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Sample records for antibody-based time-resolved fluorescence

  1. Time-resolved fluorescence microscopy.

    PubMed

    Suhling, Klaus; French, Paul M W; Phillips, David

    2005-01-01

    In fluorescence microscopy, the fluorescence emission can be characterised not only by intensity and position, but also by lifetime, polarization and wavelength. Fluorescence lifetime imaging (FLIM) can report on photophysical events that are difficult or impossible to observe by fluorescence intensity imaging, and time-resolved fluorescence anisotropy imaging (TR-FAIM) can measure the rotational mobility of a fluorophore in its environment. We compare different FLIM methods: a chief advantage of wide-field time-gating and phase modulation methods is the speed of acquisition whereas for time-correlated single photon counting (TCSPC) based confocal scanning it is accuracy in the fluorescence decay. FLIM has been used to image interactions between proteins such as receptor oligomerisation and to reveal protein phosphorylation by detecting fluorescence resonance energy transfer (FRET). In addition, FLIM can also probe the local environment of fluorophores, reporting, for example, on the local pH, refractive index, ion or oxygen concentration without the need for ratiometric measurements.

  2. Time-Resolved Fluorescence Assays.

    PubMed

    Ma, Chen-Ting; Sergienko, Eduard A

    2016-01-01

    Fluorescence-based detection techniques are popular in high throughput screening due to sensitivity and cost-effectiveness. Four commonly used techniques exist, each with distinct characteristics. Fluorescence intensity assays are the simplest to run, but suffer the most from signal interference. Fluorescence polarization assays show less interference from the compounds or the instrument, but require a design that results in change of fluorophore-containing moiety size and usually have narrow assay signal window. Fluorescence resonance energy transfer (FRET) is commonly used for detecting protein-protein interactions and is constrained not by the sizes of binding partners, but rather by the distance between fluorophores. Time-resolved fluorescence resonance energy transfer (TR-FRET), an advanced modification of FRET approach utilizes special fluorophores with long-lived fluorescence and earns its place near the top of fluorescent techniques list by its performance and robustness, characterized by larger assay window and minimized compound spectral interference. TR-FRET technology can be applied in biochemical or cell-based in vitro assays with ease. It is commonly used to detect modulation of protein-protein interactions and in detection of products of biochemical reactions and cellular activities. PMID:27316992

  3. Time-resolved fluorescence anisotropy imaging.

    PubMed

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

    2014-01-01

    Fluorescence can be characterized by its intensity, position, wavelength, lifetime, and polarization. The more of these features are acquired in a single measurement, the more can be learned about the sample, i.e., the microenvironment of the fluorescence probe. Polarization-resolved fluorescence lifetime imaging-time-resolved fluorescence anisotropy imaging, TR-FAIM-allows mapping of viscosity or binding or of homo-FRET which can indicate dimerization or generally oligomerization.

  4. Time-resolved fluorescence: 1996-1998

    PubMed

    Kricka; Stanley

    1999-01-01

    Luminescence continues to provide comprehensive literature surveys which will be published in most issues. These are a continuation of the literature surveys begun in 1986 in the Journal of Bioluminescence and Chemiluminescence which, up until 1998, encompassed more than 6000 references cited by year or specialized topic. With this newly named journal these searches are expanding to reflect the journal's wider scope. In future we will cover all fundamental and applied aspects of biological and chemical luminescence and include not only bioluminescence and chemiluminescence but also fluorescence, time resolved fluorescence, electrochemiluminescence, phosphorescence, sonoluminescence, lyoluminescence and triboluminescence. The compilers would be pleased to receive any comments from the readership. Copyright 1999 John Wiley & Sons, Ltd. PMID:10398560

  5. Time-resolved fluorescence decay measurements for flowing particles

    DOEpatents

    Deka, Chiranjit; Steinkamp, John A.

    1999-01-01

    Time-resolved fluorescence decay measurements for flowing particles. An apparatus and method for the measurement and analysis of fluorescence for individual cells and particles in flow are described, wherein the rapid measurement capabilities of flow cytometry and the robust measurement and analysis procedures of time-domain fluorescence lifetime spectroscopy are combined. A pulse-modulated cw laser is employed for excitation of the particles. The characteristics and the repetition rate of the excitation pulses can be readily adjusted to accommodate for fluorescence decays having a wide range of lifetimes.

  6. Time-resolved fluorescence decay measurements for flowing particles

    DOEpatents

    Deka, C.; Steinkamp, J.A.

    1999-06-01

    Time-resolved fluorescence decay measurements are disclosed for flowing particles. An apparatus and method for the measurement and analysis of fluorescence for individual cells and particles in flow are described, wherein the rapid measurement capabilities of flow cytometry and the robust measurement and analysis procedures of time-domain fluorescence lifetime spectroscopy are combined. A pulse-modulated CW laser is employed for excitation of the particles. The characteristics and the repetition rate of the excitation pulses can be readily adjusted to accommodate for fluorescence decays having a wide range of lifetimes. 12 figs.

  7. Time resolved imaging microscopy. Phosphorescence and delayed fluorescence imaging.

    PubMed Central

    Marriott, G; Clegg, R M; Arndt-Jovin, D J; Jovin, T M

    1991-01-01

    An optical microscope capable of measuring time resolved luminescence (phosphorescence and delayed fluorescence) images has been developed. The technique employs two phase-locked mechanical choppers and a slow-scan scientific CCD camera attached to a normal fluorescence microscope. The sample is illuminated by a periodic train of light pulses and the image is recorded within a defined time interval after the end of each excitation period. The time resolution discriminates completely against light scattering, reflection, autofluorescence, and extraneous prompt fluorescence, which ordinarily decrease contrast in normal fluorescence microscopy measurements. Time resolved image microscopy produces a high contrast image and particular structures can be emphasized by displaying a new parameter, the ratio of the phosphorescence to fluorescence. Objects differing in luminescence decay rates are easily resolved. The lifetime of the long lived luminescence can be measured at each pixel of the microscope image by analyzing a series of images that differ by a variable time delay. The distribution of luminescence decay rates is displayed directly as an image. Several examples demonstrate the utility of the instrument and the complementarity it offers to conventional fluorescence microscopy. Images FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 5 FIGURE 6 PMID:1723311

  8. Time-resolved fluorescence spectroscopy of spinach chloroplast.

    PubMed

    Yu, W; Pellegrino, F; Alfano, R R

    1977-04-11

    Picosecond fluorescent kinetics and time-resolved spectra of spinach chloroplast were measured at room temperature and low temperatures. The measurement is conducted with 530 nm excitation at an average intensity of 2-10(14) photons/cm2, pluse and at a pulse separation of 6 ns for the 100 pulses used. The 685 nm fluorescent kinetics was found to decay with two components, a fast component with a 56 ps lifetime, and a slow component with a 220 ps lifetime. The 730 nm fluorescent kinetics at room temperature is a single exponential decay with a 100 ps lifetime. The 730 nm fluorescence lifetime was found to increase by a factor of 6 when the temperature was lowered from room temperature to 90 K, while the 685 and 695 nm fluorescent kinetics were unchanged. The time-resolved spectra data obtained within 10 ps after excitation is consistent with the kinetic data reported here. A two-level fluorescence scheme is proposed to explain the kinetics. The effect of excitation with high light intensity and multiple pulses is discussed.

  9. Time-resolved fluorescence lifetime for cutaneous melanoma detection

    PubMed Central

    Pires, Layla; Nogueira, Marcelo Saito; Pratavieira, Sebastião; Moriyama, Lilian Tan; Kurachi, Cristina

    2014-01-01

    Melanoma is the most aggressive skin cancer type. It is characterized by pigmented lesions with high tissue invasion and metastatic potential. The early detection of melanoma is extremely important to improve patient prognosis and survival rate, since it can progress to the deadly metastatic stage. Presently, the melanoma diagnosis is based on the clinical analysis of the macroscopic lesion characteristics such as shape, color, borders following the ABCD rules. The aim of this study is to evaluate the time-resolved fluorescence lifetime of NADH and FAD molecules to detect cutaneous melanoma in an experimental in vivo model. Forty-two lesions were analyzed and the data was classified using linear discriminant analysis, a sensitivity of 99.4%, specificity of 97.4% and accuracy of 98.4% were achieved. These results show the potential of this fluorescence spectroscopy for melanoma detection. PMID:25401022

  10. Time-Resolved Synchronous Fluorescence for Biomedical Diagnosis

    PubMed Central

    Zhang, Xiaofeng; Fales, Andrew; Vo-Dinh, Tuan

    2015-01-01

    This article presents our most recent advances in synchronous fluorescence (SF) methodology for biomedical diagnostics. The SF method is characterized by simultaneously scanning both the excitation and emission wavelengths while keeping a constant wavelength interval between them. Compared to conventional fluorescence spectroscopy, the SF method simplifies the emission spectrum while enabling greater selectivity, and has been successfully used to detect subtle differences in the fluorescence emission signatures of biochemical species in cells and tissues. The SF method can be used in imaging to analyze dysplastic cells in vitro and tissue in vivo. Based on the SF method, here we demonstrate the feasibility of a time-resolved synchronous fluorescence (TRSF) method, which incorporates the intrinsic fluorescent decay characteristics of the fluorophores. Our prototype TRSF system has clearly shown its advantage in spectro-temporal separation of the fluorophores that were otherwise difficult to spectrally separate in SF spectroscopy. We envision that our previously-tested SF imaging and the newly-developed TRSF methods will combine their proven diagnostic potentials in cancer diagnosis to further improve the efficacy of SF-based biomedical diagnostics. PMID:26404289

  11. Time-resolved fluorescence anisotropies in mixed surfactant solutions

    SciTech Connect

    McCarroll, M.E.; Joly, A.G.; Wang, Z.; Friedrich, D.M.; Wandruszka, R. von

    1999-10-01

    Time-resolved fluorescence anisotropy decays of solutions of Triton X-114 (TX-114) with various amounts of sodium dodecyl sulfate (SDS) were measured using the emission both from the surfactant itself and from added perylene. In the former case, the monomer and aggregate species of the surfactant were spectroscopically isolated and were shown to have significantly different rotational correlation times. The rotational diffusion of perylene in micellar TX-114 with small amounts of added SDS appeared to have a component with a very short correlation time. The anisotropy decay curves showed the existence of limiting anisotropies (r{sub {infinity}}), indicating hindered probe rotation in the micellar environment. At higher SDS concentrations, the fast-decaying component slowed down and the limiting anisotropy decreased substantially, suggesting some migration of the probe to the interior of the micelle.

  12. Picosecond time-resolved fluorescence spectroscopy of phytochrome and stentorin

    NASA Astrophysics Data System (ADS)

    Song, Pill-Soon

    1991-05-01

    Phytochrome is a tetrapyrrole chromoprotein. It serves as a sensitive photosensor for red lightmediated gene expression and other developmental/morphological responses in plants. In this paper photochemical dynamics of the phytochrome molecule have been described in terms of photoisomerization of the tetrapyrrole chromophore in its singlet excited state and subsequent thermal processes in the Pr Pfr phototransformation of phytochrome. Stentorin acts as the photosensor molecule in the ciliate Stentor coeruleus. This unicellular protozoan is most sensitive to red light (610-620 urn). Stentor also senses the direction of light propagation as evidenced by their light-avoiding and negative phototactic swimming behaviors. This aneural photosensory phenomenon is triggered by the photoreceptor stentorin. The possible involvement of a light-induced transient proton release from the photoreceptor as the primary mechanism of light-signal processing has been discussed on the basis of picosecond fluorescence decays and time-resolved fluorescence spectra of stentorin in solution. An initial sensory signal generated by the primary photoprocess of stentorin then triggers subsequent transduction steps that include calcium ion influx from the extracellular medium. Calcium ion influx from the extracellular medium to the cytosol causes the Stentor cell to reverse its ciliary beating and subsequently steer away from the light trap. II.

  13. Thermally activated delayed fluorescence of fluorescein derivative for time-resolved and confocal fluorescence imaging.

    PubMed

    Xiong, Xiaoqing; Song, Fengling; Wang, Jingyun; Zhang, Yukang; Xue, Yingying; Sun, Liangliang; Jiang, Na; Gao, Pan; Tian, Lu; Peng, Xiaojun

    2014-07-01

    Compared with fluorescence imaging utilizing fluorophores whose lifetimes are in the order of nanoseconds, time-resolved fluorescence microscopy has more advantages in monitoring target fluorescence. In this work, compound DCF-MPYM, which is based on a fluorescein derivative, showed long-lived luminescence (22.11 μs in deaerated ethanol) and was used in time-resolved fluorescence imaging in living cells. Both nanosecond time-resolved transient difference absorption spectra and time-correlated single-photon counting (TCSPC) were employed to explain the long lifetime of the compound, which is rare in pure organic fluorophores without rare earth metals and heavy atoms. A mechanism of thermally activated delayed fluorescence (TADF) that considers the long wavelength fluorescence, large Stokes shift, and long-lived triplet state of DCF-MPYM was proposed. The energy gap (ΔEST) of DCF-MPYM between the singlet and triplet state was determined to be 28.36 meV by the decay rate of DF as a function of temperature. The ΔE(ST) was small enough to allow efficient intersystem crossing (ISC) and reverse ISC, leading to efficient TADF at room temperature. The straightforward synthesis of DCF-MPYM and wide availability of its starting materials contribute to the excellent potential of the compound to replace luminescent lanthanide complexes in future time-resolved imaging technologies.

  14. Time-resolved fluorescence study of all-trans-retinal

    NASA Astrophysics Data System (ADS)

    Erez, Yuval; Presiado, Itay; Gepshtein, Rinat; Simkovitch, Ron; Huppert, Dan

    2014-11-01

    UV-vis steady-state and time-resolved emission techniques were employed to study the ultrafast relaxation path of all-trans-retinal. We found that the steady-state emission spectrum consists mainly of two bands that we assign to the allowed transition from the ? state and the forbidden transition from the ?(ππ*) state. The time-resolved emission signal is dependent on the excitation wavelength, and is composed of three decay components. The short-time component of less than 80 fs, irrespective of the solvent, is assigned to the transition from the ? state. The intermediate-time decay component is assigned to the transition from the ?(ππ*) state, depends on the solvent's polarity and not on the existence of hydrogen bonds between the solute and the solvent or the viscosity of the latter. It has a lifetime of ~1 ps in polar solvents, and of 0.6 and 0.4 ps in the non-polar solvents n-octane and cyclohexane, respectively.

  15. Polar plot representation of time-resolved fluorescence.

    PubMed

    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. Monitoring tissue metabolism via time-resolved laser fluorescence

    NASA Astrophysics Data System (ADS)

    Maerz, Holger K.; Buchholz, Rainer; Emmrich, Frank; Fink, Frank; Geddes, Clive L.; Pfeifer, Lutz; Raabe, Ferdinand; Marx, Uwe

    1999-05-01

    Most assays for drug screening are monitoring the metabolism of cells by detecting the NADH content, which symbolize its metabolic activity, indirectly. Nowadays, the performance of a LASER enables us to monitor the metabolic state of mammalian cells directly and on-line by using time-resolved autofluorescence detection. Therefore, we developed in combination with tissue engineering, an assay for monitoring minor toxic effects of volatile organic compounds (VOC), which are accused of inducing Sick Building Syndrome (SBS). Furthermore, we used the Laserfluoroscope (LF) for pharmacological studies on human bone marrow in vitro with special interest in chemotherapy simulation. In cancer research and therapy, the effect of chemostatica in vitro in the so-called oncobiogram is being tested; up to now without great success. However, it showed among other things that tissue structure plays a vital role. Consequently, we succeeded in simulating a chemotherapy in vitro on human bone marrow. Furthermore, after tumor ektomy we were able to distinguish between tumoric and its surrounding healthy tissue by using the LF. With its sensitive detection of metabolic changes in tissues the LF enables a wide range of applications in biotechnology, e.g. for quality control in artificial organ engineering or biocompatability testing.

  17. Time-resolved fluorescence spectroscopy for chemical sensors

    NASA Astrophysics Data System (ADS)

    Draxler, Sonja; Lippitsch, Max E.

    1996-07-01

    A family of sensors is presented with fluorescence decay-time measurements used as the sensing technique. The concept is to take a single fluorophore with a suitably long fluorescence decay time as the basic building block for numerous different sensors. Analyte recognition can be performed by different functional groups that are necessary for selective interaction with the analyte. To achieve this, the principle of excited-state electron transfer is applied with pyrene as the fluorophore. Therefore the same instrumentation based on a small, ambient air-nitrogen laser and solid-state electronics can be used to measure different analytes, for example, oxygen, pH, carbon dioxide, potassium, ammonium, lead, cadmium, zinc, and phosphate.

  18. Determining biomolecular structures by time-resolved fluorescence

    NASA Astrophysics Data System (ADS)

    Rolinski, Olaf J.; Hernandez-Santana, Aaron; Graham, Duncan

    2006-09-01

    We demonstrate a new fluorescence resonance energy transfer (FRET) based approach to determine the donor-acceptor distributions and apply it to two model molecular systems: double stranded DNA labeled with Hoechst 33258 and FAM, and perylene randomly surrounded by cobalt ions in a bulk solution. The approach makes some generic assumptions regarding the FRET kinetics, but no a priori assumptions regarding the distribution function.

  19. Excitation emission and time-resolved fluorescence spectroscopy of selected varnishes used in historical musical instruments.

    PubMed

    Nevin, Austin; Echard, Jean-Philippe; Thoury, Mathieu; Comelli, Daniela; Valentini, Gianluca; Cubeddu, Rinaldo

    2009-11-15

    The analysis of various varnishes from different origins, which are commonly found on historical musical instruments was carried out for the first time with both fluorescence excitation emission spectroscopy and laser-induced time-resolved fluorescence spectroscopy. Samples studied include varnishes prepared using shellac, and selected diterpenoid and triterpenoid resins from plants, and mixtures of these materials. Fluorescence excitation emission spectra have been collected from films of naturally aged varnishes. In parallel, time-resolved fluorescence spectroscopy of varnishes provides means for discriminating between short- (less than 2.0 ns) and long-lived (greater than 7.5 ns) fluorescence emissions in each of these complex materials. Results suggest that complementary use of the two non destructive techniques allows a better understanding of the main fluorophores responsible for the emission in shellac, and further provides means for distinguishing the main classes of other varnishes based on differences in fluorescence lifetime behaviour. Spectrofluorimetric data and time resolved spectra presented here may form the basis for the interpretation of results from future in situ fluorescence examination and time resolved fluorescence imaging of varnished musical instruments.

  20. Time-resolved laser-induced fluorescence study on dyes used in DNA sequencing

    SciTech Connect

    Chang, Kaisyang; Force, R.K. )

    1993-01-01

    Research on the time-resolved fluorescence of fluorescein isothiocyanate, NBD, tetramethylrhodamine isothiocyanate, and Texas Red - the dyes used for fluorescence-based DNA sequencing - is described. Mean fluorescence lifetiems in both aqueous buffer solution and 5.3%T, 4.8%C polyacrylamide gel were determined as a function of excitation wave-lengths at 337, 470, and 550 nm and were found to be 3.5, 1.1, 2.5, and 4.3 ns; the detection limits are 10, 200, 200 and 200 amol for FITC, NBD, TEMR, and T. Red, respectively. Comparisons of fluorescence parameters between the conjugated dyes and the free dyes are also reported. Results on the optimization of the excitation source wavelengths to improve sensitivity and reduce background scattering in polyacrylamide gel are also reported. Time-resolved fluorescence was successfully applied to resolve spectral overlapping of emissions in both solution and in polyacrylamide gel. 12 refs., 6 figs., 1 tab.

  1. Classification of aortic atherosclerotic lesions with time-resolved fluorescence spectroscopy

    NASA Astrophysics Data System (ADS)

    Maarek, Jean-Michel I.; Marcu, Laura; Grundfest, Warren S.; Fishbein, Michael C.

    1999-07-01

    In this study, we examine the possibility of differentiating between classes of atherosclerotic lesions based on time- resolved fluorescence spectroscopy and we compare the performance of classification schemes that use either the time-resolved spectra or only the intensity spectra. Transient fluorescence emissions induced by pulsed nitrogen laser excitation was measured on 87 excised samples of human aorta. The samples were classified histologically using the AHA classification Predictor variables derived from the time-resolved spectra included the spectral intensities at 360-510 nm and parameters of a biexponential fit of the fluorescence impulse response function. Stepwise discriminant analysis using these predict variables showed that a few predictor variables sufficed to correctly classify 89 percent of the samples. Excluding the time- dependent decay and using only the spectral intensities, the percentage of correctly classified cases was significantly lower: 51 percent. These results establish that time- resolved fluorescence spectroscopy markedly improved on the performance of steady-state fluorescence spectroscopy for fine classification of atherosclerotic lesions.

  2. Plastique: A synchrotron radiation beamline for time resolved fluorescence in the frequency domain

    NASA Astrophysics Data System (ADS)

    De Stasio, Gelsomina; Zema, N.; Antonangeli, F.; Savoia, A.; Parasassi, T.; Rosato, N.

    1991-06-01

    PLASTIQUE is the only synchrotron radiation beamline in the world that performs time resolved fluorescence experiments in frequency domain. These experiments are extremely valuable sources of information on the structure and dynamics of molecules. We describe the beamline and some initial data.

  3. Validation of a time-resolved fluorescence spectroscopy apparatus in a rabbit atherosclerosis model

    NASA Astrophysics Data System (ADS)

    Fang, Qiyin; Jo, Javier A.; Papaioannou, Thanassis; Dorafshar, Amir; Reil, Todd; Qiao, Jian-Hua; Fishbein, Michael C.; Freischlag, Julie A.; Marcu, Laura

    2004-07-01

    Time-resolved laser-induced fluorescence spectroscopy (tr-LIFS) has been studied as a potential tool for in vivo diagnosis of atherosclerotic lesions. This study is to evaluate the potential of a compact fiber-optics based tr-LIFS instrument developed in our laboratory for in vivo analysis of atherosclerotic plaque composition. Time-resolved fluorescence spectroscopy studies were performed in vivo on fifteen New Zealand White rabbits (atherosclerotic: N=8, control: N=7). Time-resolved fluorescence spectra were acquired (range: 360-600 nm, increment: 5 nm, total acquisition time: 65 s) from normal aorta wall and lesions in the abdominal aorta. Data were analyzed in terms of fluorescence emission spectra and wavelength specific lifetimes. Following trichrome staining, tissue specimens were analyzed histopathologically in terms of intima/media thickness and biochemical composition (collagen, elastin, foam cells, and etc). Based on intimal thickness, the lesions were divided into thin and thick lesions. Each group was further separated into two categories: collagen rich lesions and foam cell rich lesions based on their biochemical composition. The obtained spectral and time domain fluorescence signatures were subsequently correlated to the histopathological findings. The results have shown that time-domain fluorescence spectral features can be used in vivo to separate atherosclerotic lesions from normal aorta wall as well discrimination within certain types of lesions.

  4. Use of Time-Resolved Fluorescence to Monitor Bioactive Compounds in Plant Based Foodstuffs.

    PubMed

    Lemos, M Adília; Sárniková, Katarína; Bot, Francesca; Anese, Monica; Hungerford, Graham

    2015-06-26

    The study of compounds that exhibit antioxidant activity has recently received much interest in the food industry because of their potential health benefits. Most of these compounds are plant based, such as polyphenolics and carotenoids, and there is a need to monitor them from the field through processing and into the body. Ideally, a monitoring technique should be non-invasive with the potential for remote capabilities. The application of the phenomenon of fluorescence has proved to be well suited, as many plant associated compounds exhibit fluorescence. The photophysical behaviour of fluorescent molecules is also highly dependent on their microenvironment, making them suitable probes to monitor changes in pH, viscosity and polarity, for example. Time-resolved fluorescence techniques have recently come to the fore, as they offer the ability to obtain more information, coupled with the fact that the fluorescence lifetime is an absolute measure, while steady state just provides relative and average information. In this work, we will present illustrative time-resolved measurements, rather than a comprehensive review, to show the potential of time-resolved fluorescence applied to the study of bioactive substances. The aim is to help assess if any changes occur in their form, going from extraction via storage and cooking to the interaction with serum albumin, a principal blood transport protein.

  5. Use of Time-Resolved Fluorescence to Monitor Bioactive Compounds in Plant Based Foodstuffs

    PubMed Central

    Lemos, M. Adília; Sárniková, Katarína; Bot, Francesca; Anese, Monica; Hungerford, Graham

    2015-01-01

    The study of compounds that exhibit antioxidant activity has recently received much interest in the food industry because of their potential health benefits. Most of these compounds are plant based, such as polyphenolics and carotenoids, and there is a need to monitor them from the field through processing and into the body. Ideally, a monitoring technique should be non-invasive with the potential for remote capabilities. The application of the phenomenon of fluorescence has proved to be well suited, as many plant associated compounds exhibit fluorescence. The photophysical behaviour of fluorescent molecules is also highly dependent on their microenvironment, making them suitable probes to monitor changes in pH, viscosity and polarity, for example. Time-resolved fluorescence techniques have recently come to the fore, as they offer the ability to obtain more information, coupled with the fact that the fluorescence lifetime is an absolute measure, while steady state just provides relative and average information. In this work, we will present illustrative time-resolved measurements, rather than a comprehensive review, to show the potential of time-resolved fluorescence applied to the study of bioactive substances. The aim is to help assess if any changes occur in their form, going from extraction via storage and cooking to the interaction with serum albumin, a principal blood transport protein. PMID:26132136

  6. Locating and classifying fluorescent tags behind turbid layers using time-resolved inversion.

    PubMed

    Satat, Guy; Heshmat, Barmak; Barsi, Christopher; Raviv, Dan; Chen, Ou; Bawendi, Moungi G; Raskar, Ramesh

    2015-04-13

    The use of fluorescent probes and the recovery of their lifetimes allow for significant advances in many imaging systems, in particular, medical imaging systems. Here we propose and experimentally demonstrate reconstructing the locations and lifetimes of fluorescent markers hidden behind a turbid layer. This opens the door to various applications for non-invasive diagnosis, analysis, flowmetry and inspection. The method is based on a time-resolved measurement that captures information about both fluorescence lifetime and spatial position of the probes. To reconstruct the scene, the method relies on a sparse optimization framework to invert time-resolved measurements. This wide-angle technique does not rely on coherence, and does not require the probes to be directly in line of sight of the camera, making it potentially suitable for long-range imaging.

  7. A fluorescence LIDAR sensor for hyper-spectral time-resolved remote sensing and mapping.

    PubMed

    Palombi, Lorenzo; Alderighi, Daniele; Cecchi, Giovanna; Raimondi, Valentina; Toci, Guido; Lognoli, David

    2013-06-17

    In this work we present a LIDAR sensor devised for the acquisition of time resolved laser induced fluorescence spectra. The gating time for the acquisition of the fluorescence spectra can be sequentially delayed in order to achieve fluorescence data that are resolved both in the spectral and temporal domains. The sensor can provide sub-nanometric spectral resolution and nanosecond time resolution. The sensor has also imaging capabilities by means of a computer-controlled motorized steering mirror featuring a biaxial angular scanning with 200 μradiant angular resolution. The measurement can be repeated for each point of a geometric grid in order to collect a hyper-spectral time-resolved map of an extended target.

  8. A fluorescence LIDAR sensor for hyper-spectral time-resolved remote sensing and mapping.

    PubMed

    Palombi, Lorenzo; Alderighi, Daniele; Cecchi, Giovanna; Raimondi, Valentina; Toci, Guido; Lognoli, David

    2013-06-17

    In this work we present a LIDAR sensor devised for the acquisition of time resolved laser induced fluorescence spectra. The gating time for the acquisition of the fluorescence spectra can be sequentially delayed in order to achieve fluorescence data that are resolved both in the spectral and temporal domains. The sensor can provide sub-nanometric spectral resolution and nanosecond time resolution. The sensor has also imaging capabilities by means of a computer-controlled motorized steering mirror featuring a biaxial angular scanning with 200 μradiant angular resolution. The measurement can be repeated for each point of a geometric grid in order to collect a hyper-spectral time-resolved map of an extended target. PMID:23787661

  9. Tubulin equilibrium unfolding followed by time-resolved fluorescence and fluorescence correlation spectroscopy

    PubMed Central

    Sánchez, Susana A.; Brunet, Juan E.; Jameson, David M.; Lagos, Rosalba; Monasterio, Octavio

    2004-01-01

    The pathway for the in vitro equilibrium unfolding of the tubulin heterodimer by guanidinium chloride (GdmCl) has been studied using several spectroscopic techniques, specifically circular dichroism (CD), two-photon Fluorescence Correlation Spectroscopy (FCS), and time-resolved fluorescence, including lifetime and dynamic polarization. The results show that tubulin unfolding is characterized by distinct processes that occur in different GdmCl concentration ranges. From 0 to 0.5 M GdmCl, a slight alteration of the tubulin heterodimer occurs, as evidenced by a small, but reproducible increase in the rotational correlation time of the protein and a sharp decrease in the secondary structure monitored by CD. In the range 0.5–1.5 M GdmCl, significant decreases in the steady-state anisotropy and average lifetime of the intrinsic tryptophan fluorescence occur, as well as a decrease in the rotational correlation time, from 48 to 26 nsec. In the same GdmCl range, the number of protein molecules (labeled with Alexa 488), as determined by two-photon FCS measurements, increases by a factor of two, indicating dissociation of the tubulin dimer into monomers. From 1.5 to 4 M GdmCl, these monomers unfold, as evidenced by the continual decrease in the tryptophan steady-state anisotropy, average lifetime, and rotational correlation time, concomitant with secondary structural changes. These results help to elucidate the unfolding pathway of the tubulin heterodimer and demonstrate the value of FCS measurements in studies on oligomeric protein systems. PMID:14691224

  10. Halide (Cl(super -)) Quenching of Quinine Sulfate Fluorescence: A Time-Resolved Fluorescence Experiment for Physical Chemistry

    ERIC Educational Resources Information Center

    Gutow, Jonathan H.

    2005-01-01

    The time-resolved fluorescence experiment investigating the halide quenching of fluorescence from quinine sulfate in water is described. The objectives of the experiment include reinforcing student understanding of the kinetics of competing pathways, making connections with microscopic theories of kinetics through comparison of experimental and…

  11. Multiplexed analysis using time-resolved near-IR fluorescence for the detection of genomic material

    NASA Astrophysics Data System (ADS)

    Stryjewski, Wieslaw J.; Soper, Steven A.; Lassiter, Suzzane; Davis, Lloyd M.

    2002-06-01

    While fluorescence continues to be an important tool in genomics, new challenges are being encountered due to increased efforts toward miniaturization reducing detection volumes and the need for screening multiple targets simultaneously. We have initiated work on developing time- resolved near-IR fluorescence as an additional tool for the multiplexed analyses of DNA, either for sequencing or mutation detection. We have fabricated simple and compact time-resolved fluorescence microscopes for reading fluorescence from electrophoresis or DNA microarrays. These microscopes consist of solid-state diode lasers and diode detectors and due to their compact size, the optical components and laser head can be mounted on high-speed micro-translational stages to read fluorescence from either multi-channel capillary electrophoresis instruments or micro fabricated DNA sorting devices. The detector is configured in a time-correlated single photon counting format to allow acquisition of fluorescence lifetimes on-the-fly during data acquisition in the limit of low counting statistics. In multiplexed analyses, lifetime discrimination serves as a method for dye-reporter identification using only a single readout channel. Also, coupled to multi-color systems, lifetime identification can significantly increase the number of probes monitored in a single instrument. In this work, near-IR fluorescence, including dye-labels and hardware, will be discussed as well as the implementation of near-IR fluorescence in DNA sequencing using slab gel electrophoresis and DNA microarrays.

  12. A 0.18-µm CMOS Array Sensor for Integrated Time-Resolved Fluorescence Detection

    PubMed Central

    Huang, Ta-chien D.; Sorgenfrei, Sebastian; Gong, Ping; Levicky, Rastislav; Shepard, Kenneth L.

    2010-01-01

    This paper describes the design of an active, integrated CMOS sensor array for fluorescence applications which enables time-gated, time-resolved fluorescence spectroscopy. The 64-by-64 array is sensitive to photon densities as low as 8.8 × 106 photons/cm2 with 64-point averaging and, through a differential pixel design, has a measured impulse response of better than 800 ps. Applications include both active microarrays and high-frame-rate imagers for fluorescence lifetime imaging microscopy. PMID:20436922

  13. CMOS Time-Resolved, Contact, and Multispectral Fluorescence Imaging for DNA Molecular Diagnostics

    PubMed Central

    Guo, Nan; Cheung, Ka Wai; Wong, Hiu Tung; Ho, Derek

    2014-01-01

    Instrumental limitations such as bulkiness and high cost prevent the fluorescence technique from becoming ubiquitous for point-of-care deoxyribonucleic acid (DNA) detection and other in-field molecular diagnostics applications. The complimentary metal-oxide-semiconductor (CMOS) technology, as benefited from process scaling, provides several advanced capabilities such as high integration density, high-resolution signal processing, and low power consumption, enabling sensitive, integrated, and low-cost fluorescence analytical platforms. In this paper, CMOS time-resolved, contact, and multispectral imaging are reviewed. Recently reported CMOS fluorescence analysis microsystem prototypes are surveyed to highlight the present state of the art. PMID:25365460

  14. Feasibility analysis of an epidermal glucose sensor based on time-resolved fluorescence

    NASA Astrophysics Data System (ADS)

    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.

  15. Time-Resolved Fluorescence in Lipid Bilayers: Selected Applications and Advantages over Steady State

    PubMed Central

    Amaro, Mariana; Šachl, Radek; Jurkiewicz, Piotr; Coutinho, Ana; Prieto, Manuel; Hof, Martin

    2014-01-01

    Fluorescence methods are versatile tools for obtaining dynamic and topological information about biomembranes because the molecular interactions taking place in lipid membranes frequently occur on the same timescale as fluorescence emission. The fluorescence intensity decay, in particular, is a powerful reporter of the molecular environment of a fluorophore. The fluorescence lifetime can be sensitive to the local polarity, hydration, viscosity, and/or presence of fluorescence quenchers/energy acceptors within several nanometers of the vicinity of a fluorophore. Illustrative examples of how time-resolved fluorescence measurements can provide more valuable and detailed information about a system than the time-integrated (steady-state) approach will be presented in this review: 1), determination of membrane polarity and mobility using time-dependent spectral shifts; 2), identification of submicroscopic domains by fluorescence lifetime imaging microscopy; 3), elucidation of membrane leakage mechanisms from dye self-quenching assays; and 4), evaluation of nanodomain sizes by time-resolved Förster resonance energy transfer measurements. PMID:25517142

  16. BHHST: An improved lanthanide chelate for time-resolved fluorescence applications

    NASA Astrophysics Data System (ADS)

    Connally, Russell; Jin, Dayong; Piper, James

    2005-04-01

    The detection of the waterborne pathogens Giardia lamblia and Cryptosporidium parvum in environmental water bodies requires concentration of large volumes of water due to the low dose required for infection. The highly concentrated (10,000-fold) water sample is often rich in strongly autofluorescent algae, organic debris and mineral particles that can obscure immunofluorescently labeled (oo)cysts during analysis. Time-resolved fluorescence techniques exploit the long fluorescence lifetimes of lanthanide chelates (ms) to differentiate target fluorescence from background autofluorescence (ns). Relatively simple instrumentation can be used to enhance the signal-to-noise ratio (S/N) of labelled target. Time-resolved fluorescence techniques exploit the large difference in lifetime by briefly exciting fluorescence from the sample using a pulsed excitation source. Capture of the resulting fluorescence emission is delayed until the more rapidly decaying autofluorescence has faded beyond detection, whereon the much stronger and slower fading emission from labelled target is collected. BHHCT is a tetradentate beta-diketone chelate that is activated to bind with protein (antibody) as the chlorosulfonate. The high activity of this residue makes conjugations difficult to control and can lead to the formation of unstable immunoconjugates. To overcome these limitations a 5-atom hydrophylic molecular tether was attached to BHHCT via the chlorosulfonate and the BHHCT derivative was then activated to bind to proteins as the succinimide. The new compound (BHHST) could be prepared in high purity and was far more stable than the chlorosulfonate on storage. A high activity immunocojugate was prepared against Cryptosporidium that yielded an 8-fold increase in SNR using a lab-built time-resolved fluorescence microscope.

  17. Time-resolvable fluorescent conjugates for the detection of pathogens in environmental samples containing autofluorescent material

    NASA Astrophysics Data System (ADS)

    Connally, Russell; Veal, Duncan; Piper, James A.

