the...

  7. Remote UV Fluorescence Lifetime Spectrometer Project

    Data.gov (United States)

    National Aeronautics and Space Administration — In-situ studies of the rocks, minerals, and soil on the Moon's surface provide a wealth of information during field geology and the mining phase for planetary...

  8. 利用基于扫描相机的荧光寿命成像显微技术研究细胞周期%Study on Cell Cycle Using Fluorescence Lifetime Imaging Microscopic System Based on a Streak Camera

    Institute of Scientific and Technical Information of China (English)

    王岩; 赵羚伶; 陈同生; 许改霞; Artem Pliss; Tymish Y.Ohulchanskyy; Paras N.Prasad; 屈军乐; 牛憨笨

    2011-01-01

    The fluorescence lifetime of the HeLa cells which are transfected with green fluorescent proteins during the cell cycle is investigated using fluorescence lifetime imaging microscopic system based on a streak camera.Experimental results show that fluorescence lifetime of HeLa cells is between 2.50 ns and 3.00 ns during different processes of cell cycle. The fluorescence lifetime of the mitosis cell drops to 2.82 ns from 2.86 ns within one hour,and it further drops to 2.78 ns from 2.82 ns during the roughly eight hours of the pre-DNA-synthetic phase (Glphase). The difference of the fluorescence lifetime implies that the macromolecular concentration in the nucleoplasm of the cell nucleus changes throughout the cell cycle, the study of which is significant to the understanding of macromolecular processes, kinetics and concentrations in the nucleoplasm of the cell nucleus throughout the cell cycle, as well as the regulation of cell cycles.%利用基于扫描相机的荧光寿命成像显微系统,以细胞周期为模型,研究转染绿色荧光蛋白的HeLa细胞的荧光寿命.结果表明,处于周期内不同进程的细胞的荧光寿命为2.50~3.00 ns.处于分裂期的细胞的荧光寿命在1 h内从2.86 ns下降到2.82 ns;在DNA合成前期的8 h内,荧光寿命从2.82 ns下降到2.78 ns.荧光寿命的差异反映了细胞周期中核浆内大分子浓度的变化,对了解细胞周期的分子机制有一定的意义.

  9. Principles of fluorescence techniques

    CERN Document Server

    2016-01-01

    Fluorescence techniques are being used and applied increasingly in academics and industry. The Principles of Fluorescence Techniques course will outline the basic concepts of fluorescence techniques and the successful utilization of the currently available commercial instrumentation. The course is designed for students who utilize fluorescence techniques and instrumentation and for researchers and industrial scientists who wish to deepen their knowledge of fluorescence applications. Key scientists in the field will deliver theoretical lectures. The lectures will be complemented by the direct utilization of steady-state and lifetime fluorescence instrumentation and confocal microscopy for FLIM and FRET applications provided by leading companies.

  10. Standard test method for analysis of uranium and thorium in soils by energy dispersive X-Ray fluorescence spectroscopy

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2011-01-01

    1.1 This test method covers the energy dispersive X-ray fluorescence (EDXRF) spectrochemical analysis of trace levels of uranium and thorium in soils. Any sample matrix that differs from the general ground soil composition used for calibration (that is, fertilizer or a sample of mostly rock) would have to be calibrated separately to determine the effect of the different matrix composition. 1.2 The analysis is performed after an initial drying and grinding of the sample, and the results are reported on a dry basis. The sample preparation technique used incorporates into the sample any rocks and organic material present in the soil. This test method of sample preparation differs from other techniques that involve tumbling and sieving the sample. 1.3 Linear calibration is performed over a concentration range from 20 to 1000 μg per gram for uranium and thorium. 1.4 The values stated in SI units are to be regarded as the standard. The inch-pound units in parentheses are for information only. 1.5 This standard...

  11. High efficiency fluorescent tubes. An efficient solution for the economical lighting of industrial and tertiary buildings; Les tubes fluorescents haut rendement. Une solution performante pour l'eclairage economique des locaux indutriels et tertiaires

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2003-07-01

    Fluorescent lamps are low pressure mercury vapor lamps (discharge lamps). The high efficiency fluorescent tubes have a greater lighting efficiency thanks to the integration of tri-chromatic powders emitting in the three fundamental colors (red, green, blue). Their useful lifetime is exceptionally long (90% of their initial flux after 12000 hours of use), and they require a lower maintenance with respect to the traditional fluorescent tubes. This document presents the characteristics and advantages of high efficiency fluorescent tubes for professional use and their cost and performance with respect to standard fluorescent tubes. (J.S.)

  12. LEDs for fluorescence microscopy

    NARCIS (Netherlands)

    Young, I.T.; Garini, Y.; Dietrich, H.R.C.; Van Oel, W.; Liqui Lung, G.

    2004-01-01

    Traditional light sources for fluorescence microscopy have been mercury lamps, xenon lamps, and lasers. These sources have been essential in the development of fluorescence microscopy but each can have serious disadvantages: lack of near monochromaticity, heat generation, cost, lifetime of the light

  13. MIQE précis: Practical implementation of minimum standard guidelines for fluorescence-based quantitative real-time PCR experiments

    NARCIS (Netherlands)

    Bustin, S.A.; Beaulieu, J.F.; Huggett, J.; Jaggi, R.; Kibenge, F.S.; Olsvik, P.A.; Penning, L.C.; Toegel, S.

    2010-01-01

    MIQE précis: Practical implementation of minimum standard guidelines for fluorescence-based quantitative real-time PCR experiments Stephen A Bustin1 , Jean-François Beaulieu2 , Jim Huggett3 , Rolf Jaggi4 , Frederick SB Kibenge5 , Pål A Olsvik6 , Louis C Penning7 and Stefan Toegel8 1 Centre for Diges

  14. Determining the Performance of Fluorescence Molecular Imaging Devices Using Traceable Working Standards With SI Units of Radiance.

    Science.gov (United States)

    Zhu, Banghe; Rasmussen, John C; Litorja, Maritoni; Sevick-Muraca, Eva M

    2016-03-01

    To date, no emerging preclinical or clinical near-infrared fluorescence (NIRF) imaging devices for noninvasive and/or surgical guidance have their performances validated on working standards with SI units of radiance that enable comparison or quantitative quality assurance. In this work, we developed and deployed a methodology to calibrate a stable, solid phantom for emission radiance with International System of Units (SI) units of mW ·sr(-1) ·cm(-2) for use in characterizing the measurement sensitivity of ICCD and IsCMOS detection, signal-to-noise ratio, and contrast. In addition, at calibrated radiances, we assess transverse and lateral resolution of ICCD and IsCMOS camera systems. The methodology allowed demonstration of superior SNR of the ICCD over the IsCMOS technology and superior resolution of the IsCMOS over the ICCD approach. Contrast depended upon the camera settings (binning and integration time) and gain of intensifier. Finally, because the architecture of CMOS and CCD camera systems results in vastly different performance, we comment on the utility of these technologies for small animal imaging as well as clinical applications for noninvasive and surgical guidance.

  15. Experimental Investigation of Excited-State Lifetimes in Atomic Ytterbium

    Energy Technology Data Exchange (ETDEWEB)

    Bowers, C.J.; Budker, D.; Commins, E.D.; DeMille, D.; Freedman, S.J.; Nguyen, A.-T.; Shang, S.-Q.; /UC, Berkeley; Zolotorev, M.; /SLAC

    2011-11-15

    Lifetimes of 21 excited states in atomic Yb were measured using time-resolved fluorescence detection following pulsed laser excitation. The lifetime of the 4f{sup 14}5d6s {sup 3}D{sub 1} state, which is of particular importance for a proposed study of parity nonconservation in atoms, was measured to be 380(30) ns.

  16. Octyl gallate: An antioxidant demonstrating selective and sensitive fluorescent property.

    Science.gov (United States)

    Wang, Qing; Zhang, Yongkui; Li, Hui

    2017-03-15

    Octyl gallate (OG) is an internationally recognized antioxidant that demonstrates selective and sensitive fluorescent property. The fluorescence of OG can be selectively enhanced in the presence of human serum albumin (HSA) and bovine serum albumin (BSA). The specific structures of HSA and BSA provided the basic conditions for fluorescence enhancement. OG yielded approximately 49- and 11-fold increments in emission intensity in the presence of HSA and BSA at a molar ratio of 1:1, respectively. The lifetimes of HSA and BSA correspondingly decreased. A Förster resonance energy transfer phenomenon occurred during interaction between OG and HSA or BSA. Our in-depth investigation of OG-HSA interaction showed that formation of a stable complex was an important prerequisite to efficiently enhance the fluorescence of OG. The selective and sensitive fluorescent property of OG can possibly be used to determine OG concentration via the standard addition method, which must be performed under certain conditions.

  17. Modelling lifetime data with multivariate Tweedie distribution

    Science.gov (United States)

    Nor, Siti Rohani Mohd; Yusof, Fadhilah; Bahar, Arifah

    2017-05-01

    This study aims to measure the dependence between individual lifetimes by applying multivariate Tweedie distribution to the lifetime data. Dependence between lifetimes incorporated in the mortality model is a new form of idea that gives significant impact on the risk of the annuity portfolio which is actually against the idea of standard actuarial methods that assumes independent between lifetimes. Hence, this paper applies Tweedie family distribution to the portfolio of lifetimes to induce the dependence between lives. Tweedie distribution is chosen since it contains symmetric and non-symmetric, as well as light-tailed and heavy-tailed distributions. Parameter estimation is modified in order to fit the Tweedie distribution to the data. This procedure is developed by using method of moments. In addition, the comparison stage is made to check for the adequacy between the observed mortality and expected mortality. Finally, the importance of including systematic mortality risk in the model is justified by the Pearson's chi-squared test.

  18. Computing Battery Lifetime Distributions

    NARCIS (Netherlands)

    Cloth, L.; Haverkort, Boudewijn R.H.M.; Jongerden, M.R.

    The usage of mobile devices like cell phones, navigation systems, or laptop computers, is limited by the lifetime of the included batteries. This lifetime depends naturally on the rate at which energy is consumed, however, it also depends on the usage pattern of the battery. Continuous drawing of a

  19. Computing Battery Lifetime Distributions

    NARCIS (Netherlands)

    Cloth, Lucia; Jongerden, Marijn R.; Haverkort, Boudewijn R.

    2007-01-01

    The usage of mobile devices like cell phones, navigation systems, or laptop computers, is limited by the lifetime of the included batteries. This lifetime depends naturally on the rate at which energy is consumed, however, it also depends on the usage pattern of the battery. Continuous drawing of a

  20. Lifetime of Mechanical Equipment

    Energy Technology Data Exchange (ETDEWEB)

    Leland, K.

    1999-07-01

    The gas plant at Kaarstoe was built as part of the Statpipe gas transport system and went on stream in 1985. In 1993 another line was routed from the Sleipner field to carry condensate, and the plant was extended accordingly. Today heavy additional supply- and export lines are under construction, and the plant is extended more than ever. The main role of the factory is to separate the raw gas into commercial products and to pump or ship it to the markets. The site covers a large number of well-known mechanical equipment. This presentation deals with piping, mechanical and structural disciplines. The lifetime of mechanical equipment is often difficult to predict as it depends on many factors, and the subject is complex. Mechanical equipment has been kept in-house, which provides detailed knowledge of the stages from a new to a 14 years old plant. The production regularity has always been very high, as required. The standard of the equipment is well kept, support systems are efficient, and human improvisation is extremely valuable.

  1. Phasor approaches simplify the analysis of tryptophan fluorescence data in protein denaturation studies

    NARCIS (Netherlands)

    Bader, A.N.; Visser, N.V.; Amerongen, van H.; Visser, A.J.W.G.

    2014-01-01

    The intrinsic fluorescence of tryptophan is frequently used to investigate the structure of proteins. The analysis of tryptophan fluorescence data is challenging: fluorescence (anisotropy) decays typically have multiple lifetime (correlation time) components and fluorescence spectra are broad and ex

  2. Lifetime of organic photovoltaics

    DEFF Research Database (Denmark)

    Corazza, Michael; Krebs, Frederik C; Gevorgyan, Suren A.

    2015-01-01

    A comprehensive outdoor study of polymer solar cells and modules for duration of one year was conducted. Different sample geometries and encapsulations were employed in order to study the spread in the lifetimes. The study is a complimentary report to previous work that focused on indoor ageing...... tests. Comparison of the indoor and outdoor lifetimes was performed by means of the o-diagram, which constitutes the initial steps towards establishing a method for predicting the lifetime of an organic photovoltaic device under real operational conditions based on a selection of accelerated indoor...

  3. Design of a compact, low-price, lifetime measuring instrument

    Science.gov (United States)

    Draxler, Sonja; Lippitsch, Max E.; Moeller, Reinhard; Tafeit, Erwin

    1994-08-01

    The technical requirements for a small, rugged, and moderately- priced device for measuring fluorescence lifetimes have been investigated. The suitability and performance of various lifetime measuring schemes were compared. Based on these investigations a compact time-domain instrument was developed allowing measurement of fluorescence decays with a time resolution well below 1 ns. A semiconductor laser (frequency-doubled, if necessary) is used as a light source. Detection is done with a miniaturized photomultiplier. In favorable cases measurement of a fluorescent decay curve is accomplished within less than one minute.

  4. Lifetime-based sensing:  influence of the microenvironment.

    Science.gov (United States)

    Draxler, S; Lippitsch, M E

    1996-03-01

    The influence of the microenvironment on the fluorescence behavior of indicator molecules is investigated. A model is developed to describe the fluorescence decay of indicator molecules in a nonuniform medium. Its consequences for fluorescence lifetime-based chemical sensors are discussed and verified in two examples, namely, a pH sensor using a pyrene compound in a hydrogel and a ruthenium complex for oxygen sensing embedded in a polystyrene membrane.

  5. Measuring Lifetime Poverty

    OpenAIRE

    Michael Hoy; Buhong Zheng

    2008-01-01

    This paper presents an axiomatic framework for measuring life time poverty over multiple periods. For an individual, we argue that lifetime poverty is influenced by both the snapshot poverty of each period and the poverty level of the "permanent" lifetime consumption; it is also influenced by how poverty spells are distributed over the life time. Two obvious candidates for aggregation are to aggregate over time and then across individuals, or vice versa. For a society, we consider a path-inde...

  6. Gated Detection Measurements of Phosphorescence Lifetimes

    Directory of Open Access Journals (Sweden)

    Yordan Kostov

    2004-10-01

    Full Text Available A low-cost, gated system for measurements of phosphorescence lifetimes is presented. An extensive description of the system operating principles and metrological characteristics is given. Remarkably, the system operates without optical filtering of the LED excitation source. A description of a practical system is also given and its performance is discussed. Because the device effectively suppresses high-level background fluorescence and scattered light, it is expected to find wide-spread application in bioprocess, environmental and biomedical fields.

  7. The Number of Accumulated Photons and the Quality of Stimulated Emission Depletion Lifetime Images

    Energy Technology Data Exchange (ETDEWEB)

    Syed, Aleem [Ames Laboratory; Lesoine, Michael D [Ames Laboratory; Bhattacharjee, Ujjal [Ames Laboratory; Petrich, Jacob W [Ames Laboratory; Smith, Emily A [Ames Laboratory

    2014-03-03

    Time binning is used to increase the number of photon counts in the peak channel of stimulated emission depletion (STED) fluorescence lifetime decay curves to determine how it affects the resulting lifetime image. The fluorescence lifetime of the fluorophore, Alexa Fluor 594 phalloidin, bound to F-actin is probed in cultured S2 cells at a spatial resolution of ~40 nm. This corresponds to a tenfold smaller probe volume compared to confocal imaging, and a reduced number of photons contributing to the signal. Pixel-by-pixel fluorescence lifetime measurements and error analysis show that an average of 40 ± 30 photon counts in the peak channel with a signal-to-noise ratio of 20 is enough to calculate a reliable fluorescence lifetime from a single exponential fluorescence decay. No heterogeneity in the actin cytoskeleton in different regions of the cultured cells was measured in the 40- to 400-nm spatial regime.

  8. Lifetime Measurement for 6snp Rydberg States of Barium

    Institute of Scientific and Technical Information of China (English)

    SHEN Li; WANG Lei; YANG Hai-Feng; LIU Xiao-Jun; LIU Hong-Ping

    2011-01-01

    @@ We present a simple and efficient method for measuring the atomic lifetimes in order of tens of microseconds and demonstrate it in the lifetime determination of barium Rydberg states.This method extracts the lifetime information from the time-of-flight spectrum directly, which is much more efficient than other methods such as the time-delayed field ionization and the traditional laser induced fluorescence.The lifetimes determined with our method for barium Rydberg 6snp(n=37-59)series are well coincident with the values deduced from the absolute oscillator strengths of barium which were given in the literature [J.Phys.B 14(1981)4489, 29(1996)655]on experiments.%We present a simple and efficient method for measuring the atomic lifetimes in order of tens of microseconds and demonstrate it in the lifetime determination of barium Rydberg states. This method extracts the lifetime information from the time-of-flight spectrum directly, which is much more efficient than other methods such as the time-delayed field ionization and the traditional laser induced fluorescence. The lifetimes determined with our method for barium Rydberg 6snp (n=37-59) series are well coincident with the values deduced from the absolute oscillator strengths of barium which were given in the literature [J. Phys. B 14 (1981) 4489, 29 (1996) 655] onexperiments.

  9. Charm Lifetimes and Mixing

    CERN Document Server

    Cheung, H W K

    2002-01-01

    A review of the latest results on charm lifetimes and D-mixing is presented. The e+e- collider experiments are now able to measure charm lifetimes quite precisely, however comparisons with the latest results from fixed-target experiments show that possible systematic effects could be evident. The new D-mixing results from the B-factories have changed the picture that is emerging. Although the new world averaged value of y_CP is now consistent with zero, there is still a very interesting and favoured scenario if the strong phase difference between the Doubly-Cabibbo-suppressed and the Cabibbo-flavoured D0 -> Kpi decay is large.

  10. Our Allotted Lifetimes

    Science.gov (United States)

    Gould, Stephen Jay

    1977-01-01

    It is suggested that measured by the internal clock of heartbeats or breathing, all mammals live a similar lifespan. This is based on the fact that mammals, regardless of size, breathe about 200 million times in their lifetime at a rate of 1 breath for every 4 heartbeats. (AJ)

  11. Design and evaluation of a device for fast multispectral time-resolved fluorescence spectroscopy and imaging

    Energy Technology Data Exchange (ETDEWEB)

    Yankelevich, Diego R. [Department of Electrical and Computer Engineering, University of California, 3101 Kemper Hall, Davis, California 95616 (United States); Department of Biomedical Engineering, University of California, 451 Health Sciences Drive, Davis, California 95616 (United States); Ma, Dinglong; Liu, Jing; Sun, Yang; Sun, Yinghua; Bec, Julien; Marcu, Laura, E-mail: lmarcu@ucdavis.edu [Department of Biomedical Engineering, University of California, 451 Health Sciences Drive, Davis, California 95616 (United States); Elson, Daniel S. [Hamlyn Centre for Robotic Surgery, Department of Surgery and Cancer, Imperial College London, Exhibition Road, London SW7 2AZ (United Kingdom)

    2014-03-15

    The application of time-resolved fluorescence spectroscopy (TRFS) to in vivo tissue diagnosis requires a method for fast acquisition of fluorescence decay profiles in multiple spectral bands. This study focusses on development of a clinically compatible fiber-optic based multispectral TRFS (ms-TRFS) system together with validation of its accuracy and precision for fluorescence lifetime measurements. It also presents the expansion of this technique into an imaging spectroscopy method. A tandem array of dichroic beamsplitters and filters was used to record TRFS decay profiles at four distinct spectral bands where biological tissue typically presents fluorescence emission maxima, namely, 390, 452, 542, and 629 nm. Each emission channel was temporally separated by using transmission delays through 200 μm diameter multimode optical fibers of 1, 10, 19, and 28 m lengths. A Laguerre-expansion deconvolution algorithm was used to compensate for modal dispersion inherent to large diameter optical fibers and the finite bandwidth of detectors and digitizers. The system was found to be highly efficient and fast requiring a few nano-Joule of laser pulse energy and <1 ms per point measurement, respectively, for the detection of tissue autofluorescent components. Organic and biological chromophores with lifetimes that spanned a 0.8–7 ns range were used for system validation, and the measured lifetimes from the organic fluorophores deviated by less than 10% from values reported in the literature. Multi-spectral lifetime images of organic dye solutions contained in glass capillary tubes were recorded by raster scanning the single fiber probe in a 2D plane to validate the system as an imaging tool. The lifetime measurement variability was measured indicating that the system provides reproducible results with a standard deviation smaller than 50 ps. The ms-TRFS is a compact apparatus that makes possible the fast, accurate, and precise multispectral time-resolved fluorescence

  12. Role of Indocyanine Green in Sentinel Node Mapping in Gynecologic Cancer: Is Fluorescence Imaging the New Standard?

    Science.gov (United States)

    Darin, María Cecilia; Gómez-Hidalgo, Natalia Rodriguez; Westin, Shannon N; Soliman, Pamela T; Escobar, Pedro F; Frumovitz, Michael; Ramirez, Pedro T

    2016-02-01

    Sentinel lymph node biopsy has proven safe and feasible in a number of gynecologic cancers such as vulvar cancer, cervical cancer, and endometrial cancer. The proposed aim of lymphatic mapping and sentinel node identification is to decrease the associated morbidity of a complete lymphadenectomy, particularly the rate of lymphedema, while also increasing the detection of small tumor deposits in the node. Different tracers have been shown to be useful, including technetium-99 and blue dye, with a detection reported in 66% to 86%. Recently, there has been increasing interest in the use of fluorescent dies such as indocyanine green (ICG). In this report we provide a review of the existing literature regarding the use of ICG in cervical or endometrial cancer with the goal to provide details on its utility and compare it with other tracers.

  13. Determination of the Free Neutron Lifetime

    CERN Document Server

    Bowman, J David; Clayton, S M; Dewey, M S; Fomin, N; Grammer, K B; Greene, G L; Huffman, P R; Holley, A T; Jones, G L; Liu, C -Y; Makela, M; Mendenhall, M P; Morris, C L; Mulholland, J; Nollett, K M; Pattie,, R W; Penttila, S; Ramsey-Musolf, M; Salvat, D J; Saunders, A; Seestrom, S J; Snow, W M; Steyerl, A; Wietfeldt, F E; Young, A R; Yue, A T

    2014-01-01

    We present the status of current US experimental efforts to measure the lifetime of the free neutron by the "beam" and "bottle" methods. BBN nucleosynthesis models require accurate measurements with 1 second uncertainties, which are currently feasible. For tests of physics beyond the standard model, future efforts will need to achieve uncertainties well below 1 second. We outline paths achieve both.

  14. A Lifetime Prediction Method for LEDs Considering Real Mission Profiles

    OpenAIRE

    Qu, Xiaohui; Wang, Huai; Zhan, Xiaoqing; Blaabjerg, Frede; Chung, Henry Shu-Hung

    2017-01-01

    The Light-Emitting Diode (LED) has become a very promising alternative lighting source with the advantages of longer lifetime and higher efficiency than traditional ones. The lifetime prediction of LEDs is important to guide the LED system designers to fulfill the design specifications and to benchmark the cost-competitiveness of different lighting technologies. However, the existing lifetime data released by LED manufacturers or standard organizations are usually applicable only for some spe...

  15. Positronium lifetime in polymers

    CERN Document Server

    Camacho, Abel

    2004-01-01

    A model describing the relationship between the ortho--positronium lifetime and the volume of a void, located in a synthetic zeolite, is analyzed. Our idea, which allows us to take into account the effects of temperature, comprises the introduction of a non--hermitian term in the Hamiltonian, which accounts for the annihilation of the ortho--positronium. The predictions of the present model are also confronted against an already known experimental result.

  16. Prompt Neutron Lifetime for the NBSR Reactor

    Energy Technology Data Exchange (ETDEWEB)

    Hanson, A.L.; Diamond, D.

    2012-06-24

    In preparation for the proposed conversion of the National Institute of Standards and Technology (NIST) research reactor (NBSR) from high-enriched uranium (HEU) to low-enriched uranium (LEU) fuel, certain point kinetics parameters must be calculated. We report here values of the prompt neutron lifetime that have been calculated using three independent methods. All three sets of calculations demonstrate that the prompt neutron lifetime is shorter for the LEU fuel when compared to the HEU fuel and longer for the equilibrium end-of-cycle (EOC) condition when compared to the equilibrium startup (SU) condition for both the HEU and LEU fuels.

  17. The neutron lifetime experiment PENeLOPE

    Energy Technology Data Exchange (ETDEWEB)

    Schreyer, Wolfgang [Technische Universitaet Muenchen (Germany); Collaboration: PENeLOPE-Collaboration

    2015-07-01

    The neutron lifetime τ{sub n}=880.3±1.1 s is an important parameter in the Standard Model of particle physics and in Big Bang cosmology. Several systematic corrections of previously published results reduced the PDG world average by several σ in the last years and call for a new experiment with complementary systematics. The experiment PENeLOPE, currently under construction at the Physik-Department of Technische Universitaet Muenchen, aims to determine the neutron lifetime with a precision of 0.1 s. It will trap ultra-cold neutrons in a magneto-gravitational trap using a large superconducting magnet and will measure their lifetime by both neutron counting and online proton detection. This presentation gives an overview over the latest developments of the experiment.