    2003-07-01

    Water is routinely monitored for environmental pathogens such a Cryptosporidium and Giardia using immunofluorescence microscopy (IFM). Autofluorescence can greatly diminish an operators capacity to resolve labeled pathogens from non-specific background. Naturally fluorescing components (autofluorophores) encountered in biological samples typically have fluorescent lifetimes (τ) of less than 100 nanoseconds and their emissions may be excluded through use of time-resolved fluorescence microscopy (TRFM). TRFM relies on the large differences in τ between autofluorescent molecules and long-lived lanthanide chelates. In TRFM, targets labeled with a time-resolvable fluorescent immunoconjugate are excited by an intense (UV) light pulse. A short delay is imposed to permit the decay of autofluorescence before capture of luminescence from the excited chelate using an image intensified CCD camera. In our experience, autofluorescence can be reduced to insignificant levels with a consequent 30-fold increase in target visibility using TRFM techniques. We report conjugation of a novel europium chelate to a monoclonal antibody specific for Giardia lamblia and use of the immunoconjugate for TRFM studies. Initial attempts to conjugate the same chelate to a monoclonal antibody directed against Cryptosporidium parvum led to poorly fluorescent constructs that were prone to denature and precipitate. We successfully conjugated BHHCT to anti-mouse polyvalent immunoglobulin and used this construct to overcome the difficulties in direct labeling of the anti-Cryptosporidium antibody. Both Giardia and Cryptosporidium were labeled using the anti-mouse protocol with a subsequent 20-fold and 6.6-fold suppression of autofluorescence respectively. A rapid protocol for conjugating and purifying the immunoconjugate was found and methods of quantifying the fluorescence to protein ratio determined. Performance of our TRFM was dependent on the quality and brightness of the immunoconjugate and

  18. Molecular diffusivity measurement through an alumina membrane using time-resolved fluorescence imaging

    NASA Astrophysics Data System (ADS)

    Kennard, Raymond; DeSisto, William J.; Mason, Michael D.

    2010-11-01

    We present a simple fluorescence imaging method for measuring the time-resolved concentration of a fluorescent molecule diffusing through an anodic alumina membrane with a pore diameter of 20 nm. From the concentration breakthrough curve, the molecular diffusivity of the fluorophore was extracted. The experimentally determined diffusivity was three orders of magnitude lower than reported bulk values. Due to the relative simplicity and ease of use, this method can be applied to provide fundamental information for biomolecular separations applications. One feature of this method is the high sensitivity at intercellular volumes broadening its application to drug delivery and controlled cell growth.

  19. Time-resolved and steady-state fluorescence spectroscopy from bacteria subjected to bactericidal agents

    NASA Astrophysics Data System (ADS)

    Katz, Alvin; Alimova, Alexandra; Siddique, Masood; Savage, Howard E.; Shah, Mahendra; Rosen, Richard; Alfano, Robert

    2004-03-01

    The time-resolved and steady-state changes in fluorescence were investigated from one spore-forming (Bacillus subtilis) and four non-spore forming (Escherichia coli, Staphylococcus aureus, Enterococcus faecalis, and Pseudomonas aeruginosa) bacteria subjected to different bactericidal agents. The bactericidal agents were sodium hypochlorite (bleach) hydrogen peroxide, formaldehyde, and UV light exposure. Application of sodium hypochlorite resulted in an almost total lose of fluorescence signal and large decrease in the optical density of the bacterial suspension. Addition of hydrogen peroxide resulted in a 35% decrease in emission intensity fom the Sa and an 85-95% decrease for the other bacteria. Ultraviolet light exposure resulted in a 5-35% decrease in the emission intensity of the tryptophan band. The addition of formaldehyde to the bacteria did not result in significant changes in the steady-state emission intensity, but did shift the tryptophan emission peak position to shorter wavelengths by 3 to 5 nm. Time-resolved fluorescence measurements showed that the fluorescence lifetime of tryptophan in the bacteria could not be described by a single exponential decay, and was similar to that of tryptophan in neutral aqueous solution. Upon addition of formaldehyde to the Gram positive bacteria (Bs and Sa) the strength of the short lifetime component increased dramatically, while for the Gram negative bacteria, a smaller increase was observed. These fluorescence changes reflect the different mechanisms of the bactericidal agents and may provide a useful tool to monitor the effectiveness of disinfectants.

  20. Time-resolved fluorescence monitoring of cholesterol in peripheral blood mononuclear cells

    NASA Astrophysics Data System (ADS)

    Martinakova, Z.; Horilova, J.; Lajdova, I.; Marcek Chorvatova, A.

    2014-12-01

    Precise evaluation of intracellular cholesterol distribution is crucial for improving diagnostics of diseased states associated with cholesterol alteration. Time-resolved fluorescence techniques are tested for non-invasive investigation of cholesterol in living cells. Fluorescent probe NBD attached to cholesterol was employed to evaluate cholesterol distribution in peripheral blood mononuclear cells (PBMC) isolated from the human blood. Fluorescence Lifetime Imaging Microscopy (FLIM) was successfully applied to simultaneously monitor the spatial distribution and the timeresolved characteristics of the NBD-cholesterol fluorescence in PBMC. Gathered data are the first step in the development of a new perspective non-invasive diagnostic method for evaluation of cholesterol modifications in diseases associated with disorders of lipid metabolism.

  1. [Discrimination of Crude Oil Samples Using Laser-Induced Time-Resolved Fluorescence Spectroscopy].

    PubMed

    Han, Xiao-shuang; Liu, De-qing; Luan, Xiao-ning; Guo, Jin-jia; Liu, Yong-xin; Zheng, Rong-er

    2016-02-01

    The Laser-induced fluorescence spectra combined with pattern recognition method has been widely applied in discrimination of different spilled oil, such as diesel, gasoline, and crude oil. However, traditional three-dimension fluorescence analysis method, which is not adapted to requirement of field detection, is limited to laboratory investigatio ns. The development of oil identification method for field detection is significant to quick response and operation of oil spill. In this paper, a new method based on laser-induced time-resolved fluorescence combined with support vector machine (SVM) model was introduced to discriminate crude oil samples. In this method, time-resolved spectra data was descended into two dimensions with selecting appropriate range in time and wavelength domains respectively to form a SVM data base. It is found that the classification accurate rate increased with an appropriate selection. With a selected range from 54 to 74 ns in time domain, the classification accurate rate has been increased from 83.3% (without selection) to 88.1%. With a selected wavelength range of 387.00~608.87 nm, the classification accurate rate of suspect oil was improved from 84% (without selection) to 100%. Since the detection delay of fluorescence lidar fluctuates due to wave and platform swing, the identification method with optimizing in both time and wavelength domains could offer a better flexibility for field applications. It is hoped that the developed method could provide some useful reference with data reduction for classification of suspect crude oil in the future development.

  2. Time-resolved tryptophan fluorescence in photosynthetic reaction centers from Rhodobacter sphaeroides

    NASA Technical Reports Server (NTRS)

    Godik, V. I.; Blankenship, R. E.; Causgrove, T. P.; Woodbury, N.

    1993-01-01

    Tryptophan fluorescence of reaction centers isolated from Rhodobacter sphaeroides, both stationary and time-resolved, was studied. Fluorescence kinetics were found to fit best a sum of four discrete exponential components. Half of the initial amplitude was due to a component with a lifetime of congruent to 60 ps, belonging to Trp residues, capable of efficient transfer of excitation energy to bacteriochlorophyll molecules of the reaction center. The three other components seem to be emitted by Trp ground-state conformers, unable to participate in such a transfer. Under the influence of intense actinic light, photooxidizing the reaction centers, the yield of stationary fluorescence diminished by congruent to 1.5 times, while the number of the kinetic components and their life times remained practically unchanged. Possible implications of the observed effects for the primary photosynthesis events are considered.

  3. Cucurbiturils: molecular nanocapsules for time-resolved fluorescence-based assays.

    PubMed

    Marquez, Cesar; Huang, Fang; Nau, Werner M

    2004-03-01

    A new fluorescent host-guest system based on the inclusion of the fluorophore 2,3-diazabicyclo[2.2.2]oct-2-ene (DBO) into the cavity of the molecular container compound cucurbit[7]uril (CB7) has been designed which possesses an exceedingly long-lived emission (690 ns in aerated water). The large binding constant of (4 +/- 1) x 10(5) M(-1) along with the resistance of the CB7.DBO complex toward external fluorescence quenchers allow the use of CB7 as an enhancer in time-resolved fluorescence-based assays, e.g., to screen enzyme activity or inhibition by using DBO-labeled peptides as substrates. The response of CB7.DBO to different environmental conditions and possible quenchers are described.

  4. Drug/protein interactions studied by time-resolved fluorescence spectroscopy

    NASA Astrophysics Data System (ADS)

    Gustavsson, Thomas; Markovitsi, Dimitra; Vayá, Ignacio; Bonancía, Paula; Jiménez, M. C.; Miranda, Miguel A.

    2014-09-01

    We report here on a recent time-resolved fluorescence study [1] of the interaction between flurbiprofen (FBP), a chiral non-steroidal anti-inflammatory drug, and human serum albumin (HSA), the main transport protein in the human body. We compare the results obtained for the drug-protein complex with those of various covalently linked flurbiprofentryptophan dyads having well-defined geometries. In all cases stereoselective dynamic fluorescence quenching is observed, varying greatly from one system to another. In addition, the fluorescence anisotropy decays also display a clear stereoselectivity. For the drug-protein complexes, this can be interpreted in terms of the protein microenvironment playing a significant role in the conformational relaxation of FBP, which is more restricted in the case of the (R)- enantiomer.

  5. Efficient signal processing for time-resolved fluorescence detection of nitrogen-vacancy spins in diamond

    NASA Astrophysics Data System (ADS)

    Gupta, A.; Hacquebard, L.; Childress, L.

    2016-03-01

    Room-temperature fluorescence detection of the nitrogen-vacancy center electronic spin typically has low signal to noise, requiring long experiments to reveal an averaged signal. Here, we present a simple approach to analysis of time-resolved fluorescence data that permits an improvement in measurement precision through signal processing alone. Applying our technique to experimental data reveals an improvement in signal to noise equivalent to a 14% increase in photon collection efficiency. We further explore the dependence of the signal to noise ratio on excitation power, and analyze our results using a rate equation model. Our results provide a rubric for optimizing fluorescence spin detection, which has direct implications for improving precision of nitrogen-vacancy-based sensors.

  6. Revealing the photophysics of gold-nanobeacons via time-resolved fluorescence spectroscopy.

    PubMed

    Wei, Guoke; Simionesie, Dorin; Sefcik, Jan; Sutter, Jens U; Xue, Qingjiang; Yu, Jun; Wang, Jinliang; Birch, David J S; Chen, Yu

    2015-12-15

    We demonstrate that time-resolved fluorescence spectroscopy is a powerful tool to investigate the conformation states of hairpin DNA on the surface of gold nanoparticles (AuNPs) and energy transfer processes in Au-nanobeacons. Long-range fluorescence quenching of Cy5 by AuNPs has been found to be in good agreement with electrodynamics modeling. Moreover, time-correlated single-photon counting (TCSPC) is shown to be promising for real-time monitoring of the hybridization kinetics of Au-nanobeacons, with up to 60% increase in decay time component and 300% increase in component fluorescence fraction observed. Our results also indicate the importance of the stem and spacer designs for the performance of Au-nanobeacons. PMID:26670500

  7. Advanced Time-Resolved Fluorescence Microscopy Techniques for the Investigation of Peptide Self-Assembly

    NASA Astrophysics Data System (ADS)

    Anthony, Neil R.

    The ubiquitous cross beta sheet peptide motif is implicated in numerous neurodegenerative diseases while at the same time offers remarkable potential for constructing isomorphic high-performance bionanomaterials. Despite an emerging understanding of the complex folding landscape of cross beta structures in determining disease etiology and final structure, we lack knowledge of the critical initial stages of nucleation and growth. In this dissertation, I advance our understanding of these key stages in the cross-beta nucleation and growth pathways using cutting-edge microscopy techniques. In addition, I present a new combined time-resolved fluorescence analysis technique with the potential to advance our current understanding of subtle molecular level interactions that play a pivotal role in peptide self-assembly. Using the central nucleating core of Alzheimer's Amyloid-beta protein, Abeta(16 22), as a model system, utilizing electron, time-resolved, and non-linear microscopy, I capture the initial and transient nucleation stages of peptide assembly into the cross beta motif. In addition, I have characterized the nucleation pathway, from monomer to paracrystalline nanotubes in terms of morphology and fluorescence lifetime, corroborating the predicted desolvation process that occurs prior to cross-beta nucleation. Concurrently, I have identified unique heterogeneous cross beta domains contained within individual nanotube structures, which have potential bionanomaterials applications. Finally, I describe a combined fluorescence theory and analysis technique that dramatically increases the sensitivity of current time-resolved techniques. Together these studies demonstrate the potential for advanced microscopy techniques in the identification and characterization of the cross-beta folding pathway, which will further our understanding of both amyloidogenesis and bionanomaterials.

  8. Time-resolved fluorescence studies of nucleotide flipping by restriction enzymes.

    PubMed

    Neely, Robert K; Tamulaitis, Gintautas; Chen, Kai; Kubala, Marta; Siksnys, Virginijus; Jones, Anita C

    2009-11-01

    Restriction enzymes Ecl18kI, PspGI and EcoRII-C, specific for interrupted 5-bp target sequences, flip the central base pair of these sequences into their protein pockets to facilitate sequence recognition and adjust the DNA cleavage pattern. We have used time-resolved fluorescence spectroscopy of 2-aminopurine-labelled DNA in complex with each of these enzymes in solution to explore the nucleotide flipping mechanism and to obtain a detailed picture of the molecular environment of the extrahelical bases. We also report the first study of the 7-bp cutter, PfoI, whose recognition sequence (T/CCNGGA) overlaps with that of the Ecl18kI-type enzymes, and for which the crystal structure is unknown. The time-resolved fluorescence experiments reveal that PfoI also uses base flipping as part of its DNA recognition mechanism and that the extrahelical bases are captured by PfoI in binding pockets whose structures are quite different to those of the structurally characterized enzymes Ecl18kI, PspGI and EcoRII-C. The fluorescence decay parameters of all the enzyme-DNA complexes are interpreted to provide insight into the mechanisms used by these four restriction enzymes to flip and recognize bases and the relationship between nucleotide flipping and DNA cleavage.

  9. Viability of Saccharomyces cerevisiae incorporated within silica and polysaccharide hosts monitored via time-resolved fluorescence.

    PubMed

    Holmes-Smith, A Sheila; Hollas, Alexis C; McLoskey, David; Hungerford, Graham

    2013-12-01

    The viability of Saccharomyces cerevisiae in biocompatible polymers under different growth conditions and studied using time-resolved fluorescence techniques is presented. Two fluorophores, the viscosity sensitive probe 4-(4-(dimethylamino)styryl)-N-methyl-pyridiniumiodine (DASPMI) and the yeast viability stain 2-chloro-4-(2,3-dihydro-3-methyl-(benzo-1,3-thiazol-2-yl)-methylidene)-1-phenylquinolinium iodide (FUN-1) are used to elucidate information on the incorporated yeast cell viability. Variations in cell viscosity, which are indicative of the cell state, were obtained using DASPMI. Prior to observing FUN-1 in yeast cells using fluorescence lifetime imaging, its photophysics in solution and heterogeneous media were investigated. Time-resolved emission spectra were measured and analysed to associate lifetimes to the spectral emission. Preliminary results show that monitoring the fluorescence lifetime of FUN-1 may give a useful insight into cellular metabolism. The results indicate that both fluorophores may be used to monitor the entrapped yeast cell viability, which is important for in vitro studies and applications, such as that in the biofuel industry, where Saccharomyces cerevisiae are required to remain active in high ethanol environments.

  10. Use of time-resolved fluorescence spectroscopy to evaluate diagnostic value of collagen degradation products.

    PubMed

    Sikora, Joanna; Cyrankiewicz, Michał; Wybranowski, Tomasz; Ziomkowska, Blanka; Ośmiałowski, Borys; Obońska, Ewa; Augustyńska, Beata; Kruszewski, Stefan; Kubica, Jacek

    2015-05-01

    The concentration of collagen degradation products (CDPs) may reflect the process of left ventricular remodeling (LVR). The aim of this study was to evaluate the potential diagnostic usefulness of time-resolved fluorescence spectroscopy (TRFS) in assessment of CDPs. The preliminary experiment was designed to establish if CDPs’ characteristics might be visible by mean fluorescence lifetime (FLT) in determined conditions. The in vitro model of CDPs was prepared by conducting the hydrolysis of type III collagen. The FLT of samples was measured by the time-resolved spectrometer Life Spec II with the subnanosecond pulsed 360-nm EPLED diode. The FLTs were obtained by deconvolution analysis of the data using a multiexponential model of fluorescence decay. In order to determine the limit of traceability of CDPs, a comparison of different collagen/plasma ratio in samples was performed. The results of our study showed that the increase of added plasma to hydrolyzed collagen extended the mean FLT. Thus, the diagnosis of LVR based on measurements using TRFS is possible. However, it is important to point out the experiment was preliminary and further investigation in this field of research is crucial. PMID:25764396

  11. Use of time-resolved fluorescence spectroscopy to evaluate diagnostic value of collagen degradation products

    NASA Astrophysics Data System (ADS)

    Sikora, Joanna; Cyrankiewicz, Michał; Wybranowski, Tomasz; Ziomkowska, Blanka; Ośmiałowski, Borys; Obońska, Ewa; Augustyńska, Beata; Kruszewski, Stefan; Kubica, Jacek

    2015-05-01

    The concentration of collagen degradation products (CDPs) may reflect the process of left ventricular remodeling (LVR). The aim of this study was to evaluate the potential diagnostic usefulness of time-resolved fluorescence spectroscopy (TRFS) in assessment of CDPs. The preliminary experiment was designed to establish if CDPs' characteristics might be visible by mean fluorescence lifetime (FLT) in determined conditions. The in vitro model of CDPs was prepared by conducting the hydrolysis of type III collagen. The FLT of samples was measured by the time-resolved spectrometer Life Spec II with the subnanosecond pulsed 360-nm EPLED diode. The FLTs were obtained by deconvolution analysis of the data using a multiexponential model of fluorescence decay. In order to determine the limit of traceability of CDPs, a comparison of different collagen/plasma ratio in samples was performed. The results of our study showed that the increase of added plasma to hydrolyzed collagen extended the mean FLT. Thus, the diagnosis of LVR based on measurements using TRFS is possible. However, it is important to point out the experiment was preliminary and further investigation in this field of research is crucial.

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

    PubMed Central

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

    2010-01-01

    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. PMID:21057657

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

    PubMed

    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.

  14. Noninvasive multimodal evaluation of bioengineered cartilage constructs combining time-resolved fluorescence and ultrasound imaging.

    PubMed

    Fite, Brett Z; Decaris, Martin; Sun, Yinghua; Sun, Yang; Lam, Adrian; Ho, Clark K L; Leach, J Kent; Marcu, Laura

    2011-04-01

    A multimodal diagnostic system that integrates time-resolved fluorescence spectroscopy, fluorescence lifetime imaging microscopy, and ultrasound backscatter microscopy is evaluated here as a potential tool for assessing changes in engineered tissue composition and microstructure nondestructively and noninvasively. The development of techniques capable of monitoring the quality of engineered tissue, determined by extracellular matrix (ECM) content, before implantation would alleviate the need for destructive assays over multiple time points and advance the widespread development and clinical application of engineered tissues. Using a prototype system combining time-resolved fluorescence spectroscopy, FLIM, and UBM, we measured changes in ECM content occurring during chondrogenic differentiation of equine adipose stem cells on 3D biodegradable matrices. The optical and ultrasound results were validated against those acquired via conventional techniques, including collagen II immunohistochemistry, picrosirius red staining, and measurement of construct stiffness. Current results confirm the ability of this multimodal approach to follow the progression of tissue maturation along the chondrogenic lineage by monitoring ECM production (namely, collagen type II) and by detecting resulting changes in mechanical properties of tissue constructs. Although this study was directed toward monitoring chondrogenic tissue maturation, these data demonstrate the feasibility of this approach for multiple applications toward engineering other tissues, including bone and vascular grafts. PMID:21303258

  15. Time-resolved imaging of fluorescent inclusions in optically turbid medium — phantom study

    NASA Astrophysics Data System (ADS)

    Kacprzak, M.; Liebert, A.; Sawosz, P.; Żołek, N.; Milej, D.; Maniewski, R.

    2010-03-01

    We present results of application of a time-resolved optical system for imaging of fluorescence excited in an inclusion containing indocyanine green (ICG), and located in optically turbid medium. The developed imaging system enabled simultaneous acquisition of fluorescence and diffusive reflectance. Eight independent time-resolved measurement channels based on time-correlated single photon counting technique were applied. In four of these channels, used for the fluorescence detection, sets of filters were applied in order to block the excitation light. Fast optomechanical switches allowed us to illuminate sequentially nine different spots on the surface of the studied object and finally 4×4 pixels maps at excitation and emission wavelengths were obtained. A liquid phantom used in this study consists of the fish tank filed with a solution ofmilk and water with black ink added to obtain optical properties in the range of the optical properties typical for the living tissue. A gel ball of a diameter of 5 mm with precisely controlled concentration of ICG was immersed in the liquid. The measurements were performed for inclusion located at different depths and for various ICG concentrations in the gel ball and in the surrounding liquid. The recorded distributions of times of arrival (DTA) of fluorescence photons and times of flight (DTOF) of diffusely reflected photons were analyzed by calculation of their statistical moments. We observed specific changes in moments of the measured DTAs as a function of depth of immersion of the fluorescent inclusion in the medium. We noted also that the changes of moments depend significantly on concentration of the dye in the fluorescence inclusion as well as in the surrounding liquid.

  16. Microscopic Study of Glass Transition: Time-Resolved Fluorescence Measurements of Doped Dye Molecules

    NASA Astrophysics Data System (ADS)

    Nakatsuka, H.; Ye, J. Y.; Hattori, T.; Maruyama, Y.; Ishikawa, M.

    The microscopic dynamics of several monomeric and polymeric glass formers has been investigated by the time-resolved fluorescence measurement of doped malachite green molecules in a wide temperature range. For monomers and a polymer without side chains, beside a kink around the calorimetric glass transition temperature Tg, another crossover at Tc about 30 - 50 K above Tg has been clearly observed, which is in agreement with the prediction of the mode-coupling theory. On the other hand, for the complex polymers with side chains, although we could not distinguish any singularities above Tg, we observed another kink below Tg, which can be attributed to the side-chain motions.

  17. Probing Ternary Complex Equilibria of Crown Ether Ligands by Time-Resolved Fluorescence Spectroscopy

    PubMed Central

    2015-01-01

    Ternary complex formation with solvent molecules and other adventitious ligands may compromise the performance of metal-ion-selective fluorescent probes. As Ca(II) can accommodate more than 6 donors in the first coordination sphere, commonly used crown ether ligands are prone to ternary complex formation with this cation. The steric strain imposed by auxiliary ligands, however, may result in an ensemble of rapidly equilibrating coordination species with varying degrees of interaction between the cation and the specific donor atoms mediating the fluorescence response, thus diminishing the change in fluorescence properties upon Ca(II) binding. To explore the influence of ligand architecture on these equilibria, we tethered two structurally distinct aza-15-crown-5 ligands to pyrazoline fluorophores as reporters. Due to ultrafast photoinduced electron-transfer (PET) quenching of the fluorophore by the ligand moiety, the fluorescence decay profile directly reflects the species composition in the ground state. By adjusting the PET driving force through electronic tuning of the pyrazoline fluorophores, we were able to differentiate between species with only subtle variations in PET donor abilities. Concluding from a global analysis of the corresponding fluorescence decay profiles, the coordination species composition was indeed strongly dependent on the ligand architecture. Altogether, the combination of time-resolved fluorescence spectroscopy with selective tuning of the PET driving force represents an effective analytical tool to study dynamic coordination equilibria and thus to optimize ligand architectures for the design of high-contrast cation-responsive fluorescence switches. PMID:25313708

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

    PubMed Central

    Yankelevich, Diego R.; Ma, Dinglong; Liu, Jing; Sun, Yang; Sun, Yinghua; Bec, Julien; Elson, Daniel S.; Marcu, Laura

    2014-01-01

    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

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

    PubMed

    Yankelevich, Diego R; Ma, Dinglong; Liu, Jing; Sun, Yang; Sun, Yinghua; Bec, Julien; Elson, Daniel S; Marcu, Laura

    2014-03-01

    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 lifetime

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

    SciTech Connect

    Yankelevich, Diego R.; Ma, Dinglong; Liu, Jing; Sun, Yang; Sun, Yinghua; Bec, Julien; Marcu, Laura; Elson, Daniel S.

    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

  1. Time-resolved fluorescence study of interaction of the monoclonal anticoproporphyrin antibodies and (Pt-)coproporphyrin

    NASA Astrophysics Data System (ADS)

    Lobanov, Oleg I.; Savitsky, Alexander P.; Nelen, Mary; Demcheva, Marina V.; Yatsimirsky, Anatoly K.; Korppi-Tommola, Jouko E.

    1995-02-01

    Mechanisms of ligand binding by monoclonal anti-coproporphyrin antibodies are studied by steady-state and time-resolved fluorescence spectroscopy by use of a picosecond laser system. The antibodies quench the coproporphyrin (CP) fluorescence, but the CP fluorescence spectra show a strong shift of maxima at high concentrations of antibodies (Ab) or their Fab fragment. This can be explained by a special type of Ab or Fab dimerization. Fluorescence decays of CP are measured at different concentrations of Ab and different pH values. The following deconvolution procedure based on the non-linear least squares method reveals a two- exponential character of the fluorescence decay. Data obtained by polarized fluorescence spectroscopy allows us to attribute the slow component (16 ns) to the free CP and the fast one (2.5 ns) to the CP-Ab complex. The obtained dependence of the ratio of amplitudes of the fast and slow components on the ratio of concentrations of CP and Ab at different pH values makes it possible to register separately the binding at different centers within the Ab and investigation of the effects of cooperation in the Ab molecule. The following experiments with Pt-coproporphyrin reveal the ability of Ab to reduce quenching of long-lived phosphorescence of the ligand.

  2. The binding of Curcuma longa extract with bovine serum albumin monitored via time-resolved fluorescence.

    PubMed

    Lemos, M Adília; Hungerford, Graham

    2013-01-01

    Turmeric (Curcuma longa L.) is obtained from the rhizome of the Zingberaceae family and has a long history as an ingredient in cooking. It has been used as a dye and recently research has concentrated on its possible health benefits, specifically because of its antioxidant activity. The principal compound that is responsible for this activity is curcumin, which is present with the other curcuminoids; demethoxycurcumin and bisdemethoxycurcumin. Curcumin exhibits fluorescence and its photophysics are markedly affected by the polarity, hydrogen bonding and pH. This provides a means to examine its interaction with proteins, which is important if its potential health role is to be fully investigated. In this work, we monitor the binding kinetics using time-resolved fluorescence measurements, enabled by the use of low dead time electronics coupled with a high repetition rate excitation source and time-resolved emission spectra of the extracted curcuminoids upon interaction with bovine serum albumin. From these measurements the decay-associated spectra of the different lifetime components were obtained, which is consistent with reports of more than one binding site. Monitoring changes in these spectra with increasing temperature also allows for the denaturing of the serum albumin to be inferred.

  3. Multimodal imaging of vascular grafts using time-resolved fluorescence and ultrasound

    NASA Astrophysics Data System (ADS)

    Fatakdawala, Hussain; Griffiths, Leigh G.; Wong, Maelene L.; Humphrey, Sterling; Marcu, Laura

    2015-02-01

    The translation of engineered tissues into clinic requires robust monitoring of tissue development, both in vitro and in vivo. Traditional methods for the same are destructive, inefficient in time and cost and do not allow time-lapse measurements from the same sample or animal. This study reports on the ability of time-resolved fluorescence and ultrasound measurements for non-destructive characterization of explanted tissue engineered vascular grafts. Results show that TRFS and FLIm are able to assess alterations in luminal composition namely elastin, collagen and cellular (hyperplasia) content via changes in fluorescence lifetime values between normal and grafted tissue. These observations are complemented by structural changes observed in UBM pertaining to graft integration and intimal thickness over the grafted region. These results encourage the future application of a catheter-based technique that combines these imaging modalities for non-destructive characterization of vascular grafts in vivo.

  4. Development of time resolved fluorescence resonance energy transfer-based assay for FXR antagonist discovery.

    PubMed

    Yu, Donna D; Lin, Wenwei; Chen, Taosheng; Forman, Barry M

    2013-07-15

    FXR (farnesoid X receptor, NRIH4), a nuclear receptor, plays a major role in the control of cholesterol metabolism. FXR ligands have been investigated in preclinical studies for targeted therapy against metabolic diseases, but have shown limitations. Therefore, there is a need for new agonist or antagonist ligands of FXR, both for potential clinical applications, as well as to further elucidate its biological functions. Here we describe the use of the X-ray crystal structure of FXR complexed with the potent small molecule agonist GW4064 to design and synthesize a novel fluorescent, high-affinity probe (DY246) for time resolved fluorescence resonance energy transfer (TR-FRET) assays. We then used the TR-FRET assay for high throughput screening of a library of over 5000 bioactive compounds. From this library, we identified 13 compounds that act as putative FXR transcriptional antagonists.

  5. Time-resolved spectroscopy of the probe fluorescence in the study of human blood protein dynamic structure on SR beam

    NASA Astrophysics Data System (ADS)

    Dobretsov, G. E.; Kurek, N. K.; Syrejshchikova, T. I.; Yakimenko, M. N.; Clarke, D. T.; Jones, G. R.; Munro, I. H.

    2000-06-01

    Time-resolved spectroscopy on the SRS of the Daresbury Laboratory was used for the study of the human serum lipoproteins and human blood albumins with fluorescent probes K-37 and K-35, developed in Russia. The probe K-37 was found sensitive to the difference in dynamic properties of the lipid objects. Two sets of the parameters were used for the description of lipid dynamic structure: (1) time-resolved fluorescence spectra and (2) time-resolved fluorescence depolarization as a function of rotational mobility of lipid molecules. Each measured dynamic parameter reflected the monotonous changes of dynamic properties in the range: lipid spheres-very low density lipoproteins-low density lipoproteins-high density lipoproteins-phospholipid liposomes. The range is characterized by the increase of the ratio polar/ nonpolar lipids. Thus, time-resolved fluorescence could be used to detect some structural modifications in lipoproteins related to atherosclerosis and subsequent cardiovascular diseases development.

  6. Near-infrared time-resolved fluorescence lifetime determinations in poly(methylmethacrylate) microchip electrophoresis devices.

    PubMed

    Llopis, Shawn D; Stryjewski, Wieslaw; Soper, Steven A

    2004-11-01

    High aspect-ratio microstructures were hot-embossed in polymer substrates with a molding tool fabricated using lithography/electroplating/forming (LIGA). The resulting devices were used for the electrophoretic separation of oligonucleotides labeled with near-infrared (near-IR) dyes. Near-IR time-resolved fluorescence was used as an identification method for the labeling dyes. The detection apparatus consisted of a pulsed laser diode operating at 680 nm, a single-photon avalanche diode, an integrated microscope, and a PC-board incorporating time-correlated single photon counting electronics. Investigation of the optical quality and amount of autofluorescence generated from different polymer substrates was carried out in the near-IR region for determining compatibility with time-resolved fluorescence. Our results revealed that of several poly(methylmethacrylate)(PMMA) substrates, brand Plexiglas offered minimal replication errors in the embossed features using appropriate embossing conditions with low background fluorescence contributions to the observed decay. Near-IR dye-labeled oligonucleotides were separated to determine the applicability of fluorescence lifetime discrimination between Cy5.5 (tauf = 930 ps) and IRD700 (tauf = 851 ps) labeling dyes during the microchip separation. These dyes were used to label T-fragments (thymine) of an M13mp18 ssDNA template. The DNA ladders were electrophoresed at 130 V/cm in a 4% linear polyacrylamide gel (LPA) gel matrix in a 9.5 cm long serpentine channel heated to 50 degrees C. The electropherogram revealed that the lifetimes could be accurately read well beyond 450 bases, although single-base pair resolution in the electropherogram was difficult to achieve due to potential solute-wall interactions in the polymer microdevice or the electroosmotic flow (EOF) properties of the device. The relative standard deviations secured for individual bands in the electropherogram were similar to those obtained using capillary gel

  7. A Novel Europium Chelate Coated Nanosphere for Time-Resolved Fluorescence Immunoassay.

    PubMed

    Shen, Yifeng; Xu, Shaohan; He, Donghua

    2015-01-01

    A novel europium ligand 2,2',2'',2'''-(4,7-diphenyl-1,10-phenanthroline-2,9-diyl) bis (methylene) bis (azanetriyl) tetra acetic acid (BC-EDTA) was synthesized and characterized. It shows an emission spectrum peak at 610 nm when it is excited at 360 nm, with a large Stock shift (250 nm). It is covalently coated on the surface of a bare silica nanosphere containi free amino groups, using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-Hydroxysuccinimide. We also observed an interesting phenomenon that when BC-EDTA is labeled with a silica nanosphere, the chelate shows different excitation spectrum peaks of about 295 nm. We speculate that the carboxyl has a significant influence on its excitation spectrum. The BC-EDTA/Eu3+coated nanosphere could be used as a fluorescent probe for time-resolved fluorescence immunoassay. We labeled the antibody with the fluorescent nanosphere to develop a nanosphere based hepatitis B surface antigen as a time-resolved fluorescence immunoassay reagent, which is very easy to operate and eliminates potential contamination of Eu3+ contained in the environment. The analytical and functional sensitivities are 0.0037 μg/L and 0.08 μg/L (S/N≥2.0) respectively. The detection range is 0.08-166.67 μg/L, which is much wider than that of ELISA (0.2-5 μg/L). It is comparable to the commercial dissociation-enhanced lanthanide fluoro-immunoassay system (DELFIA) reagents (0.2-145 μg/L). We propose that it can fulfill clinical applications.

  8. A Novel Europium Chelate Coated Nanosphere for Time-Resolved Fluorescence Immunoassay

    PubMed Central

    Shen, Yifeng; Xu, Shaohan; He, Donghua

    2015-01-01

    A novel europium ligand 2, 2’, 2’’, 2’’’-(4, 7-diphenyl-1, 10-phenanthroline-2, 9-diyl) bis (methylene) bis (azanetriyl) tetra acetic acid (BC-EDTA) was synthesized and characterized. It shows an emission spectrum peak at 610 nm when it is excited at 360 nm, with a large Stock shift (250 nm). It is covalently coated on the surface of a bare silica nanosphere containi free amino groups, using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-Hydroxysuccinimide. We also observed an interesting phenomenon that when BC-EDTA is labeled with a silica nanosphere, the chelate shows different excitation spectrum peaks of about 295 nm. We speculate that the carboxyl has a significant influence on its excitation spectrum. The BC-EDTA/Eu3+coated nanosphere could be used as a fluorescent probe for time-resolved fluorescence immunoassay. We labeled the antibody with the fluorescent nanosphere to develop a nanosphere based hepatitis B surface antigen as a time-resolved fluorescence immunoassay reagent, which is very easy to operate and eliminates potential contamination of Eu3+ contained in the environment. The analytical and functional sensitivities are 0.0037 μg/L and 0.08 μg/L (S/N≥2.0) respectively. The detection range is 0.08-166.67 μg/L, which is much wider than that of ELISA (0.2-5μg/L). It is comparable to the commercial dissociation-enhanced lanthanide fluoro-immunoassay system (DELFIA) reagents (0.2-145μg/L). We propose that it can fulfill clinical applications. PMID:26056826

  9. [Discrimination of Crude Oil Samples Using Laser-Induced Time-Resolved Fluorescence Spectroscopy].

    PubMed

    Han, Xiao-shuang; Liu, De-qing; Luan, Xiao-ning; Guo, Jin-jia; Liu, Yong-xin; Zheng, Rong-er

    2016-02-01

    The Laser-induced fluorescence spectra combined with pattern recognition method has been widely applied in discrimination of different spilled oil, such as diesel, gasoline, and crude oil. However, traditional three-dimension fluorescence analysis method, which is not adapted to requirement of field detection, is limited to laboratory investigatio ns. The development of oil identification method for field detection is significant to quick response and operation of oil spill. In this paper, a new method based on laser-induced time-resolved fluorescence combined with support vector machine (SVM) model was introduced to discriminate crude oil samples. In this method, time-resolved spectra data was descended into two dimensions with selecting appropriate range in time and wavelength domains respectively to form a SVM data base. It is found that the classification accurate rate increased with an appropriate selection. With a selected range from 54 to 74 ns in time domain, the classification accurate rate has been increased from 83.3% (without selection) to 88.1%. With a selected wavelength range of 387.00~608.87 nm, the classification accurate rate of suspect oil was improved from 84% (without selection) to 100%. Since the detection delay of fluorescence lidar fluctuates due to wave and platform swing, the identification method with optimizing in both time and wavelength domains could offer a better flexibility for field applications. It is hoped that the developed method could provide some useful reference with data reduction for classification of suspect crude oil in the future development. PMID:27209747

  10. Time-resolved fluorescence polarization spectroscopy of visible and near infrared dyes in picosecond dynamics

    NASA Astrophysics Data System (ADS)

    Pu, Yang; Alfano, Robert R.