  18. The Sprint to Lifetime Sports

    Science.gov (United States)

    Ernst, Leonard

    1973-01-01

    Describes the trend in high school physical education programs toward lifetime sports, defined by the author as physical activities that will serve the interests of students for a lifetime. Included are a special report on program costs and a model of a performance-based lifetime sports program. (Author/DN)

  19. A lifetime prediction method for LEDs considering mission profiles

    DEFF Research Database (Denmark)

    Qu, Xiaohui; Wang, Huai; Zhan, Xiaoqing

    2016-01-01

    Light-Emitting Diodes (LEDs) has become a very promising alternative lighting source with the advantages of longer lifetime and higher efficiency than traditional ones. The lifetime prediction of LEDs is important to guide the LED system designers to fulfill the design specifications...... and to benchmark the cost-competitiveness of different lighting technologies. The existing lifetime data released by LED manufacturers or standard organizations are usually applicable only for specific temperature and current levels. Significant lifetime discrepancies may be observed in field operations due...... available from accelerated degradation testing. It identifies also the key variables (e.g., heat sink parameters and lifetime-matching of LED drivers) that can be designed to achieve a specified lifetime and reliability level. Two case studies of an indoor residential lighting and an outdoor street lighting...

  20. Measuring Luminescence Lifetime With Help of a DSP

    Science.gov (United States)

    Danielson, J. D. S.

    2009-01-01

    An instrument for measuring the lifetime of luminescence (fluorescence or phosphorescence) includes a digital signal processor (DSP) as the primary means of control, generation of excitation signals, and analysis of response signals. The DSP hardware in the present instrument makes it possible to switch among a variety of operating modes by making changes in software only.

  1. Lifetimes and HQE

    CERN Document Server

    Lenz, Alexander

    2014-01-01

    Kolya Uraltsev was one of the inventors of the Heavy Quark Expansion (HQE), that describes inclusive weak decays of hadrons containing heavy quarks and in particular lifetimes. Besides giving a pedagogic introduction to the subject, we review the development and the current status of the HQE, which just recently passed several non-trivial experimental tests with an unprecedented precision. In view of many new experimental results for lifetimes of heavy hadrons, we also update several theory predictions: $\\tau (B^+) / \\tau (B_d) = 1.04^{+0.05}_{-0.01} \\pm 0.02 \\pm 0.01$, $\\tau (B_s) / \\tau (B_d) = 1.001 \\pm 0.002$, $\\tau (\\Lambda_b)/ \\tau (B_d) = 0.935 \\pm 0.054$ and $\\bar {\\tau} (\\Xi_b^0) / \\bar{\\tau} (\\Xi_b^+) = 0.95 \\pm 0.06$. The theoretical precision is currently strongly limited by the unknown size of the non-perturbative matrix elements of four-quark operators, which could be determined with lattice simulations.

  2. Interlaboratory comparison of positron and positronium lifetimes in polymers

    DEFF Research Database (Denmark)

    Wastlund, C.; Eldrup, Morten Mostgaard; Maurer, F.H.J.

    1998-01-01

    A comparison of the results of a series of positron annihilation lifetime measurements performed in 12 laboratories is presented. The measurements were conducted on three different polymer samples, all prepared in one laboratory under standard conditions. The objective of the work was to gain...... insight into the variation in derived positron and positronium lifetimes and intensities measured in the different laboratories on identical specimens. Lifetime data were collected at room temperature by each laboratory following their own standard measurement and data evaluation procedures. The polymers...

  3. Experimental and theoretical lifetimes and transition probabilities in Sb I

    CERN Document Server

    Hartman, Henrik; Engström, Lars; Lundberg, Hans; Palmeri, Patrick; Quinet, Pascal; Biémont, Emile; 10.1103/PhysRevA.82.052512

    2010-01-01

    We present experimental atomic lifetimes for 12 levels in Sb I, out of which seven are reported for the first time. The levels belong to the 5p$^2$($^3$P)6s $^{2}$P, $^{4}$P and 5p$^2$($^3$P)5d $^{4}$P, $^{4}$F and $^{2}$F terms. The lifetimes were measured using time-resolved laser-induced fluorescence. In addition, we report new calculations of transition probabilities in Sb I using a Multiconfigurational Dirac-Hartree-Fock method. The physical model being tested through comparisons between theoretical and experimental lifetimes for 5d and 6s levels. The lifetimes of the 5d $^4$F$_{3/2, 5/2, 7/2}$ levels (19.5, 7.8 and 54 ns, respectively) depend strongly on the $J$-value. This is explained by different degrees of level mixing for the different levels in the $^4$F term.

  4. Lifetime-weighted photoacoustic imaging

    Science.gov (United States)

    Forbrich, A.; Shao, P.; Shi, W.; Zemp, Roger J.

    2016-12-01

    Photoacoustic (PA) imaging has been utilized to quantify the lifetime profile of exogenous agents using a series of pump-probe pulses with a varying time delay; however, current techniques typically lead to long acquisition times which are sensitive to motion and cause absorption or photobleaching. We introduce a technique called lifetime-weighted imaging, which uses only three laser pulses to preferentially weight signals from chromophores with long lifetimes (including exogenous contrast agents with triplet excited states such as methylene blue and porphyrins) while nulling chromophores with short picosecond- to nanosecond-scale lifetimes (including hemoglobin). This technique detects the PA signal from a probe pulse either with or without a pump pulse. By subtracting the probe-only signal from the pump-present probe signal, we effectively eliminate signals from chromophores with short lifetimes while preserving PA signals from chromophores with long-lifetimes. We demonstrate the oxygen-dependent lifetime of both methylene blue and porphyrin-lipids and demonstrate both ground-state recovery and excited-state lifetime-weighted imaging. Lifetime-weighted PA imaging may have applications in many molecular imaging application including: photodynamic therapy dosimetry guidance and oxygen sensing.

  5. Energy Savings Lifetimes and Persistence

    Energy Technology Data Exchange (ETDEWEB)

    Hoffman, Ian M. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Schiller, Steven R. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Todd, Annika [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Billingsley, Megan A. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Goldman, Charles A. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Schwartz, Lisa C. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

    2016-02-01

    This technical brief explains the concepts of energy savings lifetimes and savings persistence and discusses how program administrators use these factors to calculate savings for efficiency measures, programs and portfolios. Savings lifetime is the length of time that one or more energy efficiency measures or activities save energy, and savings persistence is the change in savings throughout the functional life of a given efficiency measure or activity. Savings lifetimes are essential for assessing the lifecycle benefits and cost effectiveness of efficiency activities and for forecasting loads in resource planning. The brief also provides estimates of savings lifetimes derived from a national collection of costs and savings for electric efficiency programs and portfolios.

  6. Copulas Between Wealth and Lifetime

    Institute of Scientific and Technical Information of China (English)

    YE Dongyan

    2009-01-01

    The life insurance industry is very interested in how a person's lifetime is related to his wealth with financial advisors interested in how even a person's portfolio choice affects his lifetime. This paper presents a statistical analysis combined with intuitive relationships between lifetime and wealth. Key properties of this relationship are given and then various copulas are analyzed to see whether they have these properties. Other advantages and disadvantages of these copulas for describing the dependence are stated. The results show that some copulas are not appropriate for relating lifetime and wealth, including the Gaussian family.

  7. The Atmospheric Lifetime Experiment. II - Calibration

    Science.gov (United States)

    Rasmussen, R. A.; Lovelock, J. E.

    1983-10-01

    The calibration standards used in the Atmospheric Lifetime Experiment (ALE) for CFCl3, CF2Cl2, CH3CCl3, and CCl4 are described. This includes the preparation of the primary standards by static dilution and their propagation and stability for the period 1977-1982. Two independent assessments of the absolute concentrations of the primary standards used to initiate the ALE measurements in 1977-1978 are reported. For consistency in the ALE program the values assigned to the primary standards and subsequent working standards used in the field were not altered during the experiment when results of better estimates of the original concentration values were obtained. Rather, the appropriate factors by which the ALE mixing ratios for a given species should be multiplied to obtain the best estimate of the current concentration of a given species, are provided.

  8. Fluorescence antibunching microscopy

    CERN Document Server

    Schwartz, Osip

    2011-01-01

    Breaking the diffraction limit in microscopy by utilizing quantum properties of light has been the goal of intense research in the recent years. We propose a quantum superresolution technique based on non-classical emission statistics of fluorescent markers, routinely used as contrast labels for bio-imaging. The technique can be readily implemented using standard fluorescence microscopy equipment.

  9. Lifetime Improvement by Battery Scheduling

    NARCIS (Netherlands)

    Jongerden, M.R.; Schmitt, Jens B.; Haverkort, Boudewijn R.H.M.

    2012-01-01

    The use of mobile devices is often limited by the lifetime of their batteries. For devices that have multiple batteries or that have the option to connect an extra battery, battery scheduling, thereby exploiting the recovery properties of the batteries, can help to extend the system lifetime. Due to

  10. Lifetime improvement by battery scheduling

    NARCIS (Netherlands)

    Jongerden, M.R.; Haverkort, Boudewijn R.H.M.

    2011-01-01

    The use of mobile devices is often limited by the lifetime of its battery. For devices that have multiple batteries or that have the option to connect an extra battery, battery scheduling, thereby exploiting the recovery properties of the batteries, can help to extend the system lifetime. Due to the

  11. Lifetime Improvement by Battery Scheduling

    NARCIS (Netherlands)

    Jongerden, M.R.; Schmitt, Jens B.; Haverkort, Boudewijn R.H.M.

    The use of mobile devices is often limited by the lifetime of their batteries. For devices that have multiple batteries or that have the option to connect an extra battery, battery scheduling, thereby exploiting the recovery properties of the batteries, can help to extend the system lifetime. Due to

  12. Lifetime improvement by battery scheduling

    NARCIS (Netherlands)

    Jongerden, M.R.; Haverkort, Boudewijn R.H.M.

    The use of mobile devices is often limited by the lifetime of its battery. For devices that have multiple batteries or that have the option to connect an extra battery, battery scheduling, thereby exploiting the recovery properties of the batteries, can help to extend the system lifetime. Due to the

  13. Radiative Lifetime Measurements of Even-Parity Levels of Singly Ionized Erbium

    Institute of Scientific and Technical Information of China (English)

    XU Huai-Liang(徐淮良); JIANG Hong-Mei(蒋红梅); LIU Qian(刘倩); JIANG Zhan-Kui(蒋占魁); S.Svanberg

    2004-01-01

    Radiative lifetime measurements were performed by time-resolved laser-induced fluorescence (LIF) technique for eight even-parity levels of the astrophysically importantion Er+ over the energy range from 33753 to 55317 cm-1.Free erbium ions were generated by a laser-induced plasma. A narrow bandwidth UV laser pulse (1 ns) was employed to populate selectively the short-lived upper levels, and the lifetime value were evaluated from the time-resolved fluorescence signals. The lifetimes reported fall in the range of 3-35 ns with the experimental accuracy 5-8%.

  14. Lifetimes, branching fractions, and oscillator strengths of doubly ionized tungsten

    Energy Technology Data Exchange (ETDEWEB)

    Schultz-Johanning, M.; Schnabel, R.; Kock, M. [Inst. fuer Atom- and Molekuelphysik, Abt. Plasmaphysik, Univ. Hannover (Germany); Kling, R. [Inst. fuer Atom- and Molekuelphysik, Abt. Plasmaphysik, Univ. Hannover (Germany); National Inst. of Standards and Technology, Gaithersburg, MD (United States); Li, Z.; Lundberg, H. [Dept. of Physics, Lund Inst. of Tech. (Sweden); Johansson, S. [Atomic Spectroscopy, Dept. of Physics, Lund (Sweden)

    2001-05-01

    A first small set of W III oscillator strengths has been obtained from combined lifetime and branching fraction measurements. The branching fractions in the wavelength region of 154-334 nm were measured with a Penning discharge and a Fourier transform spectrometer. Three levels have been calibrated and absolute scales with lifetimes measured with the time-resolved laser-induced fluorescence technique. The f-values derived have uncertainties of about 8% at best. A comparison with Cowan-code calculations is given since no other data are available in the literature. (orig.)

  15. Radiative lifetimes of odd-parity levels in Nb I

    Science.gov (United States)

    Mukund, Sheo; Bhattacharyya, Soumen; Yarlagadda, Suresh; Nakhate, S. G.

    2015-11-01

    Radiative lifetimes are reported for 37 odd-parity energy levels of neutral niobium (Nb I), out of which 33 have been measured for the first time. The levels belong to electronic configurations 4d35s5p and 4d45p between 18,790 and 35,730 cm-1. The time-resolved laser-induced fluorescence spectroscopy technique was employed. The Nb atoms were generated in a free-jet by laser vaporization of niobium metal. Lifetime values reported in this work fall in the range 12-340 ns and are accurate to ±10%.

  16. DBD dyes as fluorescent probes for sensing lipophilic environments.

    Science.gov (United States)

    Wawrzinek, Robert; Wessig, Pablo; Möllnitz, Kristian; Nikolaus, Jörg; Schwarzer, Roland; Müller, Peter; Herrmann, Andreas

    2012-09-01

    Small fluorescent organic molecules based on [1,3]dioxolo[4,5-f][1,3]benzodioxole (DBD) could be used as probes for lipophillic microenvironments in aqueous solutions by indicating the critical micelles concentration of detergents and staining cell organelles. Their fluorescence lifetime decreases drastically by the amount of water in their direct environment. Therefore they are potential probes for fluorescence lifetime imaging microscopy (FLIM).

  17. New Detectors to Explore the Lifetime Frontier

    CERN Document Server

    Chou, John Paul; Lubatti, H J

    2016-01-01

    Long-lived particles (LLPs) are a common feature in many beyond the Standard Model theories, including supersymmetry, and are generically produced in exotic Higgs decays. Unfortunately, no existing or proposed search strategy will be able to probe neutral LLPs with lifetimes near the limit set by Big Bang Nucleosynthesis (BBN), $c \\tau \\lesssim 10^7 - 10^8$ m. We propose the MATHUSLA surface detector concept (MAssive Timing Hodoscope for Ultra Stable neutraL pArticles), which can be implemented cost-effectively and in time for the high luminosity LHC upgrade to find such ultra-long-lived particles (ULLPs), whether produced in exotic Higgs decays or more general production modes. We also advocate for a dedicated LLP detector at a future 100 TeV collider, where a modestly sized underground design can discover ULLPs with lifetimes at the BBN limit produced in sub-percent level exotic Higgs decays

  18. Lifetime measurement of trapped staus using ATLAS

    CERN Document Server

    Sibley, Logan

    I study the creation of long-lived staus at a 14 TeV centre of mass energy in proton-proton collisions at the LHC using both the ATLAS and ACME detectors. The ATLAS overburden or underburden, or even ATLAS itself, may trap the semi-stable staus at that place where they will remain until the time at which they decay, where the stau lifetime ranges between seven days and one year. Using a novel method, one may count the number of muons and pions originating from the stau decay using the standard ATLAS cosmic ray trigger. Using an idealized detector model, I find that this method can lead to measurements of the stau lifetime and SUSY cross-section to within statistical uncertainties of 6% and 1% of their actual values, respectively.

  19. New parameters influencing hydraulic runner lifetime

    Science.gov (United States)

    Sabourin, M.; Thibault, D.; Bouffard, D. A.; Lévesque, M.

    2010-08-01

    Traditionally, hydraulic runner mechanical design is based on calculation of static stresses. Today, validation of hydraulic runner design in terms of reliability requires taking into account the fatigue effect of dynamics loads. A damage tolerant approach based on fracture mechanics is the method chosen by Alstom and Hydro-Québec to study fatigue damage in runners. This requires a careful examination of all factors influencing material fatigue behavior. Such material behavior depends mainly on the chemical composition, microstructure and thermal history of the component, and on the resulting residual stresses. Measurement of fracture mechanics properties of various steels have demonstrated that runner lifetime can be significantly altered by differences in the manufacturing process, although remaining in accordance with agreed practices and standards such as ASTM. Carbon content and heat treatment are suspected to influence fatigue lifetime. This will have to be investigated by continuing the current research.

  20. Predictive Models of Li-ion Battery Lifetime

    Energy Technology Data Exchange (ETDEWEB)

    Smith, Kandler; Wood, Eric; Santhanagopalan, Shriram; Kim, Gi-heon; Shi, Ying; Pesaran, Ahmad

    2015-06-15

    It remains an open question how best to predict real-world battery lifetime based on accelerated calendar and cycle aging data from the laboratory. Multiple degradation mechanisms due to (electro)chemical, thermal, and mechanical coupled phenomena influence Li-ion battery lifetime, each with different dependence on time, cycling and thermal environment. The standardization of life predictive models would benefit the industry by reducing test time and streamlining development of system controls.

  1. Lifetime costs of cerebral palsy

    DEFF Research Database (Denmark)

    Kruse, Marie; Michelsen, Susan Ishøy; Flachs, Esben Meulengracht

    2009-01-01

    This study quantified the lifetime costs of cerebral palsy (CP) in a register-based setting. It was the first study outside the US to assess the lifetime costs of CP. The lifetime costs attributable to CP were divided into three categories: health care costs, productivity costs, and social costs....... The population analyzed was retrieved from the Danish Cerebral Palsy Register, which covers the eastern part of the country and has registered about half of the Danish population of individuals with CP since 1950. For this study we analyzed 2367 individuals with CP, who were born in 1930 to 2000 and were alive...

  2. Fluorescence lifetime imaging of induced pluripotent stem cells

    Science.gov (United States)

    Uchugonova, Aisada; Batista, Ana; König, Karsten

    2014-02-01

    The multiphoton FLIM tomograph MPTflex with its flexible scan head, articulated arm, and the tunable femtosecond laser source was employed to study cell monolayers and 3D cell clusters. FLIM was performed with 250 ps temporal resolution and submicron special resolution using time-correlated single photon counting. The autofluorescence based on NAD(P)H and flavins/flavoproteins has been measured in mouse embryonic fibroblasts, induced pluripotent stem cells (iPS cells) originated from mouse embryonic fibroblasts and non-proliferative mouse embryonic fibroblasts.

  3. Fluorescence amplification by electrochemically deposited silver nanowires with fractal architecture.

    Science.gov (United States)

    Goldys, Ewa M; Drozdowicz-Tomsia, Krystyna; Xie, Fang; Shtoyko, Tanya; Matveeva, Eva; Gryczynski, Ignacy; Gryczynski, Zygmunt

    2007-10-10

    Electrochemically deposited silver structures with nanowires 50-100 nm in diameter show high fluorescence amplification and strongly reduced fluorescence lifetimes. Both quantities depend on the structure thickness. With increasing thickness the fluorescence amplification proportionally increases and the fluorescence lifetime decreases. This thickness dependence is caused by fluorophore interaction with a system of plasmon excitations in coupled nanowires extending over micrometer size regions. Thus the amplification is attributed to a combination of extended structure area and strong plasmonic coupling between nanowires which also help to radiatively scatter the fluorescence emission.

  4. Multiphoton autofluorescence lifetime imaging of induced pluripotent stem cells

    Science.gov (United States)

    Uchugonova, Aisada

    2017-06-01

    The multiphoton fluorescence lifetime imaging tomograph MPTflex with its flexible 360-deg scan head, articulated arm, and tunable femtosecond laser source was employed to study induced pluripotent stem cell (iPS) cultures. Autofluorescence (AF) lifetime imaging was performed with 250-ps temporal resolution and submicron spatial resolution using time-correlated single-photon counting. The two-photon excited AF was based on the metabolic coenzymes NAD(P)H and flavin adenine dinucleotide/flavoproteins. iPS cells generated from mouse embryonic fibroblasts (MEFs) and cocultured with growth-arrested MEFs as feeder cells have been studied. Significant differences on AF lifetime signatures were identified between iPS and feeder cells as well as between their differentiating counterparts.

  5. Interaction of fluorescence dyes with 5-fluorouracil: A photoinduced electron transfer study in bulk and biologically relevant water

    Science.gov (United States)

    Kuchlyan, Jagannath; Banik, Debasis; Kundu, Niloy; Roy, Arpita; Sarkar, Nilmoni

    2014-10-01

    The interactions of widely used chemotherapeutic drug, 5-fluorouracil (5FU) with coumarin dyes have been investigated for the first time using steady-state and time-resolved fluorescence spectroscopic measurements. The fluorescence quenching along with the decrease in lifetimes of excited state of coumarin derivatives with gradual addition of 5FU is explained by photoinduced electron transfer (PET) mechanism. Our studies were performed in bulk water and confined water of AOT (aerosol OT) reverse micelle to investigate the effect of confinement on PET dynamics. The feasibility of PET reaction for coumarin-5FU systems is investigated calculating the standard free energy changes using the Rehm-Weller equation.

  6. Technical support document: Energy efficiency standards for consumer products: Room air conditioners, water heaters, direct heating equipment, mobile home furnaces, kitchen ranges and ovens, pool heaters, fluorescent lamp ballasts and television sets. Volume 1, Methodology

    Energy Technology Data Exchange (ETDEWEB)

    1993-11-01

    The Energy Policy and Conservation Act (P.L. 94-163), as amended, establishes energy conservation standards for 12 of the 13 types of consumer products specifically covered by the Act. The legislation requires the Department of Energy (DOE) to consider new or amended standards for these and other types of products at specified times. DOE is currently considering amending standards for seven types of products: water heaters, direct heating equipment, mobile home furnaces, pool heaters, room air conditioners, kitchen ranges and ovens (including microwave ovens), and fluorescent light ballasts and is considering establishing standards for television sets. This Technical Support Document presents the methodology, data, and results from the analysis of the energy and economic impacts of the proposed standards. This volume presents a general description of the analytic approach, including the structure of the major models.

  7. Final report on reliability and lifetime prediction.

    Energy Technology Data Exchange (ETDEWEB)

    Gillen, Kenneth T; Wise, Jonathan; Jones, Gary D.; Causa, Al G.; Terrill, Edward R.; Borowczak, Marc

    2012-12-01

    This document highlights the important results obtained from the subtask of the Goodyear CRADA devoted to better understanding reliability of tires and to developing better lifetime prediction methods. The overall objective was to establish the chemical and physical basis for the degradation of tires using standard as well as unique models and experimental techniques. Of particular interest was the potential application of our unique modulus profiling apparatus for assessing tire properties and for following tire degradation. During the course of this complex investigation, extensive relevant information was generated, including experimental results, data analyses and development of models and instruments. Detailed descriptions of the findings are included in this report.

  8. Measurement of the B_s^0→K^+K^- lifetime relative to the B_d^0→K^+π^- lifetime

    CERN Document Server

    Cliff, H; Kerzel, U

    2009-01-01

    The study of B decays to charmless charged hadrons offers an opportunity to improve our understanding CP violation and to search for New Physics beyond the Standard Model. We present an analysis to make a measurement of the B_s^0→K^+K^- lifetime relative to B_d^0 lifetime which removes systematic bias introduced to the lifetime by distance of flight based selections.

  9. Dynamic Cluster Head for Lifetime Efficiency in WSN

    Institute of Scientific and Technical Information of China (English)

    Hesham Abusaimeh; Shuang-Hua Yang

    2009-01-01

    Saving energy and increasing network lifetime are significant challenges in wireless sensor networks (WSNs).In this paper,we propose a mechanism to distribute the responsibility of cluster-heads among the wireless sensor nodes in the same cluster based on the ZigBee standard,which is the latest WSN standard.ZigBee supports ad hoc on-demand vector (AODV) and cluster-tree routing protocols in its routing layer. However,none of these protocols considers the energy level of the nodes in the network establishing process or in the data routing process. The cluster-tree routing protocol supports single or multi-cluster networks. However,each single cluster in the multi-cluster network has only one node acting as a cluster head. These cluster-heads are fixed in each cluster during the network lifetime.Consequently,using these cluster-heads will cause them to die quickly,and the entire linked nodes to these cluster-heads will be disconnected from the main network.Therefore,the proposed technique to distribute the role of the cluster head among the wireless sensor nodes in the same cluster is vital to increase the lifetime of the network.Our proposed technique is better in terms of performance than the original structure of these protocols.It has increased the lifetime of the wireless sensor nodes,and increased the lifetime of the WSN by around 50% of the original network lifetime.

  10. Effects of short term changes in the blood glucose level on the autofluorescence lifetime of the human retina in healthy volunteers

    Science.gov (United States)

    Klemm, Matthias; Nagel, Edgar; Schweitzer, Dietrich; Schramm, Stefan; Haueisen, Jens

    2016-03-01

    Purpose: Fluorescence lifetime imaging ophthalmoscopy (FLIO) provides in vivo metabolic mapping of the ocular fundus. Changes in FLIO have been found in e.g. diabetes patients. The influence of short term metabolic changes caused by blood glucose level changes on is unknown. Aim of this work is the detection of short-term changes in fundus autofluorescence lifetime during an oral glucose tolerance test. Methods: FLIO was performed in 10 healthy volunteers (29+/-4 years, fasting for 12h) using a scanning laser ophthalmoscope (30° fundus, 34μm resolution, excitation with 473nm diode laser with 70 ps pulses at 80 MHz repetition rate, detection in two spectral channels 500-560nm (ch1) and 560-720nm (ch2) using the timecorrelated single photon counting method). The blood glucose level (BGL) was measured by an Accu-Chek® Aviva self-monitoring device. Before and after a glucose drink (300ml solution, containing 75g of glucose (Accu-Chek® Dextrose O.G.T.), BGL and FLIO were measured every 15min. The FLIMX software package was applied to compute the average fluorescence lifetime τ on the inner ring of the ETDRS grid using a modified 3-exponential approach. Results: The results are given as mean +/- standard deviation over all volunteers in ch1. Baseline measurement: BGL: 5.3+/-0.4 mmol/l, τ1: 49+/-6ps. A significant reduction (α=5% Wilcoxon rank-sum test) in τ1 is detected after 15min (BGL: 8.4+/-1.1 mmol/l, τ1: 44+/-5ps) and after 90min (BGL: 6.3+/-1.4 mmol/l, τ1: 41+/-5ps). Results of ch2 show smaller reductions in the fluorescence lifetimes over time.

  11. A Lifetime Prediction Method for LEDs Considering Real Mission Profiles

    DEFF Research Database (Denmark)

    Qu, Xiaohui; Wang, Huai; Zhan, Xiaoqing

    2017-01-01

    The Light-Emitting Diode (LED) has become a very promising alternative lighting source with the advantages of longer lifetime and higher efficiency than traditional ones. The lifetime prediction of LEDs is important to guide the LED system designers to fulfill the design specifications...... and to benchmark the cost-competitiveness of different lighting technologies. However, the existing lifetime data released by LED manufacturers or standard organizations are usually applicable only for some specific temperature and current levels. Significant lifetime discrepancies may be seen in the field...... properties of the life data available from accelerated degradation testing. The electrical and thermal characteristics of LEDs are measured by a T3Ster system, used for the electro-thermal modeling. It also identifies key variables (e.g., heat sink parameters) that can be designed to achieve a specified...

  12. 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.

  13. RADIATIVE LIFETIMES OF V I AND V II

    Energy Technology Data Exchange (ETDEWEB)

    Den Hartog, E. A.; Lawler, J. E.; Wood, M. P., E-mail: eadenhar@wisc.edu, E-mail: jelawler@wisc.edu, E-mail: mpwood@wisc.edu [Department of Physics, University of Wisconsin, Madison, WI 53706 (United States)

    2014-11-01

    New radiative lifetimes are reported for 168 levels of V I ranging in energy from 18086 cm{sup –1} to 47702 cm{sup –1}, and for 31 levels of V II ranging in energy from 34593 cm{sup –1} to 47420 cm{sup –1}. These lifetimes are measured using time-resolved laser-induced fluorescence on a slow atomic/ionic beam as part of an ongoing study of the radiative properties of the iron group elements. All but two of the V II lifetimes have been measured before using modern laser-based methods, but a large fraction of the V I lifetimes are reported here for the first time. Comparison to earlier measurements is discussed. These new lifetimes are, for the most part, accurate to ±5%. They will be combined with branching fraction measurements to produce a large set of transition probabilities for V I and V II which are needed by the astrophysics community for stellar abundance determinations.

  14. Fluorescence spectroscopy

    DEFF Research Database (Denmark)

    Bagatolli, Luis

    2016-01-01

    Fluorescence spectroscopy is a powerful experimental tool used by scientists from many disciplines. During the last decades there have been important developments on distinct fluorescence methods, particularly those related to the study of biological phenomena. This chapter discusses...

  15. Polar plot representation of time-resolved fluorescence.

    Science.gov (United States)

    Eichorst, John Paul; Wen Teng, Kai; Clegg, Robert M

    2014-01-01

    Measuring changes in a molecule's fluorescence emission is a common technique to study complex biological systems such as cells and tissues. Although the steady-state fluorescence intensity is frequently used, measuring the average amount of time that a molecule spends in the excited state (the fluorescence lifetime) reveals more detailed information about its local environment. The lifetime is measured in the time domain by detecting directly the decay of fluorescence following excitation by short pulse of light. The lifetime can also be measured in the frequency domain by recording the phase and amplitude of oscillation in the emitted fluorescence of the sample in response to repetitively modulated excitation light. In either the time or frequency domain, the analysis of data to extract lifetimes can be computationally intensive. For example, a variety of iterative fitting algorithms already exist to determine lifetimes from samples that contain multiple fluorescing species. However, recently a method of analysis referred to as the polar plot (or phasor plot) is a graphical tool that projects the time-dependent features of the sample's fluorescence in either the time or frequency domain into the Cartesian plane to characterize the sample's lifetime. The coordinate transformations of the polar plot require only the raw data, and hence, there are no uncertainties from extensive corrections or time-consuming fitting in this analysis. In this chapter, the history and mathematical background of the polar plot will be presented along with examples that highlight how it can be used in both cuvette-based and imaging applications.

  16. Fluorescent nanoparticles for intracellular sensing: A review

    Energy Technology Data Exchange (ETDEWEB)

    Ruedas-Rama, Maria J., E-mail: mjruedas@ugr.esmailto [Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada (Spain); Walters, Jamie D. [Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, UK CB2 1QT (United Kingdom); Orte, Angel [Department of Physical Chemistry, Faculty of Pharmacy, University of Granada, Campus Cartuja, 18071, Granada (Spain); Hall, Elizabeth A.H., E-mail: lisa.hall@biotech.cam.ac.uk [Department of Chemical Engineering and Biotechnology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1QT (United Kingdom)

    2012-11-02

    Highlights: Black-Right-Pointing-Pointer Analytical applications of fluorescent nanoparticles (NPs) in intracellular sensing. Black-Right-Pointing-Pointer Critical review on performance of QDots, metal NPs, silica NPs, and polymer NPs. Black-Right-Pointing-Pointer Highlighted potential of fluorescence lifetime imaging microscopy (FLIM). - Abstract: Fluorescent nanoparticles (NPs), including semiconductor NPs (Quantum Dots), metal NPs, silica NPs, polymer NPs, etc., have been a major focus of research and development during the past decade. The fluorescent nanoparticles show unique chemical and optical properties, such as brighter fluorescence, higher photostability and higher biocompatibility, compared to classical fluorescent organic dyes. Moreover, the nanoparticles can also act as multivalent scaffolds for the realization of supramolecular assemblies, since their high surface to volume ratio allow distinct spatial domains to be functionalized, which can provide a versatile synthetic platform for the implementation of different sensing schemes. Their excellent properties make them one of the most useful tools that chemistry has supplied to biomedical research, enabling the intracellular monitoring of many different species for medical and biological purposes. In this review, we focus on the developments and analytical applications of fluorescent nanoparticles in chemical and biological sensing within the intracellular environment. The review also points out the great potential of fluorescent NPs for fluorescence lifetime imaging microscopy (FLIM). Finally, we also give an overview of the current methods for delivering of fluorescent NPs into cells, where critically examine the benefits and liabilities of each strategy.

  17. Metal-induced fluorescence properties of three-dimensionally ordered macroporous silver inverse opal platforms

    Science.gov (United States)

    Chae, Weon-Sik; Lee, Myung-Jin; Kim, Kisun; Hyun, Jerome K.; Jeon, Seokwoo

    2016-02-01

    This study examined the metal-induced fluorescence properties of three-dimensionally ordered macroporous silver inverse opal (IO) films. Electrochemically synthesized silver IO films with a micrometer cavity exhibited notable fluorescence enhancement at the silver frame, and a decrease in fluorescence lifetime. Numerical calculations supported the observations of a higher fluorescence efficiency at the frame than in the cavity.

  18. Positron lifetime in polycrystalline gadolinium

    Energy Technology Data Exchange (ETDEWEB)

    Gomez, A.M.; Serna, J. (Universidad Complutense de Madrid (Spain). Dept. de Fisica del Estado Solido)

    1984-06-16

    Positron lifetimes are measured on polycrystalline gadolinium between 15 and 25 /sup 0/C taking into account the microstructure of the specimens, especially the grain sizes of untreated or annealed sheets. Results show the existence of a trapping effect of positrons in Gd due to different trapping centers such as point defects, dislocations, grain boundaries, and other defects.

  19. Measurement of Charm Meson Lifetimes

    Energy Technology Data Exchange (ETDEWEB)

    Bonvicini, G.; Cinabro, D.; Greene, R.; Perera, L.P.; Zhou, G.J. [Wayne State University, Detroit, Michigan 48202 (United States); Chan, S.; Eigen, G.; Lipeles, E.; Schmidtler, M.; Shapiro, A.; Sun, W.M.; Urheim, J.; Weinstein, A.J.; Wuerthwein, F. [California Institute of Technology, Pasadena, California 91125 (United States); Jaffe, D.E.; Masek, G.; Paar, H.P.; Potter, E.M.; Prell, S.; Sharma, V. [University of California, San Diego, La Jolla, California 92093 (United States); Asner, D.M.; Eppich, A.; Gronberg, J.; Hill, T.S.; Korte, C.M.; Lange, D.J.; Morrison, R.J.; Nelson, H.N.; Nelson, T.K.; Roberts, D.; Tajima, H. [University of California, Santa Barbara, California 93106 (United States); Behrens, B.H.; Ford, W.T.; Gritsan, A.; Krieg, H.; Roy, J.; Smith, J.G. [University of Colorado, Boulder, Colorado 80309-0390 (United States); Alexander, J.P.; Baker, R.; Bebek, C.; Berger, B.E.; Berkelman, K.; Boisvert, V.; Cassel, D.G.; Crowcroft, D.S.; Dickson, M.; von Dombrowski, S.; Drell, P.S.; Dumas, D.J.; Ecklund, K.M.; Ehrlich, R.; Foland, A.D.; Gaidarev, P.; Gibbons, L.; Gittelman, B.; Gray, S.W.; Hartill, D.L.; Heltsley, B.K.; Henderson, S.; Hopman, P.I.; Katayama, N.; Kreinick, D.L.; Lee, T.; Liu, Y.; Meyer, T.O.; Mistry, N.B.; Ng, C.R.; Nordberg, E.; Ogg, M.; Patterson, J.R.; Peterson, D.; Riley, D.; Soffer, A.; Thayer, J.G.; Thies, P.G.; Valant-Spaight, B.; Warburton, A.; Ward, C. [Cornell University, Ithaca, New York 14853 (United States); Athanas, M.; Avery, P.; Jones, C.D.; Lohner, M.; Prescott, C.; Rubiera, A.I.; Yelton, J.; Zheng, J. [University of Florida, Gainesville, Florida 32611 (United States); Brandenburg, G.; Briere, R.A.; Ershov, A.; Gao, Y.S.; Kim, D.Y.; Wilson, R. [Harvard University, Cambridge, Massachusetts 02138 (United States); Browder, T.E.; Li, Y.; Rodriguez, J.L.; Yamamoto, H. [University of Hawaii at Manoa, Honolulu, Hawaii 96822 (United States); Bergfeld, T.; Eisenstein, B.I.; Ernst, J.; Gladding, G.E.; Gollin, G.D; and others

    1999-06-01

    We report measurements of the D{sup 0} , D{sup +} , and D{sup +}{sub s} meson lifetimes using 3.7 fb{sup {minus}1} of e{sup +}e{sup {minus}} annihilation data collected near the {Upsilon}(4S) resonance with the CLEO detector. The measured lifetimes of the D{sup 0} , D{sup +} , and D{sup +}{sub s} mesons are 408.5{plus_minus}4.1{sup +3.5}{sub {minus}3.4} fs , 1033.6{plus_minus}22.1{sup +9.9}{sub {minus}12.7} fs , and 486.3{plus_minus}15.0{sup +4.9}{sub {minus}5.1} fs . The precisions of these lifetimes are comparable to those of the best previous measurements, and the systematic errors are very different. In a single experiment we find that the ratio of the D{sup +}{sub s} and D{sup 0} lifetimes is 1.19{plus_minus}0.04 . {copyright} {ital 1999} {ital The American Physical Society}

  20. Fluorescence properties of meso-tetrafurylporphyrins

    Indian Academy of Sciences (India)

    Iti Gupta; M Ravikanth

    2005-03-01

    Fluorescence properties of meso-tetrafurylporphyrins with N4, N3S and N2S2 porphyrin cores are studied by both steady-state and time-resolved fluorescence techniques and compared with the corresponding meso-tetraarylporphyrins. The study shows that the replacement of six-membered aryl groups with five-membered furyl groups at meso-positions alter the fluorescence properties considerably, as reflected in the large red shifts and broadening of fluorescence bands with reduction in quantum yields and singlet excited-state lifetimes. However, zinc(II) derivatives of meso-tetrafurylporphyrin and mesotetraarylporphyrin did not show significant differences in their emission properties.

  1. Second-order multivariate models for the processing of standard-addition synchronous fluorescence-pH data. Application to the analysis of salicylic acid and its major metabolite in human urine.

    Science.gov (United States)

    Pagani, Ariana P; Ibañez, Gabriela A

    2014-05-01

    In the present work, we describe the determination of salicylic acid and its major metabolite, salicyluric acid, in spiked human urine samples, using synchronous fluorescence spectra measured in a flow-injection system with a double pH gradient. Because the fluorescent urine background constitutes a potentially interfering signal, it becomes necessary to achieve the second-order advantage. Moreover, due to significant changes in the signal of the analytes in the presence of the urine matrix, mainly for salicyluric acid, standard addition was required in order to obtain appropriate quantifications. Several second-order multivariate calibration models were evaluated for this purpose: PARAFAC and MCR-ALS in two different modes, and PLS/RBL.

  2. Anthracene Fluorescence Quenching by a Tetrakis (Ketocarboxamide Cavitand

    Directory of Open Access Journals (Sweden)

    Tibor Zoltan Janosi

    2014-01-01

    Full Text Available Quenching of both fluorescence lifetime and fluorescence intensity of anthracene was investigated in the presence of a newly derived tetrakis (ketocarboxamide cavitand at various concentrations. Time-correlated single photon counting method was applied for the lifetime measurements. A clear correlation between the fluorescence lifetime of anthracene as a function of cavitand concentration in dimethylformamide solution was observed. The bimolecular collisional quenching constant was derived from the decrease of lifetime. Fluorescence intensity was measured in the emission wavelength region around 400 nm as a result of excitation at 280 nm. Effective quenching was observed in the presence of the cavitand. The obtained Stern-Volmer plot displayed upward curvature. The results did not follow even extended Stern-Volmer behavior, often used to describe deviations from static bimolecular quenching. To explain our results we adopted the Smoluchowski model and obtained a reasonable estimate for the molecular radius of the cavitand in solution.

  3. Exits in order: How crowding affects particle lifetimes

    Science.gov (United States)

    Penington, Catherine J.; Baker, Ruth E.; Simpson, Matthew J.

    2016-06-01

    Diffusive processes are often represented using stochastic random walk frameworks. The amount of time taken for an individual in a random walk to intersect with an absorbing boundary is a fundamental property that is often referred to as the particle lifetime, or the first passage time. The mean lifetime of particles in a random walk model of diffusion is related to the amount of time required for the diffusive process to reach a steady state. Mathematical analysis describing the mean lifetime of particles in a standard model of diffusion without crowding is well known. However, the lifetime of agents in a random walk with crowding has received much less attention. Since many applications of diffusion in biology and biophysics include crowding effects, here we study a discrete model of diffusion that incorporates crowding. Using simulations, we show that crowding has a dramatic effect on agent lifetimes, and we derive an approximate expression for the mean agent lifetime that includes crowding effects. Our expression matches simulation results very well, and highlights the importance of crowding effects that are sometimes overlooked.

  4. Fluorescence properties of porcine odorant binding protein Trp 16 residue

    Energy Technology Data Exchange (ETDEWEB)

    Albani, Jihad Rene, E-mail: Jihad-Rene.Albani@univ-lille1.f [Laboratoire de Biophysique Moleculaire, Universite des Sciences et Technologies de Lille, F-59655 Villeneuve d' Ascq Cedex (France)

    2010-11-15

    Summary: The present work deals with fluorescence studies of adult porcine odorant binding protein at pH=7.5. At this pH, the protein is a dimer, each monomer contains one tryptophan residue. Our results show that tryptophan residue displays significant motions and emits with three fluorescence lifetimes. Decay associated spectra showed that the three lifetime's components emanate from sub-structures surrounded by the same microenvironment.

  5. A New Lifetime Distribution with Bathtube and Unimodal Hazard Function

    Science.gov (United States)

    Barriga, Gladys D. C.; Louzada-Neto, Francisco; Cancho, Vicente G.

    2008-11-01

    In this paper we propose a new lifetime distribution which accommodate bathtub-shaped, unimodal, increasing and decreasing hazard function. Some special particular cases are derived, including the standard Weibull distribution. Maximum likelihood estimation is considered for estimate the tree parameters present in the model. The methodology is illustrated in a real data set on industrial devices on a lite test.

  6. Tomographic lifetime imaging using combined early- and late-arriving photons.

    Science.gov (United States)

    Hou, Steven S; Rice, William L; Bacskai, Brian J; Kumar, Anand T N

    2014-03-01

    We present a novel, hybrid approach for time domain fluorescence tomography that efficiently combines lifetime multiplexing using late-arriving or asymptotic photons, with the high spatial resolution capability of early photon tomography. We also show that a decay amplitude-based asymptotic approach is superior to direct inversion of late-arriving photons for tomographic lifetime imaging within turbid media. The hybrid reconstruction approach is experimentally shown to recover fluorescent inclusions separated as close as 1.4 mm, with improved resolution and reduced cross talk compared to just using early photons or the asymptotic approach alone.

  7. Angular distributions as lifetime probes

    Energy Technology Data Exchange (ETDEWEB)

    Dror, Jeff Asaf; Grossman, Yuval [Department of Physics, LEPP, Cornell University,Ithaca, NY 14853 (United States)

    2014-06-27

    If new TeV scale particles are discovered, it will be important to determine their width. There is, however, a problematic region, where the width is too small to be determined directly, and too large to generate a secondary vertex. For a collection of colored, spin polarized particles, hadronization depolarizes the particles prior to their decay. The amount of depolarization can be used to probe the lifetime in the problematic region. In this paper we apply this method to a realistic scenario of a top-like particle that can be produced at the LHC. We study how depolarization affects the angular distributions of the decay products and derive an equation for the distributions that is sensitive to the lifetime.

  8. Angular Distributions as Lifetime Probes

    CERN Document Server

    Dror, Jeff Asaf

    2013-01-01

    If new TeV scale particles are discovered, it will be important to determine their width. There is, however, a problematic region, where the width is too small to be determined directly, and too large to generate a secondary vertex. For a collection of colored, spin polarized particles, hadronization depolarizes the particles prior to their decay. The amount of depolarization can be used to probe the lifetime in the problematic region. In this paper we apply this method to a realistic scenario of a top-like particle that can be produced at the LHC. We study how depolarization affects the angular distributions of the decay products and derive an equation for the distributions that is sensitive to the lifetime.

  9. Cosmological constraints on the neutron lifetime

    CERN Document Server

    Salvati, Laura; Consiglio, Rossella; Melchiorri, Alessandro

    2015-01-01

    We derive new constraints on the neutron lifetime based on the recent Planck 2015 observations of temperature and polarization anisotropies of the CMB. Under the assumption of standard Big Bang Nucleosynthesis, we show that Planck data constrains the neutron lifetime to $\\tau_n=(907 \\pm 69) \\, [\\text{s}]$ at $68 \\%$ c.l.. Moreover, by including the direct measurements of primordial Helium abundance of Izotov et al. 2014 and Mucciarelli et al. 2014, we show that cosmological data provide the stringent constraint $\\tau_n=(905.7 \\pm 7.8) \\, [\\text{s}]$. This value is in tension with the most recent experimental value of $\\tau_n^{\\text{bottle}}=(879.6 \\pm 0.8) \\, [\\text{s}]$ provided by the "bottle method" based on Ultra Cold Neutrons, but in agreement with the experimental value of $\\tau_n^{\\text{beam}}=(888.0 \\pm 2.1) \\, [\\text{s}]$ based on the "beam method". Future CMB surveys as COrE+, in combination with a weak lensing survey as EUCLID, could constrain the neutron life time up to a $\\sim 6$ s precision.

  10. New detectors to explore the lifetime frontier

    Directory of Open Access Journals (Sweden)

    John Paul Chou

    2017-04-01

    Full Text Available Long-lived particles (LLPs are a common feature in many beyond the Standard Model theories, including supersymmetry, and are generically produced in exotic Higgs decays. Unfortunately, no existing or proposed search strategy will be able to observe the decay of non-hadronic electrically neutral LLPs with masses above ∼ GeV and lifetimes near the limit set by Big Bang Nucleosynthesis (BBN, cτ≲107–108 m. We propose the MATHUSLA surface detector concept (MAssive Timing Hodoscope for Ultra Stable neutraL pArticles, which can be implemented with existing technology and in time for the high luminosity LHC upgrade to find such ultra-long-lived particles (ULLPs, whether produced in exotic Higgs decays or more general production modes. We also advocate a dedicated LLP detector at a future 100 TeV collider, where a modestly sized underground design can discover ULLPs with lifetimes at the BBN limit produced in sub-percent level exotic Higgs decays.

  11. Introduction to fluorescence

    CERN Document Server

    Jameson, David M

    2014-01-01

    "An essential contribution to educating scientists in the principles of fluorescence. It will also be an important addition to the libraries of practitioners applying the principles of molecular fluorescence."-Ken Jacobson, Kenan Distinguished Professor of Cell Biology and Physiology, University of North Carolina at Chapel Hill"An exquisite compendium of fluorescence and its applications in biochemistry enriched by a very exciting historical perspective. This book will become a standard text for graduate students and other scientists."-Drs. Zygmunt (Karol) Gryczynski and Ignacy Gryczynski, University of North Texas Health Science Center"… truly a masterwork, combining clarity, precision, and good humor. The reader, novice or expert, will be pleased with the text and will not stop reading. It is a formidable account of the fluorescence field, which has impacted the life sciences so considerably in the last 60 years."-Jerson L. Silva, M.D., Ph.D., Professor and Director, National Institute of Science and Tech...

  12. Lifetime of MCP-PMTs

    Science.gov (United States)

    Lehmann, A.; Britting, A.; Eyrich, W.; Pfaffinger, M.; Uhlig, F.; Belias, A.; Dzhygadlo, R.; Gerhardt, A.; Götzen, K.; Kalicy, G.; Krebs, M.; Lehmann, D.; Nerling, F.; Patsyuk, M.; Peters, K.; Schepers, G.; Schmitt, L.; Schwarz, C.; Schwiening, J.; Traxler, M.; Zühlsdorf, M.; Düren, M.; Etzelmüller, E.; Föhl, K.; Hayrapetyan, A.; Kröck, B.; Merle, O.; Rieke, J.; Schmidt, M.; Cowie, E.; Keri, T.; Achenbach, P.; Cardinali, M.; Hoek, M.; Lauth, W.; Schlimme, S.; Sfienti, C.; Thiel, M.

    2016-05-01

    The hadron identification in the PANDA experiment at FAIR will be done with DIRC detectors. Because of design and space reasons the sensors of the DIRCs have to be placed inside the strong magnetic field of the solenoid. As the favored photon sensors microchannel-plate photomultipliers (MCP-PMTs) were identified. However, these devices showed serious aging problems until very recently, which manifest themselves by a fast degrading quantum efficiency (QE) of the photo cathode (PC). This is mainly due to feedback ions from the residual gas. In this paper we discuss the recently accomplished huge improvements in the lifetime of MCP-PMTs. With innovative countermeasures applied to the MCP-PMTs in the attempt to reduce the aging effects the manufacturers were able to increase the lifetime of MCP-PMT prototypes by almost two orders of magnitude compared to the former commercially available devices. Our group has studied the aging of MCP-PMTs for more than four years by simultaneously illuminating different types of lifetime-enhanced MCP-PMTs at the same photon rate. Gain, dark count rate, and QE as a function of the wavelength and the PC surface were measured in regular time intervals and studied in dependence of the integrated anode charge. We observe that MCP-PMTs treated with an atomic layer deposition (ALD) technique are by far the best devices available now. A lifetime of up to 10 C/cm2 integrated anode charge was reached with these sensors. This is sufficient for both PANDA DIRCs.

  13. Projected Lifetime Healthcare Costs Associated with HIV Infection

    DEFF Research Database (Denmark)

    Nakagawa, Fumiyo; Miners, Alec; Smith, Colette J

    2015-01-01

    computer simulation model to project the distribution of lifetime outcomes and costs of men-who-have-sex-with-men (MSM) infected with HIV in 2013 aged 30, over 10,000 simulations. We assumed a resource-rich setting with no loss to follow-up, and that standards and costs of healthcare management remain...... had been infected in 2013, then future lifetime costs relating to HIV care is likely to be in excess of £ 1 billion. It is imperative for investment into prevention programmes to be continued or scaled-up in settings with good access to HIV care services. Costs would be reduced considerably with use...

  14. Estimation of multiexponential fluorescence decay parameters using compressive sensing.

    Science.gov (United States)

    Yang, Sejung; Lee, Joohyun; Lee, Youmin; Lee, Minyung; Lee, Byung-Uk

    2015-09-01

    Fluorescence lifetime imaging microscopy (FLIM) is a microscopic imaging technique to present an image of fluorophore lifetimes. It circumvents the problems of typical imaging methods such as intensity attenuation from depth since a lifetime is independent of the excitation intensity or fluorophore concentration. The lifetime is estimated from the time sequence of photon counts observed with signal-dependent noise, which has a Poisson distribution. Conventional methods usually estimate single or biexponential decay parameters. However, a lifetime component has a distribution or width, because the lifetime depends on macromolecular conformation or inhomogeneity. We present a novel algorithm based on a sparse representation which can estimate the distribution of lifetime. We verify the enhanced performance through simulations and experiments.