    2015-03-01

    Near-infrared (NIR) dyes absorb and emit light within the range from 700 to 900 nm have several benefits in biological studies for one- and/or two-photon excitation for deeper penetration of tissues. These molecules undergo vibrational and rotational motion in the relaxation of the excited electronic states, Due to the less than ideal anisotropy behavior of NIR dyes stemming from the fluorophores elongated structures and short fluorescence lifetime in picosecond range, no significant efforts have been made to recognize the theory of these dyes in time-resolved polarization dynamics. In this study, the depolarization of the fluorescence due to emission from rotational deactivation in solution will be measured with the excitation of a linearly polarized femtosecond laser pulse and a streak camera. The theory, experiment and application of the ultrafast fluorescence polarization dynamics and anisotropy are illustrated with examples of two of the most important medical based dyes. One is NIR dye, namely Indocyanine Green (ICG) and is compared with Fluorescein which is in visible range with much longer lifetime. A set of first-order linear differential equations was developed to model fluorescence polarization dynamics of NIR dye in picosecond range. Using this model, the important parameters of ultrafast polarization spectroscopy were identified: risetime, initial time, fluorescence lifetime, and rotation times.

  11. Time resolved laser induced fluorescence measurements: Considerations when using Nd:YAG based system

    NASA Astrophysics Data System (ADS)

    Rabasovic, Maja S.; Sevic, Dragutin; Terzic, Mira; Marinkovic, Bratislav P.

    2012-05-01

    Time-resolved laser-induced fluorescence (TR-LIF) and the laser induced breakdown spectroscopy (LIBS) have been shown to be methods which are fast and sensitive to provide information about the constituents in analyzed samples. TR-LIF and LIBS have similar hardware requirements. In this paper, we analyze some characteristics of TR-LIF/LIBS system implemented in our laboratory, considering the fact that the excitation part of the system is based on Nd:YAG laser and Optical Parametric Oscillator (OPO). The laser is more than powerful enough (365 mJ at 1064 nm, variable OPO output >5 mJ) for LIBS, but somehow slow (the length of fundamental laser harmonic output pulse is about 5 ns) for fluorescence measurements in our present area of interest, namely plants and food products. Fortunately, the pulse length of tunable OPO output (320-475 nm) is less then 1 ns, so by means of a correct deconvolution procedure it is possible to measure the fluorescence lifetimes in the range as small as a few nanoseconds. The fluorescence detection part of our system is based on picosecond streak camera. Using the fluorescent dyes (Rhodamine B and Fluorescein) ethanol solutions we verified the analyzing capabilities of our TR-LIF system.

  12. Time-resolved fluorescence spectroscopy for intraoperative assistance of thyroid surgery

    NASA Astrophysics Data System (ADS)

    Bachmann, L.; Brandao, M. P.; Iwakura, R.; Basilio, F. S.; Haleplian, K.; Ito, A. S.; Conti de Freitas, L. C.

    2016-03-01

    Searching for new methods to provide information of biochemical composition and structure is critical to improve the prognosis of thyroid diseases. The use of time-resolved fluorescence techniques to detect biochemical composition and tissue structure alterations could help develop a portable, minimally invasive, and non-destructive method to assist during surgical procedures. This research looks for employ a fluorescence technique based on lifetime measurements to differentiate healthy and benign lesions from malignant thyroid tissue. We employ a wide range of excitation and chose a more appropriate region for this work: 298-300 nm; and the fluorescence decay was measured at 340-450 nm. We observed fluorescence lifetimes at 340 nm emission of 0.80+/-0.26 and 3.94+/-0.47 ns for healthy tissue; 0.90+/-0.24 and 4.05+/-0.46 ns for benign lesions; and 1.21+/-0.14 and 4.63+/-0.25 ns for malignant lesions. For 450 nm emissions, we obtain lifetimes of 0.25+/-0.18 and 3.99+/-0.39 ns for healthy tissue, 0.24+/-0.17 and 4.20+/-0.48 ns for benign lesions, 0.33+/-0.32 and 4.55+/-0.55 ns for malignant lesions. We successfully demonstrated that fluorescence lifetimes at 340 nm emission can differentiate between thyroid malignant and healthy/benign tissues.

  13. A time-resolved fluorescence study of electronic excitation energy transport in concentrated dye solutions

    NASA Astrophysics Data System (ADS)

    Scully, A. D.; Matsumoto, A.; Hirayama, S.

    1991-11-01

    Singlet-state radiative and nonradiative energy transport among randomly distributed donor and acceptor molecules in solutions of ethylene glycol has been investigated by using time-resolved fluorescence spectroscopy. Radiative energy transfer in moderately concentrated solutions of rhodamine 6G contained in a 1 cm pathlength cuvette results in nonexponential fluorescence decay curves which can be described in terms of the relative fluorescence emission yield of the ith transfer process, φ i, the average number of transfer steps, < n >, and the apparent fluorescence lifetime, <τ >. The fluorescence decay profiles measured from thin films (⩽ 20 μm) of solutions of rhodamine 6G donors in the presence of malachite green acceptors are also nonexponential and at low donor concentrations (1.0 × 10 -4 M) the decay curves can be described by using Förster's dipole-dipole model for nonradiative energy transfer, where the value for the critical transfer distance, R0, is calculated to be (5.9±0.1) nm. At higher donor concentrations (3.0 × 10 -3 M) the Förster model is inappropriate. However, the model proposed by Loring, Anderson and Fayer (LAF), which allows for the effects of nonradiative energy transfer among donors, provides excellent fits to the experimentally determined fluorescence decay curves for this donor concentration and results in a value of R0 for nonradiative energy transfer between rhodamine 6G chromophores of (5.8±0.1) nm. The LAF model also provides a satisfactory description of the kinetics of the quenching by rhodamine 6G dimers of the fluorescence from thin films of highly concentrated solutions of rhodamine 6G. The value of R0 for nonradiative energy transfer from monomer to dimer and the equilibrium constant for rhodamine 6G dimerization are calculated to be (3.2±0.1) nm and 19 M -1, respectively.

  14. Time-resolved remote Raman and fluorescence spectrometers for planetary exploration

    NASA Astrophysics Data System (ADS)

    Sharma, Shiv K.; Misra, Anupam K.; Acosta, Tayro E.; Lucey, Paul G.

    2012-06-01

    At the University of Hawaii, we have developed compact time-resolved (TR) Raman, and fluorescence spectrometers suitable for planetary exploration under NASA's Mars Instrument Development Program. The compact Raman and fluorescence spectrometers consist of custom miniature spectrographs based on volume holographic gratings, and custom miniature intensified CCD cameras. These spectrographs have been interfaced with a regular 50 mm camera lens as well as with a three and a half inch diameter telescope for remotely interrogating minerals, water, water-ice and dry ice. Using a small frequency-doubled Nd:YAG pulsed laser (35 mJ/pulse, 20 Hz) and 50 mm camera lens, TRRaman and LINF spectra of minerals, and bio-minerals can be measured within 30 s under super-critical CO2, and with 3.5-inch telescope these samples can be interrogated to 50 m radial distance during day time and nighttime. The fluorescence spectrograph is capable of measuring TR- laser-induced fluorescence excited with 355 nm laser in the spectral range 400-800 nm spectral range. The TR-fluorescence spectra allow measurement of LINF from rare-earths and transition-metal ions in time domain, and also assist in differentiating between abiogenic minerals from organic and biogenic materials based on the fluorescence lifetime. Biological materials are also identified from their characteristic short-lived (<10 ns) laser-induced fluorescence lifetime. These instruments will play important role in planetary exploration especially in NASA's future Mars Sample Return Mission, and lander and rover missions.

  15. Applications of time-resolved laser fluorescence spectroscopy to the environmental biogeochemistry of actinides.

    PubMed

    Collins, Richard N; Saito, Takumi; Aoyagi, Noboru; Payne, Timothy E; Kimura, Takaumi; Waite, T David

    2011-01-01

    Time-resolved laser fluorescence spectroscopy (TRLFS) is a useful means of identifying certain actinide species resulting from various biogeochemical processes. In general, TRLFS differentiates chemical species of a fluorescent metal ion through analysis of different excitation and emission spectra and decay lifetimes. Although this spectroscopic technique has largely been applied to the analysis of actinide and lanthanide ions having fluorescence decay lifetimes on the order of microseconds, such as UO , Cm, and Eu, continuing development of ultra-fast and cryogenic TRLFS systems offers the possibility to obtain speciation information on metal ions having room-temperature fluorescence decay lifetimes on the order of nanoseconds to picoseconds. The main advantage of TRLFS over other advanced spectroscopic techniques is the ability to determine in situ metal speciation at environmentally relevant micromolar to picomolar concentrations. In the context of environmental biogeochemistry, TRLFS has principally been applied to studies of (i) metal speciation in aqueous and solid phases and (ii) the coordination environment of metal ions sorbed to mineral and bacterial surfaces. In this review, the principles of TRLFS are described, and the literature reporting the application of this methodology to the speciation of actinides in systems of biogeochemical interest is assessed. Significant developments in TRLFS methodology and advanced data analysis are highlighted, and we outline how these developments have the potential to further our mechanistic understanding of actinide biogeochemistry.

  16. Energy transfer in Anabaena variabilis filaments under nitrogen depletion, studied by time-resolved fluorescence.

    PubMed

    Onishi, Aya; Aikawa, Shimpei; Kondo, Akihiko; Akimoto, Seiji

    2015-08-01

    Some filamentous cyanobacteria (including Anabaena) differentiate into heterocysts under nitrogen-depleted conditions. During differentiation, the phycobiliproteins and photosystem II in the heterocysts are gradually degraded. Nitrogen depletion induces changes in the pigment composition of both vegetative cells and heterocysts, which affect the excitation energy transfer processes. To investigate the changes in excitation energy transfer processes of Anabaena variabilis filaments grown in standard medium (BG11) and a nitrogen-free medium (BG110), we measured their steady-state absorption spectra, steady-state fluorescence spectra, and time-resolved fluorescence spectra (TRFS) at 77 K. TRFS were measured with a picosecond time-correlated single photon counting system. The pigment compositions of the filaments grown in BG110 changed throughout the growth period; the relative phycocyanin levels monotonically decreased, whereas the relative carotenoid (Car) levels decreased and then recovered to their initial value (at day 0), with formation of lower-energy Cars. Nitrogen starvation also altered the fluorescence kinetics of PSI; the fluorescence maximum of TRFS immediately after excitation occurred at 735, 740, and 730 nm after 4, 8, and 15 days growth in BG110, respectively. Based on these results, we discuss the excitation energy transfer dynamics of A. variabilis filaments under the nitrogen-depleted condition throughout the growth period. PMID:25596847

  17. Fluorescence Dynamics in the Endoplasmic Reticulum of a Live Cell: Time-Resolved Confocal Microscopy.

    PubMed

    Ghosh, Shirsendu; Nandi, Somen; Ghosh, Catherine; Bhattacharyya, Kankan

    2016-09-19

    Fluorescence dynamics in the endoplasmic reticulum (ER) of a live non-cancer lung cell (WI38) and a lung cancer cell (A549) are studied by using time-resolved confocal microscopy. To selectively study the organelle, ER, we have used an ER-Tracker dye. From the emission maximum (λmaxem) of the ER-Tracker dye, polarity (i.e. dielectric constant, ϵ) in the ER region of the cells (≈500 nm in WI38 and ≈510 nm in A549) is estimated to be similar to that of chloroform (λmaxem =506 nm, ϵ≈5). The red shift by 10 nm in λmaxem in the cancer cell (A549) suggests a slightly higher polarity compared to the non-cancer cell (WI38). The fluorescence intensity of the ER-Tracker dye exhibits prolonged intermittent oscillations on a timescale of 2-6 seconds for the cancer cell (A549). For the non-cancer cell (WI38), such fluorescence oscillations are much less prominent. The marked fluorescence intensity oscillations in the cancer cell are attributed to enhanced calcium oscillations. The average solvent relaxation time (<τs >) of the ER region in the lung cancer cell (A549, 250±50 ps) is about four times faster than that in the non-cancer cell (WI38, 1000±50 ps).

  18. Lasing dynamics study by femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy

    NASA Astrophysics Data System (ADS)

    Wei, Dang; Qing, Liao; Peng-Cheng, Mao; Hong-Bing, Fu; Yu-Xiang, Weng

    2016-05-01

    Femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy (FNOPAS) is a versatile technique with advantages of high sensitivity, broad detection bandwidth, and intrinsic spectrum correction function. These advantages should benefit the study of coherent emission, such as measurement of lasing dynamics. In this letter, the FNOPAS was used to trace the lasing process in Rhodamine 6G (R6G) solution and organic semiconductor nano-wires. High-quality transient emission spectra and lasing dynamic traces were acquired, which demonstrates the applicability of FNOPAS in the study of lasing dynamics. Our work extends the application scope of the FNOPAS technique. Project supported by the National Natural Science Foundation of China (Grant Nos. 20925313 and 21503066), the Innovation Program of Chinese Academy of Sciences (Grant No. KJCX2-YW-W25), the Postdoctoral Project of Hebei University, China, and the Project of Science and Technology Bureau of Baoding City, China (Grant No. 15ZG029).

  19. A CMOS Time-Resolved Fluorescence Lifetime Analysis Micro-System

    PubMed Central

    Rae, Bruce R.; Muir, Keith R.; Gong, Zheng; McKendry, Jonathan; Girkin, John M.; Gu, Erdan; Renshaw, David; Dawson, Martin D.; Henderson, Robert K.

    2009-01-01

    We describe a CMOS-based micro-system for time-resolved fluorescence lifetime analysis. It comprises a 16 × 4 array of single-photon avalanche diodes (SPADs) fabricated in 0.35 μm high-voltage CMOS technology with in-pixel time-gated photon counting circuitry and a second device incorporating an 8 × 8 AlInGaN blue micro-pixellated light-emitting diode (micro-LED) array bump-bonded to an equivalent array of LED drivers realized in a standard low-voltage 0.35 μm CMOS technology, capable of producing excitation pulses with a width of 777 ps (FWHM). This system replaces instrumentation based on lasers, photomultiplier tubes, bulk optics and discrete electronics with a PC-based micro-system. Demonstrator lifetime measurements of colloidal quantum dot and Rhodamine samples are presented. PMID:22291564

  20. Development of a Rapid Insulin Assay by Homogenous Time-Resolved Fluorescence.

    PubMed

    Farino, Zachary J; Morgenstern, Travis J; Vallaghe, Julie; Gregor, Nathalie; Donthamsetti, Prashant; Harris, Paul E; Pierre, Nicolas; Freyberg, Robin; Charrier-Savournin, Fabienne; Javitch, Jonathan A; Freyberg, Zachary

    2016-01-01

    Direct measurement of insulin is critical for basic and clinical studies of insulin secretion. However, current methods are expensive and time-consuming. We developed an insulin assay based on homogenous time-resolved fluorescence that is significantly more rapid and cost-effective than current commonly used approaches. This assay was applied effectively to an insulin secreting cell line, INS-1E cells, as well as pancreatic islets, allowing us to validate the assay by elucidating mechanisms by which dopamine regulates insulin release. We found that dopamine functioned as a significant negative modulator of glucose-stimulated insulin secretion. Further, we showed that bromocriptine, a known dopamine D2/D3 receptor agonist and newly approved drug used for treatment of type II diabetes mellitus, also decreased glucose-stimulated insulin secretion in islets to levels comparable to those caused by dopamine treatment. PMID:26849707

  1. Time-resolved laser fluorescence spectroscopy of UO2(CO3)3(4-).

    PubMed

    Jung, E C; Cho, H-R; Baik, M H; Kim, H; Cha, W

    2015-11-21

    The objective of the present study is to examine the luminescence characteristics of UO2(CO3)3(4-) in detail using time-resolved laser fluorescence spectroscopy. The peak wavelengths and lifetime of UO2(CO3)3(4-) were determined at room temperature using the two excitation laser wavelengths of 266 and 448 nm. The peak wavelengths in the luminescence spectrum exhibited hypsochromic shifts compared with those of UO2(2+). The lifetime determined from several samples containing various uranium concentrations was 8.9 ± 0.8 ns. Explanations for the hindrance to the observation of the luminescence spectrum of UO2(CO3)3(4-) in previous investigations are discussed. The representative experimental parameters, which might interrupt the measurement of weak luminescence, are the insertion delay time of the detection device, the overlapped luminescence of the background materials and the primary inner filter effect in the sample solution.

  2. Time resolved fluorescence from parity mixed rotational energy levels - Collisions vs electric field effects

    NASA Astrophysics Data System (ADS)

    Mandich, M. L.; Gaebe, C. E.; Gottscho, R. A.

    1985-10-01

    Moore et al. (1984) have described a method for the in situ and nonintrusive measurement of plasma electric fields by a method involving the excitation of a parity or Lambda doublet of the polar diatomic molecule BCl. Three approximations are made in deriving a theoretical relationship between field strength and the forbidden to allowed line intensity ratio. One approximation is related to the neglect of collisional transfer, while another is based on the neglect of coherent phenomena, such as quantum beats between the mixed parity levels. New experimental evidence is provided, and it is shown that the latter approximation is not always justified. The last assumption is the neglect of hyperfine structure effects on field-dependent line intensities and polarizations. Hyperfine effects are accounted for in a phenomenological fashion which is justified empirically. Attention is given to both time-resolved and time-integrated fluorescence measurements from parity-mixed energy levels in the polar diatomic molecule BCl.

  3. Development of a Rapid Insulin Assay by Homogenous Time-Resolved Fluorescence

    PubMed Central

    Vallaghe, Julie; Gregor, Nathalie; Donthamsetti, Prashant; Harris, Paul E.; Pierre, Nicolas; Freyberg, Robin; Charrier-Savournin, Fabienne; Javitch, Jonathan A.; Freyberg, Zachary

    2016-01-01

    Direct measurement of insulin is critical for basic and clinical studies of insulin secretion. However, current methods are expensive and time-consuming. We developed an insulin assay based on homogenous time-resolved fluorescence that is significantly more rapid and cost-effective than current commonly used approaches. This assay was applied effectively to an insulin secreting cell line, INS-1E cells, as well as pancreatic islets, allowing us to validate the assay by elucidating mechanisms by which dopamine regulates insulin release. We found that dopamine functioned as a significant negative modulator of glucose-stimulated insulin secretion. Further, we showed that bromocriptine, a known dopamine D2/D3 receptor agonist and newly approved drug used for treatment of type II diabetes mellitus, also decreased glucose-stimulated insulin secretion in islets to levels comparable to those caused by dopamine treatment. PMID:26849707

  4. Time-resolved laser fluorescence spectroscopy of UO2(CO3)3(4-).

    PubMed

    Jung, E C; Cho, H-R; Baik, M H; Kim, H; Cha, W

    2015-11-21

    The objective of the present study is to examine the luminescence characteristics of UO2(CO3)3(4-) in detail using time-resolved laser fluorescence spectroscopy. The peak wavelengths and lifetime of UO2(CO3)3(4-) were determined at room temperature using the two excitation laser wavelengths of 266 and 448 nm. The peak wavelengths in the luminescence spectrum exhibited hypsochromic shifts compared with those of UO2(2+). The lifetime determined from several samples containing various uranium concentrations was 8.9 ± 0.8 ns. Explanations for the hindrance to the observation of the luminescence spectrum of UO2(CO3)3(4-) in previous investigations are discussed. The representative experimental parameters, which might interrupt the measurement of weak luminescence, are the insertion delay time of the detection device, the overlapped luminescence of the background materials and the primary inner filter effect in the sample solution. PMID:26460936

  5. Applications of Phasors to In Vitro Time-Resolved Fluorescence Measurements

    PubMed Central

    Štefl, Martin; James, Nicholas G.; Ross, Justin A.; Jameson, David M.

    2010-01-01

    The phasor method of treating fluorescence lifetime data provides a facile and convenient approach to characterize lifetime heterogeneity and to detect the presence of excited state reactions, such as solvent relaxation and Förster Resonance Energy Transfer. The method utilizes a plot of M sin(Φ) versus M cos(Φ), where M is the modulation ratio and Φ is the phase angle taken from frequency domain fluorometry. A principle advantage of the phasor method is that it provides a model-less approach to time-resolved data, amenable to visual inspection. Although the phasor approach has been recently applied to Fluorescence Lifetime Imaging Microscopy it has not been extensively utilized for cuvette studies. In the present study we explore the applications of the method to in vitro samples. The phasors of binary and ternary mixtures of fluorescent dyes demonstrates the utility of the method for investigating complex mixtures. Data from excited state reactions, such as dipolar relaxation in membrane and protein systems and also energy transfer from the tryptophan residue to the chromophore in EGFP, are also presented. PMID:21078290

  6. Non-contact characterization of bacteria by time-resolved fluorescence

    NASA Astrophysics Data System (ADS)

    Bouchard, Alain; Frechette, Julie; Long, William F.; Vernon, Marcia; Cormier, Jean-Francois; Vallee, Real; Mafu, Akier A.; Lemay, Marie-Josee

    2004-07-01

    Accurate real-time methods for the detection of pathogenic microorganisms in the agri-food industry would represent an improvement over standard methods of analysis. We are currently developing a non-contact, scanning optical system for the detection of bacteria on meat surfaces based on fluorescence lifetime and intensity measurements. The system detects autofluorescent light emitted by the naturally occurring fluorophores in bacteria. Potential expected advantages of this system include accurate and efficient 2D real-time mapping of bacterial contamination of surfaces, and elimination of sample-to-sample cross-contamination. Furthermore, as the technique only requires minimal sample preparation and handling, the chemical properties of the specimen are preserved. This article presents the preliminary results obtained from a time-resolved fluorescence imaging system for the characterization of a non-pathogenic gram-negative bacteria, Pseudomonas fluorescens. Additionally we present a particular application of the system of interest to the agri-food industry, demonstrating its potential as a real-time macroscopic imaging system for mapping bacterial contamination on meat surfaces. Initial results indicate that the combination of fluorescence lifetime and intensity measurements provides a means for characterizing biological media and for detecting microorganisms on surfaces.

  7. Time-resolved fluorescence-detected magnetic resonance and fluorescence studies of trialkylamines irradiated by pulse radiolysis in alkane solvents

    SciTech Connect

    Lefkowitz, S.M.; Trifunac, A.D.

    1984-01-05

    Time-resolved fluorescence-detected magnetic resonance (FDMR) studies of irradiated alkane solutions of trialkylamines and scintillators reveal the EPR spectra of the trialkylaminium radicals, formed by scavenging solvent radical cations. A qualitative kinetic analysis indicates that the growth of the triethylaminium radical (TEA/sup +/-) FDMR signal occurs on similar time scales in both n-hexane and cyclohexane, suggesting that, in cyclohexane, TEA/sup +/- is formed by scavenging the lower mobility ''trapped'' cyclohexane radical cations. Fluorescence results indicate that TEA quenches both scintillator fluorescence and total FDMR intensities to a greater extent than is expected from amine scavenging of solvent holes. TEA also exhibits an intense, relatively long-lived fluorescence which is apparently not produced by radical ion recombination or energy transfer. 6 figures

  8. A homogeneous time-resolved fluorescence resonance energy transfer assay for phosphatidylserine exposure on apoptotic cells.

    PubMed

    Gasser, Jean-Philippe; Hehl, Michaela; Millward, Thomas A

    2009-01-01

    A simple, "mix-and-measure" microplate assay for phosphatidylserine (PtdSer) exposure on the surface of apoptotic cells is described. The assay exploits the fact that annexin V, a protein with high affinity and specificity for PtdSer, forms trimers and higher order oligomers on binding to membranes containing PtdSer. The transition from soluble monomer to cell-bound oligomer is detected using time-resolved fluorescence resonance energy transfer from europium chelate-labeled annexin V to Cy5-labeled annexin V. PtdSer detection is achieved by a single addition of a reagent mix containing labeled annexins and calcium ions directly to cell cultures in a 96-well plate, followed by a brief incubation before fluorescence measurement. The assay can be used to quantify PtdSer exposure on both suspension cells and adherent cells in situ. This method is simpler and faster than existing annexin V binding assays based on flow cytometry or microscopy, and it yields precise data with Z' values of 0.6-0.7. PMID:18835236

  9. Tryptophan dynamics of the FK506 binding protein: time-resolved fluorescence and simulations.

    PubMed Central

    Silva, N D; Prendergast, F G

    1996-01-01

    The FK506-binding protein (FKBP12) is important in the immunosuppressant action of FK506 and rapamycin. We have investigated Trp side chain dynamics in FKBP12, with and without a bound immunosuppressant, by measuring the Trp time-resolved fluorescence anisotropy decay r(t). The r(t) for W59 in aqueous uncomplexed FKBP12 at 20 degrees C is well described by a single exponential with a recovered initial anisotropy, r(eff)o, of 0.192 and an overall rotational correlation time for the protein, phi p, of 4.7 ns; r(eff)o = 0.214 and phi p = 4.2 ns for the FKBP12/FK506 complex. Using an expression for the order parameter squared, namely S2 = r(eff)o/rTo, where rTo is the vitrified steady-state excitation anisotropy, we recovered an S2 of 0.75 for W59 fluorescence in uncomplexed FKBP12 and S2 approximately equal to 1 in the FKBP12/FK506 complex. Results obtained for the FKBP12/rapamycin complex are similar to those found for the FKBP12/FK506 complex. Minimum perturbation mapping simulations were performed on the free and complexed forms of FKBP12 and the results were generally in agreement with the experimental data. Images FIGURE 5 FIGURE 6 PMID:8785272

  10. Characterization of powellite-based solid solutions by site-selective time resolved laser fluorescence spectroscopy.

    PubMed

    Schmidt, Moritz; Heck, Stephanie; Bosbach, Dirk; Ganschow, Steffen; Walther, Clemens; Stumpf, Thorsten

    2013-06-21

    We present a comprehensive study of the solid solution system Ca2(MoO4)2-NaGd(MoO4)2 on the molecular scale, by means of site-selective time resolved laser fluorescence spectroscopy (TRLFS). Eu(3+) is used as a trace fluorescent probe, homogeneously substituting for Gd(3+) in the solid solution crystal structure. Site-selective TRLFS of a series of polycrystalline samples covering the whole composition range of the solid solution series from 10% substitution of Ca(2+) to the NaGd end-member reveals it to be homogeneous throughout the whole range. The trivalent ions are incorporated into the powellite structure in only one coordination environment, which exhibits a very strong ligand-metal interaction. Polarization-dependent measurements of a single crystal of NaGd(Eu)(MoO4)2 identify the coordination geometry to be of C2v point symmetry. The S4 symmetry of the Ca site within the powellite lattice can be transformed into C2v assuming minor motion in the first coordination sphere.

  11. Light adaptation of the unicellular red alga, Cyanidioschyzon merolae, probed by time-resolved fluorescence spectroscopy.

    PubMed

    Ueno, Yoshifumi; Aikawa, Shimpei; Kondo, Akihiko; Akimoto, Seiji

    2015-08-01

    Photosynthetic organisms change the quantity and/or quality of their pigment-protein complexes and the interactions among these complexes in response to light conditions. In the present study, we analyzed light adaptation of the unicellular red alga Cyanidioschyzon merolae, whose pigment composition is similar to that of cyanobacteria because its phycobilisomes (PBS) lack phycoerythrin. C. merolae were grown under different light qualities, and their responses were measured by steady-state absorption, steady-state fluorescence, and picosecond time-resolved fluorescence spectroscopies. Cells were cultivated under four monochromatic light-emitting diodes (blue, green, yellow, and red), and changes in pigment composition and energy transfer were observed. Cells grown under blue and green light increased their relative phycocyanin levels compared with cells cultured under white light. Energy-transfer processes to photosystem I (PSI) were sensitive to yellow and red light. The contribution of direct energy transfer from PBS to PSI increased only under yellow light, while red light induced a reduction in energy transfer from photosystem II to PSI and an increase in energy transfer from light-harvesting chlorophyll protein complex I to PSI. Differences in pigment composition, growth, and energy transfer under different light qualities are discussed. PMID:25577254

  12. Full time-resolved diffuse fluorescence tomography accelerated with parallelized Fourier-series truncated diffusion approximation.

    PubMed

    Yi, Xi; Wang, Bingyuan; Wan, Wenbo; Wang, Yihan; Zhang, Yanqi; Zhao, Huijuan; Gao, Feng

    2015-05-01

    Of the three measurement schemes established for diffuse fluorescence tomography (DFT), the time-domain scheme is well known to provide the richest information about the distribution of the targeting fluorophore in living tissues. However, the explicit use of the full time-resolved data usually leads to a considerably lengthy time for image reconstruction, limiting its applications to three-dimensional or small-volume imaging. To cope with the adversity, we propose herein a computationally efficient scheme for DFT image reconstruction where the time-dependent photon density is expanded to a Fourier-series and calculated by solving the independent frequency-domain diffusion equations at multiple sampling frequencies with the support of a combined multicore CPU-based coarse-grain and multithread GPU-based fine-grain parallelization strategy. With such a parallelized Fourier-series truncated diffusion approximation, both the time- and frequency-domain inversion procedures are developed and validated for their effectiveness and accuracy using simulative and phantom experiments. The results show that the proposed method can generate reconstructions comparable to the explicit time-domain scheme, with significantly reduced computational time.

  13. Validation and evaluation of a novel time-resolved laser-induced fluorescence technique.

    PubMed

    Durot, C J; Gallimore, A D; Smith, T B

    2014-01-01

    We present a novel technique to measure time-resolved laser-induced fluorescence signals in plasma sources that have a relatively constant Fourier spectrum of oscillations in steady-state operation, but are not periodically pulsed, e.g., Hall thrusters. The technique uses laser modulation of the order of MHz and recovers signal via a combination of band-pass filtering, phase-sensitive detection, and averaging over estimated transfer functions calculated for many different cycles of the oscillation. Periodic discharge current oscillations were imposed on a hollow cathode. Measurements were validated by comparison with independent measurements from a lock-in amplifier and by comparing the results of the transfer function average to an independent analysis technique triggering averaging over many oscillation cycles in the time domain. The performance of the new technique is analyzed and compared to prior techniques, and it is shown that this new technique has a niche in measurements where the analog photomultiplier signal has a nonwhite noise spectral density and cycles of oscillation are not sufficiently repeatable to allow for reliable triggering or a meaningful average waveform in the time domain.

  14. Lasing dynamics study by femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy

    NASA Astrophysics Data System (ADS)

    Wei, Dang; Qing, Liao; Peng-Cheng, Mao; Hong-Bing, Fu; Yu-Xiang, Weng

    2016-05-01

    Femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy (FNOPAS) is a versatile technique with advantages of high sensitivity, broad detection bandwidth, and intrinsic spectrum correction function. These advantages should benefit the study of coherent emission, such as measurement of lasing dynamics. In this letter, the FNOPAS was used to trace the lasing process in Rhodamine 6G (R6G) solution and organic semiconductor nano-wires. High-quality transient emission spectra and lasing dynamic traces were acquired, which demonstrates the applicability of FNOPAS in the study of lasing dynamics. Our work extends the application scope of the FNOPAS technique. Project supported by the National Natural Science Foundation of China (Grant Nos. 20925313 and 21503066), the Innovation Program of Chinese Academy of Sciences (Grant No. KJCX2-YW-W25), the Postdoctoral Project of Hebei University, China, and the Project of Science and Technology Bureau of Baoding City, China (Grant No. 15ZG029).

  15. Simultaneous detection of sulfamethazine and sulfaquinoxaline using a dual-label time-resolved fluorescence immunoassay.

    PubMed

    Le, Tao; Yan, Peifeng; Liu, Jin; Wei, Shu

    2013-01-01

    A dual-label time-resolved fluoroimmunoassay (TRFIA) was introduced for the simultaneous quantification of sulfamethazine (SM2) and sulfaquinoxaline (SQX). Lanthanide (Eu(3+) and Sm(3+))-labelled antibodies were used because lanthanides have higher stabilities and narrower emission spectra than most fluorescent dyes. The sensitivity of the TRFIA for SM2 was 0.02 ng ml(-1), and the average recoveries and the intra- and inter-assay CVs were 77.2-107.6%, 5.4-10.5%, and 6.0-11.2%, respectively. The sensitivity of the TRFIA for SQX was 0.04 ng ml(-1); and the average recoveries and the intra- and inter-assay CVs were 74.1-102.8%, 4.6-10.9%, and 8.7-11.2%, respectively. The method was used to analyse chicken tissue and egg samples, and the results agreed well with the results of HPLC and enzyme-linked immunosorbent assay (ELISA) analyses, with correlation coefficients (R(2)) of 0.9415-0.9724. The TRFIA developed is a simple, fast and sensitive method for the high-throughput simultaneous screening of SM2 and SQX in edible animal tissues. PMID:23782396

  16. Conformational transitions in the calcium adenosinetriphosphatase studied by time-resolved fluorescence resonance energy transfer.

    PubMed

    Birmachu, W; Nisswandt, F L; Thomas, D D

    1989-05-01

    We have used time-resolved fluorescence to study proposed conformational transitions in the Ca-ATPase in skeletal sarcoplasmic reticulum (SR). Resonance energy transfer was used to measure distances between the binding sites of 5-[[2-[(iodoacetyl)amino]ethyl]amino]naphthalene-1-sulfonic acid (IAEDANS) and fluorescein 5-isothiocyanate (FITC) as a function of conditions proposed to affect the enzyme's conformation. When 1.0 +/- 0.15 IAEDANS is bound per Ca-ATPase, most (76 +/- 4%) of the probes have an excited-state lifetime (tau) of 18.6 +/- 0.5 ns, and the remainder have a lifetime of 2.5 +/- 0.9 ns. When FITC is bound to a specific site on each IAEDANS-labeled enzyme, most of the long-lifetime component is quenched into two short-lifetime components, indicating energy transfer that corresponds to two donor-acceptor distances. About one-third of the quenched population has a lifetime tau = 11.1 +/- 2.5 ns, corresponding to a transfer efficiency E = 0.40 +/- 0.07 and a donor-acceptor distance R1 = 52 +/- 3 A. The remaining two-thirds exhibit lifetimes in the range of 1.2-4.2 ns, corresponding to a second distance 31 A less than or equal to R2 less than or equal to 40 A. Addition of Ca2+ (in the micromolar to millimolar range), or vanadate (to produce a phosphoenzyme analogue), had no effect on the donor-acceptor distances. Addition of decavanadate results in the quenching of IAEDANS fluorescence but has no effect on the energy-transfer distance.(ABSTRACT TRUNCATED AT 250 WORDS)

  17. Structure and function of the photoreceptor stentorins in Stentor coeruleus. II. Primary photoprocess and picosecond time-resolved fluorescence.

    PubMed

    Song, P S; Kim, I H; Florell, S; Tamai, N; Yamazaki, T; Yamazaki, I

    1990-08-01

    Stentorin serves as the photoreceptor for the photophobic and negative phototactic responses in Stentor coeruleus. Two forms of the stentorin have been isolated and purified. The strongly fluorescent form, stentorin I at pH 7.8, exhibited nearly exponential fluorescence decay monitored at 620 nm, having two comparable lifetime decay components of 2.53 ns (47%) and 5.95 ns (53%). Stentorin I showed no significant time-resolved fluorescence emission spectra in the picosecond-nanosecond time scales. The weakly fluorescent form, stentorin II, exhibited an ultrafast fluorescence decay component (10 ps) at an emission wavelength of 630 nm and pH 7.8. The amplitudes of the multi-component fluorescence in stentorin II were found to be emission wavelength-dependent. Furthermore, the fluorescence emission spectrum was time-resolvable in the picosecond time scales. Effects of pH and pD on the fluorescence decay kinetics and time-resolved spectra of stentorins I and II have also been investigated. Results are suggestive of proton dissociation as a primary photoprocess from the excited state of stentorin II. PMID:2378902

  18. Steady-State and Time-Resolved Studies into the Origin of the Intrinsic Fluorescence of G-Quadruplexes.

    PubMed

    Sherlock, Madeline E; Rumble, Christopher A; Kwok, Chun Kit; Breffke, Jens; Maroncelli, Mark; Bevilacqua, Philip C

    2016-06-16

    Stretches of guanines in DNA and RNA can fold into guanine quadruplex structures (GQSs). These structures protect telomeres in DNA and regulate gene expression in RNA. GQSs have an intrinsic fluorescence that is sensitive to different parameters, including loop sequence and length. However, the dependence of GQS fluorescence on solution and sequence parameters and the origin of this fluorescence are poorly understood. Herein we examine effects of dangling nucleotides and cosolute conditions on GQS fluorescence using both steady-state and time-resolved fluorescence spectroscopy. The quantum yield of dGGGTGGGTGGGTGGG, termed "dG3T", is found to be modest at ∼2 × 10(-3). Nevertheless, dG3T and its variants are significantly brighter than the common nucleic acid fluorophore 2-aminopurine (2AP) largely due to their sizable extinction coefficients. Dangling 5'-end nucleotides generally reduce emission and blue-shift the resultant spectrum, whereas dangling 3'-end nucleotides slightly enhance fluorescence, particularly on the red side of the emission band. Time-resolved fluorescence decays are broadly distributed in time and require three exponential components for accurate fits. Time-resolved emission spectra suggest the presence of two emitting populations centered at ∼330 and ∼390 nm, with the redder component being a well-defined long-lived (∼1 ns) entity. Insights into GQS fluorescence obtained here should be useful in designing brighter intrinsic RNA and DNA quadruplexes for use in label-free biotechnological applications.