  15. Minority carrier lifetime in indium phosphide

    Science.gov (United States)

    Jenkins, Phillip; Landis, Geoffrey A.; Weinberg, Irving; Kneisel, Keith

    1991-01-01

    Transient photoluminescence is used to measure the minority carrier lifetime on n-type and p-type InP wafers. The measurements show that unprocessed InP wafers have very high minority carrier lifetimes. Lifetimes of 200 ns and 700 ns were observed for lightly-doped p- and n-type material respectively. Lifetimes over 5 ns were found in heavily doped n-type material.

  16. Quantitative frequency-domain fluorescence spectroscopy in tissues and tissue-like media

    Science.gov (United States)

    Cerussi, Albert Edward

    1999-09-01

    In the never-ending quest for improved medical technology at lower cost, modern near-infrared optical spectroscopy offers the possibility of inexpensive technology for quantitative and non-invasive diagnoses. Hemoglobin is the dominant chromophore in the 700-900 nm spectral region and as such it allows for the optical assessment of hemoglobin concentration and tissue oxygenation by absorption spectroscopy. However, there are many other important physiologically relevant compounds or physiological states that cannot be effectively sensed via optical methods because of poor optical contrast. In such cases, contrast enhancements are required. Fluorescence spectroscopy is an attractive component of optical tissue spectroscopy. Exogenous fluorophores, as well as some endogenous ones, may furnish the desperately needed sensitivity and specificity that is lacking in near-infrared optical tissue spectroscopy. The main focus of this thesis was to investigate the generation and propagation of fluorescence photons inside tissues and tissue-like media (i.e., scattering dominated media). The standard concepts of fluorescence spectroscopy have been incorporated into a diffusion-based picture that is sometimes referred to as photon migration. The novelty of this work lies in the successful quantitative recovery of fluorescence lifetimes, absolute fluorescence quantum yields, fluorophore concentrations, emission spectra, and both scattering and absorption coefficients at the emission wavelength from a tissue-like medium. All of these parameters are sensitive to the fluorophore local environment and hence are indicators of the tissue's physiological state. One application demonstrating the capabilities of frequency-domain lifetime spectroscopy in tissue-like media is a study of the binding of ethidium bromide to bovine leukocytes in fresh milk. Ethidium bromide is a fluorescent dye that is commonly used to label DNA, and hence visualize chromosomes in cells. The lifetime of

  17. Predicting polarizabilities and lifetimes of excitons on conjugated polymer chains

    Science.gov (United States)

    van der Horst, J.-W.; Bobbert, P. A.; de Jong, P. H. L.; Michels, M. A. J.; Siebbeles, L. D. A.; Warman, J. M.; Gelinck, G. H.; Brocks, G.

    2001-02-01

    The properties of excitons on three different conjugated polymers in solution are investigated both experimentally and theoretically. The theoretical description of the excitons is obtained by solving the electron-hole Bethe-Salpeter equation (BSE) for the polymers, starting from a calculation within density-functional theory. The calculated radiative lifetimes and polarizabilities of the excitons are compared with experimental results from time-resolved fluorescence decay and flash-photolysis microwave conductivity measurements. The quantitative agreement demonstrates the predictive power of the theoretical approach.

  18. Personality, IQ, and Lifetime Earnings

    DEFF Research Database (Denmark)

    Gensowski, Miriam

    2014-01-01

    Talented individuals are seen as drivers of long-term growth, but how do they realize their full potential? In this paper, I show that lifetime earnings of high-IQ men and women are substantially influenced by their personality traits, in addition to intelligence and education. Personality traits......, as identified in a factor model, significantly affect earnings, but not for young workers. The effects are furthermore heterogeneous by educational attainment. For women, personality traits do not affect family earnings in the same way as own earnings. Personality and IQ also influence earnings indirectly...

  19. Estimation of luminescence lifetime in frequency domain

    Institute of Scientific and Technical Information of China (English)

    Zhang Fu-Jun; Xu Zheng; Zhao Su-Ling; Lou Zhi-Dong; Yang Sheng-Yi; Xu Xu-Rong

    2006-01-01

    Absorption is the origin of luminescence. But it must be noticed that the lifetime of luminescence might reversely influence the rate of absorption. In this paper, it is reported that the luminescence intensity of copper and manganese changes with the driving frequency at constant voltage. The variation of luminescent intensity depends only on the lifetime of luminescence but not on the type of quenching or other factors. Generally the rate of absorption is dominantly determined by the material property and the lifetime of luminescence centres, the absorption of shorter lifetime centre will be larger than that of the longer lifetime centre at the same excited condition.

  20. Carrier lifetimes in thin-film photovoltaics

    Science.gov (United States)

    Baek, Dohyun

    2015-09-01

    The carrier lifetimes in thin-film solar cells are reviewed and discussed. Shockley-Read-Hall recombination is dominant at low carrier density, Auger recombination is dominant under a high injection condition and high carrier density, and surface recombination is dominant under any conditions. Because the surface photovoltage technique is insensitive to the surface condition, it is useful for bulk lifetime measurements. The photoconductance decay technique measures the effective recombination lifetime. The time-resolved photoluminescence technique is very useful for measuring thin-film semiconductor or solar-cell materials lifetime, because the sample is thin, other techniques are not suitable for measuring the lifetime. Many papers have provided time-resolved photoluminescence (TRPL) lifetimes for copper-indium-gallium-selenide (CIGS) and CdTe thin-film solar cell. The TRPL lifetime strongly depends on open-circuit voltage and conversion efficiency; however, the TRPL life time is insensitive to the short-circuit current.

  1. Standard test method for determination of bromine and chlorine in UF6 and uranyl nitrate by X-Ray fluorescence (XRF) spectroscopy

    CERN Document Server

    American Society for Testing and Materials. Philadelphia

    2001-01-01

    1.1 This method covers the determination of bromine (Br) and chlorine (Cl) in uranium hexafluoride (UF6) and uranyl nitrate solution. The method as written covers the determination of bromine in UF6 over the concentration range of 0.2 to 8 μg/g, uranium basis. The chlorine in UF6 can be determined over the range of 4 to 160 μg/g, uranium basis. Higher concentrations may be covered by appropriate dilutions. The detection limit for Br is 0.2 μg/g uranium basis and for Cl is 4 μg/g uranium basis. 1.2 This standard may involve hazardous materials, operations and equipment. This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use.

  2. Lifetime Measurement of 26O

    Science.gov (United States)

    Redpath, Thomas; MoNA Collaboration

    2017-01-01

    An interesting property of some neutron-unbound systems is true two-neutron emission where the neutrons are emitted simultaneously as opposed to a sequential decay through an intermediate state. Since neutrons are only affected by the angular momentum barrier, the timescale for this process is much shorter than for two proton emission which is dominated by the Coulomb barrier. One such case is 26O where a very low decay energy was measured and the two valence neutrons are expected to occupy d-wave orbitals. Also, the ground state of 25O is located 700 keV higher. In a first experiment, the MoNA collaboration extracted a lifetime of 4 .5-1 . 5 + 1 . 1 (stat) +/- 3(syst) ps with a confidence level of 82%. Recently, an experiment dedicated to measuring the 26O lifetime in order to improve the confidence level of the measurement was performed at NSCL. The experiment utilized a newly developed segmented target which increased the statistics without degrading the resolution. Preliminary results will be presented. NSF PHY-1002511, DOE-NNSA DE-NA0000979.

  3. Corrosion Preventive Compounds Lifetime Testing

    Science.gov (United States)

    Hale, Stephanie M.; Kammerer, Catherine C.; Copp, Tracy L.

    2007-01-01

    Lifetime Testing of Corrosion Preventive Compounds (CPCs) was performed to quantify performance in the various environments to which the Space Shuttle Orbiter is exposed during a flight cycle. Three CPCs are approved for use on the Orbiter: RD Calcium Grease, Dinitrol AV-30, and Braycote 601 EF. These CPCs have been rigorously tested to prove that they mitigate corrosion in typical environments, but little information is available on how they perform in the unique combination of the coastal environment at the launch pad, the vacuum of low-earth orbit, and the extreme heat of reentry. Currently, there is no lifetime or reapplication schedule established for these compounds that is based on this combination of environmental conditions. Aluminum 2024 coupons were coated with the three CPCs and exposed to conditions that simulate the environments to which the Orbiter is exposed. Uncoated Aluminum 2024 coupons were exposed to the environmental conditions as a control. Visual inspection and Electro- Impedance Spectroscopy (EIS) were performed on the samples in order to determine the effectiveness of the CPCs. The samples were processed through five mission life cycles or until the visual inspection revealed the initiation of corrosion and EIS indicated severe degradation of the coating.

  4. Carrier Bulk-Lifetime Measurements

    Directory of Open Access Journals (Sweden)

    M. Solcansky

    2012-01-01

    Full Text Available For the measurement of the minority carrier bulk-lifetime the characterization method MW-PCD is used, where the result of measurement is the effective carrier lifetime, which is very dependent on the surface recombination velocity and therefore on the quality of a silicon surface passivation. This work deals with an examination of a different solution types for the chemical passivation of a silicon surface. Various solutions are tested on silicon wafers for their consequent comparison. The main purpose is to find optimal solution, which suits the requirements of a time stability and start-up velocity of passivation, reproducibility of the measurements and a possibility of a perfect cleaning of a passivating solution remains from a silicon surface, so that the parameters of a measured silicon wafer will not worsen and there will not be any contamination of the other wafers series in the production after a repetitive return of the measured wafer into the production process. The cleaning process itself is also a subject of a development.

  5. A scalable assessment of Plasmodium falciparum transmission in the standard membrane-feeding assay, using transgenic parasites expressing green fluorescent protein-luciferase

    NARCIS (Netherlands)

    Stone, W.J.R.; Churcher, T.S.; Graumans, W.; Gemert, G.J.A. van; Vos, M.W.; Lanke, K.H.W.; Vegte-Bolmer, M.G. van de; Siebelink-Stoter, R.; Dechering, K.J.; Vaughan, A.M.; Camargo, N.; Kappe, S.H.; Sauerwein, R.W.; Bousema, T.

    2014-01-01

    BACKGROUND: The development of drugs and vaccines to reduce malaria transmission is an important part of eradication plans. The transmission-reducing activity (TRA) of these agents is currently determined in the standard membrane-feeding assay (SMFA), based on subjective microscopy-based readouts an

  6. Personality, IQ, and Lifetime Earnings

    DEFF Research Database (Denmark)

    Gensowski, Miriam

    2014-01-01

    , as identified in a factor model, significantly affect earnings, but not for young workers. The effects are furthermore heterogeneous by educational attainment. For women, personality traits do not affect family earnings in the same way as own earnings. Personality and IQ also influence earnings indirectly...... through education, which has sizeable positive rates of return for men in this sample. Women’s returns to education past a bachelor’s degree are lowered through worse marriage prospects, which offset gains to education in terms of own earnings. The causal effect of education is identified through matching......Talented individuals are seen as drivers of long-term growth, but how do they realize their full potential? In this paper, I show that lifetime earnings of high-IQ men and women are substantially influenced by their personality traits, in addition to intelligence and education. Personality traits...

  7. Experimentally Measured Radiative Lifetimes and Oscillator Strengths in Neutral Vanadium

    Science.gov (United States)

    Holmes, C. E.; Pickering, J. C.; Ruffoni, M. P.; Blackwell-Whitehead, R.; Nilsson, H.; Engström, L.; Hartman, H.; Lundberg, H.; Belmonte, M. T.

    2016-06-01

    We report a new study of the V i atom using a combination of time-resolved laser-induced fluorescence and Fourier transform spectroscopy that contains newly measured radiative lifetimes for 25 levels between 24,648 cm-1 and 37,518 cm-1 and oscillator strengths for 208 lines between 3040 and 20000 Å from 39 upper energy levels. Thirteen of these oscillator strengths have not been reported previously. This work was conducted independently of the recent studies of neutral vanadium lifetimes and oscillator strengths carried out by Den Hartog et al. and Lawler et al., and thus serves as a means to verify those measurements. Where our data overlap with their data, we generally find extremely good agreement in both level lifetimes and oscillator strengths. However, we also find evidence that Lawler et al. have systematically underestimated oscillator strengths for lines in the region of 9000 ± 100 Å. We suggest a correction of 0.18 ± 0.03 dex for these values to bring them into agreement with our results and those of Whaling et al. We also report new measurements of hyperfine structure splitting factors for three odd levels of V i lying between 24,700 and 28,400 cm-1.

  8. The comparison of the WHO standard rabies immunoglobulin and the national standard Human rabies immunogiobulin used in the rapid fluorescent focus inhibition test (RFFIT)%狂犬病抗血清WHO标准品和国家标准品用于RFFIT方法的比较

    Institute of Scientific and Technical Information of China (English)

    于鹏程; 申辛欣; 吕新军; 单虎; 唐青

    2010-01-01

    目的 比较狂犬病快速荧光灶抑制试验(RFFIT)方法中狂犬病免疫球蛋白WHO标准品和狂犬患者免疫球蛋白国家标准品为参照所得结果的差异.方法 在同一次RFFIT试验中同时设置WHO标准品参照和国家标准品参照,同时测定12份待测人血清,比较两种标准品所产生荧光灶的百分比;按照中和抗体滴度计算公式计算不同标准品参照所得12份待测血清抗体滴度,比较其差异.结果 结果显示WHO标准品和国家标准品的50%感染量均出现在第5孔和第6孔之间,但国家标准品荧光灶百分比低于WHO标准品;以WHO标准品做参照所得到的同一份血清滴度略高于国家标准品.结论 WHO标准品和国家标准品在RFFIT方法中存在差异,但不影响结果判定.%Objective Compare the difference of the results referred to the WHO standard rabies immunoglobulin and the national standard human rabies immunoglobulin used in the rapid fluorescent focus inhibition test (RFFIT).Methods Setting the WHO standard immunoglobulin and the national standard immunoglobulin in the same system and testing 12 human serum at the same time.Compare the fluorescence percentage of the two different standard immunoglobulin;compare the 12 serum results calculated from the two different standard immunoglobulin used the calculation formula of neutralization antibody titer.Results The Results display that the 50% percent of the two standard immunoglobulin are all between the fifth and the sixth well,but the percentage of the national standard immunoglobulin is lower than the WHO one.The same testserum result calculated from the WHO standard immunoglobulin is little higher than the national one.Conclusion There is difference in the WHO standard immunoglobulin and the national one,but there is no influence in the results.

  9. A new technique for the deposition of standard solutions in total reflection X-ray fluorescence spectrometry (TXRF) using pico-droplets generated by inkjet printers and its applicability for aerosol analysis with SR-TXRF

    Energy Technology Data Exchange (ETDEWEB)

    Fittschen, U.E.A. [University of Hamburg, Department of Chemistry, University of Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg (Germany)]. E-mail: ursula.fittschen@chemie.uni-hamburg.de; Hauschild, S. [University of Hamburg, Department of Chemistry, University of Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg (Germany); Amberger, M.A. [University of Hamburg, Department of Chemistry, University of Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg (Germany); Lammel, G. [Max Planck Institute for Meteorology, Bundesstrasse 53, 20146 Hamburg (Germany); Streli, C. [Atominstitut, Vienna University of Technology, Stadionallee 2, 1020 Vienna (Austria); Foerster, S. [University of Hamburg, Department of Chemistry, University of Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg (Germany); Wobrauschek, P. [Atominstitut, Vienna University of Technology, Stadionallee 2, 1020 Vienna (Austria); Jokubonis, C. [Atominstitut, Vienna University of Technology, Stadionallee 2, 1020 Vienna (Austria); Pepponi, G. [ITC-irst, Via Sommarive 18, 38050 Povo (Trento) (Italy); Falkenberg, G. [Hamburger Synchrotronstrahlungslabor at Deutsches Elektronen-Synchrotron DESY, Notkestr. 85, 22603 Hamburg (Germany); Broekaert, J.A.C. [University of Hamburg, Department of Chemistry, University of Hamburg, Martin-Luther-King-Platz 6, 20146 Hamburg (Germany)

    2006-11-15

    A new technique for the deposition of standard solutions on particulate aerosol samples using pico-droplets for elemental determinations with total reflection X-ray fluorescence spectrometry (TXRF) is described. It enables short analysis times without influencing the sample structure and avoids time consuming scanning of the sample with the exciting beam in SR-TXRF analysis. Droplets of picoliter volume ({approx} 5-130 pL) were generated with commercially available and slightly modified inkjet printers operated with popular image processing software. The size of the dried droplets on surfaces of different polarity namely silicone coated and untreated quartz reflectors, was determined for five different printer types and ten different cartridge types. The results show that droplets generated by inkjet printers are between 50 and 200 {mu}m in diameter (corresponding to volumes of 5 to 130 pL) depending on the cartridge type, which is smaller than the width of the synchrotron beam used in the experiments (< 1 mm at an energy of 17 keV at the beamline L at HASYLAB, Hamburg). The precision of the printing of a certain amount of a single element standard solution was found to be comparable to aliquoting with micropipettes in TXRF, where for 2.5 ng of cobalt relative standard deviations of 12% are found. However, it could be shown that the printing of simple patterns is possible, which is important when structured samples have to be analysed.

  10. Memory nearly on a spring: a mean first passage time approach to memory lifetimes.

    Science.gov (United States)

    Elliott, Terry

    2014-09-01

    We study memory lifetimes in a perceptron-based framework with binary synapses, using the mean first passage time for the perceptron's total input to fall below firing threshold to define memory lifetimes. Working with the simplest memory-related model of synaptic plasticity, we may obtain exact results for memory lifetimes or, working in the continuum limit, good analytical approximations that afford either much qualitative insight or extremely good quantitative agreement. In one particular limit, we find that memory dynamics reduce to the well-understood Ornstein-Uhlenbeck process. We show that asymptotically, the lifetimes of memories grow logarithmically in the number of synapses when the perceptron's firing threshold is zero, reproducing standard results from signal-to-noise ratio analyses. However, this is only an asymptotically valid result, and we show that extending its application outside the range of its validity leads to a massive overestimate of the minimum number of synapses required for successful memory encoding. In the case that the perceptron's firing threshold is positive, we find the remarkable result that memory lifetimes are strictly bounded from above. Asymptotically, the dependence of memory lifetimes on the number of synapses drops out entirely, and this asymptotic result provides a strict upper bound on memory lifetimes away from this asymptotic regime. The classic logarithmic growth of memory lifetimes in the simplest, palimpsest memories is therefore untypical and nongeneric: memory lifetimes are typically strictly bounded from above.

  11. Laser-Stimulated Fluorescence in Paleontology

    OpenAIRE

    Kaye, Thomas G.; Falk, Amanda R.; Michael Pittman; Sereno, Paul C.; Martin, Larry D.; Burnham, David A.; Enpu Gong; Xing Xu; Yinan Wang

    2015-01-01

    Fluorescence using ultraviolet (UV) light has seen increased use as a tool in paleontology over the last decade. Laser-stimulated fluorescence (LSF) is a next generation technique that is emerging as a way to fluoresce paleontological specimens that remain dark under typical UV. A laser's ability to concentrate very high flux rates both at the macroscopic and microscopic levels results in specimens fluorescing in ways a standard UV bulb cannot induce. Presented here are five paleontological c...

  12. Changes in job stability - evidence from lifetime job histories

    OpenAIRE

    Rokkanen, Miikka; Uusitalo, Roope

    2010-01-01

    We use individual-level panel data spanning over 42 years from the pension records to evaluate changes in job stability in Finland between 1963 and 2004. Compared with previous research on job stability we cover much longer period and for some cohorts observe the entire lifetime job histories. These data allow us to study job stability using standard duration models instead of simply examining changes in elapsed tenure. We find that hazard of job loss increased during the recession years in t...

  13. Computing lifetimes for battery-powered devices

    OpenAIRE

    Jongerden, Marijn; Haverkort, Boudewijn

    2010-01-01

    The battery lifetime of mobile devices depends on the usage pattern of the battery, next to the discharge rate and the battery capacity. Therefore, it is important to include the usage pattern in battery lifetime computations. We do this by combining a stochastic workload, modeled as a continuous-time Markov model, with a well-known battery model. For this combined model, we provide new algorithms to efficiently compute the expected lifetime and the distribution and expected value of the deli...

  14. Systems and methods for circuit lifetime evaluation

    Science.gov (United States)

    Heaps, Timothy L. (Inventor); Sheldon, Douglas J. (Inventor); Bowerman, Paul N. (Inventor); Everline, Chester J. (Inventor); Shalom, Eddy (Inventor); Rasmussen, Robert D. (Inventor)

    2013-01-01

    Systems and methods for estimating the lifetime of an electrical system in accordance with embodiments of the invention are disclosed. One embodiment of the invention includes iteratively performing Worst Case Analysis (WCA) on a system design with respect to different system lifetimes using a computer to determine the lifetime at which the worst case performance of the system indicates the system will pass with zero margin or fail within a predetermined margin for error given the environment experienced by the system during its lifetime. In addition, performing WCA on a system with respect to a specific system lifetime includes identifying subcircuits within the system, performing Extreme Value Analysis (EVA) with respect to each subcircuit to determine whether the subcircuit fails EVA for the specific system lifetime, when the subcircuit passes EVA, determining that the subcircuit does not fail WCA for the specified system lifetime, when a subcircuit fails EVA performing at least one additional WCA process that provides a tighter bound on the WCA than EVA to determine whether the subcircuit fails WCA for the specified system lifetime, determining that the system passes WCA with respect to the specific system lifetime when all subcircuits pass WCA, and determining that the system fails WCA when at least one subcircuit fails WCA.

  15. Lifetime Extension Report: Progress on the SAVY-4000 Lifetime Extension Program

    Energy Technology Data Exchange (ETDEWEB)

    Welch, Cynthia F. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Smith, Paul Herrick [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Weis, Eric M. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Blair, Michael W. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Stone, Timothy Amos [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Veirs, Douglas Kirk [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Manufacturing Engineering and Technology; Reeves, Kirk Patrick [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Karns, Tristan [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Oka, Jude M. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Keller, Jennie [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Meincke, Linda Jeanne [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Torres, Joseph Angelo [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Herman, Matthew Joseph [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Weaver, Brian Phillip [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Computer, Computational, and Statistical Sciences. Statistical Sciences; Adams, Jillian Cathleen [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Trautschold, Olivia Carol [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials

    2016-09-20

    The 3-year accelerated aging study of the SAVY-4000 O-ring shows very little evidence of significant degradation to samples subjected to aggressive elevated temperature and radiation conditions. Whole container thermal aging studies followed by helium leakage testing and compression set measurements were used to establish an estimate for a failure criterion for O-ring compression set of ≥65 %. The whole container aging studies further show that the air flow and efficiency functions of the filter do not degrade significantly after thermal aging. However, the degradation of the water resistant function leads to water penetration failure after four months at 210°C, but does not cause failure after 10 months at 120°C (130°C is the maximum operating temperature for the PTFE membrane). The thermal aging data for O-ring compression set do not meet the assumptions of standard time-temperature superposition analysis for accelerated aging studies. Instead, the data suggest that multiple degradation mechanisms are operative, with a reversible mechanism operative at low aging temperatures and an irreversible mechanism dominating at high aging temperatures. To distinguish between these mechanisms, we have measured compression set after allowing the sample to physically relax, thereby minimizing the effect of the reversible mechanism. The resulting data were analyzed using two distinct mathematical methods to obtain a lifetime estimate based on chemical degradation alone. Both methods support a lifetime estimate of greater than 150 years at 80°C. Although the role of the reversible mechanism is not fully understood, it is clear that the contribution to the total compression set is small in comparison to that due to the chemical degradation mechanism. To better understand the chemical degradation mechanism, thermally aged O-ring samples have been characterized by Fourier Transform Infrared (FTIR), Electron Paramagnetic Resonance (EPR), Gel Permeation Chromatography (GPC

  16. Lifetime Extension Report: Progress on the SAVY-4000 Lifetime Extension Program

    Energy Technology Data Exchange (ETDEWEB)

    Welch, Cynthia F. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Smith, Paul Herrick [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Weis, Eric M. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Blair, Michael W. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Stone, Timothy Amos [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Veirs, Douglas Kirk [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Manufacturing Engineering and Technology; Reeves, Kirk Patrick [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Karns, Tristan [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Oka, Jude M. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Nuclear Process Infrastructure; Keller, Jennie [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Meincke, Linda Jeanne [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Torres, Joseph Angelo [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Herman, Matthew Joseph [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Weaver, Brian Phillip [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Computer, Computational, and Statistical Sciences. Statistical Sciences; Adams, Jillian Cathleen [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials; Trautschold, Olivia Carol [Los Alamos National Lab. (LANL), Los Alamos, NM (United States). Materials Science and Technology. Engineered Materials

    2016-09-20

    The 3-year accelerated aging study of the SAVY-4000 O-ring shows very little evidence of significant degradation to samples subjected to aggressive elevated temperature and radiation conditions. Whole container thermal aging studies followed by helium leakage testing and compression set measurements were used to establish an estimate for a failure criterion for O-ring compression set of ≥65 %. The whole container aging studies further show that the air flow and efficiency functions of the filter do not degrade significantly after thermal aging. However, the degradation of the water-resistant function leads to water penetration failure after four months at 210°C, but does not cause failure after 10 months at 120°C (130°C is the maximum operating temperature for the PTFE membrane). The thermal aging data for O-ring compression set do not meet the assumptions of standard time-temperature superposition analysis for accelerated aging studies. Instead, the data suggest that multiple degradation mechanisms are operative, with a reversible mechanism operative at low aging temperatures and an irreversible mechanism dominating at high aging temperatures. To distinguish between these mechanisms, we have measured compression set after allowing the sample to physically relax, thereby minimizing the effect of the reversible mechanism. The resulting data were analyzed using two distinct mathematical methods to obtain a lifetime estimate based on chemical degradation alone. Both methods support a lifetime estimate of greater than 150 years at 80°C. Although the role of the reversible mechanism is not fully understood, it is clear that the contribution to the total compression set is small in comparison to that due to the chemical degradation mechanism. To better understand the chemical degradation mechanism, thermally aged O-ring samples have been characterized by Fourier Transform Infrared (FTIR), Electron Paramagnetic Resonance (EPR), Gel Permeation Chromatography (GPC

  17. Experimental study of multi-photon contamination on the measurement of fluorescent decay time

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    In the measurement of fluorescent lifetime based on time correlation-single photon counting technique by means of TAC, due to the contamination of multi-photons a deviation of fluorescent lifetime measured from the expected value is experimentally studied. A correction function instead of a simple exponential function is used to fit the experiment data. The validation of the correction function is checked using the experimental data of several test samples: YAP, NaI(T1) and LSO. The results show that the correction function well fits the data and the reasonable fluorescent lifetimes are obtained.