  19. Detection of rhodopsin dimerization in situ by PIE-FCCS, a time-resolved fluorescence spectroscopy.

    PubMed

    Smith, Adam W

    2015-01-01

    Rhodopsin self-associates in the plasma membrane. At low concentrations, the interactions are consistent with a monomer-dimer equilibrium (Comar et al., J Am Chem Soc 136(23):8342-8349, 2014). At high concentrations in native tissue, higher-order clusters have been observed (Fotiadis et al., Nature 421:127-128, 2003). The physiological role of rhodopsin dimerization is still being investigated, but it is clear that a quantitative assessment is essential to determining the function of rhodopsin clusters in vision. To quantify rhodopsin interactions, I will outline the theory and methodology of a specialized time-resolved fluorescence spectroscopy for measuring membrane protein-protein interactions called pulsed-interleaved excitation fluorescence cross-correlation spectroscopy (PIE-FCCS). The strength of this technique is its ability to quantify rhodopsin interactions in situ (i.e., a live cell plasma membrane). There are two reasons for restricting the scope to live cell membranes. First, the compositional heterogeneity of the plasma membrane creates a complex milieu with thousands of lipid, protein, and carbohydrate species. This makes it difficult to infer quaternary interactions from detergent solubilized samples or construct a model phospholipid bilayer that recapitulates all of the interactions present in native membranes. Second, organizational structure and dynamics is a key feature of the plasma membrane, and fixation techniques like formaldehyde cross-linking and vitrification will modulate the interactions. PIE-FCCS is based on two-color fluorescence imaging with time-correlated single-photon counting (TCSPC) (Becker et al., Rev Sci Instrum 70:1835-1841, 1999). By time-tagging every detected photon, the data can be analyzed as a fluorescence intensity distribution, fluorescence lifetime histogram, or fluorescence (cross-)correlation spectra (FCS/FCCS) (Becker, Advanced time-correlated single-photon counting techniques, Springer, Berlin, 2005). These

  20. Time-Resolved Fluorescent Immunochromatography of Aflatoxin B1 in Soybean Sauce: A Rapid and Sensitive Quantitative Analysis.

    PubMed

    Wang, Du; Zhang, Zhaowei; Li, Peiwu; Zhang, Qi; Zhang, Wen

    2016-01-01

    Rapid and quantitative sensing of aflatoxin B1 with high sensitivity and specificity has drawn increased attention of studies investigating soybean sauce. A sensitive and rapid quantitative immunochromatographic sensing method was developed for the detection of aflatoxin B1 based on time-resolved fluorescence. It combines the advantages of time-resolved fluorescent sensing and immunochromatography. The dynamic range of a competitive and portable immunoassay was 0.3-10.0 µg·kg(-1), with a limit of detection (LOD) of 0.1 µg·kg(-1) and recoveries of 87.2%-114.3%, within 10 min. The results showed good correlation (R² > 0.99) between time-resolved fluorescent immunochromatographic strip test and high performance liquid chromatography (HPLC). Soybean sauce samples analyzed using time-resolved fluorescent immunochromatographic strip test revealed that 64.2% of samples contained aflatoxin B1 at levels ranging from 0.31 to 12.5 µg·kg(-1). The strip test is a rapid, sensitive, quantitative, and cost-effective on-site screening technique in food safety analysis. PMID:27428975

  1. Time-Resolved Fluorescent Immunochromatography of Aflatoxin B1 in Soybean Sauce: A Rapid and Sensitive Quantitative Analysis

    PubMed Central

    Wang, Du; Zhang, Zhaowei; Li, Peiwu; Zhang, Qi; Zhang, Wen

    2016-01-01

    Rapid and quantitative sensing of aflatoxin B1 with high sensitivity and specificity has drawn increased attention of studies investigating soybean sauce. A sensitive and rapid quantitative immunochromatographic sensing method was developed for the detection of aflatoxin B1 based on time-resolved fluorescence. It combines the advantages of time-resolved fluorescent sensing and immunochromatography. The dynamic range of a competitive and portable immunoassay was 0.3–10.0 µg·kg−1, with a limit of detection (LOD) of 0.1 µg·kg−1 and recoveries of 87.2%–114.3%, within 10 min. The results showed good correlation (R2 > 0.99) between time-resolved fluorescent immunochromatographic strip test and high performance liquid chromatography (HPLC). Soybean sauce samples analyzed using time-resolved fluorescent immunochromatographic strip test revealed that 64.2% of samples contained aflatoxin B1 at levels ranging from 0.31 to 12.5 µg·kg−1. The strip test is a rapid, sensitive, quantitative, and cost-effective on-site screening technique in food safety analysis. PMID:27428975

  2. Dynamics and Flexibility of Human Aromatase Probed by FTIR and Time Resolved Fluorescence Spectroscopy

    PubMed Central

    Sadeghi, Sheila J.; Castrignanò, Silvia; Mei, Giampiero; Di Venere, Almerinda; Nicolai, Eleonora; Allegra, Paola; Gilardi, Gianfranco

    2013-01-01

    Human aromatase (CYP19A1) is a steroidogenic cytochrome P450 converting androgens into estrogens. No ligand-free crystal structure of the enzyme is available to date. The crystal structure in complex with the substrate androstenedione and the steroidal inhibitor exemestane shows a very compact conformation of the enzyme, leaving unanswered questions on the conformational changes that must occur to allow access of the ligand to the active site. As H/D exchange kinetics followed by FTIR spectroscopy can provide information on the conformational changes in proteins where solvent accessibility is affected, here the amide I region was used to measure the exchange rates of the different elements of the secondary structure for aromatase in the ligand-free form and in the presence of the substrate androstenedione and the inhibitor anastrozole. Biphasic exponential functions were found to fit the H/D exchange data collected as a function of time. Two exchange rates were assigned to two populations of protons present in different flexible regions of the protein. The addition of the substrate androstenedione and the inhibitor anastrozole lowers the H/D exchange rates of the α-helices of the enzyme when compared to the ligand-free form. Furthermore, the presence of the inhibitor anastrozole lowers exchange rate constant (k1) for β-sheets from 0.22±0.06 min−1 for the inhibitor-bound enzyme to 0.12±0.02 min−1 for the free protein. Dynamics effects localised in helix F were studied by time resolved fluorescence. The data demonstrate that the fluorescence lifetime component associated to Trp224 emission undergoes a shift toward longer lifetimes (from ≈5.0 to ≈5.5 ns) when the substrate or the inhibitor are present, suggesting slower dynamics in the presence of ligands. Together the results are consistent with different degrees of flexibility of the access channel and therefore different conformations adopted by the enzyme in the free, substrate- and inhibitor

  3. Highly sensitive detection of human papillomavirus type 16 DNA using time-resolved fluorescence microscopy and long lifetime probes

    NASA Astrophysics Data System (ADS)

    Wang, Xue F.; Periasamy, Ammasi; Wodnicki, Pawel; Siadat-Pajouh, M.; Herman, Brian

    1995-04-01

    We have been interested in the role of Human Papillomavirus (HPV) in cervical cancer and its diagnosis; to that end we have been developing microscopic imaging and fluorescent in situ hybridization (FISH) techniques to genotype and quantitate the amount of HPV present at a single cell level in cervical PAP smears. However, we have found that low levels of HPV DNA are difficult to detect accurately because theoretically obtainable sensitivity is never achieved due to nonspecific autofluorescence, fixative induced fluorescence of cells and tissues, and autofluorescence of the optical components in the microscopic system. In addition, the absorption stains used for PAP smears are intensely autofluorescent. Autofluorescence is a rapidly decaying process with lifetimes in the range of 1-100 nsec, whereas phosphorescence and delayed fluorescence have lifetimes in the range of 1 microsecond(s) ec-10 msec. The ability to discriminate between specific fluorescence and autofluorescence in the time-domain has improved the sensitivity of diagnostic test such that they perform comparably to, or even more sensitive than radioisotopic assays. We have developed a novel time-resolved fluorescence microscope to improve the sensitivity of detection of specific molecules of interest in slide based specimens. This time-resolved fluorescence microscope is based on our recently developed fluorescence lifetime imaging microscopy (FILM) in conjunction with the use of long lifetime fluorescent labels. By using fluorescence in situ hybridization and the long lifetime probe (europium), we have demonstrated the utility of this technique for detection of HPV DNA in cervicovaginal cells. Our results indicate that the use of time-resolved fluorescence microscopy and long lifetime probes increases the sensitivity of detection by removing autofluorescence and will thus lead to improved early diagnosis of cervical cancer. Since the highly sensitive detection of DNA in clinical samples using

  4. Nanosecond time-resolved emission spectroscopy of a fluorescence probe adsorbed to L-alpha-egg lecithin vesicles.

    PubMed Central

    Easter, J H; DeToma, R P; Brand, L

    1976-01-01

    Nanosecond time-resolved emission spectra (TRES) are fluorescence emission spectra obtained at discrete times during the fluorescence decay. The complete data-set obtainable is a surface representing the intensity at all wavelengths and times during the emission decay time. When 2-p-toluidinonaphthalene-6-sulfonate (2,6 p-TNS) is adsorbed to egg lecithin vesicles, an excited-state reaction associated with energetic changes of the emitting species occurs on the nanosecond time scale. Convolution of the fluorescence decay with the excitation response introduces an artifact in the time-dependent spectra. A precedure is described by which this artifact can be eliminated. The data for the generation of time-resolved emission spectra are obtained with a computer-interfaced instrument based on the single-photon counting method. PMID:945086

  5. Comparison of beetroot extracts originating from several sites using time-resolved laser-induced fluorescence spectroscopy

    NASA Astrophysics Data System (ADS)

    Rabasović, M. S.; Šević, D.; Terzić, M.; Marinković, B. P.

    2012-05-01

    Beetroot (Beta vulgaris) juice contains a large number of fluorophores which can fluoresce. There is a growing interest in beetroot extracts analysis. In contrast, there is only limited information about beetroot obtained without sample preparation and/or extraction of components from the sample. In this work, we continue our previous study (Rabasović et al 2009 Acta Phys. Pol. A 116 570-2), analyzing and comparing beetroot extracts from several sites, using the time-resolved laser-induced fluorescence technique to measure the fluorescence of samples at different excitation wavelengths (340-470 nm) and for different sample dilutions.

  6. Time-Resolved Fluorescence Studies Of Ph Effects On The Conformation Of Troponin C

    NASA Astrophysics Data System (ADS)

    Wang, Chien-Kao; Liao, Ronglihi; Cheung, Herbert C.

    1988-06-01

    Using time-resolved nanosecond fluorescence spectroscopy, we investigated the conformational changes of skeletal and cardiac troponin C. A thiol specific fluorescence probe N-(iodoacety1)-N'-(5-sulfo-1-naphthyl)- ethylenediamine (IAEDANS) was attached to cysteine 98 of skeletal troponin C (STnC) and 2-(4'-iodoacetamido-anilino)-naphthalene-6-sulfonic acid (IAANS) was linked to cysteine 35 and 84 of cardiac troponin C (CTnC). With excitation at 340 nm for STnC-IAEDANS and at 335 nm for CTnC-IAANS, apo-STnC and apo-CTnC exhibited biexponential decay kinetics. At 20°C and neutral pH, the following lifetimes were observed: (1) apo-STnC, 9 and 16 ns, and (2) apo-CTnC, 2.3 and 7 ns. The long lived component of the emission in STnC-IAEDANS comprised ~ 61% of observed intensity, however, the corresponding component in CTnC contributed only. A decrease of pH from 7.2 to 5.2 induced an increase of the lifetimes 20% (STnC) and 10% (CTnC). These results suggest that Cys-98 of STnC and Cys-35 and Cys-84 of CTnC became less quenched by their neighboring residues at low pH. Addition of guanidine hydrochloride to STnC resulted in a decrease of ~30% of both lifetimes. The lifetimes increased slightly when the temperature was lowered. Variation of solution viscosity by addition of sucrose did not affect the long component of the lifetimes of STnC. However, the short component did sense a viscosity effect. These results suggest that there was likely a chromophore heterogeneity which may arise from differences in conformation, environment, and or ionization of the excited state of the chromophore. At 20°C and neutral pH, two rotational correlation times were observed: (1) apo-STnC, - 1.2 ns, .2 1 11.3 ns, and (2) apo-CTnC, .~ 0.6 ns, .2 1 13.6 ns. The short rotational correlation times likely reflect rapid motions of the chromophores covalently attached to the side chain of the cysteine residues, and the long correlation times reflect the overall protein motions. The .2 values

  7. A Vinblastine Fluorescent Probe for Pregnane X Receptor in a Time-Resolved Fluorescence Resonance Energy Transfer Assay

    PubMed Central

    Lin, Wenwei; Chen, Taosheng

    2013-01-01

    The pregnane X receptor (PXR) regulates the metabolism and excretion of xenobiotics and endobiotics by regulating the expression of drug-metabolizing enzymes and transporters. The unique structure of PXR allows the binding of many drugs and drug leads to it, possibly causing undesired drug-drug interactions. Therefore, it is crucial to evaluate whether lead compounds bind to PXR. Fluorescence-based assays are preferred because of their sensitivity and non-radioactive nature. One fluorescent PXR probe is currently commercially available; however, because its chemical structure is not publicly disclosed, it is not optimal for studying ligand-PXR interactions. Here we report the characterization of BODIPY FL Vinblastine, generated by labeling vinblastine with the fluorophore 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene (BODIPY FL), as a high-affinity ligand for human PXR with a Kd value of 673 nM. We provide evidence that BODIPY FL Vinblastine is a unique chemical entity different from either vinblastine or the fluorophore 4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene in its function as a high-affinity human PXR ligand. We describe a BODIPY FL Vinblastine-based human PXR Time-Resolved Fluorescence Resonance Energy Transfer assay, which was used to successfully test a panel of human PXR ligands. The BODIPY FL Vinblastine–based biochemical assay is suitable for high-throughput screening to evaluate whether lead compounds bind to PXR. PMID:24044991

  8. Computational modeling of time-resolved fluorescence transport in turbid media for non-invasive clinical diagnostics

    NASA Astrophysics Data System (ADS)

    Vishwanath, Karthik

    Fluorescence spectroscopy and imaging methods, including fluorescence lifetime sensing, are being developed for a variety of non-invasive clinical diagnostic procedures, including applications to early cancer diagnosis. Here, both the theoretical developments and experimental validations of a versatile, numerical Monte Carlo code that models photon migration in turbid media to include simulations of time-resolved fluorescence transport are presented. The developed numerical model was used to study, for the first time, the dependence of time-resolved fluorescence signals emanating from turbid media on the optical transport coefficients, fluorophore properties and source-detector configurations in single-layered turbid media as well as more complex multi-layered turbid media. The numerical codes presented here can be adapted to model a wide range of experimental techniques measuring the optical responses of biological tissues to laser irradiation and are demonstrated here for two specific applications (a) to model time-resolved fluorescence dynamics in human colon tissues and (b) to extract the frequency-dependent optical responses of a model adult human head to an incident laser-source whose intensity was harmonically modulated i.e. simulating frequency-domain measurements. Specifically, measurements of time-resolved fluorescence decays from a previous clinical study aimed toward detecting differences in tissue pathologies in patients undergoing gastro-intestinal endoscopy were simulated using the Monte Carlo model and results demonstrated that variations in tissue optical transport coefficients (absorption and scattering) alone could not account for the fluorescence decay differences detected between tissue pathologies in vivo. However, variations in fluorescence decay time as large as those detected clinically between normal and pre-malignant tissues (of 2 ns) could be accounted for by simulated variations in tissue morphology or biochemistry while intrinsic

  9. Time-resolved fluorescence of thioredoxin single-tryptophan mutants: modeling experimental results with minimum perturbation mapping

    NASA Astrophysics Data System (ADS)

    Silva, Norberto D., Jr.; Haydock, Christopher; Prendergast, Franklyn G.

    1994-08-01

    The time-resolved fluorescence decay of single tryptophan (Trp) proteins is typically described using either a distribution of lifetimes or a sum of two or more exponential terms. A possible interpretation for this fluorescence decay heterogeneity is the existence of different isomeric conformations of Trp about its (chi) +1) and (chi) +2) dihedral angles. Are multiple Trp conformations compatible with the remainder of the protein in its crystallographic configuration or do they require repacking of neighbor side chains? It is conceivable that isomers of the neighbor side chains interconvert slowly on the fluorescence timescale and contribute additional lifetime components to the fluorescence intensity. We have explored this possibility by performing minimum perturbation mapping simulations of Trp 28 and Trp 31 in thioredoxin (TRX) using CHARMm 22. Mappings of Trp 29 and Trp 31 give the TRX Trp residue energy landscape as a function of (chi) +1) and (chi) +2) dihedral angles. Time-resolved fluorescence intensity and anisotropy decay of mutant TRX (W28F and W31F) are measured and interpreted in light of the above simulations. Relevant observables, like order parameters and isomerization rates, can be derived from the minimum perturbation maps and compared with experiment.

  10. Multispectral fluorescence lifetime imaging of feces-contaminated apples by time-resolved laser-induced fluorescence imaging system with tunable excitation wavelengths

    NASA Astrophysics Data System (ADS)

    Kim, Moon S.; Cho, Byoung-Kwan; Lefcourt, Alan M.; Chen, Yud-Ren; Kang, Sukwon

    2008-04-01

    We recently developed a time-resolved multispectral laser-induced fluorescence (LIF) imaging system capable of tunable wavelengths in the visible region for sample excitation and nanosecond-scale characterizations of fluorescence responses (lifetime imaging). Time-dependent fluorescence decay characteristics and fluorescence lifetime imaging of apples artificially contaminated with a range of diluted cow feces were investigated at 670 and 685 nm emission bands obtained by 418, 530, and 630 nm excitations. The results demonstrated that a 670 nm emission with a 418 nm excitation provided the greatest difference in time-dependent fluorescence responses between the apples and feces-treated spots. The versatilities of the time-resolved LIF imaging system, including fluorescence lifetime imaging of a relatively large biological object in a multispectral excitation-emission wavelength domain, were demonstrated.

  11. Time-resolved fluorescence for breast cancer detection using an octreotate-indocyanine green derivative dye conjugate

    NASA Astrophysics Data System (ADS)

    Sordillo, Laura A.; Das, B. B.; Pu, Yang; Liang, Kexian; Milione, Giovanni; Sordillo, Peter P.; Achilefu, Sam; Alfano, R. R.

    2013-03-01

    Time-resolved fluorescence was used to investigate malignant and normal adjacent breast tissues stained with a conjugate of indocyanine green and octreotate. A marked increase in fluorescence lifetime intensity was seen in the breast cancer sample compared to the normal sample. The fluorescent lifetimes were also investigated and showed similar fluorescence decay curves in stained malignant and normal breast tissue. These results confirm that somatostatin receptors occur on human breast carcinomas, suggest that the presence of somatostatin receptors should be investigated as a marker of breast cancer aggressiveness, and suggest that this conjugate might be used to detect the presence of residual breast cancer after surgery, allowing better assessment of tumor margins and reducing the need for second or repeat biopsies in selected patients. These results may also provide clues for designing future treatment options for breast cancer patients.

  12. 340 nm pulsed UV LED system for europium-based time-resolved fluorescence detection of immunoassays.

    PubMed

    Rodenko, Olga; Fodgaard, Henrik; Tidemand-Lichtenberg, Peter; Petersen, Paul Michael; Pedersen, Christian

    2016-09-19

    We report on the design, development and investigation of an optical system based on UV light emitting diode (LED) excitation at 340 nm for time-resolved fluorescence detection of immunoassays. The system was tested to measure cardiac marker Troponin I with a concentration of 200 ng/L in immunoassay. The signal-to-noise ratio was comparable to state-of-the-art Xenon flash lamp based unit with equal excitation energy and without overdriving the LED. We performed a comparative study of the flash lamp and the LED based system and discussed temporal, spatial, and spectral features of the LED excitation for time-resolved fluorimetry. Optimization of the suggested key parameters of the LED promises significant increase of the signal-to-noise ratio and hence of the sensitivity of immunoassay systems. PMID:27661948

  13. 340 nm pulsed UV LED system for europium-based time-resolved fluorescence detection of immunoassays.

    PubMed

    Rodenko, Olga; Fodgaard, Henrik; Tidemand-Lichtenberg, Peter; Petersen, Paul Michael; Pedersen, Christian

    2016-09-19

    We report on the design, development and investigation of an optical system based on UV light emitting diode (LED) excitation at 340 nm for time-resolved fluorescence detection of immunoassays. The system was tested to measure cardiac marker Troponin I with a concentration of 200 ng/L in immunoassay. The signal-to-noise ratio was comparable to state-of-the-art Xenon flash lamp based unit with equal excitation energy and without overdriving the LED. We performed a comparative study of the flash lamp and the LED based system and discussed temporal, spatial, and spectral features of the LED excitation for time-resolved fluorimetry. Optimization of the suggested key parameters of the LED promises significant increase of the signal-to-noise ratio and hence of the sensitivity of immunoassay systems.

  14. Ns-scale time-resolved laser induced fluorescence imaging for detection of fecal contamination on apples

    NASA Astrophysics Data System (ADS)

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

    2004-11-01

    Our laboratory has been utilizing fluorescence techniques as a potential means for detection of quality and wholesomeness of food products. A system with a short pulse light source such as a laser coupled with a gated detector can be used to harvest fluorescence in ambient light conditions from biological samples with relatively low fluorescence yields. We present a versatile multispectral laser-induced fluorescence (LIF) imaging system capable of ns-scale time resolved fluorescence. The system is equipped with a tunable pulse laser system that emits in the visible range from 410 nm to 690 nm. Ns-scale, time-dependent multispectral fluorescence emissions of apples contaminated with a range of diluted cow feces were acquired. Four spectral bands, F670, F680, F685 and F730, centered near the emission peak wavelengths of the major constituents responsible for the red fluorescence emissions from apples artificially contaminated with cow feces were examined to determine a suitable single red fluorescence band and optimal ns-gate window for detection of fecal contamination on apples. The results based on the ns decay curves showed that 670 nm with 10 nm full width at half maximum (FWHM) at a gate-delay of 4 ns from the laser excitation peak provided the greatest differences in time-dependent fluorescence responses between feces contaminated spots and apples surfaces.

  15. Energy transfer in the chlorophyll f-containing cyanobacterium, Halomicronema hongdechloris, analyzed by time-resolved fluorescence spectroscopies.

    PubMed

    Akimoto, Seiji; Shinoda, Toshiyuki; Chen, Min; Allakhverdiev, Suleyman I; Tomo, Tatsuya

    2015-08-01

    We prepared thylakoid membranes from Halomicronema hongdechloris cells grown under white fluorescent light or light from far-red (740 nm) light-emitting diodes, and observed their energy-transfer processes shortly after light excitation. Excitation-relaxation processes were examined by steady-state and time-resolved fluorescence spectroscopies. Two time-resolved fluorescence techniques were used: time-correlated single photon counting and fluorescence up-conversion methods. The thylakoids from the cells grown under white light contained chlorophyll (Chl) a of different energies, but were devoid of Chl f. At room temperature, the excitation energy was equilibrated among the Chl a pools with a time constant of 6.6 ps. Conversely, the thylakoids from the cells grown under far-red light possessed both Chl a and Chl f. Two energy-transfer pathways from Chl a to Chl f were identified with time constants of 1.3 and 5.0 ps, and the excitation energy was equilibrated between the Chl a and Chl f pools at room temperature. We also examined the energy-transfer pathways from phycobilisome to the two photosystems under white-light cultivation.

  16. Time-resolved and steady-state fluorescence spectroscopy for the assessment of skin photoaging process

    NASA Astrophysics Data System (ADS)

    D´Almeida, Camila de Paula; Campos, Carolina; Saito Nogueira, Marcelo; Pratavieira, Sebastião.; Kurachi, Cristina

    2015-06-01

    pathology. The optical properties of these intrinsic fluorophores respond to the microenvironment and the metabolic status, thus making fluorescence spectroscopy a valuable tool to study the conditions of biological tissues. The purpose of this study is to investigate the hairless mice skin metabolic changes during the photoaging process through lifetime and fluorescence measurements targeting NADH and FAD. Two lasers centered at 378 nm and 445 nm, respectively, perform excitation of NADH and FAD. The fluorescence acquisition is carried out at mice dorsal and ventral regions throughout the photoaging protocol and aging process. Differences in fluorescence and lifetime data between young and photoaged mice measurements were observed. The endogenous fluorescence spectrum of photoaged dorsal skin showed an increase compared to young and aged skin. Lifetime of bound NADH and free FAD presented an increase in the first week that continued until the end of the protocol. Aging process is being investigated to complement the information obtained from fluorescence data and lifetime of photoaging process.

  17. Time-resolved multicolor two-photon excitation fluorescence microscopy of cells and tissues

    NASA Astrophysics Data System (ADS)

    Zheng, Wei

    2014-11-01

    Multilabeling which maps the distribution of different targets is an indispensable technique in many biochemical and biophysical studies. Two-photon excitation fluorescence (TPEF) microscopy of endogenous fluorophores combining with conventional fluorescence labeling techniques such as genetically encoded fluorescent protein (FP) and fluorescent dyes staining could be a powerful tool for imaging living cells. However, the challenge is that the excitation and emission wavelength of these endogenous fluorophores and fluorescent labels are very different. A multi-color ultrafast source is required for the excitation of multiple fluorescence molecules. In this study, we developed a two-photon imaging system with excitations from the pump femtosecond laser and the selected supercontinuum generated from a photonic crystal fiber (PCF). Multiple endogenous fluorophores, fluorescent proteins and fluorescent dyes were excited in their optimal wavelengths simultaneously. A time- and spectral-resolved detection system was used to record the TPEF signals. This detection technique separated the TPEF signals from multiple sources in time and wavelength domains. Cellular organelles such as nucleus, mitochondria, microtubule and endoplasmic reticulum, were clearly revealed in the TPEF images. The simultaneous imaging of multiple fluorophores of cells will greatly aid the study of sub-cellular compartments and protein localization.

  18. Correlative Time-Resolved Fluorescence Microscopy To Assess Antibiotic Diffusion-Reaction in Biofilms

    PubMed Central

    Daddi Oubekka, S.; Briandet, R.; Fontaine-Aupart, M.-P.

    2012-01-01

    The failure of antibiotics to inactivate in vivo pathogens organized in biofilms has been shown to trigger chronic infections. In addition to mechanisms involving specific genetic or physiological cell properties, antibiotic sorption and/or reaction with biofilm components may lessen the antibiotic bioavailability and consequently decrease their efficiency. To assess locally and accurately the antibiotic diffusion-reaction, we used for the first time a set of advanced fluorescence microscopic tools (fluorescence recovery after photobleaching, fluorescence correlation spectroscopy, and fluorescence lifetime imaging) that offer a spatiotemporal resolution not available with the commonly used time-lapse confocal imaging method. This set of techniques was used to characterize the dynamics of fluorescently labeled vancomycin in biofilms formed by two Staphylococcus aureus human isolates. We demonstrate that, at therapeutic concentrations of vancomycin, the biofilm matrix was not an obstacle to the diffusion-reaction of the antibiotic that can reach all cells through the biostructure. PMID:22450986

  19. Two-Dimensional Subpicosecond Time-Resolved Fluorescence Anisotropy: Optical Kerr-Gating with a Dynamic Polarization Excitation.

    NASA Astrophysics Data System (ADS)

    Fujiwara, Takashige; Romano, Natalie C.; Modarelli, David A.; Lim, Edward C.

    2013-06-01

    With an advent of ultrafast lasers, a number of applications are widely adopted to probe photophysical and photochemical properties of a molecule that occurs in an ultrafast (femtosecond to picosecond) time scale. Intramolecular charge transfer (ICT) or proton transfer in photoexcited electron donor-acceptor (EDA) molecules, for instance, has been a topic of very extensive time-resolved studies for several decades. Time-evolution of an anisotropic property can track dipole orientations or conformational changes in their photoexcited molecular systems, which is of extreme importance to examine its structure and excited-state dynamics rather than probing an isotropic "population change".With this respect, we recently developed a subpicosecond time-resolved 2-D fluorescence anisotropy (TRFA) in which method implements a dynamic alternation of laser polarizations to excite a sample using a photoelastic modulator (PEM). In the combination of an ultrafast optical shutter (Kerr-gating) and a spectrograph that is coupled with a CCD, two signal phases so-obtained dynamically, I_{∥}( t, λ) and I_{⊥}( t, λ), provide a 2-D mapped information on both a wide range for spectra and time-resolved kinetics of photoexcited molecules of interest. From the definition of an anisotropy 2-D TRFA, r (t, λ), is given instantly and even more reliably at a single measurement. In this paper we will present benchmark tests of some target samples to establish performance of TRFA.

  20. Understanding THz and IR signals beneath time-resolved fluorescence from excited-state ab initio dynamics.

    PubMed

    Petrone, Alessio; Donati, Greta; Caruso, Pasquale; Rega, Nadia

    2014-10-22

    The detailed interpretation of time-resolved spectroscopic signals in terms of the molecular rearrangement during a photoreaction or a photophysical event is one of the most important challenges of both experimental and theoretical chemistry. Here we simulate a time-resolved fluorescence spectrum of a dye in aqueous solution, the N-methyl-6-oxyquinolinium betaine, and analyze it in terms of far IR and THz frequency contributions, providing a direct connection to specific molecular motions. To obtain this result, we build up an innovative and general approach based on excited state ab-initio molecular dynamics and a wavelet-based time-dependent frequency analysis of nonstationary signals. We obtain a nice agreement with key parameters of the solvent dynamics, such as the total Stokes shift and the Stokes shift relaxation times. As an important finding, we observe a strong change of specific solute-solvent interactions upon the electronic excitation, with the migration of about 1.5 water molecules from the first solvation shell toward the bulk. In spite of this event, the Stokes shift dynamics is ruled by collective solvent motions in the THz and far IR, which guide and modulate the strong rearrangement of the dye microsolvation. By the relaxation of THz and IR contributions to the emission signal, we can follow and understand in detail the molecularity of the process. The protocol presented here is, in principle, transferable to other time-resolved spectroscopic techniques. PMID:25243826

  1. Transient Absorption and Time-Resolved Fluorescence Studies of Solvated Ruthenium Di-Bipyridine Pseudo-Halide Complexes

    NASA Astrophysics Data System (ADS)

    Compton, R.; Weidinger, D.; Owrutsky, J. C.

    2012-06-01

    Time-resolved IR and fluorescence measurements were performed to probe the vibrational and electronic properties, respectively, of ruthenium di-bipyridine pseudo-halide (Ru(Bpy){_2}(X){_2} (where X = CN, N{_3} or NCS)) complexes. Vibrational energy relaxation (VER) times were determined for the complexes dissolved in dimethyl sulfoxide (DMSO) with a trend in VER time of NCS > CN > N{_3}. A similar trend and comparable absolute rates for NCS- and N3- were previously observed by our group and others for simple inorganic anions in solution, suggesting a minimal contribution due to complexation. Measurements of the VER time of the CN complex in various solvents provide VER times in ethanol (42.3 ps) and DMSO (53.3 ps), which shows that protic solvents promote the relaxation. Time-resolved fluorescence measurements indicate a strong ligand dependence, with a factor of five decrease in the excited electronic state decay time from the CN (215 ns) to the NCS (39 ns) complex. A solvent dependence of the CN complex reveals a nearly 3-fold increase in the fluorescence decay time from acetonitrile (70 ns) to DMSO (215 ns).

  2. Time-resolved hyperspectral single-pixel camera implementation for compressive wide-field fluorescence lifetime imaging

    NASA Astrophysics Data System (ADS)

    Pian, Qi; Yao, Ruoyang; Intes, Xavier

    2016-03-01

    Single-pixel imaging based on compressive sensing theory has been a highlighted technique in the biomedical imaging field for many years. This interest has been driven by the possibility of performing microscopic or macroscopic imaging based on low-cost detector arrays, increased SNR (signal-to-noise ratio) in the acquired data sets and the ability to perform high quality image reconstruction with compressed data sets by exploiting signal sparsity. In this work, we present our recent work in implementing this technique to perform time domain fluorescence-labeled investigations in preclinical settings. More precisely, we report on our time-resolved hyperspectral single-pixel camera for fast, wide-field mapping of molecular labels and lifetime-based quantification. The hyperspectral single-pixel camera implements a DMD (Digital micro-mirror device) to generate optical masks for modulating the illumination field before it is delivered onto the sample and focuses the emission light signals into a multi-anode hyperspectral time-resolved PMT (Photomultiplier tube) to acquire spatial, temporal and spectral information enriched 4-D data sets. Fluorescence dyes with lifetime and spectral contrast are embedded in well plates and thin tissues. L-1 norm based regularization or the least square method, is applied to solve the underdetermined inverse problem during image reconstruction. These experimental results prove the possibility of fast, wide-field mapping of fluorescent labels with lifetime and spectral contrast in thin media.

  3. Steady-state and time-resolved fluorescence studies of stripped Borage oil.

    PubMed

    Smyk, Bogdan; Amarowicz, Ryszard; Szabelski, Mariusz; Gryczynski, Ignacy; Gryczynski, Zygmunt

    2009-07-30

    In this study we explored the spectroscopic properties of Borage oil, particularly the use of fluorescence techniques to investigate the presence of conjugated fatty acids (CFAs). This research has important health and dietary applications. The absorption and fluorescence spectra of different CFAs and Borage oil in ethanol were measured. Time-domain fluorescence was employed to establish the life times of the samples. We found that Borage oil contains 1.2x10(-3) mol L(-1) of alpha-eleostearic acid or its isomer (i.e., a conjugated triene), 1.6x10(-4) mol L(-1) of cis-parinaric acid (i.e., a conjugated tetraene) and 1.1x10(-5) mol L(-1) of c-COPA (i.e., a conjugated pentaene). Because of the three-exponential fluorescence intensity decay for Borage oil, other fatty acids with a four conjugated double bond system could not be excluded. PMID:19523559

  4. Time-resolved fluorescence study of electron transfer in a model peptide system

    NASA Astrophysics Data System (ADS)

    Donald, Fiona; Hungerford, Graham; Moore, Barry D.; Birch, David J. S.

    1994-08-01

    At present there is a great deal of interest in the study of the transference of energy in biological systems. For example, electron transfer is of major importance in many synthetic and biological processes and in nature is mediated by proteins. Information regarding this process is therefore useful in leading to a greater understanding of phenomena such as photosynthesis and respiration. Previous work on protein systems has shown the electron transfer process to be complex to analyze because of the presence of competing pathways. This has led to the use of model systems to simplify the kinetics. We have synthesized novel model systems using peptides containing both a fluorescent methoxy- naphthalene donor and a dicyanoethylene group as a potential electron acceptor and observed fluorescence quenching for both dipeptide and oligopeptide systems. Biexponential fluorescence decay behavior was observed for all donor acceptor systems, with an increase in the amount of the shorter fluorescence decay component on increasing temperature.

  5. Steady-state and time-resolved fluorescence studies of stripped Borage oil.

    PubMed

    Smyk, Bogdan; Amarowicz, Ryszard; Szabelski, Mariusz; Gryczynski, Ignacy; Gryczynski, Zygmunt

    2009-07-30

    In this study we explored the spectroscopic properties of Borage oil, particularly the use of fluorescence techniques to investigate the presence of conjugated fatty acids (CFAs). This research has important health and dietary applications. The absorption and fluorescence spectra of different CFAs and Borage oil in ethanol were measured. Time-domain fluorescence was employed to establish the life times of the samples. We found that Borage oil contains 1.2x10(-3) mol L(-1) of alpha-eleostearic acid or its isomer (i.e., a conjugated triene), 1.6x10(-4) mol L(-1) of cis-parinaric acid (i.e., a conjugated tetraene) and 1.1x10(-5) mol L(-1) of c-COPA (i.e., a conjugated pentaene). Because of the three-exponential fluorescence intensity decay for Borage oil, other fatty acids with a four conjugated double bond system could not be excluded.