  18. Fluorescence kinetics of Trp-Trp dipeptide and its derivatives in water via ultrafast fluorescence spectroscopy.

    Science.gov (United States)

    Jia, Menghui; Yi, Hua; Chang, Mengfang; Cao, Xiaodan; Li, Lei; Zhou, Zhongneng; Pan, Haifeng; Chen, Yan; Zhang, Sanjun; Xu, Jianhua

    2015-08-01

    Ultrafast fluorescence dynamics of Tryptophan-Tryptophan (Trp-Trp/Trp2) dipeptide and its derivatives in water have been investigated using a picosecond resolved time correlated single photon counting (TCSPC) apparatus together with a femtosecond resolved upconversion spectrophotofluorometer. The fluorescence decay profiles at multiple wavelengths were fitted by a global analysis technique. Nanosecond fluorescence kinetics of Trp2, N-tert-butyl carbonyl oxygen-N'-aldehyde group-l-tryptophan-l-tryptophan (NBTrp2), l-tryptophan-l-tryptophan methyl ester (Trp2Me), and N-acetyl-l-tryptophan-l-tryptophan methyl ester (NATrp2Me) exhibit multi-exponential decays with the average lifetimes of 1.99, 3.04, 0.72 and 1.22ns, respectively. Due to the intramolecular interaction between two Trp residues, the "water relaxation" lifetime was observed around 4ps, and it is noticed that Trp2 and its derivatives also exhibit a new decay with a lifetime of ∼100ps, while single-Trp fluorescence decay in dipeptides/proteins shows 20-30ps. The intramolecular interaction lifetime constants of Trp2, NBTrp2, Trp2Me and NATrp2Me were then calculated to be 3.64, 0.93, 11.52 and 2.40ns, respectively. Candidate mechanisms (including heterogeneity, solvent relaxation, quasi static self-quenching or ET/PT quenching) have been discussed.

  19. Autofluorescence lifetime variation in the cuticle of the bedbug Cimex lectularius.

    Science.gov (United States)

    Reinhardt, Klaus; Breunig, Hans Georg; König, Karsten

    2017-01-01

    The decay time of the fluorescence of excited molecules, called fluorescence lifetime, can provide information about the cuticle composition additionally to widely used spectral characteristics. We compared autofluorescence lifetimes of different cuticle regions in the copulatory organ of females of the bedbug, Cimex lectularius. After two-photon excitation at 720 nm, regions recently characterised as being rich in resilin showed a longer bimodal distribution of the mean autofluorescence lifetime τm (tau-m) at 0.4 ns and 1.0-1.5 ns, while resilin-poor sites exhibited a unimodal pattern with a peak around 0.8 ns. The mean lifetime, and particularly its second component, can be useful to distinguish resilin-rich from resilin-poor parts of the cuticle. The few existing literature data suggest that chitin is unlikely responsible for the main autofluorescent component observed in the resilin-poor areas in our study and that melanin requires further scrutiny. Autofluorescence lifetime measurements can help to characterise properties of the arthropod cuticle, especially when coupled with multiphoton excitation to allow for deeper tissue penetration.

  20. Cosmology in Mr. Tompkins' Lifetime

    Science.gov (United States)

    Lindner, Rudi Paul

    2016-01-01

    Mr. Tompkins, the hero of George Gamow's most famous book, was born in the first decade of the twentieth century and lived until its end. A bank clerk by day, Mr. Tompkins had wide-ranging interests, and his curiosity led him to popular scientific presentations, and these in turn brought him a long and happy marriage to Maud, the daughter of a professor of physics. His lifetime offers an appropriate framework for a meditation on the history of cosmology during the century in which cosmology became a scientific enterprise. As it happens, Mr. Tompkins' first exposure to cosmology, in which he observed both the expansion and contraction of an oscillating universe in 1939, happened during the long night of relativity, the generation in which relativity specialists became few and, like the galaxies, far between. This talk will consider the heyday of early relativistic cosmology from 1917 to 1935, the causes and consequences of the "long night" from 1935 until 1963, and the renaissance of cosmology, which, occurring as it did upon the retirement of Mr. Tompkins, afforded him great pleasure in his later years.

  1. The Lifetime of Axion Stars

    CERN Document Server

    Eby, Joshua; Wijewardhana, L C R

    2015-01-01

    We investigate the decay of condensates of scalars in a field theory defined by $V({\\cal A})=m^2 f^2 [1-\\cos({\\cal A}/f)]$, where $m$ and $f$ are the mass and decay constant of the scalar field. An example of such a theory is that of the axion, in which case the condensates are called axion stars. The axion field, $\\cal A$, is self adjoint. As a result the axion number is not an absolutely conserved quantity. Therefore, axion stars are not stable and have finite lifetimes. Bound axions, localized on the volume of the star, have a coordinate uncertainty $\\Delta x \\sim R \\sim 1/(m_a \\Delta)$, where $R$ is the radius of the star and $\\Delta = \\sqrt{1-E_0^2/m_a^2}$. Here $m_a$ and $E_0$ are the mass and the ground state energy of the bound axion. Then the momentum distribution of axions has a width of $\\Delta p \\sim m_a\\Delta$. At strong binding, $\\Delta={\\cal O}(1)$, bound axions can easily transfer a sufficient amount of momentum to create and emit a free axion, leading to fast decay of the star with a transiti...

  2. Fluorescent microspheres

    Science.gov (United States)

    Rembaum, A.

    1978-01-01

    Latex particles with attached antibodies have potential biochemical and environmental applications. Human red blood cells and lymphocytes have been labeled with fluorescent microspheres by either direct or indirect immunological technique. Immunolatex spheres can also be used for detecting and localizing specific cell surface receptors. Hormones and toxins may also be bondable.

  3. Microscopic imaging of intracellular calcium in live cells using lifetime-based ratiometric measurements of Oregon Green BAPTA-1.

    Science.gov (United States)

    Lattarulo, Carli; Thyssen, Diana; Kuchibholta, Kishore V; Hyman, Bradley T; Bacskaiq, Brian J

    2011-01-01

    Calcium is a ubiquitous intracellular messenger that has important functions in normal neuronal function. The pathology of Alzheimer's disease has been shown to alter calcium homeostasis in neurons and astrocytes. Several calcium dye indicators are available to measure intracellular calcium within cells, including Oregon Green BAPTA-1 (OGB-1). Using fluorescence lifetime imaging microscopy, we adapted this single wavelength calcium dye into a ratiometric dye to allow quantitative imaging of cellular calcium. We used this approach for in vitro calibrations, single-cell microscopy, high-throughput imaging in automated plate readers, and in single cells in the intact living brain. While OGB is a commonly used fluorescent dye for imaging calcium qualitatively, there are distinct advantages to using a ratiometric approach, which allows quantitative determinations of calcium that are independent of dye concentration. Taking advantage of the distinct lifetime contrast of the calcium-free and calcium-bound forms of OGB, we used time-domain lifetime measurements to generate calibration curves for OGB lifetime ratios as a function of calcium concentration. In summary, we demonstrate approaches using commercially available tools to measure calcium concentrations in live cells at multiple scales using lifetime contrast. These approaches are broadly applicable to other fluorescent readouts that exhibit lifetime contrast and serve as powerful alternatives to spectral or intensity readouts in multiplexing experiments.

  4. Baselines for Lifetime of Organic Solar Cells

    DEFF Research Database (Denmark)

    Gevorgyan, Suren; Espinosa Martinez, Nieves; Ciammaruchi, Laura

    2016-01-01

    The process of accurately gauging lifetime improvements in organic photovoltaics (OPVs) or other similar emerging technologies, such as perovskites solar cells is still a major challenge. The presented work is part of a larger effort of developing a worldwide database of lifetimes that can help e...

  5. Computing lifetimes for battery-powered devices

    NARCIS (Netherlands)

    Jongerden, M.R.; Haverkort, Boudewijn R.H.M.

    The battery lifetime of mobile devices depends on the usage pattern of the battery, next to the discharge rate and the battery capacity. Therefore, it is important to include the usage pattern in battery lifetime computations. We do this by combining a stochastic workload, modeled as a

  6. Computing lifetimes for battery-powered devices

    NARCIS (Netherlands)

    Jongerden, Marijn; Haverkort, Boudewijn

    2010-01-01

    The battery lifetime of mobile devices depends on the usage pattern of the battery, next to the discharge rate and the battery capacity. Therefore, it is important to include the usage pattern in battery lifetime computations. We do this by combining a stochastic workload, modeled as a continuous-ti

  7. Stochastic Analysis of Orbital Lifetimes of Spacecraft

    Science.gov (United States)

    Sasamoto, Washito; Goodliff, Kandyce; Cornelius, David

    2008-01-01

    A document discusses (1) a Monte-Carlo-based methodology for probabilistic prediction and analysis of orbital lifetimes of spacecraft and (2) Orbital Lifetime Monte Carlo (OLMC)--a Fortran computer program, consisting of a previously developed long-term orbit-propagator integrated with a Monte Carlo engine. OLMC enables modeling of variances of key physical parameters that affect orbital lifetimes through the use of probability distributions. These parameters include altitude, speed, and flight-path angle at insertion into orbit; solar flux; and launch delays. The products of OLMC are predicted lifetimes (durations above specified minimum altitudes) for the number of user-specified cases. Histograms generated from such predictions can be used to determine the probabilities that spacecraft will satisfy lifetime requirements. The document discusses uncertainties that affect modeling of orbital lifetimes. Issues of repeatability, smoothness of distributions, and code run time are considered for the purpose of establishing values of code-specific parameters and number of Monte Carlo runs. Results from test cases are interpreted as demonstrating that solar-flux predictions are primary sources of variations in predicted lifetimes. Therefore, it is concluded, multiple sets of predictions should be utilized to fully characterize the lifetime range of a spacecraft.

  8. Lifetime of Organic Photovoltaics: Status and Predictions

    DEFF Research Database (Denmark)

    Gevorgyan, Suren; Madsen, Morten Vesterager; Roth, Bérenger

    2016-01-01

    The results of a meta-analysis conducted on organic photovoltaics (OPV) lifetime data reported in the literature is presented through the compilation of an extensive OPV lifetime database based on a large number of articles, followed by analysis of the large body of data. We fully reveal the prog...

  9. Explore the Lifetime Frontiers with MATHUSLA Detector

    CERN Document Server

    Zhang, Ce; CERN. Geneva. EP Department

    2017-01-01

    Many extensions of the Standard Model (SM) include particles that are neutral, weakly coupled, and long-lived that can decay to final states containing several hadronic jets. Long-lived particles (LLPs) can be detected as displaced decays from the interaction point, or missing energy if they escape. ATLAS and CMS have performed searches at the LHC and significant exclusion limits have been set in recent years. However, the current searches performed at colliders have limitations. An LLP does not interact with the detector and it is only visible once it decays. Unfortunately, no existing or proposed search strategy will be able to observe the decay of non-hadronic electrically neutral LLPs with masses above ~ GeV and lifetimes near the limit set by Big Bang Nucleosynthesis ($c\\tau \\sim 10^7-10^8 m$). Therefore, ultra-long-lived particles (ULLPs) produced at the LHC will escape the main detector with extremely high probability. MATHUSLA (MAssive Timing Hodoscope for Ultra Stable neutraL pArticles) is a surface ...

  10. Lifetime, Mixing and CPV in ATLAS

    CERN Document Server

    AUTHOR|(INSPIRE)INSPIRE-00340295; The ATLAS collaboration

    2016-01-01

    The latest results measured by the ATLAS experiment on lifetime, mixing and $CP$ violation in the $B^0$ and $B_s^0$ systems are presented. First, the measurement of the $B_s^0 \\to J/\\psi \\phi$ decay parameters using 4.9 fb$^{-1}$ and 14.3$^{-1}$ of integrated luminosity collected by the ATLAS detector at the LHC in $p p$ collisions at $\\sqrt{s} = 7$ TeV and $\\sqrt{s} = 8$ TeV respectively is given. The measured values include the $CP$-violating phase $\\phi_s$ and the width difference of the mass eigenstates $\\Delta\\Gamma_s$. The measurements for the 7 and 8 TeV data samples are combined, giving values of $\\phi_s = -0.098 \\pm 0.084~\\mbox{(stat.)} \\pm 0.040~\\mbox{(syst.)}$ and $\\Delta\\Gamma_s = 0.083 \\pm 0.011~\\mbox{(stat.)} \\pm 0.007~\\mbox{(syst.)}$. The measured values agree with the Standard Model predictions. Second, the measurement of the relative width difference $\\Delta \\Gamma_d / \\Gamma_d$ of the $B^0$--$\\bar B^0$ system at $\\sqrt{s} = 7$ TeV and $\\sqrt{s} = 8$ TeV, corresponding to an integrated lumino...

  11. Field Determination Of Ground Water Contamination Using Laser Fluorescence And Fiber Optics

    Science.gov (United States)

    Chudyk, Wayne; Pohlig, Kenneth; Wolf, Lisa; Fordiani, Rita

    1990-02-01

    Experience at over sixteen sites containing over one hundred wells has shown the feasibility of using fiber optic systems for in situ measurement of aromatic ground water contaminants. Aromatic solvents, as well as the benzene, ethylbenzene, toluene, and xylenes (BTEX) fraction of gasoline, have been detected using a prototype field instrument. Well depths have varied from 5 m to 30 m, and limits of detection at 10 m have been in the ppb range. We are routinely using two separate clear tefzel-coated optical fibers bound in a black teflon tubing for in situ sensing of aromatic organic ground water contaminants via laser-induced fluorescence. One fiber, the excitation fiber, carries the 266 nm, 15 nanosecond, laser pulse down to the sensor. The other fiber, used for detection, carries collected fluorescence plus scattered laser light back up to the surface to the detector. Optical crosstalk has been observed to occur along the entire length of the sensor tubing. This may be due to fiber fluorescence. The fiber crosstalk is eliminated by use of a 320 nm cutoff filter in the detector optics. Black tefzel-coated fibers are also commercially available which could eliminate this potential problem. Evaluation of fluorescence emission versus concentration using serial dilution of standards shows that fluorescence lifetimes are important when evaluating different concentrations as well as in evaluation of mixtures. Minimization of signal-to-noise ratios in the detector electronics involves tuning the gate width used in measuring the fluorescent pulse, in order to include the full fluorescent signal returning from the contaminants. Field tests of the modular prototype instrument have been successful in their demonstration of the feasibility of this new technology. Results at a variety of types of sites are presented, showing the flexibility of the modular approach used in the design and operation of this new instrument.

  12. The lifetime of axion stars

    Science.gov (United States)

    Eby, Joshua; Suranyi, Peter; Wijewardhana, L. C. R.

    2016-05-01

    We investigate the decay of condensates of scalars in a field theory defined by V (𝒜) = m2f2[1 -cos(𝒜/f)], where m and f are the mass and decay constant of the scalar field. An example of such a theory is that of the axion, in which case the condensates are called axion stars. The axion field, 𝒜, is self-adjoint. As a result, the axion number is not an absolutely conserved quantity. Therefore, axion stars are not stable and have finite lifetimes. Bound axions, localized on the volume of the star, have a coordinate uncertainty δx ˜ R ˜ 1/(maΔ), where R is the radius of the star and Δ = 1 - E0 2/ma 2. Here ma and E0 are the mass, and the ground state energy of the bound axion. Then the momentum distribution of axions has a width of δp ˜ maΔ. At strong binding, Δ = 𝒪(1), bound axions can easily transfer a sufficient amount of momentum to create and emit a free axion, leading to fast decay of the star with a transition rate Γ ˜ ma. However, when Δ ≪ 1, the momentum distribution is more restricted, and as shown in this paper, the transition rate for creating a free axion decreases as exp(-pδx) ˜exp(-Δ-1). Then sufficiently large, weakly bound axion stars, produced after the Big Bang, survive until the present time. We plot the region of their stability, limited by decay through axion loss and by gravitational instability, as a function of the mass of the axion and the mass of the star.

  13. Feasibility analysis of an epidermal glucose sensor based on time-resolved fluorescence

    Science.gov (United States)

    Katika, Kamal M.; Pilon, Laurent

    2007-06-01

    The goal of this study is to test the feasibility of using an embedded time-resolved fluorescence sensor for monitoring glucose concentration. Skin is modeled as a multilayer medium with each layer having its own optical properties and fluorophore absorption coefficients, lifetimes, and quantum yields obtained from the literature. It is assumed that the two main fluorophores contributing to the fluorescence at these excitation and emission wavelengths are nicotinamide adenine dinucleotide (NAD)H and collagen. The intensity distributions of excitation and fluorescent light in skin are determined by solving the transient radiative transfer equation by using the modified method of characteristics. The fluorophore lifetimes are then recovered from the simulated fluorescence decays and compared with the actual lifetimes used in the simulations. Furthermore, the effect of adding Poissonian noise to the simulated decays on recovering the lifetimes was studied. For all cases, it was found that the fluorescence lifetime of NADH could not be recovered because of its negligible contribution to the overall fluorescence signal. The other lifetimes could be recovered to within 1.3% of input values. Finally, the glucose concentrations within the skin were recovered to within 13.5% of their actual values, indicating a possibility of measuring glucose concentrations by using a time-resolved fluorescence sensor.

  14. Fluorescence detection of esophageal neoplasia

    Science.gov (United States)

    Borisova, E.; Vladimirov, B.; Avramov, L.

    2008-06-01

    White-light endoscopy is well-established and wide used modality. However, despite the many technological advances that have been occurred, conventional endoscopy is suboptimal and usually detects advanced stage lesions. The limitations of standard endoscopy initiate development of spectroscopic techniques, additional to standard endoscopic equipment. One of the most sensitive approaches is fluorescence spectroscopy of gastrointestinal mucosa for neoplasia detection. In the recent study delta-aminolevulinic acid/Protoporphyrin IX (5-ALA/PpIX) is used as fluorescent marker for dysplasia and tumor detection in esophagus. The 5-ALA is administered per os six hours before measurements at dose 20 mg/kg weight. Excitation source has max of emission at 405 nm and light is delivered by the standard light guide of the endoscopic equipment. Through endoscopic instrumental channel a fiber is applied to return information about fluorescence to microspectrometer. Spectral features observed during endoscopic investigations could be distinct as the next regions: 450-630 nm region, where tissue autofluorescence is observed; 630-710 nm region, where fluorescence of PpIX is clearly pronounced; 530-580 nm region, where minima in the autofluorescence signal are observed, related to reabsorption of blood. The lack of fluorescence peaks in the red spectral area for normal mucosa is an indication for selective accumulation of 5-ALA/PpIX only in abnormal sites Very good correlation between fluorescence signals and histology examination of the lesions investigated is achieved.

  15. DEFORMATION INFLUENCE ON A LIFETIME OF WELDING ELECTRODE TIPS

    Directory of Open Access Journals (Sweden)

    Ján Viňáš

    2009-02-01

    Full Text Available The contribution deals with the influence of welding electrode tips deformation on their lifetime. The influence of material properties, production technology and the intensity of welding electrodes load on their lifetime are presented. The electrode tips of the most used type of CuCr1Zr alloy of three basic standard shapes before and after the process of welding are evaluated. The process of welding is realized with low, middle and maximum welding parameters on programmable pneumatic spot welding machine VTS BPK 20. The influence of welding parameters on chosen material characteristics of welding tips is observed. Through the use of upsetting test, dependency of forming strength and deformation of material on used technology of welding tip production is observed.

  16. Nuclear Power Plant Lifetime Management Study (I)

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Sung Yull; Jeong, Ill Seok; Jang, Chang Heui; Song, Taek Ho; Song, Woo Young [Korea Electric Power Research Institute, Taejon (Korea, Republic of); Jin, Tae Eun [Korea Power Engineering Company Consulting and Architecture Engineers, (Korea, Republic of); Kim, Woo Chul [Korea Atomic Energy Research Institute, Taejon (Korea, Republic of)

    1996-12-31

    As the operation-year of nuclear power plant increases and finding sites for new nuclear power plant becomes harder, a comprehensive and systematic nuclear plant lifetime management(PLIM) program including life extension has to be established for stable and safe supply of electricity. A feasibility study was conducted to systematically evaluate technical, economic and regulatory aspect of plant lifetime managements and plant life extension for Kori-1 nuclear power plant. For technical evaluation of nuclear power plant, 13 major components were selected for lifetime evaluation by screening system. structure, and components(SSCs) of the plant. It was found that except reactor pressure vessel, which needs detailed integrity analysis, and low pressure turbine, which is scheduled to be replaced, 11 out of 13 major components have sufficient service life, for more than 40 years. Because domestic rules and regulations related to license renewal has not yet been written, review on the regulatory aspect of life extensions was conducted using US NRC rules and regulations. A cooperative effort with nuclear regulatory body is needed for early completion of license renewal rules and regulations. For economic evaluation of plant lifetime extension, a computer program was developed and used. It was found that 10 to 20 year of extension operation of Kori-1 nuclear power plant was proved. Based on the results, next phase of plant lifetime management program for detailed lifetime evaluation and presenting detailed implementation schedule for plant refurbishment for lifetime extension should be followed. (author). 74 refs., figs.

  17. Steady state and time-resolved fluorescence spectroscopy of quinine sulfate dication bound to sodium dodecylsulfate micelles: Fluorescent complex formation

    Energy Technology Data Exchange (ETDEWEB)

    Joshi, Sunita; Pant, Debi D., E-mail: ddpant@pilani.bits-pilani.ac.in

    2014-01-15

    Interaction of quinine sulfate dication (QSD) with anionic, sodium dodecylsulphate (SDS) surfactant has been studied at different premicellar, micellar and postmicellar concentrations in aqueous phase using steady state, time-resolved fluorescence and fluorescence anisotropy techniques. At premicellar concentrations of SDS, the decrease in absorbance, appearance of an extra fluorescence band at lower wavelengths and tri-exponential decay behavior of fluorescence, are attributed to complex formation between QSD molecules and surfactant monomers. At postmicellar concentrations the red shift in fluorescence spectrum, increase in quantum yield and increase in fluorescence lifetimes are attributed to incorporation of solute molecules to micelles. At lower concentrations of SDS, a large shift in fluorescence is observed on excitation at the red edge of absorption spectrum and this is explained in terms of distribution of ion pairs of different energies in the ground state and the observed fluorescence lifetime behavior corroborates with this model. The temporal fluorescence anisotropy decay of QSD in SDS micelles allowed determination of restriction on the motion of the fluorophore. All the different techniques used in this study reveal that the photophysics of QSD is very sensitive to the microenvironments of SDS micelles and QSD molecules reside at the water-micelle interface. -- Highlights: • Probe molecule is very sensitive to microenvironment of micelles. • Highly fluorescent ion-pair formation has been observed. • Modulated photophysics of probe molecule in micellar solutions has been observed. • Probe molecules strongly bind with micelles and reside at probe–micelle interface.

  18. Fluorescence chemosensors with pyrene and their interaction with nucleotide phosphate

    Institute of Scientific and Technical Information of China (English)

    李华平; 汪鹏飞; 吴世康

    1999-01-01

    A group of fluorescence chemosensor with pyrene, compounds (Ⅰ), (Ⅱ) and (Ⅲ), were synthesized The fluorescence spectra and the lifetime of these compounds were carefully measured. The fluorescence quenching spec tra of pyrenyl butyric acid, compounds (Ⅰ), (Ⅱ) and (Ⅲ) by different nucleotide phosphates, AMP ADP, ATP dTTP, were also recorded and studied. The quenching and the stability constants were calculated by Stern-Volmer equa tion and eq. (2), respectively. The mechanism of interaction between fluorescence chemosensor and nucleotide phos phate was didscussed based on the comparison of the results obtained with the CPK model of free molecules of these com pounds in the ground state.