  6. TOPICAL REVIEW: Time-resolved fluorescence imaging in biology and medicine

    NASA Astrophysics Data System (ADS)

    Cubeddu, R.; Comelli, D.; D'Andrea, C.; Taroni, P.; Valentini, G.

    2002-05-01

    Fluorescence lifetime imaging is a rather new and effective tool that can be used to study complex biological samples, either at microscopic or macroscopic levels. The map of the fluorescence lifetime allows one to discriminate amongst different fluorophores and to achieve valuable insights into the behaviour of emitting molecules, leading to information like local pH, oxygen concentration in cells, etc. Moreover, the distribution in space of any fluorescent marker achievable with this technique can be exploited for diagnostic purposes in medicine. After a brief introduction on the motivations for applying fluorescence lifetime imaging in biology and medicine, the basic principles of this technique will be addressed. Then, the two possible implementations of fluorescence lifetime imaging (i.e. the frequency domain and the time domain methods) will be presented. For this purpose, special attention will be devoted to practical aspects of image acquisition and processing, especially for what concerns the time domain method. Then, the analysis of the state-of-the-art systems will include a brief discussion on new concepts that have recently been introduced in this research field. Finally, two interesting applications of fluorescence lifetime imaging will be presented. The former refers to skin tumour detection and has been successfully applied in a preliminary clinical trial, the latter regards DNA chips reading and has been tested only at laboratory level, yet it has produced promising results for its future implementation in commercial systems.

  7. Time-resolved laser-induced fluorescence of normal and atherosclerotic coronary artery

    NASA Astrophysics Data System (ADS)

    Marcu, Laura; Maarek, Jean-Michel I.; Fishbein, Michael C.; Grundfest, Warren S.

    1999-07-01

    This study investigates the spectro-temporal fluorescence emission of normal and diseased coronary arteries with graded levels of atherosclerosis. Fluorescence emission of 58 excised human coronary artery samples was induced with N2 laser pulses and detected with a MCP-PMT connected to a digital oscilloscope. The samples were H and E and Movat stained and histologically classified in accordance with AHA classification. An algorithm based on Laguerre expansion of kernels was used to deconvolve the intrinsic fluorescence impulse response function from the measured transient pulse. A biexponential function depicted the fluorescence decay characteristics. We noticed 1) in spectral domain: peak fluorescence intensity was at 380 nm for normal and initial lesions samples and blue-shifted for advanced lesions; intensity at 450-480 nm decreased from approximately 65 percent peak intensity for normal samples to approximately 30 percent for Type V lesions; 2) in time domain: longer lasting emission for the advanced lesions. The decay constants varied as a function emission wavelength and lesion type. For instance the time constants for Type V lesions measured at 390 nm were significantly larger that those measured on normal arterial wall. The fast term decay contributed to a higher degree to the impulse response function for normal tissue. These results reveal that the analysis of the temporal characteristics of fluorescence can be used to differentiate between coronary lesion and normal coronary wall. The time domain information complements the spectral domain intensity data for improved differentiation between graded levels of coronary lesions.

  8. Time-resolved fluorescence-based assay for rapid detection of Escherichia coli.

    PubMed

    Kulpakko, Janne; Kopra, Kari; Hänninen, Pekka

    2015-02-01

    Fast and simple detection of pathogens is of utmost importance in health care and the food industry. In this article, a novel technology for the detection of pathogenic bacteria is presented. The technology uses lytic-specific bacteriophages and a nonspecific interaction of cellular components with a luminescent lanthanide chelate. As a proof of principle, Escherichia coli-specific T4 bacteriophage was used to infect the bacteria, and the cell lysis was detected. In the absence of E. coli, luminescent Eu(3+)-chelate complex cannot be formed and low time-resolved luminescence signal is monitored. In the presence of E. coli, increased luminescence signal is observed as the cellular contents are leached to the surrounding medium. The luminescence signal is observed as a function of the number of bacteria in the sample. The homogeneous assay can detect living E. coli in bacterial cultures and simulated urine samples within 25 min with a detection limit of 1000 or 10,000 bacterial cells/ml in buffer or urine, respectively. The detection limit is at the clinically relevant level, which indicates that the method could also be applicable to clinical settings for fast detection of urine bacteria.

  9. Time-resolved fluorescence spectroscopic study of crude petroleum oils: influence of chemical composition.

    PubMed

    Ryder, Alan G

    2004-05-01

    The fluorescence of crude petroleum oils is sensitive to changes in chemical composition and many different fluorescence methods have been used to characterize crude oils. The use of fluorescence lifetimes to quantitatively characterize oil composition has practical advantages over steady-state measurements, but there have been comparatively few studies in which the lifetime behavior is correlated with gross chemical compositional data. In this study, the fluorescence lifetimes for a series of 23 crude petroleum oils with American Petroleum Institute (API) gravities of between 10 and 50 were measured at several emission wavelengths (450-785 nm) using a 380 nm light emitting diode (LED) excitation source. It was found that the intensity average fluorescence lifetime (tau) at any emission wave-length does not correlate well with either API gravity or aromatic concentration. However, it was found that tau is strongly negatively correlated with both the polar and sulfur concentrations and positively correlated with the corrected alkane concentration. This indicates that the fluorescence behavior of crude petroleum oils is governed primarily by the concentration of quenching species. All the strong lifetime-concentration correlations are nonlinear and show a high degree of scatter, especially for medium to light oils with API gravities of between 25 and 40. The degree of scatter is greatest for oils where the concentrations (wt %) of the polar fraction is approximately 10 +/- 4%, the asphaltene component is approximately 1 +/- 0.5%, and sulfur is 0.5 +/- 0.4%. This large degree of scatter precludes the use of average fluorescence lifetime data obtained with 380 nm excitation for the accurate prediction of the common chemical compositional parameters of crude petroleum oils. PMID:15165340

  10. Novel flashlamp-based time-resolved fluorescence microscope reduces autofluorescence for 30-fold contrast enhancement in environmental samples

    NASA Astrophysics Data System (ADS)

    Connally, Russell; Veal, Duncan; Piper, James A.

    2003-07-01

    The abundance of naturally fluorescing components (autofluorophors) encountered in environmentally sourced samples can greatly hinder the detection and identification of fluorescently labeled target using fluorescence microscopy. Time-resolved fluorescence microscopy (TRFM) is a technique that reduces the effects of autofluorescence through precisely controlled time delays. Lanthanide chelates have fluorescence lifetimes many orders of magnitude greater than typical autofluorophors, and persist in their luminescence long after autofluorescence has ceased. An intense short pulse of (UV) light is used to excite fluorescence in the sample and after a short delay period the longer persisting fluorescence from the chelate is captured with an image-intensified CCD camera. The choice of pulsed excitation source for TRFM has a large impact on the price and performance of the instrument. A flashlamp with a short pulse duration was selected for our instrument because of the high spectral energy in the UV region and short pulse length. However, flash output decays with an approximate lifetime of 18μs and the TRFM requires a long-lived chelate to ensure probe fluorescence is still visible after decay of the flash plasma. We synthesized a recently reported fluorescent chelate (BHHCT) and conjugated it to a monoclonal antibody directed against the water-borne parasite Giardia lamblia. Fluorescence lifetime of the construct was determined to be 339μs +/- 14μs and provided a 45-fold enhancement of labeled Giardia over background using a gate delay of 100μs. Despite the sub-optimal decay characteristics of the light pulse, flashlamps have many advantages compared to optical chopper wheels and modulated lasers. Their low cost, lack of vibration, ease of interface and small footprint are important factors to consider in TRFM design.

  11. Single-label kinase and phosphatase assays for tyrosine phosphorylation using nanosecond time-resolved fluorescence detection.

    PubMed

    Sahoo, Harekrushna; Hennig, Andreas; Florea, Mara; Roth, Doris; Enderle, Thilo; Nau, Werner M

    2007-12-26

    The collision-induced fluorescence quenching of a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine (Dbo) by hydrogen atom abstraction from the tyrosine residue in peptide substrates was introduced as a single-labeling strategy to assay the activity of tyrosine kinases and phosphatases. The assays were tested for 12 different combinations of Dbo-labeled substrates and with the enzymes p60c-Src Src kinase, EGFR kinase, YOP protein tyrosine phosphatase, as well as acid and alkaline phosphatases, thereby demonstrating a broad application potential. The steady-state fluorescence changed by a factor of up to 7 in the course of the enzymatic reaction, which allowed for a sufficient sensitivity of continuous monitoring in steady-state experiments. The fluorescence lifetimes (and intensities) were found to be rather constant for the phosphotyrosine peptides (ca. 300 ns in aerated water), while those of the unphosphorylated peptides were as short as 40 ns (at pH 7) and 7 ns (at pH 13) as a result of intramolecular quenching. Owing to the exceptionally long fluorescence lifetime of Dbo, the assays were alternatively performed by using nanosecond time-resolved fluorescence (Nano-TRF) detection, which leads to an improved discrimination of background fluorescence and an increased sensitivity. The potential for inhibitor screening was demonstrated through the inhibition of acid and alkaline phosphatases by molybdate.

  12. Wide-field time-resolved fluorescence anisotropy imaging (TR-FAIM): Imaging the rotational mobility of a fluorophore

    NASA Astrophysics Data System (ADS)

    Siegel, J.; Suhling, K.; Lévêque-Fort, S.; Webb, S. E. D.; Davis, D. M.; Phillips, D.; Sabharwal, Y.; French, P. M. W.

    2003-01-01

    We report a picosecond time-gated fluorescence lifetime imaging (FLIM) system extended to perform time-resolved fluorescence anisotropy imaging (TR-FAIM). Upon excitation with linearly polarized laser pulses, the parallel and perpendicular components of the fluorescence emission from a sample are imaged simultaneously using a polarization-resolved imager. The imaging technique presented here quantitatively reports the rotational mobility of a fluorophore as it varies according to the local environment. In a single acquisition run it yields maps of both rotational correlation time and fluorescence lifetime as they vary across a sample. TR-FAIM has been applied to imaging standard multiwell plate samples of rhodamine 6G dissolved in methanol, ethylene glycol, trimethylene glycol, and glycerol. The observed rotational correlation times and fluorescence lifetimes, which report the local viscosity and refractive index of the local rhodamine 6G environment, respectively, are in good agreement with previously published single point measurements. By considering the linear dependence of the rotational correlation time on viscosity up to 20 cP, we are able to obtain a two-dimensional viscosity map. Wide-field maps of rotational correlation time, and therefore viscosity, have been obtained. This illustrates the potential to image the local viscosity and fluorescence lifetime distributions of fluorophore tagged proteins in cells.

  13. Global and time-resolved monitoring of crop photosynthesis with chlorophyll fluorescence

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Global monitoring of agricultural productivity is critical in a world under a continuous increase of food demand. Here we have used new spaceborne retrievals of chlorophyll fluorescence, an emission quantity intrinsically linked to photosynthesis, to derive spatially explicit photosynthetic uptake r...

  14. Time-resolved fluorescence resonance energy transfer as a versatile tool in the development of homogeneous cellular kinase assays.

    PubMed

    Saville, Lisa; Spais, Chrysanthe; Mason, Jennifer L; Albom, Mark S; Murthy, Seetha; Meyer, Sheryl L; Ator, Mark A; Angeles, Thelma S; Husten, Jean

    2012-12-01

    Homogeneous cellular assays can streamline product detection in the drug discovery process. One commercially available assay employing time-resolved fluorescence resonance energy transfer (TR-FRET) that detects phosphorylated products was used to evaluate inhibitors of the receptor tyrosine kinase AXL in a cell line expressing an AXL-green fluorescent protein fusion protein. This TR-FRET assay was modified to evaluate the phosphorylation state of the AXL family member MER in a cell line expressing MER with a V5 tag by adding a fluorescein-labeled anti-V5 antibody. This homogeneous cellular assay was further modified to evaluate the nonreceptor tyrosine kinase focal adhesion kinase (FAK) in cell lines that expressed an untagged kinase by the inclusion of a commercially available anti-FAK antibody conjugated with an acceptor dye. The methods described here can be further adapted for TR-FRET detection of other cellular kinase activities.

  15. Photosystem II does not possess a simple excitation energy funnel: time-resolved fluorescence spectroscopy meets theory.

    PubMed

    Shibata, Yutaka; Nishi, Shunsuke; Kawakami, Keisuke; Shen, Jian-Ren; Renger, Thomas

    2013-05-01

    The experimentally obtained time-resolved fluorescence spectra of photosystem II (PS II) core complexes, purified from a thermophilic cyanobacterium Thermosynechococcus vulcanus, at 5-180 K are compared with simulations. Dynamic localization effects of excitons are treated implicitly by introducing exciton domains of strongly coupled pigments. Exciton relaxations within a domain and exciton transfers between domains are treated on the basis of Redfield theory and generalized Förster theory, respectively. The excitonic couplings between the pigments are calculated by a quantum chemical/electrostatic method (Poisson-TrEsp). Starting with previously published values, a refined set of site energies of the pigments is obtained through optimization cycles of the fits of stationary optical spectra of PS II. Satisfactorily agreement between the experimental and simulated spectra is obtained for the absorption spectrum including its temperature dependence and the linear dichroism spectrum of PS II core complexes (PS II-CC). Furthermore, the refined site energies well reproduce the temperature dependence of the time-resolved fluorescence spectrum of PS II-CC, which is characterized by the emergence of a 695 nm fluorescence peak upon cooling down to 77 K and the decrease of its relative intensity upon further cooling below 77 K. The blue shift of the fluorescence band upon cooling below 77 K is explained by the existence of two red-shifted chlorophyll pools emitting at around 685 and 695 nm. The former pool is assigned to Chl45 or Chl43 in CP43 (Chl numbering according to the nomenclature of Loll et al. Nature2005, 438, 1040) while the latter is assigned to Chl29 in CP47. The 695 nm emitting chlorophyll is suggested to attract excitations from the peripheral light-harvesting complexes and might also be involved in photoprotection.

  16. Synthesis of Ag clusters in microemulsions: A time-resolved UV vis and fluorescence spectroscopy study

    NASA Astrophysics Data System (ADS)

    Ledo, Ana; Martínez, F.; López-Quintela, M. A.; Rivas, J.

    2007-09-01

    The combined use of the microemulsion technique and the kinetic control allows the preparation of small silver clusters. By using UV-vis and fluorescence spectroscopy the main stages by which the clusters grow, before the formation of nanoparticles, were elucidated. Transmission electron microscopy (TEM) and scanning tunnelling microscopy (STM) were used to further characterize the samples. Two main stages were clearly identified, which are associated with: (1) the formation of Ag n clusters with n<10, which self-aggregate into one atom high 2D nanodiscs of 3.2 nm size and (2) Ag n clusters, which self-aggregate into 3D nanostructures of 1.5 nm in size. The fluorescence properties observed with both stages show that the formed clusters are small enough to display a molecule-like behaviour.

  17. Adsorption of Eu(III) on a heterogeneous surface studied by time-resolved laser fluorescence microscopy (TRLFM).

    PubMed

    Ishida, Keisuke; Kimura, Takaumi; Saito, Takumi; Tanaka, Satoru

    2009-03-15

    Time-resolved laser fluorescence microscopy (TRLFM) is a useful tool to simultaneously investigate the intensity, location, type, and surrounding chemical environment of a fluorophore. In this study, we demonstrated the applicability of TRLFM for the adsorption of Eu(III) on a natural heterogeneous surface. Different adsorption species of Eu(III) were observed on the Makabe granite surface and its constituents (biotite, plagioclase, potassium feldspar, and quartz). Eu(III) heterogeneously adsorbed on biotite, plagioclase, and quartz and homogeneously on potassium feldspar. The histograms of the fluorescence decay rates of adsorbed Eu(III) indicated efficient quenching of Eu(III) fluorescence probably due to Eu(III)-surface interaction or the formation polynuclear hydoxo Eu(III) species on the surfaces. It was also revealed that single species of Eu(III) was observed on biotite and two species on plagioclase and potassium feldspar. The adsorption of Eu(III) on the granite surface was highly heterogeneous. The TRLFM measurements of different regions of the granite surface turned into the finding of Eu(III) with different fluorescence decay rates. Comparing with the fluorescence decay histograms of the mineral constituents, Eu(III) clearly adsorbed on the feldspar family. It was also found that Eu(III) adsorbed as an outer-sphere complex and on an altered mineral of the granite.

  18. Application of time-resolved fluorescence to the determination of metabolites.

    PubMed

    Murillo Pulgarín, J A; Alañón Molina, A; Martínez Ferreras, F

    2014-07-15

    A simple fluorescent methodology for the simultaneous determination of two major metabolites of acetylsalicylic acid--salicylic and gentisic acids--in pharmaceutical preparations and human urine is proposed. Due to the overlapping between the fluorescence spectra of both analytes, the use of the more selective fluorescence decay curves is proposed. Values of dependent instrumental variables affecting the signal-to-noise ratio were fixed with a simplex optimization procedure. A calibration matrix of thirteen standards plus two blank samples was processed using a partial least-squares (PLS) analysis. To assess the goodness of the proposed method, a prediction set of nine synthetic samples was analyzed, obtaining recovery percentages between 95% and 106%. Limits of detection, calculated by means of a new criterion, were 3.49 μg L(-1) and 1.66 μg L(-1) for salicylic and gentisic acids, respectively. The method was also tested in three pharmaceutical preparations containing salicylic acid, obtaining recovery percentages close to 100%. Finally, the simultaneous determination of both analytes in human urine samples was successfully carried out by the PLS-analysis of a matrix of thirteen standards plus five analyte blanks. Although spectra of analytes and urine overlap strongly, no extraction method neither prior separation of the analytes were needed. PMID:24662756

  19. Time-resolved imaging system for fluorescence-guided surgery with lifetime imaging capability

    NASA Astrophysics Data System (ADS)

    Powolny, F.; Homicsko, K.; Sinisi, R.; Bruschini, Claudio E.; Grigoriev, E.; Homulle, H.; Prior, John O.; Hanahan, D.; Dubikovskaya, E.; Charbon, E.

    2014-05-01

    We present a single-photon camera for fluorescence imaging, with a time resolution better than 100ps, capable of providing both intensity and lifetime images. the camera was fabricated in standard CMOS technology. With this FluoCam we show the possibility to study sub-nanosecond fluorescence mechanisms. The FluoCam was used to characterize a near-infrared probe, indocyanine green, conjugated with multimeric cyclic pentapeptide (cRGD). The fluorescent probe-conjugated was used to target and mark tumors with better specificity, in particular aiming at targeting the integrins αvβ3 and αvβ5. As a first step towards clinical studies, preliminary results obtained in-vivo are presented. The first envisioned clinical application would be image-guided surgical oncology to help the surgeon to remove tumor tissue by a better discrimination from normal tissues and also to improve the detection of metastatic lymph nodes. A further application could be the in-vivo determination of the αvβ3 and αvβ5 targets to select patients for therapy with RGD chemotherapy conjugates.

  20. Application of time-resolved fluorescence to the determination of metabolites

    NASA Astrophysics Data System (ADS)

    Murillo Pulgarín, J. A.; Alañón Molina, A.; Martínez Ferreras, F.

    2014-07-01

    A simple fluorescent methodology for the simultaneous determination of two major metabolites of acetylsalicylic acid - salicylic and gentisic acids - in pharmaceutical preparations and human urine is proposed. Due to the overlapping between the fluorescence spectra of both analytes, the use of the more selective fluorescence decay curves is proposed. Values of dependent instrumental variables affecting the signal-to-noise ratio were fixed with a simplex optimization procedure. A calibration matrix of thirteen standards plus two blank samples was processed using a partial least-squares (PLS) analysis. To assess the goodness of the proposed method, a prediction set of nine synthetic samples was analyzed, obtaining recovery percentages between 95% and 106%. Limits of detection, calculated by means of a new criterion, were 3.49 μg L-1 and 1.66 μg L-1 for salicylic and gentisic acids, respectively. The method was also tested in three pharmaceutical preparations containing salicylic acid, obtaining recovery percentages close to 100%. Finally, the simultaneous determination of both analytes in human urine samples was successfully carried out by the PLS-analysis of a matrix of thirteen standards plus five analyte blanks. Although spectra of analytes and urine overlap strongly, no extraction method neither prior separation of the analytes were needed.

  1. A space- and time-resolved single photon counting detector for fluorescence microscopy and spectroscopy

    NASA Astrophysics Data System (ADS)

    Michalet, X.; Siegmund, O. H. W.; Vallerga, J. V.; Jelinsky, P.; Millaud, J. E.; Weiss, S.

    2006-02-01

    We have recently developed a wide-field photon-counting detector having high-temporal and high-spatial resolutions and capable of high-throughput (the H33D detector). Its design is based on a 25 mm diameter multi-alkali photocathode producing one photo electron per detected photon, which are then multiplied up to 10 7 times by a 3-microchannel plate stack. The resulting electron cloud is proximity focused on a cross delay line anode, which allows determining the incident photon position with high accuracy. The imaging and fluorescence lifetime measurement performances of the H33D detector installed on a standard epifluorescence microscope will be presented. We compare them to those of standard single-molecule detectors such as single-photon avalanche photodiode (SPAD) or electron-multiplying camera using model samples (fluorescent beads, quantum dots and live cells). Finally, we discuss the design and applications of future generation of H33D detectors for single-molecule imaging and high-throughput study of biomolecular interactions.

  2. Time-resolved effects of an electric field in recombination fluorescence

    SciTech Connect

    Borovkov, V.I.; Anishchik, S.V.; Anisimov, O.A.

    1995-12-01

    Quenching of the recombination fluorescence by an external electric field was investigated in hexane, tetradecane, and aqualane solutions of p-terphenyl and 2.5-diphenyloxazole irradiated with X-rays. The kinetics of the recombination fluorescence I(E,t) was measured in a nanosecond time scale and the quenching-efficiency curves Q(E,t) = 1 - I(E,t)/I(0,t) were plotted. The dependence Q(E,t) was shown to have the specific character Q(E,t) = f(pt), where p = AE{sup 2}D/r{sub c}{sup 2}. Here A is a constant dependent on the initial-distance distribution function of the charges, E is the electric field strength, D is a mutual diffusion coefficient of the recombining ions, and r{sub c} is the Onsager radius. The quadratic dependence of the parameter p on the electric field strength was shown to be a consequence of the diffusion-controlled reaction of ion recombination.

  3. Time-resolved fluorescence of 2-aminopurine as a probe of base flipping in M.HhaI–DNA complexes

    PubMed Central

    Neely, Robert K.; Daujotyte, Dalia; Grazulis, Saulius; Magennis, Steven W.; Dryden, David T. F.; Klimašauskas, Saulius; Jones, Anita C.

    2005-01-01

    DNA base flipping is an important mechanism in molecular enzymology, but its study is limited by the lack of an accessible and reliable diagnostic technique. A series of crystalline complexes of a DNA methyltransferase, M.HhaI, and its cognate DNA, in which a fluorescent nucleobase analogue, 2-aminopurine (AP), occupies defined positions with respect the target flipped base, have been prepared and their structures determined at higher than 2 Å resolution. From time-resolved fluorescence measurements of these single crystals, we have established that the fluorescence decay function of AP shows a pronounced, characteristic response to base flipping: the loss of the very short (∼100 ps) decay component and the large increase in the amplitude of the long (∼10 ns) component. When AP is positioned at sites other than the target site, this response is not seen. Most significantly, we have shown that the same clear response is apparent when M.HhaI complexes with DNA in solution, giving an unambiguous signal of base flipping. Analysis of the AP fluorescence decay function reveals conformational heterogeneity in the DNA–enzyme complexes that cannot be discerned from the present X-ray structures. PMID:16340006

  4. Short-term light adaptation of a cyanobacterium, Synechocystis sp. PCC 6803, probed by time-resolved fluorescence spectroscopy.

    PubMed

    Akimoto, Seiji; Yokono, Makio; Yokono, Erina; Aikawa, Shimpei; Kondo, Akihiko

    2014-08-01

    In photosynthetic organisms, the interactions among pigment-protein complexes change in response to light conditions. In the present study, we analyzed the transfer of excitation energy from the phycobilisome (PBS) and photosystem (PS) II to PSI in the cyanobacterium Synechocystis sp. PCC 6803. After 20 min of dark adaptation, Synechocystis cells were illuminated for 5 min with strong light with different spectral profiles, blue, green, two kinds of red, and white light. After illumination, the energy-transfer characteristics were evaluated using steady-state fluorescence and picosecond time-resolved fluorescence spectroscopy techniques. The fluorescence rise and decay curves were analyzed by global analysis to obtain fluorescence decay-associated spectra, followed by spectral component analysis. Under illumination with strong light, the contribution of the energy transfer from the PSII to PSI (spillover) became greater, and that of the energy transfer from the PBS to PSI decreased; the former change was larger than the latter. The energy transfer pathway to PSI was sensitive to red light. We discuss the short-term adaptation of energy-transfer processes in Synechocystis under strong-light conditions.

  5. Correlation of conformational heterogeneity of the tryptophyl side chain and time-resolved fluorescence intensity decay kinetics

    NASA Astrophysics Data System (ADS)

    Laws, William R.; Ross, J. B. Alexander

    1992-04-01

    The time-resolved fluorescence properties of a tryptophan residue should be useful for probing protein structure, function, and dynamics. To date, however, the non-single exponential fluorescence intensity decay kinetics for numerous peptides and proteins having a single tryptophan residue have not been adequately explained. Many possibilities have been considered and include: (1) contributions from the 1La and 1Lb states of indole; (2) excited-state hydrogen exchange; and (3) environmental heterogeneity from (chi) 1 and (chi) 2 rotamers. In addition, it has been suggested that generally many factors contribute to the decay and a distribution of probabilities may be more appropriate. Two recent results support multiple species due to conformational heterogeneity as the major contributor to complex kinetics. First, a rotationally constrained tryptophan analogue has fluorescence intensity decay kinetics that can be described by the sum of two exponentials with amplitudes comparable to the relative populations of the two rotational isomers. Second, the multiple exponentials observed for tyrosine-containing model compounds and peptides correlate with the (chi) 1 rotamer populations independently determined by 1H NMR. We now report similar correlations between rotamer populations and fluorescence intensity decay kinetics for a tryptophan analogue of oxytocin. It appears for this compound that either (chi) 2 rotations do not appreciably alter the indole environment, (chi) 2 rotations are rapid enough to average the observed dependence, or only one of two possible (chi) 2 populations is associated with each (chi) 1 rotamer.

  6. Cellular Oxygen and Nutrient Sensing in Microgravity Using Time-Resolved Fluorescence Microscopy

    NASA Technical Reports Server (NTRS)

    Szmacinski, Henryk

    2003-01-01

    Oxygen and nutrient sensing is fundamental to the understanding of cell growth and metabolism. This requires identification of optical probes and suitable detection technology without complex calibration procedures. Under this project Microcosm developed an experimental technique that allows for simultaneous imaging of intra- and inter-cellular events. The technique consists of frequency-domain Fluorescence Lifetime Imaging Microscopy (FLIM), a set of identified oxygen and pH probes, and methods for fabrication of microsensors. Specifications for electronic and optical components of FLIM instrumentation are provided. Hardware and software were developed for data acquisition and analysis. Principles, procedures, and representative images are demonstrated. Suitable lifetime sensitive oxygen, pH, and glucose probes for intra- and extra-cellular measurements of analyte concentrations have been identified and tested. Lifetime sensing and imaging have been performed using PBS buffer, culture media, and yeast cells as a model systems. Spectral specifications, calibration curves, and probes availability are also provided in the report.

  7. Microsecond protein folding events revealed by time-resolved fluorescence resonance energy transfer in a microfluidic mixer.

    PubMed

    Jiang, Liguo; Zeng, Yan; Sun, Qiqi; Sun, Yueru; Guo, Zhihong; Qu, Jianan Y; Yao, Shuhuai

    2015-06-01

    We demonstrate the combination of the time-resolved fluorescence resonance energy transfer (tr-FRET) measurement and the ultrarapid hydrodynamic focusing microfluidic mixer. The combined technique is capable of probing the intermolecular distance change with temporal resolution at microsecond level and structural resolution at Angstrom level, and the use of two-photon excitation enables a broader exploration of FRET with spectrum from near-ultraviolet to visible wavelength. As a proof of principle, we used the coupled microfluidic laminar flow and time-resolved two-photon excitation microscopy to investigate the early folding states of Cytochrome c (cyt c) by monitoring the distance between the tryptophan (Trp-59)-heme donor-acceptor (D-A) pair. The transformation of folding states of cyt c in the early 500 μs of refolding was revealed on the microsecond time scale. For the first time, we clearly resolved the early transient state of cyt c, which is populated within the dead time of the mixer (<10 μs) and has a characteristic Trp-59-heme distance of ∼31 Å. We believe this tool can find more applications in studying the early stages of biological processes with FRET as the probe.

  8. A novel luminescent terbium-3-carboxycoumarin probe for time-resolved fluorescence sensing of pesticides methomyl, aldicarb and prometryne.

    PubMed

    Azab, Hassan A; Duerkop, Axel; Saad, E M; Awad, F K; Abd El Aal, R M; Kamel, Rasha M

    2012-11-01

    The luminescence arising from lanthanide cations offers several advantages over organic fluorescent molecules: sharp, distinctive emission bands allow for easy resolution between multiple lanthanide signals; long emission lifetimes (μs-ms) make them excellent candidates for time-resolved measurements; and high resistance to photo bleaching allow for long or repeated experiments. A time-resolved (gated) luminescence-based method for determination of pesticides methomyl, aldicarb and prometryne in microtiterplate format using the long-lived terbium-3-carboxycoumarin in 1:3 metal:ligand ratio has been developed. The limit of detection is 1.20×10(6), 5.19×10(5) and 2.74×10(6)ng L(-1) for methomyl, prometryne and aldicarb, respectively. The quantum yield (QY=0.08) of Tb(III)-3-carboxycoumarin was determined using 3-(2-benzothiazolyl)-7-diethylamino-coumarin (coumarin 6). Stern-volmer studies at different temperatures indicate that collisional quenching dominates for methomyl, aldicarb and prometryne. Binding constants were determined at 303, 308 and 313 K by using Lineweaver-Burk equation. A thermodynamic analysis showed that the reaction is spontaneous with negative ΔG. Effect of some relevant interferents on the detection of pesticides has been investigated.

  9. Nonradiative decay mechanisms of complexed indole derivatives studied by time-resolved fluorescence

    NASA Astrophysics Data System (ADS)

    Schauerte, Joseph A.; Gafni, Ari

    1990-05-01

    Ground and excited state characteristics of substituted indole derivatives reveal a sensitivity of indoles' electronic properties to the nature and location of substitutions on the indole ring. These substitutions affect both the nature of the excited electronic state and the susceptibility of this state to non-radiative decay processes. A number of mechanisms that deactivate the excited state have been identified including intersystem crossing, electron photoejection into polar solvents, and >N-H dissociation in polar solvents (see Glasser & Lami,1986) . While the >N-H group has been implicated in non-radiative decay processes in polar solvents, covalent substitutions elsewhere on the indole molecule may modulate the importance of this site in non-radiative decay mechanisms or alternatively these substitutions may introduce new deactivation mechanisms. Additionally, complexes formed between indole derivatives and β-cyclodextrin cavities show different sensitivity to excited state deactivation mechanisms dependent upon the location and nature of the covalent substitution. We have investigated the excited states of some indole derivatives substituted at position 5, para to the >N-H group on the benzyl ring, to determine the effect of such covalent substitutions on the fluorescence emission characteristics of the indole ring as well as on its susceptibility to alternate excited state decay mechanisms.

  10. Global and Time-Resolved Monitoring of Crop Photosynthesis with Chlorophyll Fluorescence

    NASA Technical Reports Server (NTRS)

    Guanter, Luis; Zhang, Yongguang; Jung, Martin; Joiner, Joanna; Voigt, Maximilian; Berry, Joseph A.; Frankenberg, Christian; Huete, Alfredo R.; Zarco-Tejada, Pablo; Lee, Jung-Eun; Moran, M. Susan; Ponce-Campos, Guillermo; Beer, Christian; Camps-Valls, Gustavo; Buchmann, Nina; Gianelle, Damiano; Klumpp, Katja; Cescatti, Alessandro; Baker, John M.; Griffis, Timothy J.

    2014-01-01

    Photosynthesis is the process by which plants harvest sunlight to produce sugars from carbon dioxide and water. It is the primary source of energy for all life on Earth; hence it is important to understand how this process responds to climate change and human impact. However, model-based estimates of gross primary production (GPP, output from photosynthesis) are highly uncertain, in particular over heavily managed agricultural areas. Recent advances in spectroscopy enable the space-based monitoring of sun-induced chlorophyll fluorescence (SIF) from terrestrial plants. Here we demonstrate that spaceborne SIF retrievals provide a direct measure of the GPP of cropland and grassland ecosystems. Such a strong link with crop photosynthesis is not evident for traditional remotely sensed vegetation indices, nor for more complex carbon cycle models. We use SIF observations to provide a global perspective on agricultural productivity. Our SIF-based crop GPP estimates are 50-75% higher than results from state-of-the-art carbon cycle models over, for example, the US Corn Belt and the Indo-Gangetic Plain, implying that current models severely underestimate the role of management. Our results indicate that SIF data can help us improve our global models for more accurate projections of agricultural productivity and climate impact on crop yields. Extension of our approach to other ecosystems, along with increased observational capabilities for SIF in the near future, holds the prospect of reducing uncertainties in the modeling of the current and future carbon cycle.

  11. Global and time-resolved monitoring of crop photosynthesis with chlorophyll fluorescence.

    PubMed

    Guanter, Luis; Zhang, Yongguang; Jung, Martin; Joiner, Joanna; Voigt, Maximilian; Berry, Joseph A; Frankenberg, Christian; Huete, Alfredo R; Zarco-Tejada, Pablo; Lee, Jung-Eun; Moran, M Susan; Ponce-Campos, Guillermo; Beer, Christian; Camps-Valls, Gustavo; Buchmann, Nina; Gianelle, Damiano; Klumpp, Katja; Cescatti, Alessandro; Baker, John M; Griffis, Timothy J

    2014-04-01

    Photosynthesis is the process by which plants harvest sunlight to produce sugars from carbon dioxide and water. It is the primary source of energy for all life on Earth; hence it is important to understand how this process responds to climate change and human impact. However, model-based estimates of gross primary production (GPP, output from photosynthesis) are highly uncertain, in particular over heavily managed agricultural areas. Recent advances in spectroscopy enable the space-based monitoring of sun-induced chlorophyll fluorescence (SIF) from terrestrial plants. Here we demonstrate that spaceborne SIF retrievals provide a direct measure of the GPP of cropland and grassland ecosystems. Such a strong link with crop photosynthesis is not evident for traditional remotely sensed vegetation indices, nor for more complex carbon cycle models. We use SIF observations to provide a global perspective on agricultural productivity. Our SIF-based crop GPP estimates are 50-75% higher than results from state-of-the-art carbon cycle models over, for example, the US Corn Belt and the Indo-Gangetic Plain, implying that current models severely underestimate the role of management. Our results indicate that SIF data can help us improve our global models for more accurate projections of agricultural productivity and climate impact on crop yields. Extension of our approach to other ecosystems, along with increased observational capabilities for SIF in the near future, holds the prospect of reducing uncertainties in the modeling of the current and future carbon cycle.

  12. Modified diglycol-amides for actinide separation: solvent extraction and time-resolved laser fluorescence spectroscopy complexation studies

    SciTech Connect

    Wilden, A.; Modolo, G.; Lange, S.; Sadowski, F.; Bosbach, D.; Beele, B.B.; Panak, P.J.; Skerencak-Frech, A.; Geist, A.; Iqbal, M.; Verboom, W.

    2013-07-01

    In this work, the back-bone of the diglycolamide-structure of the TODGA extractant was modified by adding one or two methyl groups to the central methylene carbon-atoms. The influence of these structural modifications on the extraction behavior of trivalent actinides and lanthanides and other fission products was studied in solvent extraction experiments. The addition of methyl groups to the central methylene carbon atoms leads to reduced distribution ratios, also for Sr(II). This reduced extraction efficiency for Sr(II) is beneficial for process applications, as the co-extraction of Sr(II) can be avoided, resulting in an easier process design. The use of these modified diglycol-amides in solvent extraction processes is discussed. Furthermore, the complexation of Cm(III) and Eu(III) to the ligands was studied using Time-Resolved-Laser-Fluorescence-Spectroscopy (TRLFS). The complexes were characterized by slope analysis and conditional stability constants were determined.

  13. Dynamics of Nuclear Receptor Helix-12 Switch of Transcription Activation by Modeling Time-Resolved Fluorescence Anisotropy Decays

    PubMed Central

    Batista, Mariana R.B.; Martínez, Leandro

    2013-01-01

    Nuclear hormone receptors (NRs) are major targets for pharmaceutical development. Many experiments demonstrate that their C-terminal Helix (H12) is more flexible in the ligand-binding domains (LBDs) without ligand, this increased mobility being correlated with transcription repression and human diseases. Crystal structures have been obtained in which the H12 is extended, suggesting the possibility of large amplitude H12 motions in solution. However, these structures were interpreted as possible crystallographic artifacts, and thus the microscopic nature of H12 movements is not well known. To bridge the gap between experiments and molecular models and provide a definitive picture of H12 motions in solution, extensive molecular dynamics simulations of the peroxisome proliferator-activated receptor-γ LBD, in which the H12 was bound to a fluorescent probe, were performed. A direct comparison of the modeled anisotropy decays to time-resolved fluorescence anisotropy experiments was obtained. It is shown that the decay rates are dependent on the interactions of the probe with the surface of the protein, and display little correlation with the flexibility of the H12. Nevertheless, for the probe to interact with the surface of the LBD, the H12 must be folded over the body of the LBD. Therefore, the molecular mobility of the H12 should preserve the globularity of the LBD, so that ligand binding and dissociation occur by diffusion through the surface of a compact receptor. These results advance the comprehension of both ligand-bound and ligand-free receptor structures in solution, and also guide the interpretation of time-resolved anisotropy decays from a molecular perspective, particularly by the use of simulations. PMID:24094408

  14. Use of Time-Resolved Fluorescence To Improve Sensitivity and Dynamic Range of Gel-Based Proteomics.

    PubMed

    Sandberg, AnnSofi; Buschmann, Volker; Kapusta, Peter; Erdmann, Rainer; Wheelock, Åsa M

    2016-03-15

    Limitations in the sensitivity and dynamic range of two-dimensional gel electrophoresis (2-DE) are currently hampering its utility in global proteomics and biomarker discovery applications. In the current study, we present proof-of-concept analyses showing that introducing time-resolved fluorescence in the image acquisition step of in-gel protein quantification provides a sensitive and accurate method for subtracting confounding background fluorescence at the photon level. In-gel protein detection using the minimal difference gel electrophoresis workflow showed improvements in lowest limit of quantification in terms of CyDye molecules per pixel of 330-fold in the blue-green region (Cy2) and 8000-fold in the red region (Cy5) over conventional state-of-the-art image acquisition instrumentation, here represented by the Typhoon 9400 instrument. These improvements make possible the detection of low-abundance proteins present at sub-attomolar levels, thereby representing a quantum leap for the use of gel-based proteomics in biomarker discovery. These improvements were achieved using significantly lower laser powers and overall excitation times, thereby drastically decreasing photobleaching during repeated scanning. The single-fluorochrome detection limits achieved by the cumulative time-resolved emission two-dimensional electrophoresis (CuTEDGE) technology facilitates in-depth proteomics characterization of very scarce samples, for example, primary human tissue materials collected in clinical studies. The unique information provided by high-sensitivity 2-DE, including positional shifts due to post-translational modifications, may increase the chance to detect biomarker signatures of relevance for identification of disease subphenotypes. PMID:26854653

  15. On the possibility of ephedrine detection: time-resolved fluorescence resonance energy transfer (FRET)-based approach.

    PubMed

    Varriale, Antonio; Marzullo, Vincenzo Manuel; Di Giovanni, Stefano; Scala, Andrea; Capo, Alessandro; Majoli, Adelia; Pennacchio, Angela; Staiano, Maria; D'Auria, Sabato

    2016-09-01

    Ephedrine is one of the main precursor compounds used in the illegal production of amphetamines and related drugs. Actually, conventional analytical methods such as high-performance liquid chromatography (HPLC), capillary electrophoresis (CE), and gas chromatography-mass spectrometry (GC-MS) are used for the detection of ephedrine; sadly, these methods require qualified personnel and are time-consuming and expensive. In order to overcome these problems, in recent years, different methods have been developed based on the surface plasmon resonance (SPR) and electrochemical method. In this work, we present a simple, rapid, and effective method to detect the presence of ephedrine in solution, based on competitive fluorescence resonance energy transfer (FRET) assay. The antibody anti-ephedrine and ephedrine derivative were produced and labeled respectively, with two different fluorescent probes (donor and acceptor). The change in FRET signal intensity between donor and acceptor ephedrine compounds gives the possibility of detecting ephedrine traces of at least 0.81 ± 0.04 ppm (LOD). Graphical abstract A new Time-resolved Fluorescence Resonance Energy Transfer (FRET) assay for ephedrine detection. PMID:27395357

  16. Time-resolved fluorescence imaging of slab gels for lifetime base-calling in DNA sequencing applications.

    PubMed

    Lassiter, S J; Stryjewski, W; Legendre, B L; Erdmann, R; Wahl, M; Wurm, J; Peterson, R; Middendorf, L; Soper, S A

    2000-11-01

    A compact time-resolved near-IR fluorescence imager was constructed to obtain lifetime and intensity images of DNA sequencing slab gels. The scanner consisted of a microscope body with f/1.2 relay optics onto which was mounted a pulsed diode laser (repetition rate 80 MHz, lasing wavelength 680 nm, average power 5 mW), filtering optics, and a large photoactive area (diameter 500 microns) single-photon avalanche diode that was actively quenched to provide a large dynamic operating range. The time-resolved data were processed using electronics configured in a conventional time-correlated single-photon-counting format with all of the counting hardware situated on a PC card resident on the computer bus. The microscope head produced a timing response of 450 ps (fwhm) in a scanning mode, allowing the measurement of subnano-second lifetimes. The time-resolved microscope head was placed in an automated DNA sequencer and translated across a 21-cm-wide gel plate in approximately 6 s (scan rate 3.5 cm/s) with an accumulation time per pixel of 10 ms. The sampling frequency was 0.17 Hz (duty cycle 0.0017), sufficient to prevent signal aliasing during the electrophoresis separation. Software (written in Visual Basic) allowed acquisition of both the intensity image and lifetime analysis of DNA bands migrating through the gel in real time. Using a dual-labeling (IRD700 and Cy5.5 labeling dyes)/two-lane sequencing strategy, we successfully read 670 bases of a control M13mp18 ssDNA template using lifetime identification. Comparison of the reconstructed sequence with the known sequence of the phage indicated the number of miscalls was only 2, producing an error rate of approximately 0.3% (identification accuracy 99.7%). The lifetimes were calculated using maximum likelihood estimators and allowed on-line determinations with high precision, even when short integration times were used to construct the decay profiles. Comparison of the lifetime base calling to a single

  17. Mechanisms of fluorescence decays of colloidal CdSe-CdS/ZnS quantum dots unraveled by time-resolved fluorescence measurement.

    PubMed

    Xu, Hao; Chmyrov, Volodymyr; Widengren, Jerker; Brismar, Hjalmar; Fu, Ying

    2015-11-01

    By narrowing the detection bandpass and increasing the signal-to-noise ratio in measuring the time-resolved fluorescence decay spectrum of colloidal CdSe-CdS/ZnS quantum dots (QDs), we show that directly after the photoexcitation, the fluorescence decay spectrum is characterized by a single exponential decay, which represents the energy relaxation of the photogenerated exciton from its initial high-energy state to the ground exciton state. The fluorescence decay spectrum of long decay time is in the form of β/t(2), where β is the radiative recombination time of the ground-state exciton and t is the decay time. Our findings provide us with a direct and quantitative link between fluorescence decay measurement data and fundamental photophysics of QD exciton, thereby leading to a novel way of applying colloidal QDs to study microscopic, physical and chemical processes in many fields including biomedicine. PMID:26426293

  18. Mechanisms of fluorescence decays of colloidal CdSe-CdS/ZnS quantum dots unraveled by time-resolved fluorescence measurement.

    PubMed

    Xu, Hao; Chmyrov, Volodymyr; Widengren, Jerker; Brismar, Hjalmar; Fu, Ying

    2015-11-01

    By narrowing the detection bandpass and increasing the signal-to-noise ratio in measuring the time-resolved fluorescence decay spectrum of colloidal CdSe-CdS/ZnS quantum dots (QDs), we show that directly after the photoexcitation, the fluorescence decay spectrum is characterized by a single exponential decay, which represents the energy relaxation of the photogenerated exciton from its initial high-energy state to the ground exciton state. The fluorescence decay spectrum of long decay time is in the form of β/t(2), where β is the radiative recombination time of the ground-state exciton and t is the decay time. Our findings provide us with a direct and quantitative link between fluorescence decay measurement data and fundamental photophysics of QD exciton, thereby leading to a novel way of applying colloidal QDs to study microscopic, physical and chemical processes in many fields including biomedicine.

  19. Viscosity heterogeneity inside lipid bilayers of single-component phosphatidylcholine liposomes observed with picosecond time-resolved fluorescence spectroscopy.

    PubMed

    Nojima, Yuki; Iwata, Koichi

    2014-07-24

    A number of biochemical reactions proceed inside biomembranes. Because the rate of a chemical reaction is influenced by chemical properties of the reaction field, it is important to examine the chemical properties inside the biomembranes, or lipid bilayer membranes, for understanding biochemical reactions. In this study, we estimate viscosity inside the lipid bilayers of liposomes with picosecond time-resolved fluorescence spectroscopy. trans-Stilbene is solubilized in the lipid bilayers formed by phosphatidylcholines, DSPC, DOPC, DPPC, DMPC, and DLPC, with 18, 18, 16, 14, and 12 carbon atoms in their alkyl chains, respectively, and egg-PC. Viscosity inside the lipid bilayer is estimated from the photoisomerization rate constant and from the rotational relaxation time of the first excited singlet state of trans-stilbene. The effect of the hydrocarbon chain length and temperature on viscosity is examined. The presence of two solvation environments within the lipid bilayer is indicated from the two independent estimations. One environment is 30 to 290 times more viscous than the other. Even single-component lipid bilayers are likely to have heterogeneous structures.

  20. Europium Nanospheres-Based Time-Resolved Fluorescence for Rapid and Ultrasensitive Determination of Total Aflatoxin in Feed.

    PubMed

    Wang, Du; Zhang, Zhaowei; Li, Peiwu; Zhang, Qi; Ding, Xiaoxia; Zhang, Wen

    2015-12-01

    Immunochromatographic (IC) assays are considered suitable diagnostic tools for the determination of mycotoxins. A europium nanospheres-based time-resolved fluorescence immunoassay (Eu-Nano-TRFIA), based on a monoclonal antibody and a portable TRFIA reader, was developed to determine total aflatoxin (including aflatoxins B1, B2, G1, and G2) levels in feed samples. Under optimized conditions, the Eu-Nano-TRFIA method detected total aflatoxin within 12 min. It showed good linearity (R(2) > 0.985), LOD of 0.16 μg/kg, a wide dynamic range of 0.48-30.0 μg/kg, recovery rates of 83.9-113.9%, and coefficients of variation (CVs) of 3.5-8.8%. In the 397 samples from company and livestock farms throughout China, the detection rate was 78.3%, concentrations were 0.50-145.30 μg/kg, the highest total aflatoxin content was found in cottonseed meal, and corn was found to be the most commonly contaminated feed. This method could be a powerful alternative for the rapid and ultrasensitive determination of total aflatoxin in quality control and meet the required Chinese maximum residue limits.

  1. Europium Nanospheres-Based Time-Resolved Fluorescence for Rapid and Ultrasensitive Determination of Total Aflatoxin in Feed.

    PubMed

    Wang, Du; Zhang, Zhaowei; Li, Peiwu; Zhang, Qi; Ding, Xiaoxia; Zhang, Wen

    2015-12-01

    Immunochromatographic (IC) assays are considered suitable diagnostic tools for the determination of mycotoxins. A europium nanospheres-based time-resolved fluorescence immunoassay (Eu-Nano-TRFIA), based on a monoclonal antibody and a portable TRFIA reader, was developed to determine total aflatoxin (including aflatoxins B1, B2, G1, and G2) levels in feed samples. Under optimized conditions, the Eu-Nano-TRFIA method detected total aflatoxin within 12 min. It showed good linearity (R(2) > 0.985), LOD of 0.16 μg/kg, a wide dynamic range of 0.48-30.0 μg/kg, recovery rates of 83.9-113.9%, and coefficients of variation (CVs) of 3.5-8.8%. In the 397 samples from company and livestock farms throughout China, the detection rate was 78.3%, concentrations were 0.50-145.30 μg/kg, the highest total aflatoxin content was found in cottonseed meal, and corn was found to be the most commonly contaminated feed. This method could be a powerful alternative for the rapid and ultrasensitive determination of total aflatoxin in quality control and meet the required Chinese maximum residue limits. PMID:26565941

  2. Detection of high-risk atherosclerotic lesions by time-resolved fluorescence spectroscopy based on the Laguerre deconvolution technique

    NASA Astrophysics Data System (ADS)

    Jo, J. A.; Fang, Q.; Papaioannou, T.; Qiao, J. H.; Fishbein, M. C.; Beseth, B.; Dorafshar, A. H.; Reil, T.; Baker, D.; Freischlag, J.; Marcu, L.

    2006-02-01

    This study introduces new methods of time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) data analysis for tissue characterization. These analytical methods were applied for the detection of atherosclerotic vulnerable plaques. Upon pulsed nitrogen laser (337 nm, 1 ns) excitation, TR-LIFS measurements were obtained from carotid atherosclerotic plaque specimens (57 endarteroctomy patients) at 492 distinct areas. The emission was both spectrally- (360-600 nm range at 5 nm interval) and temporally- (0.3 ns resolution) resolved using a prototype clinically compatible fiber-optic catheter TR-LIFS apparatus. The TR-LIFS measurements were subsequently analyzed using a standard multiexponential deconvolution and a recently introduced Laguerre deconvolution technique. Based on their histopathology, the lesions were classified as early (thin intima), fibrotic (collagen-rich intima), and high-risk (thin cap over necrotic core and/or inflamed intima). Stepwise linear discriminant analysis (SLDA) was applied for lesion classification. Normalized spectral intensity values and Laguerre expansion coefficients (LEC) at discrete emission wavelengths (390, 450, 500 and 550 nm) were used as features for classification. The Laguerre based SLDA classifier provided discrimination of high-risk lesions with high sensitivity (SE>81%) and specificity (SP>95%). Based on these findings, we believe that TR-LIFS information derived from the Laguerre expansion coefficients can provide a valuable additional dimension for the diagnosis of high-risk vulnerable atherosclerotic plaques.

  3. Characterization of energetic and thermalized sputtered atoms in pulsed plasma using time-resolved tunable diode-laser induced fluorescence

    SciTech Connect

    Desecures, M.; Poucques, L. de; Easwarakhanthan, T.; Bougdira, J.

    2014-11-03

    In this work, a time-resolved tunable diode-laser (DL) induced fluorescence (TR-TDLIF) method calibrated by absorption spectroscopy has been developed in order to determine atom and flux velocity distribution functions (AVDF and FVDF) of the energetic and the thermalized atoms in pulsed plasmas. The experimental set-up includes a low-frequency (∼3 Hz) and high spectral-resolution DL (∼0.005 pm), a fast rise-time pulse generator, and a high power impulse magnetron sputtering (HiPIMS) system. The induced TR-TDLIF signal is recorded every 0.5 μs with a digital oscilloscope of a second-long trace. The technique is illustrated with determining the AVDF and the FVDF of a metastable state of the sputtered neutral tungsten atoms in the HiPIMS post-discharge. Gaussian functions describing the population of the four W isotopes were used to fit the measured TR-TDLIF signal. These distribution functions provide insight into transition from the energetic to thermalized regimes from the discharge onset. This technique may be extended with appropriate DLs to probe any species with rapidly changing AVDF and FVDF in pulsed and strongly oscillating plasmas.

  4. Multi-channel lock-in amplifier assisted femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy with efficient rejection of superfluorescence background

    SciTech Connect

    Mao, Pengcheng; Wang, Zhuan; Dang, Wei; Weng, Yuxiang

    2015-12-15

    Superfluorescence appears as an intense background in femtosecond time-resolved fluorescence noncollinear optical parametric amplification spectroscopy, which severely interferes the reliable acquisition of the time-resolved fluorescence spectra especially for an optically dilute sample. Superfluorescence originates from the optical amplification of the vacuum quantum noise, which would be inevitably concomitant with the amplified fluorescence photons during the optical parametric amplification process. Here, we report the development of a femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectrometer assisted with a 32-channel lock-in amplifier for efficient rejection of the superfluorescence background. With this spectrometer, the superfluorescence background signal can be significantly reduced to 1/300–1/100 when the seeding fluorescence is modulated. An integrated 32-bundle optical fiber is used as a linear array light receiver connected to 32 photodiodes in one-to-one mode, and the photodiodes are further coupled to a home-built 32-channel synchronous digital lock-in amplifier. As an implementation, time-resolved fluorescence spectra for rhodamine 6G dye in ethanol solution at an optically dilute concentration of 10{sup −5}M excited at 510 nm with an excitation intensity of 70 nJ/pulse have been successfully recorded, and the detection limit at a pump intensity of 60 μJ/pulse was determined as about 13 photons/pulse. Concentration dependent redshift starting at 30 ps after the excitation in time-resolved fluorescence spectra of this dye has also been observed, which can be attributed to the formation of the excimer at a higher concentration, while the blueshift in the earlier time within 10 ps is attributed to the solvation process.

  5. Multi-channel lock-in amplifier assisted femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy with efficient rejection of superfluorescence background.

    PubMed

    Mao, Pengcheng; Wang, Zhuan; Dang, Wei; Weng, Yuxiang

    2015-12-01

    Superfluorescence appears as an intense background in femtosecond time-resolved fluorescence noncollinear optical parametric amplification spectroscopy, which severely interferes the reliable acquisition of the time-resolved fluorescence spectra especially for an optically dilute sample. Superfluorescence originates from the optical amplification of the vacuum quantum noise, which would be inevitably concomitant with the amplified fluorescence photons during the optical parametric amplification process. Here, we report the development of a femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectrometer assisted with a 32-channel lock-in amplifier for efficient rejection of the superfluorescence background. With this spectrometer, the superfluorescence background signal can be significantly reduced to 1/300-1/100 when the seeding fluorescence is modulated. An integrated 32-bundle optical fiber is used as a linear array light receiver connected to 32 photodiodes in one-to-one mode, and the photodiodes are further coupled to a home-built 32-channel synchronous digital lock-in amplifier. As an implementation, time-resolved fluorescence spectra for rhodamine 6G dye in ethanol solution at an optically dilute concentration of 10(-5)M excited at 510 nm with an excitation intensity of 70 nJ/pulse have been successfully recorded, and the detection limit at a pump intensity of 60 μJ/pulse was determined as about 13 photons/pulse. Concentration dependent redshift starting at 30 ps after the excitation in time-resolved fluorescence spectra of this dye has also been observed, which can be attributed to the formation of the excimer at a higher concentration, while the blueshift in the earlier time within 10 ps is attributed to the solvation process. PMID:26724012

  6. Plasma progesterone analysis by a time-resolved fluorescent antibody test to monitor estrous cycles in goats.

    PubMed

    Blaszczyk, Barbara; Stankiewicz, Tomasz; Udała, Jan; Gaczarzewicz, Dariusz

    2009-01-01

    The objective of the current study was to evaluate whether blood plasma progesterone (P(4)) measurements with a time-resolved fluorescent antibody test (TR-FAT) kit designed for humans was applicable for goats. The first experiment was designed to verify whether the concentrations of P(4) measured by TR-FAT can be used to monitor the estrous and ovarian activity in goats (n = 14). Blood samples (322) were collected, and the ovaries were scanned using ultrasonography. The second experiment was carried out on 4 goats (60 samples) and designed to compare the TR-FAT with radioimmunoassay (RIA). The time interval between the lowest concentrations of P(4) assayed by TR-FAT was 21 +/- 0.3 days and did not differ significantly from the length of the interestrous interval. The highest concentrations of P(4) were confirmed by detection of corpus luteum. During estrus, the mean concentration did not differ significantly between both methods. Significant differences were present during the luteal phases; however, the profiles of P(4) assayed by both methods followed a similar pattern. Regression analysis showed a correlation between the 2 methods (r = 0.98; r(2) = 0.96; P < 0.0001). The Bland-Altman plot showed that all averages were within the 95% limits of agreement; however, the differences between both methods tend to be greater as the average increases. The results demonstrated that the TR-FAT method can be applied to monitor estrous cycles in goats through measurements of plasma P(4) concentrations. Moreover, not only does the TR-FAT meet the requirements for safety, but it is also a method of high throughput, rapidity, and simplicity. PMID:19139505

  7. Detection of anthrax protective antigen (PA) using europium labeled anti-PA monoclonal antibody and time-resolved fluorescence.

    PubMed

    Stoddard, Robyn A; Quinn, Conrad P; Schiffer, Jarad M; Boyer, Anne E; Goldstein, Jason; Bagarozzi, Dennis A; Soroka, Stephen D; Dauphin, Leslie A; Hoffmaster, Alex R

    2014-06-01

    Inhalation anthrax is a rare but acute infectious disease following adsorption of Bacillus anthracis spores through the lungs. The disease has a high fatality rate if untreated, but early and correct diagnosis has a significant impact on case patient recovery. The early symptoms of inhalation anthrax are, however, non-specific and current anthrax diagnostics are primarily dependent upon culture and confirmatory real-time PCR. Consequently, there may be a significant delay in diagnosis and targeted treatment. Rapid, culture-independent diagnostic tests are therefore needed, particularly in the context of a large scale emergency response. The aim of this study was to evaluate the ability of monoclonal antibodies to detect anthrax toxin proteins that are secreted early in the course of B. anthracis infection using a time-resolved fluorescence (TRF) immunoassay. We selected monoclonal antibodies that could detect protective antigen (PA), as PA83 and also PA63 and LF in the lethal toxin complex. The assay reliable detection limit (RDL) was 6.63×10(-6)μM (0.551ng/ml) for PA83 and 2.51×10(-5)μM (1.58ng/ml) for PA63. Despite variable precision and accuracy of the assay, PA was detected in 9 out of 10 sera samples from anthrax confirmed case patients with cutaneous (n=7), inhalation (n=2), and gastrointestinal (n=1) disease. Anthrax Immune Globulin (AIG), which has been used in treatment of clinical anthrax, interfered with detection of PA. This study demonstrates a culture-independent method of diagnosing anthrax through the use of monoclonal antibodies to detect PA and LF in the lethal toxin complex.

  8. Structure and Stability of Pentafluoroaniline and 4-Aminononafluorobiphenyl Radical Anions: Optically Detected Electron Paramagnetic Resonance, Time-Resolved Fluorescence, Time-Resolved Magnetic Field Effect, and Quantum Chemical Study.

    PubMed

    Borovkov, Vsevolod I; Beregovaya, Irina V; Shchegoleva, Lyudmila N; Blinkova, Svetlana V; Ovchinnikov, Dmitry A; Gurskaya, Larisa Yu; Shteingarts, Vitaly D; Bagryansky, Victor A; Molin, Yuriy N

    2015-08-01

    Radical anions (RAs) are the key intermediates of the selective hydrodefluorination of polyfluoroarenes. We used the techniques of optically detected electron paramagnetic resonance (OD EPR), time-resolved fluorescence, time-resolved magnetic field effect (TR MFE), and the density functional theory to study the possibility of RAs formation from 4-aminononafluorobiphenyl (1) and pentafluoroaniline (2) and estimate their lifetimes and decay channels. To our knowledge, both RAs have not been detected earlier. We have registered the OD EPR spectrum for relatively stable in nonpolar solutions 1(-•) but failed to register the spectra for 2(-•). However, we have managed to fix the 2(-•) by the TR MFE method and obtained its hyperfine coupling constants. The lifetime of 2(-•) was found to be only a few nanoseconds. The activation energy of its decay was estimated to be 3.6 ± 0.3 kcal/mol. According to the calculation results, the short lifetime of 2(-•) is due to the RA fast fragmentation with the F(-) elimination from ortho-position to the amine group. The calculated energy barrier, 3.2 kcal/mol, is close to the experimental value. The fragmentation of 2(-•) in a nonpolar solvent is possible due to the stabilization of the incipient F(-) anion by the binding with the amine group proton.

  9. Fluorescence-suppressed time-resolved Raman spectroscopy of pharmaceuticals using complementary metal-oxide semiconductor (CMOS) single-photon avalanche diode (SPAD) detector.

    PubMed

    Rojalin, Tatu; Kurki, Lauri; Laaksonen, Timo; Viitala, Tapani; Kostamovaara, Juha; Gordon, Keith C; Galvis, Leonardo; Wachsmann-Hogiu, Sebastian; Strachan, Clare J; Yliperttula, Marjo

    2016-01-01

    In this work, we utilize a short-wavelength, 532-nm picosecond pulsed laser coupled with a time-gated complementary metal-oxide semiconductor (CMOS) single-photon avalanche diode (SPAD) detector to acquire Raman spectra of several drugs of interest. With this approach, we are able to reveal previously unseen Raman features and suppress the fluorescence background of these drugs. Compared to traditional Raman setups, the present time-resolved technique has two major improvements. First, it is possible to overcome the strong fluorescence background that usually interferes with the much weaker Raman spectra. Second, using the high photon energy excitation light source, we are able to generate a stronger Raman signal compared to traditional instruments. In addition, observations in the time domain can be performed, thus enabling new capabilities in the field of Raman and fluorescence spectroscopy. With this system, we demonstrate for the first time the possibility of recording fluorescence-suppressed Raman spectra of solid, amorphous and crystalline, and non-photoluminescent and photoluminescent drugs such as caffeine, ranitidine hydrochloride, and indomethacin (amorphous and crystalline forms). The raw data acquired by utilizing only the picosecond pulsed laser and a CMOS SPAD detector could be used for identifying the compounds directly without any data processing. Moreover, to validate the accuracy of this time-resolved technique, we present density functional theory (DFT) calculations for a widely used gastric acid inhibitor, ranitidine hydrochloride. The obtained time-resolved Raman peaks were identified based on the calculations and existing literature. Raman spectra using non-time-resolved setups with continuous-wave 785- and 532-nm excitation lasers were used as reference data. Overall, this demonstration of time-resolved Raman and fluorescence measurements with a CMOS SPAD detector shows promise in diverse areas, including fundamental chemical research, the

  10. Fluorescence-suppressed time-resolved Raman spectroscopy of pharmaceuticals using complementary metal-oxide semiconductor (CMOS) single-photon avalanche diode (SPAD) detector.

    PubMed

    Rojalin, Tatu; Kurki, Lauri; Laaksonen, Timo; Viitala, Tapani; Kostamovaara, Juha; Gordon, Keith C; Galvis, Leonardo; Wachsmann-Hogiu, Sebastian; Strachan, Clare J; Yliperttula, Marjo

    2016-01-01

    In this work, we utilize a short-wavelength, 532-nm picosecond pulsed laser coupled with a time-gated complementary metal-oxide semiconductor (CMOS) single-photon avalanche diode (SPAD) detector to acquire Raman spectra of several drugs of interest. With this approach, we are able to reveal previously unseen Raman features and suppress the fluorescence background of these drugs. Compared to traditional Raman setups, the present time-resolved technique has two major improvements. First, it is possible to overcome the strong fluorescence background that usually interferes with the much weaker Raman spectra. Second, using the high photon energy excitation light source, we are able to generate a stronger Raman signal compared to traditional instruments. In addition, observations in the time domain can be performed, thus enabling new capabilities in the field of Raman and fluorescence spectroscopy. With this system, we demonstrate for the first time the possibility of recording fluorescence-suppressed Raman spectra of solid, amorphous and crystalline, and non-photoluminescent and photoluminescent drugs such as caffeine, ranitidine hydrochloride, and indomethacin (amorphous and crystalline forms). The raw data acquired by utilizing only the picosecond pulsed laser and a CMOS SPAD detector could be used for identifying the compounds directly without any data processing. Moreover, to validate the accuracy of this time-resolved technique, we present density functional theory (DFT) calculations for a widely used gastric acid inhibitor, ranitidine hydrochloride. The obtained time-resolved Raman peaks were identified based on the calculations and existing literature. Raman spectra using non-time-resolved setups with continuous-wave 785- and 532-nm excitation lasers were used as reference data. Overall, this demonstration of time-resolved Raman and fluorescence measurements with a CMOS SPAD detector shows promise in diverse areas, including fundamental chemical research, the

  11. Isolation of Mammalian Oogonial Stem Cells by Antibody-Based Fluorescence-Activated Cell Sorting.

    PubMed

    Navaroli, Deanna M; Tilly, Jonathan L; Woods, Dori C

    2016-01-01

    The ability to isolate and subsequently culture mitotically active female germ cells from adult ovaries, referred to as either oogonial stem cells (OSCs) or adult female germline stem cells (aFGSCs), has provided a robust system to study female germ cell development under multiple experimental conditions, and in many species. Flow cytometry or fluorescence-activated cell sorting (FACS) is an integral part of many isolation and characterization protocols. Here, we provide methodological details for antibody-based flow cytometric isolation of OSCs using antibodies specific for external epitopes of the proteins Ddx4 or Ifitm3, alone or in combination with the use of fluorescent reporter mice. Beginning with sample preparation, we provide point-by-point instructions to guide researchers on how to isolate OSCs using flow cytometry. PMID:27557587

  12. Effect of ouabain on metabolic oxidative state in living cardiomyocytes evaluated by time-resolved spectroscopy of endogenous NAD(P)H fluorescence

    NASA Astrophysics Data System (ADS)

    Chorvatova, Alzbeta; Elzwiei, Fathia; Mateasik, Anton; Chorvat, Dusan

    2012-10-01

    Time-resolved spectrometry of endogenous nicotinamide dinucleotide phosphate [NAD(P)H] fluorescence is a useful method to evaluate metabolic oxidative state in living cells. Ouabain is a well-known pharmaceutical drug used in the treatment of cardiovascular disease, the effects of which on myocardial metabolism were recently demonstrated. Mechanisms implicated in these actions are still poorly understood. We investigate the effect of ouabain on the metabolic oxidative state of living cardiac cells identified by time-resolved fluorescence spectroscopy of mitochondrial NAD(P)H. Spectral unmixing is used to resolve individual NAD(P)H fluorescence components. Ouabain decreased the integral intensity of NAD(P)H fluorescence, leading to a reduced component amplitudes ratio corresponding to a change in metabolic state. We also noted that lactate/pyruvate, affecting the cytosolic NADH gradient, increased the effect of ouabain on the component amplitudes ratio. Cell oxidation levels, evaluated as the percentage of oxidized NAD(P)H, decreased exponentially with rising concentrations of the cardiac glycoside. Ouabain also stimulated the mitochondrial NADH production. Our study sheds a new light on the role that ouabain plays in the regulation of metabolic state, and presents perspective on a noninvasive, pharmaceutical approach for testing the effect of drugs on the mitochondrial metabolism by means of time-resolved fluorescence spectroscopy in living cells.

  13. Antibody-based fluorescent and fluorescent ratiometric indicators for detection of phosphotyrosine.

    PubMed

    Huynh Nhat, Kim Phuong; Watanabe, Takayoshi; Yoshikoshi, Kensuke; Hohsaka, Takahiro

    2016-08-01

    Fluorescent indicators for protein phosphorylation are very important in not only fundamental biology but also biomedical applications. In this study, we developed novel fluorescent and fluorescent ratiometric indicators for detection of phosphotyrosine (pTyr) derivatives. A single-chain antibody variable fragment (scFv) against phosphotyrosine was fluorescent-labeled by incorporation of tetramethylrhodamine (TAMRA)-linked nonnatural amino acid at the N- or C-terminus. The TAMRA-labeled scFv showed fluorescence enhancement upon addition of pTyr-containing peptides based on antigen-dependent fluorescence quenching effect on TAMRA. The TAMRA-labeled scFv was further fused with enhanced green fluorescent protein (EGFP) to generate a double-labeled scFv for pTyr. In the absence of antigen, fluorescence resonance energy transfer (FRET) occurred from EGFP to TAMRA but TAMRA was quenched. The antigen-binding removed the quenching of TAMRA while FRET occurred without altering its efficiency. As a result of the FRET and antigen-dependent fluorescence quenching effect, the double-labeled scFv exhibited fluorescence ratio enhancement upon the antigen-binding. The fluorescent and fluorescent ratiometric indicators obtained in this study will become a novel tool for analysis of protein phosphorylation. Moreover, this strategy utilizes antibody derivatives, and therefore, can be easily applied to other antigen-antibody pairs to generate fluorescent ratiometric indicators for various target molecules. PMID:26896314

  14. Monitoring changes of cellular metabolism and microviscosity in vitro based on time-resolved endogenous fluorescence and its anisotropy decay dynamics

    NASA Astrophysics Data System (ADS)

    Zheng, Wei; Li, Dong; Qu, Jianan Y.

    2010-05-01

    Reduced nicotinamide adenine dinucleotide (NADH) is a well-known metabolic coenzyme and endogenous fluorophore. In this study, we develop a system that simultaneously measures time- and wavelength-resolved fluorescence to extract free and protein-bound NADH signals from total cellular fluorescence. We analyze temporal characteristics of NADH fluorescence in a mixture of NADH and lactate dehydrogenase (LDH) as well as in living cell samples. The results show that in both the NADH/LDH mixture and cell samples, a fraction of free NADH and protein-bound components can be identified. The extracted free and bound NADH signals are confirmed by time-resolved measurement of anisotropy decay of NADH fluorescence, based on the fact that free NADH is a small fluorescent molecule with much shorter rotational diffusion time than bound NADH. The ratio of free NADH signal to bound NADH signal is very different between normal and cancer cervical epithelial cells. In addition, the ratio changes significantly when the cell samples are treated with a mitochondrial inhibitor or uncoupler, demonstrating that the method is sensitive to monitor cellular metabolic activity. Finally, we demonstrate that the microviscosity for relatively small molecules such as NADH in cells could be extracted from wavelength- and time-resolved NADH fluorescence of living cell samples.

  15. Metal binding properties of fluorescent analogues of trichogin GA IV: a conformational study by time-resolved spectroscopy and molecular mechanics investigations.

    PubMed

    Venanzi, Mariano; Bocchinfuso, Gianfranco; Gatto, Emanuela; Palleschi, Antonio; Stella, Lorenzo; Formaggio, Fernando; Toniolo, Claudio

    2009-01-01

    The metal ion binding properties of two fluorescent analogues of trichogin GA IV, which is a natural undecapeptide showing significant antimicrobial activity, were studied by circular dichroism, time-resolved optical spectroscopy, and molecular mechanics calculations. Binding of Ca(II) and Gd(III) to the peptides investigated was shown to promote a structural transition from highly helical conformations to folded structures characterized by formation of a loop that embedded the metal ion. Time-resolved spectroscopy revealed that peptide dynamics is also remarkably affected by ion binding: peptide-backbone motions slowed down to the microsecond time scale. Finally, molecular mechanics calculations emphasized the role of the central Gly5-Gly6 motif, which allowed for the twisting of the peptide segment that gave rise to the formation of the binding cavity.

  16. A novel cell-based duplex high-throughput screening assay combining fluorescent Ca(2+) measurement with homogeneous time-resolved fluorescence technology.

    PubMed

    Kiss, László; Cselenyák, Attila; Varga, Ágnes; Visegrády, András

    2016-08-15

    Cell-based assays for G-protein-coupled receptor (GPCR) activation applied in high-throughput screening (HTS) monitor various readouts for second messengers or intracellular effectors. Recently, our understanding of diverging signaling pathways downstream of receptor activation and the capability of small molecules to selectively modulate signaling routes has increased substantially, underlining the importance of selecting appropriate readouts in cellular functional screens. To minimize the rate of false negatives in large-scale screening campaigns, it is crucial to maximize the chance of a ligand being detected, and generally applicable methods for detecting multiple analytes from a single well might serve this purpose. The few assays developed so far based on multiplexed GPCR readouts are limited to only certain applications and usually rely on genetic manipulations hindering screening in native or native-like cellular systems. Here we describe a more generally applicable and HTS-compatible homogeneous assay based on the combination of fluorometric detection of [Ca(2+)] with subsequent homogeneous time-resolved fluorescence (HTRF) cAMP readout in the same well. Besides describing development and validation of the assay, using a cell line recombinantly expressing the human PTH1 receptor screening of a small library is also presented, demonstrating the robustness and HTS compatibility of the novel paradigm.

  17. A novel cell-based duplex high-throughput screening assay combining fluorescent Ca(2+) measurement with homogeneous time-resolved fluorescence technology.