  19. Fluorescent eco-particles for surface flow physics analysis

    Science.gov (United States)

    Tauro, F.; Porfiri, M.; Grimaldi, S.

    2013-03-01

    In this letter, we describe a novel methodology for fabricating inexpensive environmentally-friendly fluorescent microparticles for quantitative surface flow visualization. Particles are synthesized from natural white beeswax and a highly diluted solution of a nontoxic fluorescent red dye. Bead fluorescence exhibits a long lifetime in adverse conditions, such as exposure to weathering agents, and is enhanced by Ultra Violet radiation. The fluorescent eco-particles are integrated in a particle image velocimetry study of circular hydraulic jump to demonstrate their feasibility in tracing complex surface flows.

  20. Fluorescent eco-particles for surface flow physics analysis

    Directory of Open Access Journals (Sweden)

    F. Tauro

    2013-03-01

    Full Text Available In this letter, we describe a novel methodology for fabricating inexpensive environmentally-friendly fluorescent microparticles for quantitative surface flow visualization. Particles are synthesized from natural white beeswax and a highly diluted solution of a nontoxic fluorescent red dye. Bead fluorescence exhibits a long lifetime in adverse conditions, such as exposure to weathering agents, and is enhanced by Ultra Violet radiation. The fluorescent eco-particles are integrated in a particle image velocimetry study of circular hydraulic jump to demonstrate their feasibility in tracing complex surface flows.

  1. Models for Battery Reliability and Lifetime

    Energy Technology Data Exchange (ETDEWEB)

    Smith, K.; Wood, E.; Santhanagopalan, S.; Kim, G. H.; Neubauer, J.; Pesaran, A.

    2014-03-01

    Models describing battery degradation physics are needed to more accurately understand how battery usage and next-generation battery designs can be optimized for performance and lifetime. Such lifetime models may also reduce the cost of battery aging experiments and shorten the time required to validate battery lifetime. Models for chemical degradation and mechanical stress are reviewed. Experimental analysis of aging data from a commercial iron-phosphate lithium-ion (Li-ion) cell elucidates the relative importance of several mechanical stress-induced degradation mechanisms.

  2. Lifetimes and configuration mixing in 110Cd

    Science.gov (United States)

    Lobach, Yu. N.; Efimov, A. D.; Pasternak, A. A.

    Lifetimes of excited states in 110Cd have been measured by the Doppler shift attenuation method in the reaction (α,2nγ) at Eα= 25 MeV. Lifetime values for 8 states and lifetime limits for 3 states were obtained. The band structures of 110Cd have been interpreted in terms of a modified version of the interacting boson model (IBM + 2 q.p.). The calculations explain well the excitation energies and electromagnetic transition probabilities up to Jπ= 16+, except for the 10+1 state. The structural features are discussed in terms of collective and two quasiparticle excitations.

  3. Statistical Models and Methods for Lifetime Data

    CERN Document Server

    Lawless, Jerald F

    2011-01-01

    Praise for the First Edition"An indispensable addition to any serious collection on lifetime data analysis and . . . a valuable contribution to the statistical literature. Highly recommended . . ."-Choice"This is an important book, which will appeal to statisticians working on survival analysis problems."-Biometrics"A thorough, unified treatment of statistical models and methods used in the analysis of lifetime data . . . this is a highly competent and agreeable statistical textbook."-Statistics in MedicineThe statistical analysis of lifetime or response time data is a key tool in engineering,

  4. Fluorescence properties of crown ethers with phenylbenzothiozole pendant group

    Science.gov (United States)

    Kapoor, S.; Sapre, A. V.; Kumar, S.; Mashraqui, S. H.; Mukherjee, T.

    2005-06-01

    Photophysical characteristics of 2-phenylbenzothiazole (PBT) substituted crown ether (CRE-PBT) molecules have been investigated in various polar protic solvents. Large Stokes' and good solvatochromic shifts have been observed for CRE-PBT molecules. It is seen that in these molecules, the fluorescence lifetimes and quantum yields increase as compared to the pristine PBT molecule. Temperature-dependence studies have been carried out to understand the effect of CRE substitution and the role of C-C single bond rotation on the fluorescence quantum yield and lifetime.

  5. Fluorescence calibration method for single-particle aerosol fluorescence instruments

    Science.gov (United States)

    Shipley Robinson, Ellis; Gao, Ru-Shan; Schwarz, Joshua P.; Fahey, David W.; Perring, Anne E.

    2017-05-01

    Real-time, single-particle fluorescence instruments used to detect atmospheric bioaerosol particles are increasingly common, yet no standard fluorescence calibration method exists for this technique. This gap limits the utility of these instruments as quantitative tools and complicates comparisons between different measurement campaigns. To address this need, we have developed a method to produce size-selected particles with a known mass of fluorophore, which we use to calibrate the fluorescence detection of a Wideband Integrated Bioaerosol Sensor (WIBS-4A). We use mixed tryptophan-ammonium sulfate particles to calibrate one detector (FL1; excitation = 280 nm, emission = 310-400 nm) and pure quinine particles to calibrate the other (FL2; excitation = 280 nm, emission = 420-650 nm). The relationship between fluorescence and mass for the mixed tryptophan-ammonium sulfate particles is linear, while that for the pure quinine particles is nonlinear, likely indicating that not all of the quinine mass contributes to the observed fluorescence. Nonetheless, both materials produce a repeatable response between observed fluorescence and particle mass. This procedure allows users to set the detector gains to achieve a known absolute response, calculate the limits of detection for a given instrument, improve the repeatability of the instrumental setup, and facilitate intercomparisons between different instruments. We recommend calibration of single-particle fluorescence instruments using these methods.

  6. Metabolic Mapping of Breast Cancer with Multiphoton Spectral and Lifetime Imaging

    Science.gov (United States)

    2008-03-01

    intermediates, such as reduced Nicotinamide Adenine Dinucleotide (NADH). NADH plays a key role as a carrier of electrons and is involved in many important...approximately 1.0ns) while free NADH has been shown to have a short fluorescent lifetime of 0.4ns due to quenching of the fluorescent nicotinamide group by...Z. 341(357-377 (1965) 10. D. J. Pappajohn, R. Penneys and B. Chance, "NADH spectrofluorometry of rat skin ," J Appl Physiol 33(5), 684-687 (1972

  7. Nanostructure induced changes in lifetime and enhanced second-harmonic response of organic-plasmonic hybrids

    Energy Technology Data Exchange (ETDEWEB)

    Leißner, Till [NanoSYD, Mads Clausen Institute, University of Southern Denmark, Alsion 2, 6400 Sønderborg (Denmark); Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense (Denmark); Kostiučenko, Oksana; Rubahn, Horst-Günter; Fiutowski, Jacek, E-mail: fiutowski@mci.sdu.dk [NanoSYD, Mads Clausen Institute, University of Southern Denmark, Alsion 2, 6400 Sønderborg (Denmark); Brewer, Jonathan R. [Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, 5230 Odense (Denmark)

    2015-12-21

    In this letter we show that the optical response of organic nanofibers, grown from functionalized para-quaterphenylene molecules, can be controlled by forming organic-plasmonic hybrid systems. The interaction between nanofibers and supporting regular arrays of nanostructures leads to a strongly enhanced second harmonic response. At the same time, the fluorescence lifetime of the nanofibers is reduced from 0.32 ns for unstructured gold films to 0.22 ns for gold nanosquare arrays, demonstrating efficient organic–plasmonic interaction. To study the origin of these effects, we applied two-photon laser scanning microscopy and fluorescence lifetime imaging microscopy. These findings provide an effective approach for plasmon-enhanced second-harmonic generation at the nanoscale, which is attractive for nanophotonic circuitry.

  8. The total lifetime costs of smoking

    DEFF Research Database (Denmark)

    Rasmussen, Gitte Susanne; Prescott, Eva; Sørensen, Thorkild I A;

    2004-01-01

    Net costs of smoking in a lifetime perspective and, hence, the economic interests in antismoking policies have been questioned. It has been proposed that the health-related costs of smoking are balanced by smaller expenditure due to shorter life expectancy.......Net costs of smoking in a lifetime perspective and, hence, the economic interests in antismoking policies have been questioned. It has been proposed that the health-related costs of smoking are balanced by smaller expenditure due to shorter life expectancy....

  9. Lifetime measurement in {sup 195}Po

    Energy Technology Data Exchange (ETDEWEB)

    Grahn, T.; Page, R.D. [University of Liverpool, Department of Physics, Oliver Lodge Laboratory, Liverpool (United Kingdom); Dewald, A.; Jolie, J.; Melon, B.; Pissulla, T. [Universitaet zu Koeln, Institut fuer Kernphysik, Koeln (Germany); Greenlees, P.T.; Jakobsson, U.; Jones, P.; Julin, R.; Juutinen, S.; Ketelhut, S.; Leino, M.; Nyman, M.; Peura, P.; Rahkila, P.; Saren, J.; Scholey, C.; Sorri, J.; Uusitalo, J. [University of Jyvaeskylae, Department of Physics, P.O. Box 35, Jyvaeskylae (Finland); Kroell, T.; Kruecken, R.; Maierbeck, P. [TU Muenchen, Physik-Department E12, Garching (Germany)

    2009-03-15

    The lifetime of the 17/2{sup +} yrast state in {sup 195}Po has been measured using the recoil distance Doppler-shift technique to be {tau}=43(11) ps. The lifetime was extracted from the singles {gamma}-ray spectra obtained by using the recoil-decay tagging method. The present work provides more information of the coupling schemes, shapes and configuration mixing in neutron-deficient odd-mass Po nuclei. (orig.)

  10. Increased fluorescence intensity in CaTiO3:Pr3+ phosphor due to NH3 treatment and Nb Co-doping

    Science.gov (United States)

    Holliday, K. S.; Kohlgruber, T. A.; Tran, I. C.; Åberg, D.; Seeley, Z. M.; Bagge-Hansen, M.; Srivastava, A. M.; Cherepy, N. J.; Payne, S. A.

    2016-10-01

    Development of next generation red phosphors for commercial lighting requires understanding of how increased luminescence is achieved by various treatment strategies. In this work, we compare co-doping with Nb to NH3 treatment of CaTiO3:Pr phosphors to reveal a general mechanism responsible for the increased luminescence. The phosphors were synthesized using standard solid-state synthesis techniques and the fluorescence was characterized for potential use in fluorescent lighting, with 254 nm excitation. The lifetime of the fluorescence was determined and used to identify a change in a trap state by the co-doping of Nb5+ in the phosphor. The oxidation state of the Pr was probed by NEXAFS and revealed that both Nb5+ co-doping and NH3 treatment reduced the number of non-fluorescing Pr4+ centers. Calculations were performed to determine the energetically favorable defects. Vacuum annealing was also used to further probe the nature of the trap state. It was determined that NH3 treatments reduce the number of Pr4+ non-fluorescing centers, while Nb5+ co-doping additionally reduces the number of excess oxygen trap states that quench the fluorescence.

  11. Fluorescence of fullerene derivatives at room temperature

    Energy Technology Data Exchange (ETDEWEB)

    Lin, S.K.; Shiu, L.L.; Chien, K.M.; Luh, T.Y.; Lin, T.I. (National Taiwan Univ., Taipei (Taiwan, Province of China))

    1995-01-05

    The absorption and fluorescence spectral properties of fullerene (C[sub 60]) and its derivatives C[sub 60]C[sub 4]H[sub 6], C[sub 60]C[sub 5]H[sub 6], C[sub 60]CHCO[sub 2]Et, and C[sub 60]NCO[sub 2]Et at room temperature were investigated. Breaking the structural symmetry of C[sub 60] results in enhancing the fluorescence quantum yield 2-3-fold in some derivatives. Thus, the room temperature fluorescence of fullerene compounds could be detected more rapidly. New absorption bands and altered fluorescence spectra were observed in the derivatives. The Stokes' shifts of the derivatives are small, about 4-5 nm, compared to 68 nm for the parent compound. The time-resolved fluorescence decay study indicates that all four fullerene derivatives have a single fluorescence lifetime of ca. 1.2-1.4 as, which is about the same as that for C[sub 60] (ca. 1.3 ns). Aliphatic solvents have little influence on the absorption or fluorescence spectral profile except on the extinction coefficient whereas aromatic and polar solvents strongly interact with the fullerene derivatives, causing a peak broadening effect. 31 refs., 7 figs., 3 tabs.

  12. Lifetime of heavy hypernuclei and its implications on the weak LAMBDA N interaction

    CERN Document Server

    Cassing, W; Kamys, B; Kulessa, P; Niewodniczanski, H; Ohm, H; Pysz, K; Rudy, Z; Schult, O W B; Ströher, H

    2003-01-01

    The lifetime of the LAMBDA-hyperon in heavy hypernuclei measured in proton-Au, -Bi and -U collisions by the COSY-13 Collaboration at COSY-Juelich has been analyzed to yield tau subLAMBDA=(145+-11) ps. This value for tau subLAMBDA is compatible with the lifetime extracted from antiproton annihilation on Bi and U targets, albeit much more accurate. Theoretical models based on the meson exchange picture and assuming the validity of the phenomenological DELTA I=1/2 rule predict the lifetime of heavy hypernuclei to be significantly larger (2-3 standard deviations). Such large differences indicate that at least one of the assumptions in these models is not fulfilled. A much better reproduction of the lifetimes of heavy hypernuclei is achieved in the phase space model, if the DELTA I=1/2 rule is discarded in the nonmesonic LAMBDA decay. (orig.)

  13. The photoluminescent lifetime of polyelectrolytes in thin films formed via layer by layer self-assembly.

    Science.gov (United States)

    Reilly, Roseanne S; Smyth, Ciarán A; Rakovich, Yury P; McCabe, Eithne M

    2009-03-04

    We present results on luminescence lifetime studies of thin multilayer films of polyelectrolyte molecules produced via layer by layer (LbL) electrostatic assembly. We found that, in contrast to common assumptions, LbL films show measurable photoluminescent lifetimes with an average value of 6 ns. Scanning fluorescence lifetime imaging microscopy studies combined with steady-state photoluminescence measurements imply that this lifetime may be due to aggregation of polyelectrolyte molecules during preparation of LbL films. This conclusion has been further confirmed by atomic force microscopy (AFM). AFM images clearly show the presence of 100-200 nm high aggregates on the surface of these films. This aggregation of polyelectrolyte molecules contributes significantly to the experimentally detected luminescence decays of any light-emitting samples attached to LbL film, especially in a single molecule detection regime. To demonstrate this effect we compare photoluminescence lifetime results for CdTe quantum dots deposited on the surface of LbL polyelectrolyte films.

  14. Radiative lifetime measurements of some Tm I and Tm II levels by time-resolved laser spectroscopy

    Science.gov (United States)

    Tian, Yanshan; Wang, Xinghao; Yu, Qi; Li, Yongfan; Gao, Yang; Dai, Zhenwen

    2016-04-01

    Radiative lifetimes of 88 levels of Tm I in the energy range 22 791.176-48 547.98 cm-1 and 29 levels of Tm II in the range 27 294.79-65 612.85 cm-1 were measured by time-resolved laser-induced fluorescence spectroscopy in laser-ablation plasma. The lifetime values obtained are in the range from 15.4 to 7900 ns for Tm I and from 36.5 to 1000 ns for Tm II. To the best of our knowledge, 77 lifetimes of Tm I and 22 lifetimes of Tm II are reported for the first time. Good agreements between the present results and the previous experimental values were achieved for both Tm I and Tm II.

  15. Lifetimes of Rydberg states of Eu atoms

    Science.gov (United States)

    Jing, Hua; Ye, Shi-Wei; Dai, Chang-Jian

    2015-01-01

    The radiative lifetimes of the Eu 4f76snp (8PJ or 10PJ) Rydberg states with J = 5/2 and 11/2 are investigated with a combination of multi-step laser excitation and pulsed electric field ionization, from which their dependence on the effective principal quantum number is observed. The lifetimes of 21 states are reported along with an evaluation of their experimental uncertainty. The influence of blackbody radiation, due to the oven temperature, on the lifetime of the higher-n states is detected. The non-hydrogen behavior of the investigated states is also observed. Project supported by the National Natural Science Foundation of China (Grant No. 11174218).

  16. Improved lifetime of microchannel-plate PMTs

    Energy Technology Data Exchange (ETDEWEB)

    Lehmann, A., E-mail: lehmann@physik.uni-erlangen.de [Physikalisches Institut IV, Friedrich Alexander-University of Erlangen-Nuremberg, Erlangen (Germany); Britting, A.; Eyrich, W.; Uhlig, F. [Physikalisches Institut IV, Friedrich Alexander-University of Erlangen-Nuremberg, Erlangen (Germany); Dzhygadlo, R.; Gerhardt, A.; Götzen, K.; Höhler, R.; Kalicy, G.; Kumawat, H.; Lehmann, D.; Lewandowski, B.; Patsyuk, M.; Peters, K.; Schepers, G.; Schmitt, L.; Schwarz, C.; Schwiening, J.; Traxler, M.; Zühlsdorf, M. [GSI Helmholtzzentrum für Schwerionenforschung GmbH, Darmstadt (Germany); and others

    2014-12-01

    The charged particle identification at the PANDA experiment will be mainly performed with DIRC detectors. Because of their advantageous properties the preferred photon sensors are MCP-PMTs. However, until recently these devices showed serious aging problems which resulted in a diminishing quantum efficiency (QE) of the photo cathode. By applying innovative countermeasures against the aging causes, the manufacturers recently succeeded in drastically improving the lifetime of MCP-PMTs. Especially the application of an ALD coating technique to seal the material of the micro-channels proves very powerful and results in a lifetime of ≈6C/cm{sup 2} integrated anode charge without a substantial QE degradation for the latest PHOTONIS XP85112. This paper will present a comparative measurement of the lifetime of several older and recent MCP-PMTs demonstrating this progress.

  17. Lifetime of B hadrons from CDF

    Energy Technology Data Exchange (ETDEWEB)

    Miao, Ting; CDF Collaboration

    1996-08-01

    A review of the lifetimes of {ital B} hadrons measured by the CDF collaboration at Fermilab is presented. The data corresponds to 110 pb{sup -1} of {ital p}{ital {anti p}} collisions at {radical}s = 1.8 TeV. The inclusive {ital B} hadron lifetime is measured using a high statistics sample of {ital B} {r_arrow} {ital J}/{Psi}{Chi} decays. Species specific lifetimes of the {ital B}{sup +}, {ital B}{sup 0}, {ital B}{sup 0}{sub s}, and {Lambda}{sup 0}{sub b} are determined using both fully reconstructed decays and partially reconstructed decays consisting of a lepton associated with a charm hadron.

  18. Improved lifetime of microchannel-plate PMTs

    Science.gov (United States)

    Lehmann, A.; Britting, A.; Eyrich, W.; Uhlig, F.; Dzhygadlo, R.; Gerhardt, A.; Götzen, K.; Höhler, R.; Kalicy, G.; Kumawat, H.; Lehmann, D.; Lewandowski, B.; Patsyuk, M.; Peters, K.; Schepers, G.; Schmitt, L.; Schwarz, C.; Schwiening, J.; Traxler, M.; Zühlsdorf, M.; Dodokhov, V. Kh.; Düren, M.; Föhl, K.; Hayrapetyan, A.; Kröck, B.; Merle, O.; Rieke, J.; Cowie, E.; Keri, T.; Montgomery, R.; Rosner, G.; Achenbach, P.; Cardinali, M.; Hoek, M.; Lauth, W.; Sfienti, C.; Thiel, M.; Bühler, P.; Gruber, L.; Marton, J.; Suzuki, K.

    2014-12-01

    The charged particle identification at the PANDA experiment will be mainly performed with DIRC detectors. Because of their advantageous properties the preferred photon sensors are MCP-PMTs. However, until recently these devices showed serious aging problems which resulted in a diminishing quantum efficiency (QE) of the photo cathode. By applying innovative countermeasures against the aging causes, the manufacturers recently succeeded in drastically improving the lifetime of MCP-PMTs. Especially the application of an ALD coating technique to seal the material of the micro-channels proves very powerful and results in a lifetime of ≈ 6 C /cm2 integrated anode charge without a substantial QE degradation for the latest PHOTONIS XP85112. This paper will present a comparative measurement of the lifetime of several older and recent MCP-PMTs demonstrating this progress.

  19. Measurement of the $\\tau$ lepton lifetime

    CERN Document Server

    Buskulic, Damir; De Bonis, I; Décamp, D; Ghez, P; Goy, C; Lees, J P; Lucotte, A; Minard, M N; Odier, P; Pietrzyk, B; Ariztizabal, F; Chmeissani, M; Crespo, J M; Efthymiopoulos, I; Fernández, E; Fernández-Bosman, M; Gaitan, V; Garrido, L; Martínez, M; Orteu, S; Pacheco, A; Padilla, C; Palla, Fabrizio; Pascual, A; Perlas, J A; Sánchez, F; Teubert, F; Colaleo, A; Creanza, D; De Palma, M; Farilla, A; Gelao, G; Girone, M; Iaselli, Giuseppe; Maggi, G; Maggi, M; Marinelli, N; Natali, S; Nuzzo, S; Ranieri, A; Raso, G; Romano, F; Ruggieri, F; Selvaggi, G; Silvestris, L; Tempesta, P; Zito, G; Huang, X; Lin, J; Ouyang, Q; Wang, T; Xie, Y; Xu, R; Xue, S; Zhang, J; Zhang, L; Zhao, W; Bonvicini, G; Cattaneo, M; Comas, P; Coyle, P; Drevermann, H; Engelhardt, A; Forty, Roger W; Frank, M; Hagelberg, R; Harvey, J; Jacobsen, R; Janot, P; Jost, B; Kneringer, E; Knobloch, J; Lehraus, Ivan; Markou, C; Martin, E B; Mato, P; Minten, Adolf G; Miquel, R; Oest, T; Palazzi, P; Pater, J R; Pusztaszeri, J F; Ranjard, F; Rensing, P E; Rolandi, Luigi; Schlatter, W D; Schmelling, M; Schneider, O; Tejessy, W; Tomalin, I R; Venturi, A; Wachsmuth, H W; Wiedenmann, W; Wildish, T; Witzeling, W; Wotschack, J; Ajaltouni, Ziad J; Bardadin-Otwinowska, Maria; Barrès, A; Boyer, C; Falvard, A; Gay, P; Guicheney, C; Henrard, P; Jousset, J; Michel, B; Monteil, S; Pallin, D; Perret, P; Podlyski, F; Proriol, J; Rossignol, J M; Saadi, F; Fearnley, Tom; Hansen, J B; Hansen, J D; Hansen, J R; Hansen, P H; Nilsson, B S; Kyriakis, A; Simopoulou, Errietta; Siotis, I; Vayaki, Anna; Zachariadou, K; Blondel, A; Bonneaud, G R; Brient, J C; Bourdon, P; Passalacqua, L; Rougé, A; Rumpf, M; Tanaka, R; Valassi, Andrea; Verderi, M; Videau, H L; Candlin, D J; Parsons, M I; Focardi, E; Parrini, G; Corden, M; Delfino, M C; Georgiopoulos, C H; Jaffe, D E; Antonelli, A; Bencivenni, G; Bologna, G; Bossi, F; Campana, P; Capon, G; Chiarella, V; Felici, G; Laurelli, P; Mannocchi, G; Murtas, F; Murtas, G P; Pepé-Altarelli, M; Dorris, S J; Halley, A W; ten Have, I; Knowles, I G; Lynch, J G; Morton, W T; O'Shea, V; Raine, C; Reeves, P; Scarr, J M; Smith, K; Smith, M G; Thompson, A S; Thomson, F; Thorn, S; Turnbull, R M; Becker, U; Braun, O; Geweniger, C; Graefe, G; Hanke, P; Hepp, V; Kluge, E E; Putzer, A; Rensch, B; Schmidt, M; Sommer, J; Stenzel, H; Tittel, K; Werner, S; Wunsch, M; Beuselinck, R; Binnie, David M; Cameron, W; Colling, D J; Dornan, Peter J; Konstantinidis, N P; Moneta, L; Moutoussi, A; Nash, J; San Martin, G; Sedgbeer, J K; Stacey, A M; Dissertori, G; Girtler, P; Kuhn, D; Rudolph, G; Bowdery, C K; Brodbeck, T J; Colrain, P; Crawford, G; Finch, A J; Foster, F; Hughes, G; Sloan, Terence; Whelan, E P; Williams, M I; Galla, A; Greene, A M; Kleinknecht, K; Quast, G; Raab, J; Renk, B; Sander, H G; Wanke, R; Van Gemmeren, P; Zeitnitz, C; Aubert, Jean-Jacques; Bencheikh, A M; Benchouk, C; Bonissent, A; Bujosa, G; Calvet, D; Carr, J; Diaconu, C A; Etienne, F; Thulasidas, M; Nicod, D; Payre, P; Rousseau, D; Talby, M; Abt, I; Assmann, R W; Bauer, C; Blum, Walter; Brown, D; Dietl, H; Dydak, Friedrich; Ganis, G; Gotzhein, C; Jakobs, K; Kroha, H; Lütjens, G; Lutz, Gerhard; Männer, W; Moser, H G; Richter, R H; Rosado-Schlosser, A; Schael, S; Settles, Ronald; Seywerd, H C J; Saint-Denis, R; Wolf, G; Alemany, R; Boucrot, J; Callot, O; Cordier, A; Courault, F; Davier, M; Duflot, L; Grivaz, J F; Heusse, P; Jacquet, M; Kim, D W; Le Diberder, F R; Lefrançois, J; Lutz, A M; Musolino, G; Nikolic, I A; Park, H J; Park, I C; Schune, M H; Simion, S; Veillet, J J; Videau, I; Abbaneo, D; Azzurri, P; Bagliesi, G; Batignani, G; Bettarini, S; Bozzi, C; Calderini, G; Carpinelli, M; Ciocci, M A; Ciulli, V; Dell'Orso, R; Fantechi, R; Ferrante, I; Fidecaro, F; Foà, L; Forti, F; Giassi, A; Giorgi, M A; Gregorio, A; Ligabue, F; Lusiani, A; Marrocchesi, P S; Messineo, A; Rizzo, G; Sanguinetti, G; Sciabà, A; Spagnolo, P; Steinberger, Jack; Tenchini, Roberto; Tonelli, G; Triggiani, G; Vannini, C; Verdini, P G; Walsh, J; Betteridge, A P; Blair, G A; Bryant, L M; Cerutti, F; Gao, Y; Green, M G; Johnson, D L; Medcalf, T; Mir, L M; Perrodo, P; Strong, J A; Bertin, V; Botterill, David R; Clifft, R W; Edgecock, T R; Haywood, S; Edwards, M; Maley, P; Norton, P R; Thompson, J C; Bloch-Devaux, B; Colas, P; Emery, S; Kozanecki, Witold; Lançon, E; Lemaire, M C; Locci, E; Marx, B; Pérez, P; Rander, J; Renardy, J F; Roussarie, A; Schuller, J P; Schwindling, J; Trabelsi, A; Vallage, B; Johnson, R P; Kim, H Y; Litke, A M; McNeil, M A; Taylor, G; Beddall, A; Booth, C N; Boswell, R; Cartwright, S L; Combley, F; Dawson, I; Köksal, A; Letho, M; Newton, W M; Rankin, C; Thompson, L F; Böhrer, A; Brandt, S; Cowan, G D; Feigl, E; Grupen, Claus; Lutters, G; Minguet-Rodríguez, J A; Rivera, F; Saraiva, P; Smolik, L; Stephan, F; Apollonio, M; Bosisio, L; Della Marina, R; Giannini, G; Gobbo, B; Ragusa, F; Rothberg, J E; Wasserbaech, S R; Armstrong, S R; Bellantoni, L; Elmer, P; Feng, Z; Ferguson, D P S; Gao, Y S; González, S; Grahl, J; Harton, J L; Hayes, O J; Hu, H; McNamara, P A; Nachtman, J M; Orejudos, W; Pan, Y B; Saadi, Y; Schmitt, M; Scott, I J; Sharma, V; Turk, J; Walsh, A M; Wu Sau Lan; Wu, X; Yamartino, J M; Zheng, M; Zobernig, G

    1996-01-01

    The mean lifetime of the \\tau lepton is measured in a sample of 25700 \\tau pairs collected in 1992 with the ALEPH detector at LEP. A new analysis of the 1-1 topology events is introduced. In this analysis, the dependence of the impact parameter sum distribution on the daughter track momenta is taken into account, yielding improved precision compared to other impact parameter sum methods. Three other analyses of the one- and three-prong \\tau decays are updated with increased statistics. The measured lifetime is 293.5 \\pm 3.1 \\pm 1.7 \\fs. Including previous (1989--1991) ALEPH measurements, the combined \\tau lifetime is 293.7 \\pm 2.7 \\pm 1.6 \\fs.