    PubMed

    Kiss, László; Cselenyák, Attila; Varga, Ágnes; Visegrády, András

    2016-08-15

    Cell-based assays for G-protein-coupled receptor (GPCR) activation applied in high-throughput screening (HTS) monitor various readouts for second messengers or intracellular effectors. Recently, our understanding of diverging signaling pathways downstream of receptor activation and the capability of small molecules to selectively modulate signaling routes has increased substantially, underlining the importance of selecting appropriate readouts in cellular functional screens. To minimize the rate of false negatives in large-scale screening campaigns, it is crucial to maximize the chance of a ligand being detected, and generally applicable methods for detecting multiple analytes from a single well might serve this purpose. The few assays developed so far based on multiplexed GPCR readouts are limited to only certain applications and usually rely on genetic manipulations hindering screening in native or native-like cellular systems. Here we describe a more generally applicable and HTS-compatible homogeneous assay based on the combination of fluorometric detection of [Ca(2+)] with subsequent homogeneous time-resolved fluorescence (HTRF) cAMP readout in the same well. Besides describing development and validation of the assay, using a cell line recombinantly expressing the human PTH1 receptor screening of a small library is also presented, demonstrating the robustness and HTS compatibility of the novel paradigm. PMID:27235172

  18. A Dual Readout Assay Based on Fluorescence Polarization and Time-Resolved Fluorescence Resonance Energy Transfer to Screen for RSK1 Inhibitors.

    PubMed

    Jeong, Eun-mi; Lee, Mi Young; Lee, Jeong Hyun; Lee, Byung Ho; Oh, Kwang-Seok

    2016-01-01

    A dual readout assay based on fluorescence polarization (FP) and time-resolved fluorescence resonance energy transfer (TR-FRET) exhibits many advantages over single assay technology in terms of screening quality and efficiency. In this study, we developed a dual readout assay combining FP and TR-FRET to identify ribosomal S6 kinase 1 (RSK1) inhibitors. This dual readout assay can monitor both FP and TR-FRET signals from a single RSK1 kinase reaction by using the immobilized metal affinity for phosphochemical (IMAP)-based assay. The Z' value and signal to background (S/B) ratio were 0.85 and 4.0 using FP, and 0.79 and 10.6 using TR-FRET, which led to performance of a pilot library screening against the drug repositioning set consisting of 2320 compounds with a reasonable reproducibility. From this screening, we identified 16 compounds showing greater than 50% inhibition against RSK1 for both FP and TR-FRET; 6 compounds with greater than 50% inhibition only for FP; and 4 compounds with greater than 50% inhibition only for TR-FRET. In a cell-based functional assay to validate the hit compounds, 10 compounds identified only in a single assay had little effect on the RSK-mediated phosphorylation of liver kinase B1, whereas 5 compounds showing greater than 80% inhibition for both FP and TR-FRET reduced the phosphorylation of liver kinase B1. These results demonstrate that the dual readout assay can be used to identify hit compounds by subsequently monitoring both FP and TR-FRET signals from one RSK1 reaction. PMID:27040627

  19. An 8-hour system for Salmonella detection with immunomagnetic separation and homogeneous time-resolved fluorescence PCR.

    PubMed

    Hagren, Virve; von Lode, Piia; Syrjälä, Anniina; Korpimäki, Teemu; Tuomola, Mika; Kauko, Otto; Nurmi, Jussi

    2008-07-15

    We describe a system consisting of rapid sample enrichment and homogeneous end-point PCR analysis that enables the detection of Salmonella in various food matrices in 8 h. Sample preparation starts with 6 h enrichment step in supplemented broth, after which Salmonella cells are collected with immunomagnetic particles. The particles are washed and dispensed to ready-to-use PCR reaction vessels, which contain dried assay-specific reagents and an internal amplification control. PCR is performed with a novel instrument platform utilising the sensitive label technology of time-resolved fluorometry. Qualitative assay results are automatically interpreted and available in 45 min after sample addition. The overall accuracy, sensitivity and specificity of the Magda CA Salmonella system were 99.1%, 98.4% and 100.0%, respectively, based on the evaluation of 107 samples (beef, pork, poultry and ready-to-eat meals) artificially contaminated with sub-lethally injured Salmonella cells.

  20. Time-Resolved Fluorescence Anisotropy Study of the Interaction Between DNA and a Peptide Truncated from the p53 Protein Core Domain.

    PubMed

    Liu, Chengxuan; Liang, Gaiting; Liu, Zhen; Zu, Lily

    2014-03-01

    Time-resolved fluorescence anisotropy spectroscopy was applied to study the interaction between a peptide truncated from the binding site of tumor suppressor p53 protein and the DNAs covalently labeled with 6-carboxyfluorescein (FAM) dye. Fluorescence intensity quenching and changes of anisotropy decay lifetime were monitored when FAM labeled DNA formed complex with the peptide. The results demonstrated that the sequence of DNA could not define the binding specificity between the peptide and DNA. But the anisotropy decay of FAM can be used to examine the binding affinity of the peptide to DNA. The fluorescent dynamics of FAM can also be used to represent the rigidity of the complex formed between the peptide and DNA.

  1. An integrated logic system for time-resolved fluorescent "turn-on" detection of cysteine and histidine base on terbium (III) coordination polymer-copper (II) ensemble.

    PubMed

    Xue, Shi-Fan; Lu, Ling-Fei; Wang, Qi-Xian; Zhang, Shengqiang; Zhang, Min; Shi, Guoyue

    2016-09-01

    Cysteine (Cys) and histidine (His) both play indispensable roles in many important biological activities. An enhanced Cys level can result in Alzheimer's and cardiovascular diseases. Likewise, His plays a significant role in the growth and repair of tissues as well as in controlling the transmission of metal elements in biological bases. Therefore, it is meaningful to detect Cys and His simultaneously. In this work, a novel terbium (III) coordination polymer-Cu (II) ensemble (Tb(3+)/GMP-Cu(2+)) was proposed. Guanosine monophosphate (GMP) can self-assemble with Tb(3+) to form a supramolecular Tb(3+) coordination polymer (Tb(3+)/GMP), which can be suited as a time-resolved probe. The fluorescence of Tb(3+)/GMP would be quenched upon the addition of Cu(2+), and then the fluorescence of the as-prepared Tb(3+)/GMP-Cu(2+) ensemble would be restored again in the presence of Cys or His. By incorporating N-Ethylmaleimide and Ni(2+) as masking agents, Tb(3+)/GMP-Cu(2+) was further exploited as an integrated logic system and a specific time-resolved fluorescent "turn-on" assay for simultaneously sensing His and Cys was designed. Meanwhile it can also be used in plasma samples, showing great potential to meet the need of practical application. PMID:27343597

  2. The open, the closed, and the empty: time-resolved fluorescence spectroscopy and computational analysis of RC-LH1 complexes from Rhodopseudomonas palustris.

    PubMed

    Beyer, Sebastian R; Müller, Lars; Southall, June; Cogdell, Richard J; Ullmann, G Matthias; Köhler, Jürgen

    2015-01-29

    We studied the time-resolved fluorescence of isolated RC-LH1 complexes from Rhodopseudomonas palustris as a function of the photon fluence and the repetition rate of the excitation laser. Both parameters were varied systematically over 3 orders of magnitude. On the basis of a microstate description we developed a quantitative model for RC-LH1 and obtained very good agreement between experiments and elaborate simulations based on a global master equation approach. The model allows us to predict the relative population of RC-LH1 complexes with the special pair in the neutral state or in the oxidized state P(+) and those complexes that lack a reaction center.

  3. Time-Resolved Down-Conversion of 2-Aminopurine in a DNA Hairpin: Fluorescence Anisotropy and Solvent Effects

    NASA Astrophysics Data System (ADS)

    Tourón Touceda, Patricia; Gelot, Thomas; Crégut, Olivier; Léonard, Jérémie; Haacke, Stefan

    2013-03-01

    Femtosecond fluorescence anisotropy decay measured by type II difference frequency generation provides new insight into the local structural dynamics of ΔP(-)PBS fragments of the HIV- 1 DNA primary binding sequence, labeled with 2-aminopurine.

  4. Design of peptide substrates for nanosecond time-resolved fluorescence assays of proteases: 2,3-diazabicyclo[2.2.2]oct-2-ene as a noninvasive fluorophore.

    PubMed

    Hennig, Andreas; Florea, Mara; Roth, Doris; Enderle, Thilo; Nau, Werner M

    2007-01-15

    Fluorescence protease assays were investigated with peptide substrates containing a 2,3-diazabicyclo[2.2.2]oct-2-ene-labeled asparagine (Dbo) as a fluorescent amino acid. The special characteristic of the fluorophore Dbo is its exceedingly long fluorescence lifetime (ca. 300 ns in water under air), which allows the use of nanosecond time-resolved fluorescence (Nano-TRF) detection to efficiently suppress shorter-lived background emission. In addition, the natural amino acids tryptophan and tyrosine can be employed as intramolecular fluorescence quenchers, which facilitates substrate design. Fourteen synthetic peptide substrates (composed of 2-19 amino acids) and five enzymes (trypsin, pepsin, carboxypeptidase A, leucine aminopeptidase, and chymotrypsin) were investigated and, in all 28 examined combinations, enzymatic activity was detected by monitoring the increase in steady state fluorescence with time and determining the reaction rates as kcat/Km values, which ranged from 0.2 to 80x10(6) M-1 min-1. The results suggest an excellent compatibility of the very small and hydrophilic fluorescent probe Dbo with solid-phase peptide synthesis and the investigated proteases. For all 14 peptides the fluorescence lifetimes before and after enzymatic cleavage were measured and Nano-TRF measurements were performed in 384-well microplates. The fluorescence lifetimes of the different peptides provide the basis for the rational design of Dbo-based fluorescent substrates for protease assays. Measurements in Nano-TRF mode revealed, in addition to efficient suppression of background fluorescence, an increased differentiation between cleaved and uncleaved substrate. The Dbo-based assays can be adapted for high-throughput screening.

  5. Microviscosity in polyacrylamide gels with pendant triphenyl-methane leuco derivatives: picosecond time-resolved fluorescence study

    NASA Astrophysics Data System (ADS)

    Tamai, Naoto; Ishikawa, Masazumi; Kitamura, Noboru; Masuhara, Hiroshi

    1991-10-01

    Picosecond fluorescence dynamics of triphenylmethane dyes bonded to polyacrylamide gels before and after swelling was studied by a single-photon timing technique. Microviscosity in the gels after swelling was estimated to be 10-11 cP by examining the viscosity dependence of fluorescence dynamics of malachite green in various alcohols. The results were interpreted in terms of structured stiff water in a microcavity of the gels.

  6. Time-resolved detection of aromatic compounds on planetary surfaces by ultraviolet laser induced fluorescence and Raman spectroscopy

    NASA Astrophysics Data System (ADS)

    Eshelman, E.; Daly, M. G.; Slater, G.; Cloutis, E.

    2015-12-01

    Raman spectroscopic instruments are highly capable in the search for organics on Mars due to the potential to perform rapid and nondestructive measurements on unprepared samples. Upcoming and future Raman instruments are likely to also incorporate laser-induced fluorescence (LIF) capabilities, which can be added for modest cost and complexity. We demonstrate that it is possible to obtain sub-ns fluorescence lifetime measurements of Mars-relevant organics and minerals if a fast time-gating capability is used with an intensified detector and a short ultraviolet laser pulse. This serves a primary purpose of discriminating mineral from short-lived (less than 10 ns) organic fluorescence, considered a potential biosignature. Additionally, lifetime measurements may assist in determining if more than one fluorescing species is present and provide information concerning the molecular structure as well as the local environment. Fast time-gating is also useful at longer visible or near-IR wavelengths, as this approach increases the sensitivity of the instrument to organic material by removing the majority of the fluorescence background from the Raman signal and reducing the effect of ambient light.

  7. Time-resolved spectroscopic fluorescence imaging, transient absorption and vibrational spectroscopy of intact and photo-inhibited photosynthetic tissue.

    PubMed

    Lukins, Philip B; Rehman, Shakil; Stevens, Gregory B; George, Doaa

    2005-01-01

    Fluorescence, absorption and vibrational spectroscopic techniques were used to study spinach at the photosystem II (PS II), chloroplast and cellular levels and to determine the effects and mechanisms of ultraviolet-B (UV-B) photoinhibition of these structures. Two-photon fluorescence spectroscopic imaging of intact chloroplasts shows significant spatial variations in the component fluorescence spectra in the range 640-740 nm, indicating that the type and distribution of chlorophylls vary markedly with position in the chloroplast. The chlorophyll distributions and excitonic behaviour in chloroplasts and whole plant tissue were studied using picosecond time-gated fluorescence imaging, which also showed UV-induced kinetic changes that clearly indicate that UV-B induces both structural and excitonic uncoupling of chlorophylls within the light-harvesting complexes. Transient absorption measurements and low-frequency infrared and Raman spectroscopy show that the predominant sites of UV-B damage in PS II are at the oxygen-evolving centre (OEC) itself, as well as at specific locations near the OEC-binding sites.

  8. Excitation relaxation dynamics and energy transfer in fucoxanthin-chlorophyll a/c-protein complexes, probed by time-resolved fluorescence.

    PubMed

    Akimoto, Seiji; Teshigahara, Ayaka; Yokono, Makio; Mimuro, Mamoru; Nagao, Ryo; Tomo, Tatsuya

    2014-09-01

    In algae, light-harvesting complexes contain specific chlorophylls (Chls) and keto-carotenoids; Chl a, Chl c, and fucoxanthin (Fx) in diatoms and brown algae; Chl a, Chl c, and peridinin in photosynthetic dinoflagellates; and Chl a, Chl b, and siphonaxanthin in green algae. The Fx-Chl a/c-protein (FCP) complex from the diatom Chaetoceros gracilis contains Chl c1, Chl c2, and the keto-carotenoid, Fx, as antenna pigments, in addition to Chl a. In the present study, we investigated energy transfer in the FCP complex associated with photosystem II (FCPII) of C. gracilis. For these investigations, we analyzed time-resolved fluorescence spectra, fluorescence rise and decay curves, and time-resolved fluorescence anisotropy data. Chl a exhibited different energy forms with fluorescence peaks ranging from 677 nm to 688 nm. Fx transferred excitation energy to lower-energy Chl a with a time constant of 300fs. Chl c transferred excitation energy to Chl a with time constants of 500-600fs (intra-complex transfer), 600-700fs (intra-complex transfer), and 4-6ps (inter-complex transfer). The latter process made a greater contribution to total Chl c-to-Chl a transfer in intact cells of C. gracilis than in the isolated FCPII complexes. The lower-energy Chl a received excitation energy from Fx and transferred the energy to higher-energy Chl a. This article is part of a special issue entitled: photosynthesis research for sustainability: keys to produce clean energy.

  9. Time-resolved fluorescence of bacteriophage Pf1 DNA-binding protein. Determination of oligonucleotide and polynucleotide binding parameters.

    PubMed

    Kneale, G G; Wijnaendts van Resandt, R W

    1985-05-15

    The binding of oligonucleotides and polynucleotides to the Pf1 DNA-binding protein was followed by fluorescence spectral shift and lifetime measurements, which gave an anomalous value for the stoichiometry of binding. The anomaly was investigated in detail using fluorescence depolarisation to measure the aggregation during the titration and showed that all the fluorescence parameters are related to the specific aggregation of dimers on ligand binding. At saturation, complexes of the protein with the octanucleotide d(GCGTTGCG) and the hexadecanucleotide (dT)16 have rotational correlation times, phi, of 50 ns and 85 ns, corresponding to protein tetramers and octamers, respectively. In the presence of the tetranucleotide d(CGCA) the protein remains as the native dimer (phi = 19 ns). The titration curves could be analysed in terms of two non-equivalent binding sites, with binding constants K1 and K2. Comparison of K1 values for oligonucleotide binding leads to an estimated (single-site) intrinsic binding constant Kint approximately equal to 3 X 10(4) M-1 and a cooperativity parameter omega approximately equal to 100, in agreement with the apparent binding constant Kapp approximately equal to 3 X 10(6) M-1 for polynucleotides. Binding to the second site on the protein dimer is greatly reduced and cannot be determined accurately. The results suggest that the protein dimers bind cooperatively by lateral association along the DNA and that occupation of only one of the two DNA-binding sites of the protein dimers is sufficient to stabilize the nucleoprotein complexes.

  10. Density relaxation and particle motion characteristics in a non-ionic deep eutectic solvent (acetamide + urea): time-resolved fluorescence measurements and all-atom molecular dynamics simulations.

    PubMed

    Das, Anuradha; Das, Suman; Biswas, Ranjit

    2015-01-21

    Temperature dependent relaxation dynamics, particle motion characteristics, and heterogeneity aspects of deep eutectic solvents (DESs) made of acetamide (CH3CONH2) and urea (NH2CONH2) have been investigated by employing time-resolved fluorescence measurements and all-atom molecular dynamics simulations. Three different compositions (f) for the mixture [fCH3CONH2 + (1 - f)NH2CONH2] have been studied in a temperature range of 328-353 K which is ∼120-145 K above the measured glass transition temperatures (∼207 K) of these DESs but much lower than the individual melting temperature of either of the constituents. Steady state fluorescence emission measurements using probe solutes with sharply different lifetimes do not indicate any dependence on excitation wavelength in these metastable molten systems. Time-resolved fluorescence anisotropy measurements reveal near-hydrodynamic coupling between medium viscosity and rotation of a dissolved dipolar solute. Stokes shift dynamics have been found to be too fast to be detected by the time-resolution (∼70 ps) employed, suggesting extremely rapid medium polarization relaxation. All-atom simulations reveal Gaussian distribution for particle displacements and van Hove correlations, and significant overlap between non-Gaussian (α2) and new non-Gaussian (γ) heterogeneity parameters. In addition, no stretched exponential relaxations have been detected in the simulated wavenumber dependent acetamide dynamic structure factors. All these results are in sharp contrast to earlier observations for ionic deep eutectics with acetamide [Guchhait et al., J. Chem. Phys. 140, 104514 (2014)] and suggest a fundamental difference in interaction and dynamics between ionic and non-ionic deep eutectic solvent systems.

  11. Density relaxation and particle motion characteristics in a non-ionic deep eutectic solvent (acetamide + urea): Time-resolved fluorescence measurements and all-atom molecular dynamics simulations

    NASA Astrophysics Data System (ADS)

    Das, Anuradha; Das, Suman; Biswas, Ranjit

    2015-01-01

    Temperature dependent relaxation dynamics, particle motion characteristics, and heterogeneity aspects of deep eutectic solvents (DESs) made of acetamide (CH3CONH2) and urea (NH2CONH2) have been investigated by employing time-resolved fluorescence measurements and all-atom molecular dynamics simulations. Three different compositions (f) for the mixture [fCH3CONH2 + (1 - f)NH2CONH2] have been studied in a temperature range of 328-353 K which is ˜120-145 K above the measured glass transition temperatures (˜207 K) of these DESs but much lower than the individual melting temperature of either of the constituents. Steady state fluorescence emission measurements using probe solutes with sharply different lifetimes do not indicate any dependence on excitation wavelength in these metastable molten systems. Time-resolved fluorescence anisotropy measurements reveal near-hydrodynamic coupling between medium viscosity and rotation of a dissolved dipolar solute. Stokes shift dynamics have been found to be too fast to be detected by the time-resolution (˜70 ps) employed, suggesting extremely rapid medium polarization relaxation. All-atom simulations reveal Gaussian distribution for particle displacements and van Hove correlations, and significant overlap between non-Gaussian (α2) and new non-Gaussian (γ) heterogeneity parameters. In addition, no stretched exponential relaxations have been detected in the simulated wavenumber dependent acetamide dynamic structure factors. All these results are in sharp contrast to earlier observations for ionic deep eutectics with acetamide [Guchhait et al., J. Chem. Phys. 140, 104514 (2014)] and suggest a fundamental difference in interaction and dynamics between ionic and non-ionic deep eutectic solvent systems.

  12. Density relaxation and particle motion characteristics in a non-ionic deep eutectic solvent (acetamide + urea): Time-resolved fluorescence measurements and all-atom molecular dynamics simulations

    SciTech Connect

    Das, Anuradha; Das, Suman; Biswas, Ranjit

    2015-01-21

    Temperature dependent relaxation dynamics, particle motion characteristics, and heterogeneity aspects of deep eutectic solvents (DESs) made of acetamide (CH{sub 3}CONH{sub 2}) and urea (NH{sub 2}CONH{sub 2}) have been investigated by employing time-resolved fluorescence measurements and all-atom molecular dynamics simulations. Three different compositions (f) for the mixture [fCH{sub 3}CONH{sub 2} + (1 − f)NH{sub 2}CONH{sub 2}] have been studied in a temperature range of 328-353 K which is ∼120-145 K above the measured glass transition temperatures (∼207 K) of these DESs but much lower than the individual melting temperature of either of the constituents. Steady state fluorescence emission measurements using probe solutes with sharply different lifetimes do not indicate any dependence on excitation wavelength in these metastable molten systems. Time-resolved fluorescence anisotropy measurements reveal near-hydrodynamic coupling between medium viscosity and rotation of a dissolved dipolar solute. Stokes shift dynamics have been found to be too fast to be detected by the time-resolution (∼70 ps) employed, suggesting extremely rapid medium polarization relaxation. All-atom simulations reveal Gaussian distribution for particle displacements and van Hove correlations, and significant overlap between non-Gaussian (α{sub 2}) and new non-Gaussian (γ) heterogeneity parameters. In addition, no stretched exponential relaxations have been detected in the simulated wavenumber dependent acetamide dynamic structure factors. All these results are in sharp contrast to earlier observations for ionic deep eutectics with acetamide [Guchhait et al., J. Chem. Phys. 140, 104514 (2014)] and suggest a fundamental difference in interaction and dynamics between ionic and non-ionic deep eutectic solvent systems.

  13. Density relaxation and particle motion characteristics in a non-ionic deep eutectic solvent (acetamide + urea): time-resolved fluorescence measurements and all-atom molecular dynamics simulations.

    PubMed

    Das, Anuradha; Das, Suman; Biswas, Ranjit

    2015-01-21

    Temperature dependent relaxation dynamics, particle motion characteristics, and heterogeneity aspects of deep eutectic solvents (DESs) made of acetamide (CH3CONH2) and urea (NH2CONH2) have been investigated by employing time-resolved fluorescence measurements and all-atom molecular dynamics simulations. Three different compositions (f) for the mixture [fCH3CONH2 + (1 - f)NH2CONH2] have been studied in a temperature range of 328-353 K which is ∼120-145 K above the measured glass transition temperatures (∼207 K) of these DESs but much lower than the individual melting temperature of either of the constituents. Steady state fluorescence emission measurements using probe solutes with sharply different lifetimes do not indicate any dependence on excitation wavelength in these metastable molten systems. Time-resolved fluorescence anisotropy measurements reveal near-hydrodynamic coupling between medium viscosity and rotation of a dissolved dipolar solute. Stokes shift dynamics have been found to be too fast to be detected by the time-resolution (∼70 ps) employed, suggesting extremely rapid medium polarization relaxation. All-atom simulations reveal Gaussian distribution for particle displacements and van Hove correlations, and significant overlap between non-Gaussian (α2) and new non-Gaussian (γ) heterogeneity parameters. In addition, no stretched exponential relaxations have been detected in the simulated wavenumber dependent acetamide dynamic structure factors. All these results are in sharp contrast to earlier observations for ionic deep eutectics with acetamide [Guchhait et al., J. Chem. Phys. 140, 104514 (2014)] and suggest a fundamental difference in interaction and dynamics between ionic and non-ionic deep eutectic solvent systems. PMID:25612718

  14. Interaction of quinine sulfate with anionic micelles of sodium dodecylsulfate: A time-resolved fluorescence spectroscopy at different pH

    NASA Astrophysics Data System (ADS)

    Joshi, Sunita; Pant, Debi D.

    2015-09-01

    Photophysical behavior and rotational relaxation dynamics of quinine sulfate (QS) in anionic surfactant, sodium dodecylsulfate (SDS) at different pH have been studied using steady state and time resolved fluorescence spectroscopy. It has been observed that the cationic form of quinine sulfate (at pH 2) forms a fluorescent ion pair complex with the surfactant molecules at lower concentrations of surfactant. However, for higher concentrations of SDS, the probe molecules bind strongly with the micelles and reside at the water-micelle interface. At pH 7, QS is singly protonated in bulk aqueous solution. At lower concentrations of SDS aggregation between probe and surfactant molecules has been observed. However, for higher concentrations of SDS, an additional fluorescence peak corresponding to dicationic form of QS appears and this has been attributed to double protonation of the QS molecule in micellar solution. At pH 7, in the presence of SDS micelles, the photophysical properties of QS showed substantial changes compared to that in the bulk water solution. At pH 12, an increase in fluorescence intensity and lifetime has been observed and this has been attributed to the increase in radiative rate due to the incorporation of QS at the micelle-water interface. The local pH at micellar surface has been found different from the pH of bulk solution.

  15. Surface speciation of Eu3+ adsorbed on kaolinite by time-resolved laser fluorescence spectroscopy (TRLFS) and parallel factor analysis (PARAFAC).

    PubMed

    Ishida, Keisuke; Saito, Takumi; Aoyagi, Noboru; Kimura, Takaumi; Nagaishi, Ryuji; Nagasaki, Shinya; Tanaka, Satoru

    2012-05-15

    Time-resolved laser fluorescence spectroscopy (TRLFS) is an effective speciation technique for fluorescent metal ions and can be further extended by the parallel factor analysis (PARAFAC). The adsorption of Eu(3+) on kaolinite as well as gibbsite as a reference mineral was investigated by TRLFS together with batch adsorption measurements. The PAFAFAC modeling provided the fluorescence spectra, decay lifetimes, and relative intensity profiles of three Eu(3+) surface complexes with kaolinite; an outer-sphere (factor A) complex and two inner-sphere (factors B and C) complexes. Their intensity profiles qualitatively explained the measured adsorption of Eu(3+). Based on the TRLFS results in varied H(2)O/D(2)O media, it was shown that the outer-sphere complex exhibited more rapid fluorescence decay than Eu(3+) aquo ion, because of the energy transfer to the surface. Factor B was an inner-sphere complex, which became dominant at relatively high pH, high salt concentration and low Eu(3+) concentration. Its spectrum and lifetime were similar to those of Eu(3+) adsorbed on gibbsite, suggesting its occurrence on the edge face of the gibbsite layer of kaolinite. From the comparison with the spectra and lifetimes of crystalline or aqueous Eu(OH)(3), factor C was considered as a poly-nuclear surface complex of Eu(3+) formed at relatively high Eu(3+) concentration.

  16. Homodimerization of amyloid precursor protein at the plasma membrane: a homoFRET study by time-resolved fluorescence anisotropy imaging.

    PubMed

    Devauges, Viviane; Marquer, Catherine; Lécart, Sandrine; Cossec, Jack-Christophe; Potier, Marie-Claude; Fort, Emmanuel; Suhling, Klaus; Lévêque-Fort, Sandrine

    2012-01-01

    Classical FRET (Förster Resonance Energy Transfer) using two fluorescent labels (one for the donor and another one for the acceptor) is not efficient for studying the homodimerization of a protein as only half of the homodimers formed can be identified by this technique. We thus resorted to homoFRET detected by time-resolved Fluorescence Anisotropy IMaging (tr-FAIM). To specifically image the plasma membrane of living cells, an original combination of tr-FAIM and Total Internal Reflection Fluorescence Lifetime Imaging Microscope (TIRFLIM) was implemented. The correcting factor accounting for the depolarization due to the high numerical aperture (NA) objective, mandatory for TIRF microscopy, was quantified on fluorescein solutions and on HEK293 cells expressing enhanced Green Fluorescence Protein (eGFP). Homodimerization of Amyloid Precursor Protein (APP), a key mechanism in the etiology of Alzheimer's disease, was measured on this original set-up. We showed, both in epifluorescence and under TIRF excitation, different energy transfer rates associated with the homodimerization of wild type APP-eGFP or of a mutated APP-eGFP, which forms constitutive dimers. This original set-up thus offers promising prospects for future studies of protein homodimerization in living cells in control and pathological conditions.

  17. Modification of energy-transfer processes in the cyanobacterium, Arthrospira platensis, to adapt to light conditions, probed by time-resolved fluorescence spectroscopy.

    PubMed

    Akimoto, Seiji; Yokono, Makio; Aikawa, Shimpei; Kondo, Akihiko

    2013-11-01

    In cyanobacteria, the interactions among pigment-protein complexes are modified in response to changes in light conditions. In the present study, we analyzed excitation energy transfer from the phycobilisome and photosystem II to photosystem I in the cyanobacterium Arthrospira (Spirulina) platensis. The cells were grown under lights with different spectral profiles and under different light intensities, and the energy-transfer characteristics were evaluated using steady-state absorption, steady-state fluorescence, and picosecond time-resolved fluorescence spectroscopy techniques. The fluorescence rise and decay curves were analyzed by global analysis to obtain fluorescence decay-associated spectra. The direct energy transfer from the phycobilisome to photosystem I and energy transfer from photosystem II to photosystem I were modified depending on the light quality, light quantity, and cultivation period. However, the total amount of energy transferred to photosystem I remained constant under the different growth conditions. We discuss the differences in energy-transfer processes under different cultivation and light conditions. PMID:23605291

  18. Energy transfer in the primary stages of the photosynthetic process investigated by picosecond time resolved fluorescence spectroscopy

    NASA Astrophysics Data System (ADS)

    Pellegrino, F.

    The fate of the absorbed light energy in the primary stages of the photosynthetic process was studied. In particular, the energy transfer in the accessory pigment complex consisting of carotenoids, Chl. a and Chl. b in higher green plants and phycobiliproteins in blue-green algae were investigated. These accessory pigments are responsible for the highly efficient transfer of the excitation energy to the photochemically active reaction center traps. The risetime, decay time, fluorescence depolarization, temperature and intensity dependence of the fluoresence emission from higher green plant and algal photosystems were directly measured. Excitation was provided by single picosecond laser pulses, as well as a train of pulses at 530 nm, within an intensity range of 10 to the 12th power to 10 to the 16th power photons/sq cm per pulse.

  19. Laguerre-based method for analysis of time-resolved fluorescence data: application to in-vivo characterization and diagnosis of atherosclerotic lesions

    NASA Astrophysics Data System (ADS)

    Jo, Javier A.; Fang, Qiyin; Papaioannou, Thanassis; Baker, J. Dennis; Dorafshar, Amir; Reil, Todd; Qiao, Jianhua; Fishbein, Michael C.; Freischlag, Julie A.; Marcu, Laura

    2006-03-01

    We report the application of the Laguerre deconvolution technique (LDT) to the analysis of in-vivo time-resolved laser-induced fluorescence spectroscopy (TR-LIFS) data and the diagnosis of atherosclerotic plaques. TR-LIFS measurements were obtained in vivo from normal and atherosclerotic aortas (eight rabbits, 73 areas), and subsequently analyzed using LDT. Spectral and time-resolved features were used to develop four classification algorithms: linear discriminant analysis (LDA), stepwise LDA (SLDA), principal component analysis (PCA), and artificial neural network (ANN). Accurate deconvolution of TR-LIFS in-vivo measurements from normal and atherosclerotic arteries was provided by LDT. The derived Laguerre expansion coefficients reflected changes in the arterial biochemical composition, and provided a means to discriminate lesions rich in macrophages with high sensitivity (>85%) and specificity (>95%). Classification algorithms (SLDA and PCA) using a selected number of features with maximum discriminating power provided the best performance. This study demonstrates the potential of the LDT for in-vivo tissue diagnosis, and specifically for the detection of macrophages infiltration in atherosclerotic lesions, a key marker of plaque vulnerability.

  20. Tracking Local Conformational Changes of Ribonuclease A Using Picosecond Time-Resolved Fluorescence of the Six Tyrosine Residues

    PubMed Central

    Noronha, Melinda; Lima, João C.; Paci, Emanuele; Santos, Helena; Maçanita, António L.

    2007-01-01

    The six tyrosine residues of ribonuclease A (RNase A) are used as individual intrinsic probes for tracking local conformational changes during unfolding. The fluorescence decays of RNase A are well described by sums of three exponentials with decay times (τ1 = 1.7 ns, τ2 = 180 ps, and τ3 = 30 ps) and preexponential coefficients (A1 = 1, A2 = 1, and A3 = 4) at pH 7, 25°C. The decay times are controlled by photo-induced electron transfer from individual tyrosine residues to the nearest disulphide (–SS–), bridge, which is distance (R) dependent. We assign τ1 to Tyr-76 (R = 12.8 Å), τ2 to Tyr-115 (R = 6.9 Å), and τ3 to Tyr-25, Tyr-73, Tyr-92, and Tyr-97 (all four at R = 5.5 ± 0.3 Å) at 23°C. On the basis of this assignment, the results show that, upon thermal or chemical unfolding only Tyr-25, Tyr-92, and Tyr-76 undergo significant displacement from their nearest –SS– bridge. Despite reporting on different regions of the protein, the concordance between the transition temperatures, Tm, obtained from Tyr-76 (Tm = 59.2°C) and Tyr-25 and Tyr-92 (Tm = 58.2°C) suggests a single unfolding event in this temperature range that affects all these regions similarly. PMID:17384067

  1. Role of Coherent Low-Frequency Motion in Excited-State Proton Transfer of Green Fluorescent Protein Studied by Time-Resolved Impulsive Stimulated Raman Spectroscopy.

    PubMed

    Fujisawa, Tomotsumi; Kuramochi, Hikaru; Hosoi, Haruko; Takeuchi, Satoshi; Tahara, Tahei

    2016-03-30

    Green fluorescent protein (GFP) from jellyfish Aequorea victoria, an essential bioimaging tool, luminesces via excited-state proton transfer (ESPT) in which the phenolic proton of the p-hydroxybenzylideneimidazolinone chromophore is transferred to Glu222 through a hydrogen-bond network. In this process, the ESPT mediated by the low-frequency motion of the chromophore has been proposed. We address this issue using femtosecond time-resolved impulsive stimulated Raman spectroscopy. After coherently exciting low-frequency modes (<300 cm(-1)) in the excited state of GFP, we examined the excited-state structural evolution and the ESPT dynamics within the dephasing time of the low-frequency vibration. A clear anharmonic vibrational coupling is found between one high-frequency mode of the chromophore (phenolic CH bend) and a low-frequency mode at ∼104 cm(-1). However, the data show that this low-frequency motion does not substantially affect the ESPT dynamics. PMID:26943852

  2. Time-resolved fluorescence sensing of pesticides chlorpyrifos, crotoxyphos and endosulfan by the luminescent Eu(III)-8-allyl-3-carboxycoumarin probe.

    PubMed

    Azab, Hassan A; Khairy, Gasser M; Kamel, Rasha M

    2015-09-01

    This work describes the application of time resolved fluorescence in microtiter plates for investigating the interactions of europium-allyl-3-carboxycoumarin with pesticides chlorpyrifos, endosulfan and crotoxyphos. Stern-Volmer studies at different temperatures for chlorpyrifos and crotoxyphos shows dynamic and static quenching mechanisms respectively. Direct methods for the determination of the pesticides under investigation have been developed using the luminescence variations of the probe in solution. The detection limits are 6.53, 0.004, 3.72 μmol/L for chlorpyrifos, endosulfan, and crotoxyphos, respectively. The binding constants and thermodynamic parameters of the pesticides with probe were evaluated. A thermodynamic analysis showed that the reaction is spontaneous with negative ΔG. Effect of some relevant interferents on the detection of pesticides has been investigated. The new method was applied to the determination of the pesticides in different types of water samples (tap, mineral, and waste water).

  3. Differences in excitation energy transfer of Arthrospira platensis cells grown in seawater medium and freshwater medium, probed by time-resolved fluorescence spectroscopy

    NASA Astrophysics Data System (ADS)

    Arba, Muhammad; Aikawa, Shimpei; Niki, Kenta; Yokono, Makio; Kondo, Akihiko; Akimoto, Seiji

    2013-11-01

    Excitation energy transfer of Arthrospira platensis cells grown in f/2 medium (a high salinity medium) and SOT medium (a control) was investigated by steady-state and time-resolved spectroscopies. Growth in f/2 medium induced changes in absorption and fluorescence spectra as well as in the energy transfer pathways. Excitation energy captured by phycobilisome (PBS) was transferred directly to photosystem (PS) I, instead of being first transferred to an intermediate (PBS → PSII → PSI), as observed in SOT medium. The respiration rate increased while photosynthetic rate reduced in f/2 medium. Possible causes of the differences in light-harvesting and energy-transfer processes between the two media are discussed.