  20. Tremendously increased lifetime of MCP-PMTs

    Science.gov (United States)

    Lehmann, A.; Britting, A.; Eyrich, W.; Pfaffinger, M.; Uhlig, F.; Belias, A.; Dzhygadlo, R.; Gerhardt, A.; Götzen, K.; Kalicy, G.; Krebs, M.; Lehmann, D.; Nerling, F.; Patsyuk, M.; Peters, K.; Schepers, G.; Schmitt, L.; Schwarz, C.; Schwiening, J.; Traxler, M.; Zühlsdorf, M.; Düren, M.; Etzelmüller, E.; Föhl, K.; Hayrapetyan, A.; Kröck, B.; Merle, O.; Rieke, J.; Schmidt, M.; Cowie, E.; Keri, T.; Achenbach, P.; Cardinali, M.; Hoek, M.; Lauth, W.; Schlimme, S.; Sfienti, C.; Thiel, M.

    2017-02-01

    Microchannel plate (MCP) PMTs are very attractive photon sensors for low light level applications in strong magnetic fields. However, until recently the main drawback of MCP-PMTs was their aging behavior which manifests itself in a limited lifetime due to a rapidly decreasing quantum efficiency (QE) of the photo cathode (PC) as the integrated anode charge (IAC) increases. In the latest models of PHOTONIS, Hamamatsu, and BINP novel techniques are applied to avoid these aging effects which are supposed to be mainly caused by feedback ion impinging on the PC and damaging it. For more than four years we are running a long-term aging test with new lifetime-enhanced MCP-PMT models by simultaneously illuminating various PMTs with roughly the same photon rate. This allows a fair comparison of the lifetime of all investigated MCP-PMTs and will give some insight into the best techniques to be applied for a lifetime enhancement. In this paper the results of comprehensive aging tests will be discussed. Gain, dark count rate and QE were investigated for their dependence on the IAC. The QE was measured spectrally resolved and as a function of the position across the PC to identify regions where the damage develops first. For the best performing tubes the lifetime improvement compared to former MCP-PMTs is a factor of ∼ 50 based on an IAC of meanwhile > 10 C /cm2. This breakthrough in the lifetime of MCP-PMTs was achieved by coating the MCP pores with an atomic layer deposition (ALD) technique.

  1. B lifetimes and mixing at the Tevatron

    Energy Technology Data Exchange (ETDEWEB)

    Bedeschi, Franco; /INFN, Pisa

    2005-05-01

    The authors present recent results on b-hadron lifetimes and mixing obtained from the analysis of the data collected at the Tevatron Collider by the CDF and D0 Collaborations in the period 2002-2004. Many lifetime measurements have been updated since the Summer 2004 conferences, sometimes improving significantly the accuracy. Likewise the measurement of the B{sub d} oscillation frequency has been updated. New limits on the B{sub s} oscillation frequency have been determined using for the first time Run II data.

  2. A compact positron annihilation lifetime spectrometer

    Institute of Scientific and Technical Information of China (English)

    LI Dao-Wu; LIU Jun-Hui; ZHANG Zhi-Ming; WANG Bao-Yi; ZHANG Tian-Bao; WEI Long

    2011-01-01

    Using LYSO scintillator coupled on HAMAMATSU R9800(a fast photomultiplier)to form the small size γ-ray detectors,a compact lifetime spectrometer has been built for the positron annihilation experiments.The system time resolution FWHM=193 ps and the coincidence counting rate -8 cps/μCi were achieved.A lifetime value of 219±1 ps of positron annihilation in well annealed Si was tested,which is in agreement with the typical values published in the previous lectures.

  3. Measurement of the Bs0 lifetime

    Science.gov (United States)

    Buskulic, D.; de Bonis, I.; Decamp, D.; Ghez, P.; Goy, C.; Lees, J.-P.; Minard, M.-N.; Odier, P.; Pietrzyk, B.; Ariztizabal, F.; Comas, P.; Crespo, J. M.; Efthymiopoulos, I.; Fernandez, E.; Fernandez-Bosman, M.; Gaitan, V.; Garrido, Ll.; Martinez, M.; Mattison, T.; Orteu, S.; Pacheco, A.; Padilla, C.; Pascual, A.; Creanza, D.; de Palma, M.; Farilla, A.; Iaselli, G.; Maggi, G.; Marinelli, N.; Natali, S.; Nuzzo, S.; Ranieri, A.; Raso, G.; Romano, F.; Ruggieri, F.; Selvaggi, G.; Silvestris, L.; Tempesta, P.; Zito, G.; Chai, Y.; Hu, H.; Huang, D.; Huang, X.; Lin, J.; Wang, T.; Xie, Y.; Xu, D.; Xu, R.; Zhang, J.; Zhang, L.; Zhao, W.; Bonvicini, G.; Boudreau, J.; Casper, D.; Drevermann, H.; Forty, R. W.; Ganis, G.; Gay, C.; Girone, M.; Hagelberg, R.; Harvey, J.; Hilgart, J.; Jacobsen, R.; Jost, B.; Knobloch, J.; Lehraus, I.; Maggi, M.; Markou, C.; Mato, P.; Meinhard, H.; Minten, A.; Miquel, R.; Moser, H.-G.; Palazzi, P.; Pater, J. R.; Perlas, J. A.; Perrodo, P.; Pusztaszeri, J.-F.; Ranjard, F.; Rolandi, L.; Rothberg, J.; Ruan, T.; Saich, M.; Schlatter, D.; Schmelling, M.; Sefkow, F.; Tejessy, W.; Tomalin, I. R.; Veenhof, R.; Wachsmuth, H.; Wasserbaech, S.; Wiedenmann, W.; Wildish, T.; Witzeling, W.; Wotschack, J.; Ajaltouni, Z.; Bardadin-Otwinowska, M.; Barres, A.; Boyer, C.; Falvard, A.; Gay, P.; Guicheney, C.; Henrard, P.; Jousset, J.; Michel, B.; Montret, J.-C.; Pallin, D.; Perret, P.; Podlyski, F.; Proriol, J.; Saadi, F.; Fearnley, T.; Hansen, J. B.; Hansen, J. D.; Hansen, J. R.; Hansen, P. H.; Johnson, S. D.; Møllerud, R.; Nilsson, B. S.; Kyriakis, A.; Simopoulou, E.; Siotis, I.; Vayaki, A.; Zachariadou, K.; Badier, J.; Blondel, A.; Bonneaud, G.; Brient, J. C.; Bourdon, P.; Fouque, G.; Passalacqua, L.; Rougé, A.; Rumpf, M.; Tanaka, R.; Verderi, M.; Videau, H.; Candlin, D. J.; Parsons, M. I.; Veitch, E.; Focardi, E.; Moneta, L.; Parrini, G.; Corden, M.; Delfino, M.; Georgiopoulos, C.; Jaffe, D. E.; Levinthal, D.; Antonelli, A.; Bencivenni, G.; Bologna, G.; Bossi, F.; Campana, P.; Capon, G.; Cerutti, F.; Chiarella, V.; Felici, G.; Laurelli, P.; Mannocchi, G.; Murtas, F.; Murtas, G. P.; Pepe-Altarelli, M.; Salomone, S.; Colrain, P.; Ten Have, I.; Knowles, I. G.; Lynch, J. G.; Maitland, W.; Morton, W. T.; Raine, C.; Reeves, P.; Scarr, J. M.; Smith, K.; Smith, M. G.; Thompson, A. S.; Thorn, S.; Turnbull, R. M.; Brandl, B.; Braun, O.; Geweniger, C.; Graefe, G.; Hanke, P.; Hepp, V.; Karger, C.; Kluge, E. E.; Maumary, Y.; Putzer, A.; Rensch, B.; Stahl, A.; Tittel, K.; Wunsch, M.; Beuselinck, R.; Binnie, D. M.; Cameron, W.; Cattaneo, M.; Colling, D. J.; Dornan, P. J.; Hassard, J. F.; Lieske, N. M.; Moutoussi, A.; Nash, J.; Patton, S.; Payne, D. G.; Phillips, M. J.; San Martin, G.; Sedgbeer, J. K.; Wright, A. G.; Girtler, P.; Kuhn, D.; Rudolph, G.; Vogl, R.; Bowdery, C. K.; Brodbeck, T. J.; Finch, A. J.; Foster, F.; Hughes, G.; Jackson, D.; Keemer, N. R.; Nuttall, M.; Patel, A.; Sloan, T.; Snow, S. W.; Whelan, E. P.; Galla, A.; Greene, A. M.; Kleinknecht, K.; Raab, J.; Renk, B.; Sander, H.-G.; Schmidt, H.; Walther, S. M.; Wanke, R.; Wolf, B.; Bencheikh, A. M.; Benchouk, C.; Bonissent, A.; Calvet, D.; Carr, J.; Coyle, P.; Diaconu, C.; Drinkard, J.; Etienne, F.; Nicod, D.; Payre, P.; Ross, L.; Rousseau, D.; Schwemling, P.; Talby, M.; Adlung, S.; Assmann, R.; Bauer, C.; Blum, W.; Brown, D.; Cattaneo, P.; Dehning, B.; Dietl, H.; Dydak, F.; Frank, M.; Halley, A. W.; Jakobs, K.; Lauber, J.; Lütjens, G.; Lutz, G.; Männer, W.; Richter, R.; Schröder, J.; Schwarz, A. S.; Settles, R.; Seywerd, H.; Stierlin, U.; Stiegler, U.; Denis, R. St.; Wolf, G.; Alemany, R.; Boucrot, J.; Callot, O.; Cordier, A.; Davier, M.; Duflot, L.; Grivaz, J.-F.; Heusse, Ph.; Janot, P.; Kimfn 19, D. W.; Le Diberder, F.; Lefrançois, J.; Lutz, A.-M.; Musolino, G.; Schune, M.-H.; Veillet, J.-J.; Videau, I.; Abbaneo, D.; Bagliesi, G.; Batignani, G.; Bottigli, U.; Bozzi, C.; Calderini, G.; Carpinelli, M.; Ciocci, M. A.; Ciulli, V.; Dell'Orso, R.; Ferrante, I.; Fidecaro, F.; Foà, L.; Forti, F.; Giassi, A.; Giorgi, M. A.; Gregorio, A.; Ligabue, F.; Lusiani, A.; Mannelli, E. B.; Marrocchesi, P. S.; Messineo, A.; Palla, F.; Rizzo, G.; Sanguinetti, G.; Spagnolo, P.; Steinberger, J.; Tenchini, R.; Tonelli, G.; Triggiani, G.; Valassi, A.; Vannini, C.; Venturi, A.; Verdini, P. G.; Walsh, J.; Betteridge, A. P.; Gao, Y.; Green, M. G.; Johnson, D. L.; March, P. V.; Medcalf, T.; Mir, Ll. M.; Quazi, I. S.; Strong, J. A.; Bertin, V.; Botterill, D. R.; Clifft, R. W.; Edgecock, T. R.; Haywood, S.; Edwards, M.; Norton, P. R.; Thompson, J. C.; Bloch-Devaux, B.; Colas, P.; Duarte, H.; Emery, S.; Kozanecki, W.; Lançon, E.; Lemaire, M. C.; Locci, E.; Marx, B.; Perez, P.; Rander, J.; Renardy, J.-F.; Rosowsky, A.; Roussarie, A.; Schuller, J.-P.; Schwindling, J.; Si Mohand, D.; Vallage, B.; Johnson, R. P.; Litke, A. M.; Taylor, G.; Wear, J.; Babbage, W.; Booth, C. N.; Buttar, C.; Cartwright, S.; Combley, F.; Dawson, I.; Thompson, L. F.; Barberio, E.; Böhrer, A.; Brandt, S.; Cowan, G.; Grupen, C.; Lutters, G.; Rivera, F.; Schäfer, U.; Smolik, L.; Bosisio, L.; Della Marina, R.; Giannini, G.; Gobbo, B.; Pitis, L.; Ragusa, F.; Bellantoni, L.; Chen, W.; Conway, J. S.; Feng, Z.; Ferguson, D. P. S.; Gao, Y. S.; Grahl, J.; Harton, J. L.; Hayes, O. J.; Nachtman, J. M.; Pan, Y. B.; Saadi, Y.; Schmitt, M.; Scott, I.; Sharma, V.; Shi, Z. H.; Turk, J. D.; Walsh, A. M.; Weber, F. V.; Lan Wu, Sau; Wu, X.; Zheng, M.; Zobernig, G.; Aleph Collaboration

    1994-02-01

    The lifetime of the Bs0 has been measured in a data sample of 8890000 hadronic events recorded with the ALEPH detector at LEP. After background subtraction 30.8 ± 6.9 events are attributed to the semileptonic decay of the Bs0 to a Ds- and an opposite-sign lepton. A maximum-likelihood fit to the distribution of the proper times of these events yields a Bs0 lifetime of τBs = 1.92 -0.35+0.45 ± 0.04 ps.

  4. An approach for longer lifetime MCFCs

    Energy Technology Data Exchange (ETDEWEB)

    Matsumoto, Masaru; Tatsumi, Masahiko; Hayano, Takuro [MCFC Research Association, Tokyo (Japan)] [and others

    1996-12-31

    For entering into commercialization of MCFC power plants in the beginning of the 21st century, we will devote to research for increasing lifetime as long as 40,000 hours with cell performance decay rate of 0.25 %/1000hrs as the target in FY 1999. This paper will discuss on our approach for longer lifetime MCFCs through electrolyte-loss management and NiO precipitation management as well as micro-structural control of electrodes and matrix plates. Cell voltage decay rate will be estimated by simulation through series of experiments on accelerated conditions.

  5. A Study on Criteria for Barrel Lifetime

    Institute of Scientific and Technical Information of China (English)

    马吉胜; 郑坚; 邓辉咏

    2012-01-01

    Several criteria for barrel lifetime were summarized and discussed. Based on large amount of test data, the ad- vantages and disadvantages of the criteria were analyzed and the requirements for the easy and practical criterion were put forward. Then, a new criterion based on the radical wear at the start points of the barrel lands was proposed. The close in- terrelationship between the radical wear and interior ballistic characteristics was illuminated theoretically and experimental- ly. The research results show the great value of this criterion to solve the problem of barrel lifetime.

  6. 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.

  7. Time-resolved fluorescence analysis of the mobile flavin cofactor in -hydroxybenzoate hydroxylase

    Indian Academy of Sciences (India)

    Petra A W Van Den Berg; Koert Grever; Arie Van Hoek; Willem J H Van Berkel; Antonie J W G Visser

    2007-03-01

    Conformational heterogeneity of the FAD cofactor in -hydroxybenzoate hydroxylase (PHBH) was investigated with time-resolved polarized flavin fluorescence. For binary enzyme/substrate (analogue) complexes of wild-type PHBH and Tyr222 mutants, crystallographic studies have revealed two distinct flavin conformations; the `in’ conformation with the isoalloxazine ring located in the active site, and the `out’ conformation with the isoalloxazine ring disposed towards the protein surface. Fluorescence-lifetime analysis of these complexes revealed similar lifetime distributions for the `in’ and `out’ conformations. The reason for this is twofold. First, the active site of PHBH contains various potential fluorescence-quenching sites close to the flavin. Fluorescence analysis of uncomplexed PHBH Y222V and Y222A showed that Tyr222 is responsible for picosecond fluorescence quenching free enzyme. In addition, other potential quenching sites, including a tryptophan and two tyrosines involved in substrate binding, are located nearby. Since the shortest distance between these quenching sites and the isoalloxazine ring differs only little on average, these aromatic residues are likely to contribute to fluorescence quenching. Second, the effect of flavin conformation on the fluorescence lifetime distribution is blurred by binding of the aromatic substrates: saturation with aromatic substrates induces highly efficient fluorescence quenching. The flavin conformation is therefore only reflected in the small relative contributions of the longer lifetimes.

  8. Long-term optical stability of fluorescent solar concentrator plates

    NARCIS (Netherlands)

    Slooff, Lenneke H.; Bakker, Nicolaas J.; Sommeling, Paul M.; Büchtemann, Andreas; Wedel, Armin; Van Sark, Wilfried G J H M

    2014-01-01

    Fluorescent solar concentrators offer an alternative approach for low-cost photovoltaic energy conversion. For successful application, not only the power conversion efficiency and cost are important, but also lifetime or stability of the devices. As today's concentrator is made of polymer sheets

  9. Long-term optical stability of fluorescent solar concentrator plates

    NARCIS (Netherlands)

    Slooff, Lenneke H.; Bakker, Nicolaas J.; Sommeling, Paul M.; Büchtemann, Andreas; Wedel, Armin; Van Sark, Wilfried G J H M

    2014-01-01

    Fluorescent solar concentrators offer an alternative approach for low-cost photovoltaic energy conversion. For successful application, not only the power conversion efficiency and cost are important, but also lifetime or stability of the devices. As today's concentrator is made of polymer sheets con

  10. A new method for measuring the neutron lifetime using an in situ neutron detector

    CERN Document Server

    Morris, C L; Broussard, L J; Callahan, N B; Clayton, S M; Cude-Woods, C; Currie, S A; Ding, X; Fox, W; Hickerson, K P; Holley, A T; Komives, A; Liu, C -Y; Makela, M; Pattie, R W; Ramsey, J; Salvat, D J; Saunders, A; Seestrom, S J; Sharapov, E I; Sjue, S K; Tang, Z; Vanderwerp, J; Vogelaar, B; Walstrom, P L; Wang, Z; Wei, Wanchun; Wexler, J W; Womack, T L; Young, A R; Zeck, B A

    2016-01-01

    The neutron lifetime is important in understanding the production of light nuclei in the first minutes after the big bang and it provides basic information on the charged weak current of the standard model of particle physics. Two different methods have been used to measure the neutron lifetime: disappearance measurements using bottled ultracold neutrons and decay rate measurements using neutron beams. The best measurements using these two techniques give results that differ by nearly 4 standard deviations. In this paper we describe a new method for measuring surviving neutrons in neutron lifetime measurements using bottled ultracold neutrons that provides better characterization of systematic uncertainties and enables higher precision than previous measurement techniques. We present results obtained using our method.

  11. Smoking expands expected lifetime with musculoskeletal disease

    DEFF Research Database (Denmark)

    Brønnum-Hansen, Henrik; Juel, Knud

    2003-01-01

    By indirect estimation of mortality from smoking and life table methods we estimated expected lifetime without musculoskeletal diseases among never smokers, ex-smokers, and smokers. We found that although life expectancy of a heavy smoker is 7 years shorter than that of a never smoker, heavy...

  12. Lifetime Reliability Assessment of Concrete Slab Bridges

    DEFF Research Database (Denmark)

    Thoft-Christensen, Palle

    A procedure for lifetime assesment of the reliability of short concrete slab bridges is presented in the paper. Corrosion of the reinforcement is the deterioration mechanism used for estimating the reliability profiles for such bridges. The importance of using sensitivity measures is stressed. Fi...

  13. Smoking expands expected lifetime with musculoskeletal disease

    DEFF Research Database (Denmark)

    Brønnum-Hansen, Henrik; Juel, Knud

    2003-01-01

    By indirect estimation of mortality from smoking and life table methods we estimated expected lifetime without musculoskeletal diseases among never smokers, ex-smokers, and smokers. We found that although life expectancy of a heavy smoker is 7 years shorter than that of a never smoker, heavy...... smokers can expect to live more than 2 years longer with musculoskeletal diseases than never smokers....

  14. A compact positron annihilation lifetime spectrometer

    Institute of Scientific and Technical Information of China (English)

    李道武; 刘军辉; 章志明; 王宝义; 张天保; 魏龙

    2011-01-01

    Using LYSO scintillator coupled on HAMAMATSU R9800 (a fast photomultiplier) to form the small size γ-ray detectors, a compact lifetime spectrometer has been built for the positron annihilation experiments. The system time resolution FWHM=193 ps and the co

  15. Assessing the inequality of lifetime healthcare expenditures

    NARCIS (Netherlands)

    Wong, Albert; Boshuizen, Hendriek; Polder, Johan; Ferreira, José António

    2016-01-01

    The rise in healthcare expenditures has raised doubts about the sustainability of health systems and instigated a discussion on their design. Policy making in this field requires a proper understanding of how healthcare expenditures evolve throughout an individual's lifetime, and of how they vary

  16. Lifetime Prolonging Algorithms for Underwater Sensor Networks

    Institute of Scientific and Technical Information of China (English)

    GUO Zhong-wen; LI Zhi-wei; YU Lei

    2006-01-01

    Underwater acoustic modem technology has attained a level of maturity to support underwater acoustic sensor networks (UASNs) which are generally formed by acoustically connected sensor nodes and a surface station providing a link to an on-shore control center. While many applications require long-term monitoring of the deployment area, the battery-powered network nodes limit the lifetime of UASNs. Therefore, designing a UASN that minimizes the power consumption while maximizing lifetime becomes a very difficult task. In this paper, a method is proposed to determine the optimum number of clusters through combining an application-specific protocol architecture and underwater acoustic communication model so as to reduce the energy dissipation of UASNs. Deploying more sensor nodes which work alternately is another way to prolong the lifetime of UASNs. An algorithm is presented for selecting sensor nodes and putting them into operation in each round, ensuring the monitoring to the whole given area. The present results show that the algorithm can help prolong system lifetime remarkably when it is applied to other conventional approaches for sensor networks under the condition that the sensor node density is high.

  17. Updated measurement of the $\\tau$ lepton lifetime

    CERN Document Server

    Barate, R; Décamp, D; Ghez, P; Goy, C; Lees, J P; Lucotte, A; Minard, M N; Nief, J Y; Pietrzyk, B; Casado, M P; Chmeissani, M; Comas, P; Crespo, J M; Delfino, M C; Fernández, E; Fernández-Bosman, M; Garrido, L; Juste, A; Martínez, M; Merino, G; Miquel, R; Mir, L M; Padilla, C; Park, I C; Pascual, A; Perlas, J A; Riu, I; Sánchez, F; Teubert, F; Colaleo, A; Creanza, D; De Palma, M; Gelao, G; Iaselli, Giuseppe; Maggi, G; Maggi, M; Marinelli, N; Nuzzo, S; Ranieri, A; Raso, G; Ruggieri, F; Selvaggi, G; Silvestris, L; Tempesta, P; Tricomi, A; Zito, G; Huang, X; Lin, J; Ouyang, Q; Wang, T; Xie, Y; Xu, R; Xue, S; Zhang, J; Zhang, L; Zhao, W; Abbaneo, D; Alemany, R; Becker, U; Bazarko, A O; Bright-Thomas, P G; Cattaneo, M; Cerutti, F; Dissertori, G; Drevermann, H; Forty, Roger W; Frank, M; Hagelberg, R; Hansen, J B; Harvey, J; Janot, P; Jost, B; Kneringer, E; Knobloch, J; Lehraus, Ivan; Mato, P; Minten, Adolf G; Moneta, L; Pacheco, A; Pusztaszeri, J F; Ranjard, F; Rizzo, G; Rolandi, Luigi; Rousseau, D; Schlatter, W D; Schmitt, M; Schneider, O; Tejessy, W; Tomalin, I R; Wachsmuth, H W; Wagner, A; Ajaltouni, Ziad J; Barrès, A; Boyer, C; Falvard, A; Ferdi, C; Gay, P; Guicheney, C; Henrard, P; Jousset, J; Michel, B; Monteil, S; Montret, J C; Pallin, D; Perret, P; Podlyski, F; Proriol, J; Rosnet, P; Rossignol, J M; Fearnley, Tom; Hansen, J D; Hansen, J R; Hansen, P H; Nilsson, B S; Rensch, B; Wäänänen, A; Daskalakis, G; Kyriakis, A; Markou, C; Simopoulou, Errietta; Siotis, I; Vayaki, Anna; Blondel, A; Bonneaud, G R; Brient, J C; Bourdon, P; Rougé, A; Rumpf, M; Valassi, Andrea; Verderi, M; Videau, H L; Candlin, D J; Parsons, M I; Focardi, E; Parrini, G; Zachariadou, K; Corden, M; Georgiopoulos, C H; Jaffe, D E; Antonelli, A; Bencivenni, G; Bologna, G; Bossi, F; Campana, P; Capon, G; Casper, David William; Chiarella, V; Felici, G; Laurelli, P; Mannocchi, G; Murtas, F; Murtas, G P; Passalacqua, L; Pepé-Altarelli, M; Curtis, L; Dorris, S J; Halley, A W; Knowles, I G; Lynch, J G; O'Shea, V; Raine, C; Scarr, J M; Smith, K; Teixeira-Dias, P; Thompson, A S; Thomson, E; Thomson, F; Turnbull, R M; Buchmüller, O L; Dhamotharan, S; Geweniger, C; Graefe, G; Hanke, P; Hansper, G; Hepp, V; Kluge, E E; Putzer, A; Sommer, J; Tittel, K; Werner, S; Wunsch, M; Beuselinck, R; Binnie, David M; Cameron, W; Dornan, Peter J; Girone, M; Goodsir, S M; Martin, E B; Moutoussi, A; Nash, J; Sedgbeer, J K; Spagnolo, P; Stacey, A M; Williams, M D; Ghete, V M; Girtler, P; Kuhn, D; Rudolph, G; Betteridge, A P; Bowdery, C K; Colrain, P; Crawford, G; Finch, A J; Foster, F; Hughes, G; Jones, R W L; Sloan, Terence; Williams, M I; Galla, A; Giehl, I; Greene, A M; Hoffmann, C; Jakobs, K; Kleinknecht, K; Quast, G; Renk, B; Rohne, E; Sander, H G; Van Gemmeren, P; Zeitnitz, C; Aubert, Jean-Jacques; Benchouk, C; Bonissent, A; Bujosa, G; Carr, J; Coyle, P; Diaconu, C A; Etienne, F; Konstantinidis, N P; Leroy, O; Motsch, F; Payre, P; Talby, M; Sadouki, A; Thulasidas, M; Trabelsi, K; Aleppo, M; Antonelli, M; Ragusa, F; Berlich, R; Blum, Walter; Büscher, V; Dietl, H; Ganis, G; Gotzhein, C; Kroha, H; Lütjens, G; Lutz, Gerhard; Männer, W; Moser, H G; Richter, R H; Rosado-Schlosser, A; Schael, S; Settles, Ronald; Seywerd, H C J; Saint-Denis, R; Stenzel, H; Wiedenmann, W; Wolf, G; Boucrot, J; Callot, O; Chen, S; Choi, Y; Cordier, A; Davier, M; Duflot, L; Grivaz, J F; Heusse, P; Höcker, A; Jacholkowska, A; Jacquet, M; Kim, D W; Le Diberder, F R; Lefrançois, J; Lutz, A M; Nikolic, I A; Schune, M H; Simion, S; Tournefier, E; Veillet, J J; Videau, I; Zerwas, D; Azzurri, P; Bagliesi, G; Batignani, G; Bettarini, S; Bozzi, C; Calderini, G; Carpinelli, M; Ciocci, M A; Ciulli, V; Dell'Orso, R; Fantechi, R; Ferrante, I; Foà, L; Forti, F; Giassi, A; Giorgi, M A; Gregorio, A; Ligabue, F; Lusiani, A; Marrocchesi, P S; Messineo, A; Palla, Fabrizio; Sanguinetti, G; Sciabà, A; Steinberger, Jack; Tenchini, Roberto; Tonelli, G; Vannini, C; Venturi, A; Verdini, P G; Blair, G A; Bryant, L M; Chambers, J T; Gao, Y; Green, M G; Medcalf, T; Perrodo, P; Strong, J A; Von Wimmersperg-Töller, J H; Botterill, David R; Clifft, R W; Edgecock, T R; Haywood, S; Norton, P R; Thompson, J C; Wright, A E; Bloch-Devaux, B; Colas, P; Emery, S; Kozanecki, Witold; Lançon, E; Lemaire, M C; Locci, E; Pérez, P; Rander, J; Renardy, J F; Roussarie, A; Schuller, J P; Schwindling, J; Trabelsi, A; Vallage, B; Black, S N; Dann, J H; Johnson, R P; Kim, H Y; Litke, A M; McNeil, M A; Taylor, G; Booth, C N; Boswell, R; Brew, C A J; Cartwright, S L; Combley, F; Kelly, M S; Lehto, M H; Newton, W M; Reeve, J; Thompson, L F; Böhrer, A; Brandt, S; Cowan, G D; Grupen, Claus; Lutters, G; Saraiva, P; Smolik, L; Stephan, F; Apollonio, M; Bosisio, L; Della Marina, R; Giannini, G; Gobbo, B; Musolino, G; Pütz, J; Rothberg, J E; Wasserbaech, S R; Armstrong, S R; Charles, E; Elmer, P; Ferguson, D P S; González, S; Greening, T C; Hayes, O J; Hu, H; Jin, S; McNamara, P A; Nachtman, J M; Nielsen, J; Orejudos, W; Pan, Y B; Saadi, Y; Scott, I J; Walsh, J; Wu Sau Lan; Wu, X; Yamartino, J M; Zobernig, G

    1997-01-01

    A new measurement of the mean lifetime of the tau lepton is presented. Three different analysis methods are applied to a sample of 90000 tau pairs, collected in 1993 and 1994 with the ALEPH detector at LEP. The average of this measurement and those previously published by ALEPH is tau_tau = 290.1 +- 1.5 +- 1.1 fs.

  18. Lifetime oriented maintenance planning in the Netherlands

    NARCIS (Netherlands)

    Straub, A.

    2003-01-01

    In this paper we set up a framework for lifetime oriented maintenance planning as an outcome and input for strategic housing stock management. The maintenance planning holds maintenance activities and costs in the longer term. We consider the maintenance planning as a tool to calculate and implement

  19. Overview of Field Experience - Degradation Rates & Lifetimes

    Energy Technology Data Exchange (ETDEWEB)

    Jordan, Dirk; Kurtz, Sarah

    2015-09-14

    The way a PV module fails may depend not only on its design and the materials used in its construction, but also on the weather it experiences, the way it is mounted, and the quality control during its manufacture. This presentation gives an overview of Field Experience - what degradation rates and what lifetimes are being observed in various regions.

  20. LIFETIMES OF SUPERDEFORMED STATES IN 194PB

    NARCIS (Netherlands)

    WILLSAU, P; HUBEL, H; AZAIEZ, F; DELEPLANQUE, MA; DIAMOND, RM; KORTEN, W; MACCHIAVELLI, AO; STEPHENS, FS; KLUGE, H; HANNACHI, F; BACELAR, JC; BECKER, JA; BRINKMAN, MJ; HENRY, EA; KUHNERT, A; WANG, TF; DRAPER, JA; RUBEL, E

    1992-01-01

    Lifetimes of members of the superdeformed band in 194Pb were measured by the Doppler-shift attenuation method. Quadrupole moments around 20 eb that are constant over the whole frequency range were derived. The results rule out large centrifugal stretching effects.

  1. The College Payoff: Education, Occupations, Lifetime Earnings

    Science.gov (United States)

    Carnevale, Anthony P.; Rose, Stephen J.; Cheah, Ban

    2011-01-01

    A college degree pays off--but by just how much? In this report from the Georgetown University Center on Education and the Workforce, the authors examine just what a college degree is worth--and what else besides a degree might influence an individual's potential earnings. This report examines lifetime earnings for all education levels and…

  2. Lifetime Modeling of Thermal Barrier Coatings

    NARCIS (Netherlands)

    Hille, T.S.

    2009-01-01

    Thermal barrier coatings (TBCs) are applied in gas turbines to enhance their thermal efficiency by isolating the metallic components from the aggressive hot gas. TBC lifetime is limited by damage processes originating at internal interfaces, which may ultimately lead to delamination and spallation.

  3. Charmed particle lifetimes. [Review, six quark model

    Energy Technology Data Exchange (ETDEWEB)

    Rosner, J.L.

    1979-01-01

    Conventional estimates are reviewed for charmed particle lifetimes. Free-quark models give values of (a few) x 10/sup -13/ sec to (a few) x 10/sup -12/ sec. The shorter of these values also follows from an extrapolation based on D ..-->.. Ke/sup nu/. Possible differences among the lifetimes and production rates of D/sup 0/, D/sup +/, F/sup +/, C/sub 0//sup +/, the heavy lepton tau, and the fifth quark b are discussed. Extreme values of mixing angles in a six-quark model could extend charmed particle lifetimes by a factor of at most three from the above estimates, while shorter lifetimes than those predicted could occur for some species like D/sup 0/ or F/sup +/ if their nonleptonic decays were enhanced. The predictions are discussed in the light of some current experimental results, and it is estimated that sigma(pp ..-->.. charm) approx. = 10 ..mu..b at 400 GeV/c. 95 references.

  4. Materials Education: Opportunities over a Lifetime

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, Iver E.; Schwartz, Lyle H.; Faber, Katherine T.; Cargill III, G. Slade; Houston, Betsy

    2003-10-28

    A report, in the form of abbreviated notes, of the 17th Biennial Conference on National Materials Policy ''Materials Education: Opportunities over a Lifetime'' held May 20-21, 2002 in College Park, MD, sponsored by the Federation of Materials Societies and the University Materials Council.

  5. The nature of multiphoton fluorescence from red blood cells

    Science.gov (United States)

    Saytashev, Ilyas; Murphy, Michael; Osseiran, Sam; Spence, Dana M.; Evans, Conor L.; Dantus, Marcos

    2016-03-01

    We report on the nature of multiphoton excited fluorescence observed from human erythrocytes (red blood cells RBC's) and their "ghosts" following 800nm sub-15 fs excitation. The detected optical signal is assigned as two-photon excited fluorescence from hemoglobin. Our findings are supported by wavelength-resolved fluorescence lifetime decay measurements using time-correlated single photon counting system from RBC's, their ghosts as well as in vitro samples of various fluorophores including riboflavin, NADH, NAD(P)H, hemoglobin. We find that low-energy and short-duration pulses allow two-photon imaging of RBC's, but longer more intense pulses lead to their destruction.

  6. Steady state and time resolved fluorescence studies of azadioxatriangulenium (ADOTA) fluorophore in silica and PVA thin films

    DEFF Research Database (Denmark)

    Chib, Rahul; Raut, Sangram; Shah, Sunil

    2015-01-01

    in silica thin films and PVA films were studied by means of steady-state and time resolved fluorescence techniques. We have found that the azadioxatriangulenium entrapped in silica thin film has a wider fluorescence lifetime distribution (Lorentzian distribution), lower fluorescence efficiencies, shorter....... In contrast to the PVA matrices, the porous silica films allow restricted rotations of Azadioxatriangulenium molecules, which result in faster and complex fluorescence anisotropy decays suggesting energy migration among dye molecules....

  7. Fluorescence dynamics of interactions between polyamide PyPyPyβDp and DNA

    Institute of Scientific and Technical Information of China (English)

    ZHANG; Huijuan; WANG; Jin; WU; Yishi; YUAN; Gu; AI; Xicheng; WANG; Li

    2006-01-01

    The photophysical properties of the polyamide PyPyPyβDp (PPP) were investigated by means of steady-state absorption and fluorescence spectroscopies, as well as time-resolved fluorescence spectroscopy. It was found that the excited-state properties of PPP are very sensitive to solvents. In TKMC buffer PPP exhibited weak fluorescence with a decay time constant of 16 ps, while with the decrease of the solvent polarity PPP showed the blue-shifted peak position, increased intensity and lengthened life-time for its fluorescence behavior. In the presence of calf thymus DNA, it was observed that the fluorescence intensity was enhanced and the fluorescence lifetime increased from 16 to 32 ps for PPP, which verified that PPP bound into the minor groove of DNA duplex.

  8. Fluorescence spectroscopic studies on binding of a flavonoid antioxidant quercetin to serum albumins

    Indian Academy of Sciences (India)

    Beena Mishra; Atanu Barik; K Indira Priyadarsini; Hari Mohan

    2005-11-01

    Binding of quercetin to human serum albumin (HSA) was studied and the binding constant measured by following the red-shifted absorption spectrum of quercetin in the presence of HSA and the quenching of the intrinsic protein fluorescence in the presence of different concentrations of quercetin. Fluorescence lifetime measurements of HSA showed decrease in the average lifetimes indicating binding at a location, near the tryptophan moiety, and the possibility of fluorescence energy transfer between excited tryptophan and quercetin. Critical transfer distance () was determined, from which the mean distance between tryptophan-214 in HSA and quercetin was calculated. The above studies were also carried out with bovine serum albumin (BSA).

  9. Nanoscaled ZnO films used as enhanced substrates for fluorescence detection of dyes

    Institute of Scientific and Technical Information of China (English)

    Liu Yan-Song; Yi Fu; Ramachandram Badugu; Joseph R.Lakowicz; Xu Xiao-Liang

    2012-01-01

    The ability of nanoscaled ZnO films to enhance fluorescence was studied.We found that the fluorescence intensities of Cy5,rhodamine 6G,and fluorescein can be enhanced about 10-fold on nanoscaled ZnO films as compared to that on glass substrates.The lifetimes of all samples were measured,and no obvious change in lifetime was observed for dyes on different substrates.The mechanism for the nanoscaled ZnO film enhanced fluorescence appears to be different from that for the metal-fluorophore systems.

  10. Picosecond wide-field time-correlated single photon counting fluorescence microscopy with a delay line anode detector

    Science.gov (United States)

    Hirvonen, Liisa M.; Becker, Wolfgang; Milnes, James; Conneely, Thomas; Smietana, Stefan; Le Marois, Alix; Jagutzki, Ottmar; Suhling, Klaus

    2016-08-01

    We perform wide-field time-correlated single photon counting-based fluorescence lifetime imaging (FLIM) with a crossed delay line anode image intensifier, where the pulse propagation time yields the photon position. This microchannel plate-based detector was read out with conventional fast timing electronics and mounted on a fluorescence microscope with total internal reflection (TIR) illumination. The picosecond time resolution of this detection system combines low illumination intensity of microwatts with wide-field data collection. This is ideal for fluorescence lifetime imaging of cell membranes using TIR. We show that fluorescence lifetime images of living HeLa cells stained with membrane dye di-4-ANEPPDHQ exhibit a reduced lifetime near the coverslip in TIR compared to epifluorescence FLIM.

  11. A device for fluorescence temperature measurement based on fast fourier transform

    Institute of Scientific and Technical Information of China (English)

    WANG Dong-sheng; WANG Gui-mei; PAN Wei-wei; WANG Yu-tian

    2008-01-01

    A sapphire fiber thermal probe with Cr3+ion-doped end was grown using the laser heated pedestal method.The fluores-ccnce themal probe offers advantages of compact structure,high performance and the ability to sustain high temperature from the room temperature to 450℃.Based on the fast fourier transform(FFT),the fluorescence lifetime is obtained from the tangent function of the phase angle of the first non-zeroth item of FFT result.Compared with other traditional fitting methods,our method has advantages such as fast speed,high accuracy and being free from the influence of the base signal.The standard deviation of FFT method is about half of that of the Prony method and close to the one of the Marquardt method.In addition.since the FFT method is immunity to the background noise of the signal,the background noise analysis can be skipped.

  12. Gelation in free-radical crosslinking copolymerization; fast transient fluorescence study

    OpenAIRE

    Ö. Pekcan; D. Kaya

    2002-01-01

    The fast transient fluorescence (FTRF) technique was used to study the sol-gel phase transition in free-radical crosslinking copolymerization (FCC) in two different monomeric systems. Pyrene (Py) was used as a fluorescence probe for the in situ polymerization experiments. The fluorescence lifetimes of Py from its decay traces were measured and used to monitor the gelation process. Monomer consumption profiles were determined during gelation process using Stern-Volmer model. Gelations...

  13. Rise-time of FRET-acceptor fluorescence tracks protein folding

    OpenAIRE

    Simon Lindhoud; Adrie H. Westphal; van Mierlo, Carlo P. M.; Visser, Antonie J. W. G.; Jan Willem Borst

    2014-01-01

    Uniform labeling of proteins with fluorescent donor and acceptor dyes with an equimolar ratio is paramount for accurate determination of Förster resonance energy transfer (FRET) efficiencies. In practice, however, the labeled protein population contains donor-labeled molecules that have no corresponding acceptor. These FRET-inactive donors contaminate the donor fluorescence signal, which leads to underestimation of FRET efficiencies in conventional fluorescence intensity and lifetime-based F...

  14. Short-pulsed diode lasers as an excitation source for time-resolved fluorescence applications and confocal laser scanning microscopy in PDT

    Science.gov (United States)

    Kress, Matthias; Meier, Thomas H.; El-Tayeb, Tarek A. A.; Kemkemer, Ralf; Steiner, Rudolf W.; Rueck, Angelika C.

    2001-11-01

    This article describes a setup for subcellular time-resolved fluorescence spectroscopy and fluorescence lifetime measurements using a confocal laser scanning microscope in combination with a short pulsed diode laser for fluorescence excitation and specimen illumination. The diode laser emits pulses at 398 nm wavelength with 70 ps full width at half maximum (FWHM) duration. The diode laser can be run at a pulse repetition rate of 40 MHz down to single shot mode. For time resolved spectroscopy a spectrometer setup consisting of an Czerny Turner spectrometer and a MCP-gated and -intensified CCD camera was used. Subcellular fluorescence lifetime measurements were achieved using a time-correlated single photon counting (TCSPC) module instead of the spectrometer setup. The capability of the short pulsed diode laser for fluorescence imaging, fluorescence lifetime measurements and time-resolved spectroscopy in combination with laser scanning microscopy is demonstrated by fluorescence analysis of several photosensitizers on a single cell level.

  15. 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.

  16. Photonic mode density effects on single-molecule fluorescence blinking

    CERN Document Server

    Stefani, F D; Bocchio, N; Gaul, F; Pomozzi, A; Kreiter, M

    2006-01-01

    We investigated the influence of the photonic mode density (PMD) on the triplet dynamics of individual chromophores on a dielectric interface by comparing their response in the presence and absence of a nearby gold film. Lifetimes of the excited singlet state were evaluated in ordet to measure directly the PMD at the molecules position. Triplet state lifetimes were simultaneously determined by statistical analysis of the detection time of the fluorescence photons. The observed singlet decay rates are in agreement with the predicted PMD for molecules with different orientations. The triplet decay rate is modified in a fashion correlated to the singlet decay rate. These results show that PMD engineering can lead to an important suppression of the fluorescence, introducing a novel aspect of the physical mechanism to enhance fluorescence intensity in PMD-enhancing systems such as plasmonic devices.

  17. Quenching of chlorophyll fluorescence induced by silver nanoparticles

    Science.gov (United States)

    Queiroz, A. M.; Mezacasa, A. V.; Graciano, D. E.; Falco, W. F.; M'Peko, J.-C.; Guimarães, F. E. G.; Lawson, T.; Colbeck, I.; Oliveira, S. L.; Caires, A. R. L.

    2016-11-01

    The interaction between chlorophyll (Chl) and silver nanoparticles (AgNPs) was evaluated by analyzing the optical behavior of Chl molecules surrounded by different concentrations of AgNPs (10, 60, and 100 nm of diameter). UV-Vis absorption, steady state and time-resolved fluorescence measurements were performed for Chl in the presence and absence of these nanoparticles. AgNPs strongly suppressed the Chl fluorescence intensity at 678 nm. The Stern-Volmer constant (KSV) showed that fluorescence suppression is driven by the dynamic quenching process. In particular, KSV was nanoparticle size-dependent with an exponential decrease as a function of the nanoparticle diameter. Finally, changes in the Chl fluorescence lifetime in the presence of nanoparticles demonstrated that the fluorescence quenching may be induced by the excited electron transfer from the Chl molecules to the metal nanoparticles.

  18. Photoluminescence lifetime measurements in InP wafers

    Science.gov (United States)

    Landis, Geoffrey A.; Jenkins, Phillip; Weinberg, Irving

    1991-01-01

    A simple apparatus to measure the minority carrier lifetime in InP has been developed. The technique stimulates the sample with a short pulse of light from a diode laser and measures the photoluminescence decay to extract the minority carrier lifetime. The photoluminescence lifetime in InP as a function of doping on both n- and p-type material is examined. The results also show a marked difference in the lifetime of n-type InP and p-type InP of similar doping levels. N-type InP shows a lifetime considerably longer than the expected radiative limited lifetime.

  19. Two-photon autofluorescence lifetime and SHG imaging of healthy and diseased human corneas

    Science.gov (United States)

    Batista, Ana; Breunig, Hans Georg; Uchugonova, Aisada; Seitz, Berthold; Morgado, António Miguel; König, Karsten

    2015-03-01

    Corneal function can be drastically affected by several degenerations and dystrophies, leading to blindness. Early diagnosis of corneal disease is of major importance and it may be accomplished by monitoring changes of the metabolic state and structural organization, the first detectable pathological signs, by two-photon excitation autofluorescence lifetime and second-harmonic generation imaging. In this study, we propose to use these imaging techniques to differentiate between healthy and pathological corneas. Images were acquired using a laser-scanning microscope with a broadband sub-15 femtosecond near-infrared pulsed laser and a 16-channel photomultiplier tube detector for signal collection. This setup allows the simultaneous excitation of metabolic co-factors and to identify them based on their fluorescence spectra. We were able to discriminate between healthy and pathological corneas using two-photon excitation autofluorescence lifetime and second-harmonic generation imaging from corneal epithelium and stroma. Furthermore, differences between different pathologies were observed. Alterations in the metabolic state of corneal epithelial cells were observed using the autofluorescence lifetime of the metabolic co-factors. In the corneal stroma, we observed not only alterations in the collagen fibril structural organization but also alterations in the autofluorescence lifetime. Further tests are required as the number of pathological samples must be increased. In the future, we intend to establish a correlation between the metabolic and structural changes and the disease stage. This can be a step forward in achieving early diagnosis.

  20. Metabolism of HeLa cells revealed through autofluorescence lifetime upon infection with enterohemorrhagic Escherichia coli

    Science.gov (United States)

    Buryakina, Tatyana Yu.; Su, Pin-Tzu; Syu, Wan-Jr; Allen Chang, C.; Fan, Hsiu-Fang; Kao, Fu-Jen

    2012-10-01

    Fluorescence lifetime imaging microscopy (FLIM) is a sensitive technique in monitoring functional and conformational states of nicotinamide adenine dinucleotide reduced (NADH) and flavin adenine dinucleotide (FAD),main compounds participating in oxidative phosphorylation in cells. In this study, we have applied FLIM to characterize the metabolic changes in HeLa cells upon bacterial infection and made comparison with the results from the cells treated with staurosporine (STS), a well-known apoptosis inducer. The evolving of NADH's average autofluorescence lifetime during the 3 h after infection with enterohemorragic Escherichia coli (EHEC) or STS treatment has been observed. The ratio of the short and the long lifetime components' relative contributions of NADH increases with time, a fact indicating cellular metabolic activity, such as a decrease of oxidative phosphorylation over the course of infection, while opposite dynamics is observed in FAD. Being associated with mitochondria, FAD lifetimes and redox ratio could indicate heterogeneous mitochondrial function, microenvironment with bacterial infection, and further pathway to cell death. The redox ratios for both EHEC-infected and STS-treated HeLa cells have been observed and these observations also indicate possible apoptosis induced by bacterial infection.