  4. Conformational States of the Rapana thomasiana Hemocyanin and Its Substructures Studied by Dynamic Light Scattering and Time-Resolved Fluorescence Spectroscopy

    PubMed Central

    Georgieva, Dessislava; Schwark, Daniel; Nikolov, Peter; Idakieva, Krassimira; Parvanova, Katja; Dierks, Karsten; Genov, Nicolay; Betzel, Christian

    2005-01-01

    Hemocyanins are dioxygen-transporting proteins freely dissolved in the hemolymph of mollusks and arthropods. Dynamic light scattering and time-resolved fluorescence measurements show that the oxygenated and apo-forms of the Rapana thomasiana hemocyanin, its structural subunits RtH1 and RtH2, and those of the functional unit RtH2e, exist in different conformations. The oxygenated respiratory proteins are less compact and more asymmetric than the respective apo-forms. Different conformational states were also observed for the R. thomasiana hemocyanin in the absence and presence of an allosteric regulator. The results are in agreement with a molecular mechanism for cooperative dioxygen binding in molluscan hemocyanins including transfer of conformational changes from one functional unit to another. PMID:15533921

  5. Time-resolved fluorescence sensing of pesticides chlorpyrifos, crotoxyphos and endosulfan by the luminescent Eu(III)-8-allyl-3-carboxycoumarin probe

    NASA Astrophysics Data System (ADS)

    Azab, Hassan A.; Khairy, Gasser M.; Kamel, Rasha M.

    2015-09-01

    This work describes the application of time resolved fluorescence in microtiter plates for investigating the interactions of europium-allyl-3-carboxycoumarin with pesticides chlorpyrifos, endosulfan and crotoxyphos. Stern-Volmer studies at different temperatures for chlorpyrifos and crotoxyphos shows dynamic and static quenching mechanisms respectively. Direct methods for the determination of the pesticides under investigation have been developed using the luminescence variations of the probe in solution. The detection limits are 6.53, 0.004, 3.72 μmol/L for chlorpyrifos, endosulfan, and crotoxyphos, respectively. The binding constants and thermodynamic parameters of the pesticides with probe were evaluated. A thermodynamic analysis showed that the reaction is spontaneous with negative ΔG. Effect of some relevant interferents on the detection of pesticides has been investigated. The new method was applied to the determination of the pesticides in different types of water samples (tap, mineral, and waste water).

  6. Speciation studies on DTPA using the complementary nature of electrospray ionization mass spectrometry and time-resolved laser-induced fluorescence.

    PubMed

    Moulin, Christophe; Amekraz, Badia; Steiner, Valerie; Plancque, Gabriel; Ansoborlo, Eric

    2003-09-01

    Decorporation of radionuclides is of continuous interest in order to reduce doses in case of occupational or accidental human exposure. In the present study, insights into the non-covalent interactions that hold the well-known chelating agent DTPA (diethylenetriaminepentaacetic acid) with inorganic elements of interest, such as europium and strontium, and their ability to form stable complexes, are investigated with two spectroscopic techniques, i.e., electrospray ionization mass spectrometry (ESI-MS) and time-resolved laser-induced fluorescence (TRLIF). First investigations are on DTPA and europium alone and end with a complete study of the Eu-DTPA system. The pH variation allows one to readily investigate whether different species (protonated, hydrolyzed, etc.) exist in the pH range 2-9 and evaluate the stoichiometry and conditional stability constant for the Eu-DTPA complex. Additional experiments by ESI-MS are reported for Sr(II) in interaction with DTPA and EDTA.

  7. Interaction of Cm(III) and Am(III) with human serum transferrin studied by time-resolved laser fluorescence and EXAFS spectroscopy.

    PubMed

    Bauer, Nicole; Fröhlich, Daniel R; Panak, Petra J

    2014-05-14

    The complexation of Cm(III) with human serum transferrin was investigated in a pH range from 3.5 to 11.0 using time-resolved laser fluorescence spectroscopy (TRLFS). At pH ≥ 7.4 Cm(III) is incorporated at the Fe(III) binding site of transferrin whereas at lower pH a partially bound Cm(III) transferrin species is formed. At physiological temperature (310 K) at pH 7.4, about 70% of the partially bound and 30% of the incorporated Cm(III) transferrin species are present in solution. The Cm(III) results obtained by TRLFS are in very good agreement with Am(III) EXAFS results, confirming the incorporation of Am(III) at the Fe(III) binding site at pH 8.5.

  8. Direct on-strip analysis of size- and time-resolved aerosol impactor samples using laser induced fluorescence spectra excited at 263 and 351 nm.

    PubMed

    Wang, Chuji; Pan, Yong-Le; James, Deryck; Wetmore, Alan E; Redding, Brandon

    2014-04-11

    We report a novel atmospheric aerosol characterization technique, in which dual wavelength UV laser induced fluorescence (LIF) spectrometry marries an eight-stage rotating drum impactor (RDI), namely UV-LIF-RDI, to achieve size- and time-resolved analysis of aerosol particles on-strip. The UV-LIF-RDI technique measured LIF spectra via direct laser beam illumination onto the particles that were impacted on a RDI strip with a spatial resolution of 1.2mm, equivalent to an averaged time resolution in the aerosol sampling of 3.6 h. Excited by a 263 nm or 351 nm laser, more than 2000 LIF spectra within a 3-week aerosol collection time period were obtained from the eight individual RDI strips that collected particles in eight different sizes ranging from 0.09 to 10 μm in Djibouti. Based on the known fluorescence database from atmospheric aerosols in the US, the LIF spectra obtained from the Djibouti aerosol samples were found to be dominated by fluorescence clusters 2, 5, and 8 (peaked at 330, 370, and 475 nm) when excited at 263 nm and by fluorescence clusters 1, 2, 5, and 6 (peaked at 390 and 460 nm) when excited at 351 nm. Size- and time-dependent variations of the fluorescence spectra revealed some size and time evolution behavior of organic and biological aerosols from the atmosphere in Djibouti. Moreover, this analytical technique could locate the possible sources and chemical compositions contributing to these fluorescence clusters. Advantages, limitations, and future developments of this new aerosol analysis technique are also discussed.

  9. Direct on-strip analysis of size- and time-resolved aerosol impactor samples using laser induced fluorescence spectra excited at 263 and 351 nm.

    PubMed

    Wang, Chuji; Pan, Yong-Le; James, Deryck; Wetmore, Alan E; Redding, Brandon

    2014-04-11

    We report a novel atmospheric aerosol characterization technique, in which dual wavelength UV laser induced fluorescence (LIF) spectrometry marries an eight-stage rotating drum impactor (RDI), namely UV-LIF-RDI, to achieve size- and time-resolved analysis of aerosol particles on-strip. The UV-LIF-RDI technique measured LIF spectra via direct laser beam illumination onto the particles that were impacted on a RDI strip with a spatial resolution of 1.2mm, equivalent to an averaged time resolution in the aerosol sampling of 3.6 h. Excited by a 263 nm or 351 nm laser, more than 2000 LIF spectra within a 3-week aerosol collection time period were obtained from the eight individual RDI strips that collected particles in eight different sizes ranging from 0.09 to 10 μm in Djibouti. Based on the known fluorescence database from atmospheric aerosols in the US, the LIF spectra obtained from the Djibouti aerosol samples were found to be dominated by fluorescence clusters 2, 5, and 8 (peaked at 330, 370, and 475 nm) when excited at 263 nm and by fluorescence clusters 1, 2, 5, and 6 (peaked at 390 and 460 nm) when excited at 351 nm. Size- and time-dependent variations of the fluorescence spectra revealed some size and time evolution behavior of organic and biological aerosols from the atmosphere in Djibouti. Moreover, this analytical technique could locate the possible sources and chemical compositions contributing to these fluorescence clusters. Advantages, limitations, and future developments of this new aerosol analysis technique are also discussed. PMID:24745745

  10. Experimental support for a novel compount motion model for the time-resolved fluorescence anisotropy decay of TMA-PDH in lipid vesicle bilayers

    NASA Astrophysics Data System (ADS)

    Muller, Johan M.; van Faassen, Ernst E.; van Ginkel, Gijsbert

    1994-08-01

    Many structural studies of lipid membrane systems employ fluorescence anisotropy experiments on lipid soluble dyes that have been embedded in the lipid bilayers as optical probes. The conventional models for the interpretation of the anisotropy decay curves have a number of conceptual problems related to the form of the effective potential experienced by the probe molecules due to the interaction with the surrounding lipid environment. Therefore, a new model recently proposed by van der Sijs. (Chem. Phys. Letters 216 (1993) 559). In this paper we test this new compound model in a reanalysis of time-resolved fluorescence anisotropy experiments using TMA-DPH as a fluorescent probe. The orientational order and reorientational dynamics of TMA-DPH in small unilamellar vesicles (SUV) of POPC, DOPC, EGGPC, DLPC, EGGPG, DOPG, SQDG and DGDG was studied. We find that the new model improves the description of the underlying motional processes at short time scales and lacks certain unphysical aspects of previous models, e.g. a population of TMA-DPH probe molecules at and beyond 90° with the local bilayer normal. In contrast with previous models the compound motion model yields a good agreement between our vesicle data and previously published results from oriental lipid bilayers using ESR and AFD.

  11. Effect of Ca2+ on the Steady-State and Time-Resolved Emission Properties of the Genetically Encoded Fluorescent Sensor CatchER

    PubMed Central

    2015-01-01

    We previously designed a calcium sensor CatchER (a GFP-based Calcium sensor for detecting high concentrations in the high calcium concentration environment such as ER) with a capability for monitoring calcium ion responses in various types of cells. Calcium binding to CatchER induces the ratiometric changes in the absorption spectra, as well as an increase in fluorescence emission at 510 nm upon excitation at both 395 and 488 nm. Here, we have applied the combination of the steady-state and time-resolved optical methods and Hydrogen/Deuterium isotope exchange to understand the origin of such calcium-induced optical property changes of CatchER. We first demonstrated that calcium binding results in a 44% mean fluorescence lifetime increase of the indirectly excited anionic chromophore. Thus, CatchER is the first protein-based calcium indicator with the single fluorescent moiety to show the direct correlation between the lifetime and calcium binding. Calcium exhibits a strong inhibition on the excited-state proton transfer nonadiabatic geminate recombination in protic (vs deuteric) medium. Analysis of CatchER crystal structures and the MD simulations reveal the proton transfer mechanism in which the disrupted proton migration path in CatchER is rescued by calcium binding. Our finding provides important insights for a strategy to design calcium sensors and suggests that CatchER could be a useful probe for FLIM imaging of calcium in situ. PMID:24836743

  12. Speciation of Eu3+ bound to humic substances by time-resolved laser fluorescence spectroscopy (TRLFS) and parallel factor analysis (PARAFAC)

    NASA Astrophysics Data System (ADS)

    Lukman, Steven; Saito, Takumi; Aoyagi, Noboru; Kimura, Takaumi; Nagasaki, Shinya

    2012-07-01

    The bioavailability and toxicity of metal ions including radionuclides in the biosphere are greatly influenced by their speciation. Humic substances (HSs) are important constituents of various soil and water systems and have significant impact on the speciation and mobility of metal ions because of their high affinity to metal ions. In this study, the speciation of europium (Eu3+), a chemical homologue of trivalent actinides, with HSs collected from various origins was investigated by time-resolved laser fluorescence spectroscopy (TRLFS). The difficulties associated with the separation of the contribution of different Eu3+ species due to overlapping spectra or similar fluorescence lifetimes were addressed and mitigated by applying a multi-mode factor analysis, parallel factor analysis (PARAFAC), which resulted in the number, spectra, decay curves and relative fluorescence intensity profiles of different Eu3+ species. Subsequently, the interpretation of the Eu3+ species, was tackled by principal component analysis (PCA) and partial linear square (PLS) regression to deduce the nature of the Eu3+ species by taking into account the physicochemical properties of the HSs. Three factors corresponding to different Eu3+ species were obtained at 70 μM Eu3+ for all HSs investigated except for one humic acid. One of the factors corresponded to free Eu3+ ion interacting with HSs via diffusion. The remaining two factors were thought to be Eu3+ bound to HSs: one bound to acidic functional groups of HSs and the other to the sites of HSs influenced by the carbon backbone structures. It was also found that the latter factor exhibited strong energy transfer from the excited Eu3+ center to HSs. At lower Eu3+ concentration (10 μM), two factors having similar fluorescent characteristics to those of the second and third factors were obtained.

  13. Rotational and Translational Dynamics of Rhodamine 6G in a Pyrrolidinium Ionic Liquid: A Combined Time-Resolved Fluorescence Anisotropy Decay and NMR Study

    SciTech Connect

    Guo, Jianchang; Han, Kee Sung; Mahurin, Shannon Mark; Baker, Gary A; Hillesheim, Patrick C; Dai, Sheng; Hagaman, Edward {Ed} W; Shaw, Robert W

    2012-01-01

    NMR spectroscopy and time-resolved fluorescence anisotropy decay (TRFAD) are two of the most commonly used methods to study solute-solvent interactions. However, only a few studies have been reported to date using a combined NMR and TRFAD approach to systematically investigate the overall picture of diffusional and rotational dynamics of both the solute and solvent. In this paper, we combined NMR and TRFAD to probe fluorescent rhodamine dyes in a pyrrolidinium-based room temperature ionic liquid (RTIL), an emergent environmentally-friendly solvent type used in several energy-related applications. A specific interaction of the R6G cation and [Tf2N]- anion was identified, resulting in near-stick boundary condition rotation of R6G in this RTIL. The diffusional rates of the R6G solute and [C4mpyr][Tf2N] solvent derived from 1H NMR suggest the rates are proportional to their corresponding hydrodynamic radii. The 1H and 13C NMR studies of self-rotational dynamics of [C4mpyr][Tf2N] showed that the self-rotational correlation time of [C4mpyr]+ is 47 2 ps at 300 K. At the same temperature, we find that the correlation time for N-CH3 rotation in [C4mpyr]+ is 77 2 ps, comparable to overall molecular reorientation. This slow motion is attributed to properties of the cation structure.

  14. Complex formation of neptunium(V) with 4-hydroxy-3-methoxybenzoic acid studied by time-resolved laser-induced fluorescence spectroscopy with ultra-short laser pulses.

    PubMed

    Vulpius, D; Geipel, G; Baraniak, L; Bernhard, G

    2006-03-01

    The complex formation of neptunium(V) with 4-hydroxy-3-methoxybenzoic acid (vanillic acid) was studied by time-resolved laser-induced fluorescence spectroscopy with ultra-short laser pulses using the fluorescence properties of 4-hydroxy-3-methoxybenzoic acid. A 2:1 complex of neptunium(V) with 4-hydroxy-3-methoxybenzoic acid was found. The stability constant of this complex was determined to be logbeta(210) = 7.33 +/- 0.10 at an ionic strength of 0.1 mol/l (NaClO(4)) and at 21 degrees C. The determination of the stability constant required an investigation of the excited-state proton transfer of 4-hydroxy-3-methoxybenzoic acid over the whole pH range. It was realized that 4-hydroxy-3-methoxybenzoic acid undergoes excited-state reactions only at pH values below 5. At pH values above 5 stability constants can be determined without kinetic calculation of the proton transfer.

  15. Fulvic acid complexation of Eu(III) and Cm(III) at elevated temperatures studied by time-resolved laser fluorescence spectroscopy.

    PubMed

    Fröhlich, Daniel R; Skerencak-Frech, Andrej; Gast, Michael; Panak, Petra J

    2014-11-01

    The interaction of Eu(III) and Cm(III) with three different aquatic fulvic acids (FA) was studied as a function of the temperature (T = 20-80 °C) in 0.1 M NaCl solution by time-resolved laser fluorescence spectroscopy. The speciation of both trivalent metal ions was determined by peak deconvolution of the recorded fluorescence spectra. For each studied metal ion-FA system only one complexed species is formed under the given experimental conditions. The stability constants at 20, 40, 60 and 80 °C (log β'(T)) were determined according to the charge neutralization model. The log β' (20 °C) for the different FAs show similar values (log β(20 °C) = 5.60-6.29). The stability constants increase continuously with increasing temperature by approximately 0.3-1.0 orders of magnitude. The reaction enthalpies and entropies are derived from the integrated Van't Hoff equation. The results show that all investigated complexation reactions are endothermic and entropy-driven.

  16. A time-resolved fluorescence resonance energy transfer assay suitable for high-throughput screening for inhibitors of immunoglobulin E-receptor interactions.

    PubMed

    Kim, Beomkyu; Tarchevskaya, Svetlana S; Eggel, Alexander; Vogel, Monique; Jardetzky, Theodore S

    2012-12-15

    The interaction of immunoglobulin E (IgE) antibodies with the high-affinity receptor, FcεRI, plays a central role in initiating most allergic reactions. The IgE-receptor interaction has been targeted for treatment of allergic diseases, and many high-affinity macromolecular inhibitors have been identified. Small molecule inhibitors would offer significant advantages over current anti-IgE treatment, but no candidate compounds have been identified and fully validated. Here, we report the development of a time-resolved fluorescence resonance energy transfer (TR-FRET) assay for monitoring the IgE-receptor interaction. The TR-FRET assay measures an increase in fluorescence intensity as a donor lanthanide fluorophore is recruited into complexes of site-specific Alexa Fluor 488-labeled IgE-Fc and His-tagged FcεRIα proteins. The assay can readily monitor classic competitive inhibitors that bind either IgE-Fc or FcεRIα in equilibrium competition binding experiments. Furthermore, the TR-FRET assay can also be used to follow the kinetics of IgE-Fc-FcεRIα dissociation and identify inhibitory ligands that accelerate the dissociation of preformed complexes, as demonstrated for an engineered DARPin (designed ankyrin repeat protein) inhibitor. The TR-FRET assay is suitable for high-throughput screening (HTS), as shown by performing a pilot screen of the National Institutes of Health (NIH) Clinical Collection Library in a 384-well plate format.

  17. Novel Time-Resolved Fluorescence Europium Nanoparticle Immunoassay for Detection of Human Immunodeficiency Virus-1 Group O Viruses Using Microplate and Microchip Platforms.

    PubMed

    Haleyur Giri Setty, Mohan Kumar; Liu, Jikun; Mahtani, Prerna; Zhang, Panhe; Du, Bingchen; Ragupathy, Viswanath; Devadas, Krishnakumar; Hewlett, Indira K

    2016-06-01

    Accurate detection and quantification of HIV-1 group O viruses have been challenging for currently available HIV assays. We have developed a novel time-resolved fluorescence (TRF) europium nanoparticle immunoassay for HIV-1 group O detection using a conventional microplate enzyme-linked immunosorbent assay (ELISA) and a microchip platform. We screened several antibodies for optimal reactivity with several HIV-1 group O strains and identified antibodies that can detect all the strains of HIV-1 group O that were available for testing. The antibodies were used to develop a conventional ELISA format assay and an in-house developed europium nanoparticle-based assay for sensitivity. The method was evaluated on both microwell plate and microchip platforms. We identified two specific and sensitive antibodies among the six we screened. The antibodies, C65691 and ANT-152, were able to quantify 15 and detect all 17 group O viruses, respectively, as they were broadly cross-reactive with all HIV-1 group O strains and yielded better signals compared with other antibodies. We have developed a sensitive assay that reflects the actual viral load in group O samples by using an appropriate combination of p24 antibodies that enhance group O detection and a highly sensitive TRF-based europium nanoparticle for detection. The combination of ANT-152 and C65690M in the ratio 3:1 was able to give significantly higher signals in our europium-based assay compared with using any single antibody. PMID:26978478

  18. Development of a time-resolved fluorescence resonance energy transfer assay for cyclin-dependent kinase 4 and identification of its ATP-noncompetitive inhibitors.

    PubMed

    Lo, Mei-Chu; Ngo, Rachel; Dai, Kang; Li, Cong; Liang, Lingming; Lee, Josie; Emkey, Renee; Eksterowicz, John; Ventura, Manuel; Young, Stephen W; Xiao, Shou-Hua

    2012-02-15

    Protein kinases are recognized as important drug targets due to the pivotal roles they play in human disease. Many kinase inhibitors are ATP competitive, leading to potential problems with poor selectivity and significant loss of potency in vivo due to cellular ATP concentrations being much higher than K(m). Consequently, there has been growing interest in the development of ATP-noncompetitive inhibitors to overcome these problems. There are challenges to identifying ATP-noncompetitive inhibitors from compound library screens because ATP-noncompetitive inhibitors are often weaker and commonly excluded by potency-based hit selection criteria in favor of abundant and highly potent ATP-competitive inhibitors in screening libraries. Here we report the development of a time-resolved fluorescence resonance energy transfer (TR-FRET) assay for protein kinase cyclin-dependent kinase 4 (CDK4) and the identification of ATP-noncompetitive inhibitors by high-throughput screening after employing a strategy to favor this type of inhibitors. We also present kinetic characterization that is consistent with the proposed mode of inhibition.

  19. Design and validation of a homogeneous time-resolved fluorescence cell-based assay targeting the ligand-gated ion channel 5-HT3A.

    PubMed

    Blanc, Emilie; Wagner, Patrick; Plaisier, Fabrice; Schmitt, Martine; Durroux, Thierry; Bourguignon, Jean-Jacques; Partiseti, Michel; Dupuis, Elodie; Bihel, Frederic

    2015-09-01

    Ligand-gated ion channels (LGICs) are considered as attractive protein targets in the search for new therapeutic agents. Nowadays, this strategy involves the capability to screen large chemical libraries. We present a new Tag-lite ligand binding assay targeting LGICs on living cells. This technology combines the use of suicide enzyme tags fused to channels of interest with homogeneous time-resolved fluorescence (HTRF) as the detection readout. Using the 5-HT3 receptor as system model, we showed that the pharmacology of the HALO-5HT3 receptor was identical to that of the native receptor. After validation of the assay by using 5-HT3 agonists and antagonists of reference, a pilot screen enabled us to identify azelastine, a well-known histamine H1 antagonist, as a potent 5-HT3 antagonist. This interesting result was confirmed with electrophysiological experiments. The method described here is easy to implement and could be applicable for other LGICs, opening new ways for the screening of chemical libraries. PMID:25998104

  20. Novel Time-Resolved Fluorescence Europium Nanoparticle Immunoassay for Detection of Human Immunodeficiency Virus-1 Group O Viruses Using Microplate and Microchip Platforms.

    PubMed

    Haleyur Giri Setty, Mohan Kumar; Liu, Jikun; Mahtani, Prerna; Zhang, Panhe; Du, Bingchen; Ragupathy, Viswanath; Devadas, Krishnakumar; Hewlett, Indira K

    2016-06-01

    Accurate detection and quantification of HIV-1 group O viruses have been challenging for currently available HIV assays. We have developed a novel time-resolved fluorescence (TRF) europium nanoparticle immunoassay for HIV-1 group O detection using a conventional microplate enzyme-linked immunosorbent assay (ELISA) and a microchip platform. We screened several antibodies for optimal reactivity with several HIV-1 group O strains and identified antibodies that can detect all the strains of HIV-1 group O that were available for testing. The antibodies were used to develop a conventional ELISA format assay and an in-house developed europium nanoparticle-based assay for sensitivity. The method was evaluated on both microwell plate and microchip platforms. We identified two specific and sensitive antibodies among the six we screened. The antibodies, C65691 and ANT-152, were able to quantify 15 and detect all 17 group O viruses, respectively, as they were broadly cross-reactive with all HIV-1 group O strains and yielded better signals compared with other antibodies. We have developed a sensitive assay that reflects the actual viral load in group O samples by using an appropriate combination of p24 antibodies that enhance group O detection and a highly sensitive TRF-based europium nanoparticle for detection. The combination of ANT-152 and C65690M in the ratio 3:1 was able to give significantly higher signals in our europium-based assay compared with using any single antibody.

  1. Applications of immunomagnetic capture and time-resolved fluorescence to detect outbreak Escherichia coli O157 and Salmonella in alfalfa sprouts

    NASA Astrophysics Data System (ADS)

    Tu, Shu-I.; Gordon, Marsha; Fett, William F.; Gehring, Andrew G.; Irwin, Peter L.

    2004-03-01

    Commercially available alfalfa seeds were inoculated with low levels (~ 4 CFU/g) of pathogenic bacteria. The inoculated seeds were then allowed to sprout in sterile tap water at 22°C. After 48 hours, the irrigation water and sprouts were separately transferred to bovine heart infusion (BHI) media. The microbes in the BHI samples were allowed to grow for 4 hours at 37°C and 160 rpm. Specific immunomagnetic beads (IMB) were then applied to capture the E.coli O157 and/or Salmonella in the growth media. Separation and concentration of IMB-captured pathogens were achieved using magnetic separators. The captured E. coli O157:H7 and Salmonella spp were further tagged with europium (Eu) labeled anti-E. coli O157 antibodies and samarium (Sm) labeled anti-Salmonella antibodies, respectively. After washing, the lanthanide labels were extracted out from the complexes by specific chelators to form strongly fluorescent chelates. The specific time-resolved fluorescence (TRF) associated with Eu or Sm was measured to estimate the extent of capture of the E. coli O157 and Salmonella, respectively. The results indicated that the approach could detect E. coli O157 and Salmonella enterica from the seeds inoculated with ~ 4 CFU/g of the pathogens. Non-targeted bacteria, e.g., Aeromonas and Citrobacter exhibited essentially no cross reactivity. Since the pathogen detection from the sprouts was achieved within 6 hours, the developed methodology could be use as a rapid, sensitive and specific screening process for E. coli O157 and Salmonella enterica in this popular salad food.

  2. Sorption of Eu(III)/Cm(III) on Ca-montmorillonite and Na-illite. Part 1: Batch sorption and time-resolved laser fluorescence spectroscopy experiments

    NASA Astrophysics Data System (ADS)

    Rabung, Th.; Pierret, M. C.; Bauer, A.; Geckeis, H.; Bradbury, M. H.; Baeyens, B.

    2005-12-01

    Sorption of Cm(III) and Eu(III) at trace concentrations onto Ca-montmorillonite (SWy-1) and Na-illite (Illite du Puy) has been studied under anaerobic conditions by batch sorption experiments and time-resolved laser fluorescence spectroscopy (TRLFS). Comparison of the results from spectroscopic and batch sorption experiments with Cm and Eu indicates the existence of outer-sphere complexes at pH <4 in the experiments with Na-illite (0.25 g/L solid; 2.5 × 10 -7 mol/L Cm; 0.1 mol/L NaClO 4). In the case of Ca-montmorillonite, (0.25 g/L solid, 2.5 × 10 -7 mol/L Cm or 10 -6 mol/L Eu, 0.066 mol/L Ca(ClO 4) 2), Cm/Eu outer-sphere complexes do not form at significant levels due to the Ca 2+ competition for the clay mineral cation-exchange sites. TRLFS spectra indicate the formation of inner-sphere surface complexes at pH >5 for both clay minerals. Five H 2O/OH - molecules remain in the first metal ion coordination sphere of the sorbed Eu/Cm. Measured fluorescence lifetimes of sorbed Eu/Cm and peak deconvolution of Cm-spectra are consistent with the formation of surface complexes of the form ≡S-O-Eu/Cm(OH) x(2-x)(H 2O) 5-x. At pH ≥ 12 Cm becomes incorporated into a surface precipitate at the Ca-montmorillonite surface presumably composed of Ca(OH) 2 or calcium silicate hydrate. A dramatic shift of the fluorescence emission band by more than 20 nm and a clear increase in the fluorescence lifetime suggests the almost complete displacement of coordinated H 2O and OH -. The pH dependent Eu sorption data obtained in batch experiments are consistent with spectroscopic data on Eu and Cm within experimental uncertainties thus demonstrating the validity of Eu as a homologue for trivalent actinides. Parameterization of a two-site protolysis nonelectrostatic surface complexation and cation exchange model using the batch sorption data and spectroscopic results is discussed in Part 2 of this work.

  3. Ultrafast Time-Resolved Emission and Absorption Spectra of meso-Pyridyl Porphyrins upon Soret Band Excitation Studied by Fluorescence Up-Conversion and Transient Absorption Spectroscopy.

    PubMed

    Venkatesh, Yeduru; Venkatesan, M; Ramakrishna, B; Bangal, Prakriti Ranjan

    2016-09-01

    A comprehensive study of ultrafast molecular relaxation processes of isomeric meso-(pyridyl) porphyrins (TpyPs) has been carried out by using femtosecond time-resolved emission and absorption spectroscopic techniques upon pumping at 400 nm, Soret band (B band or S2), in 4:1 dichloromethane (DCM) and tetrahydrofuran (THF) solvent mixture. By combined studies of fluorescence up-conversion, time-correlated single photon counting, and transient absorption spectroscopic techniques, a complete model with different microscopic rate constants associated with elementary processes involved in electronic manifolds has been reported. Besides, a distinct coherent nuclear wave packet motion in Qy state is observed at low-frequency mode, ca. 26 cm(-1) region. Fluorescence up-conversion studies constitute ultrafast time-resolved emission spectra (TRES) over the whole emission range (430-710 nm) starting from S2 state to Qx state via Qy state. Careful analysis of time profiles of up-converted signals at different emission wavelengths helps to reveal detail molecular dynamics. The observed lifetimes are as indicated: A very fast decay component with 80 ± 20 fs observed at ∼435 nm is assigned to the lifetime of S2 (B) state, whereas being a rise component in the region of between 550 and 710 nm emission wavelength pertaining to Qy and Qx states, it is attributed to very fast internal conversion (IC) occurring from B → Qy and B → Qx as well. Two distinct components of Qy emission decay with ∼200-300 fs and ∼1-1.5 ps time constants are due to intramolecular vibrational redistribution (IVR) induced by solute-solvent inelastic collisions and vibrational redistribution induced by solute-solvent elastic collision, respectively. The weighted average of these two decay components is assigned as the characteristic lifetime of Qy, and it ranges between 0.3 and 0.5 ps. An additional ∼20 ± 2 ps rise component is observed in Qx emission, and it is assigned to the formation time of

  4. Ultrafast Time-Resolved Emission and Absorption Spectra of meso-Pyridyl Porphyrins upon Soret Band Excitation Studied by Fluorescence Up-Conversion and Transient Absorption Spectroscopy.

    PubMed

    Venkatesh, Yeduru; Venkatesan, M; Ramakrishna, B; Bangal, Prakriti Ranjan

    2016-09-01

    A comprehensive study of ultrafast molecular relaxation processes of isomeric meso-(pyridyl) porphyrins (TpyPs) has been carried out by using femtosecond time-resolved emission and absorption spectroscopic techniques upon pumping at 400 nm, Soret band (B band or S2), in 4:1 dichloromethane (DCM) and tetrahydrofuran (THF) solvent mixture. By combined studies of fluorescence up-conversion, time-correlated single photon counting, and transient absorption spectroscopic techniques, a complete model with different microscopic rate constants associated with elementary processes involved in electronic manifolds has been reported. Besides, a distinct coherent nuclear wave packet motion in Qy state is observed at low-frequency mode, ca. 26 cm(-1) region. Fluorescence up-conversion studies constitute ultrafast time-resolved emission spectra (TRES) over the whole emission range (430-710 nm) starting from S2 state to Qx state via Qy state. Careful analysis of time profiles of up-converted signals at different emission wavelengths helps to reveal detail molecular dynamics. The observed lifetimes are as indicated: A very fast decay component with 80 ± 20 fs observed at ∼435 nm is assigned to the lifetime of S2 (B) state, whereas being a rise component in the region of between 550 and 710 nm emission wavelength pertaining to Qy and Qx states, it is attributed to very fast internal conversion (IC) occurring from B → Qy and B → Qx as well. Two distinct components of Qy emission decay with ∼200-300 fs and ∼1-1.5 ps time constants are due to intramolecular vibrational redistribution (IVR) induced by solute-solvent inelastic collisions and vibrational redistribution induced by solute-solvent elastic collision, respectively. The weighted average of these two decay components is assigned as the characteristic lifetime of Qy, and it ranges between 0.3 and 0.5 ps. An additional ∼20 ± 2 ps rise component is observed in Qx emission, and it is assigned to the formation time of

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

    PubMed

    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.

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

    PubMed Central

    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

  7. Nuclear magnetic resonance, fluorescence correlation spectroscopy and time-resolved fluorescence anisotropy studies of intermolecular interactions in bis(1-methyl-1H-imidazol-3-ium-3-yl)dihydroborate bis(trifluoromethylsulfonyl)amide and its mixtures with various cosolvents

    NASA Astrophysics Data System (ADS)

    Sahu, Prabhat Kumar; Nanda, Raju; Seth, Sudipta; Ghosh, Arindam; Sarkar, Moloy

    2016-09-01

    Keeping in mind the potential usefulness of mixed ionic liquid (IL)-cosolvents systems in several industrial applications, intermolecular interactions between a borate-based IL, bis(1-methyl-1H-imidazol-3-ium-3-yl)dihydroborate bis(trifluoromethylsulfonyl)amide ([BIMIMDBA][TF2N]), and its binary mixtures with several molecular solvents has been investigated through NMR and fluorescence spectroscopy. Analysis of the 1H chemical shifts (δ/ppm) and translational diffusion coefficients (D) of the IL in different solvent mixtures demonstrate interplay of nonspecific (ion-dipole) and specific (hydrogen bonding) interactions in governing the properties of these mixtures. Fluorescence correlation spectroscopy (FCS) and time-resolved fluorescence anisotropy data provide evidence in favour of different IL-solvent interaction for different IL-cosolvent systems.

  8. Insight into the factors influencing the backbone dynamics of three homologous proteins, dendrotoxins I and K, and BPTI: FTIR and time-resolved fluorescence investigations.

    PubMed

    Hollecker, Michelle; Vincent, Michel; Gallay, Jacques; Ruysschaert, Jean-Marie; Goormaghtigh, Erik

    2002-12-24

    Attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, combined with hydrogen/deuterium exchange technique and time-resolved fluorescence spectroscopy, has been used to investigate the changes in structure and dynamics that underlie the thermodynamic stability differences observed for three closely homologous proteins: dendrotoxins I and K, and bovine pancreatic trypsin inhibitor (BPTI). The experiments were performed on proteins under their native state and a modified form, obtained by selective reduction of a disulfide bond at the surface of the molecule, increasing slightly the backbone flexibility without changing the average structure. The data confirmed the high local as well as global rigidity of BPTI. In protein K, the exchange process was slow during the first 2 h of exchange, presumably reflecting a compact three-dimensional conformation, and then increased rapidly, the internal amide protons of the beta-strands exchanging 10-fold faster than in BPTI or protein I. The most probable destabilizing element was identified as Pro32, in the core of the beta-sheet. Protein I was found to present a 10% more expanded volume than protein K or BPTI, and there is a possible correlation between the resulting increased flexibility of the molecule and the lower thermodynamic stability observed for this protein. Interestingly, the interior amide protons of the beta-sheet structure were found to be as protected against exchange in protein I as in BPTI, suggesting that, although globally more flexible than that of Toxin K or BPTI, the structure of Toxin I could be locally quite rigid. The structural factors suspected to be responsible for the differences in internal flexibility of the two toxins could play a significant role in determining their functional properties. PMID:12484765

  9. Time-Resolved Fluorescence Energy Transfer Measurements Between Site-Specific Probes On The Ca-Atpase Of Sarcoplasmic Reticulum In Different Enzymatic States

    NASA Astrophysics Data System (ADS)

    Squier, Thomas C.; Bigelow, Diana J.; Garcia de Ancos, Jorge; Bishop, James E.; Inesi, Giuseppe

    1988-06-01

    We have measured resonance energy transfer between two donor-acceptor pairs localized on different domains of the Ca-ATPase of sarcoplasmic reticulum in order to determine whether changes in tertiary structure accompany active calcium transport. Energy transfer was determined from both steady state intensities and time-resolved lifetimes of 5-(2-((acety1)- amino)ethyDaminonaphthalene-l-sulfonic acid (IAEDANS), specifically bound to the B tryptic fragment, using two acceptors: (1) fluorescein 5'-isothiocyanate (FITC), covalently bound at the nucleotide site, also on the B fragment, and (2) 4-dimethylaminophenylazopheny1-4'- maleimide (DABMI), bound on the Al subfragment. Neither binding of calcium to the high affinity sites nor phosphorylation by inorganic phosphate is accompanied by detectable changes in the distance between IAEDANS and FITC, suggesting that the B fragment does not undergo any large-scale (>1 A) physical distortion under these conditions. On the other hand, measurements of energy transfer from IAEDANS to the acceptor DABMI, on the Al subfragment, demonstrate that phosphorylation with inorganic phosphate or addition of pM VO4 results in increased energy transfer, that is reversible with subsequent addition of calcium. Addition of calcium to the nonphosphorylated enzyme results in no detectable change in energy transfer. The presence of the detergent lysolecithin prevents the phosphate dependent increase in fluorescence energy transfer, suggesting that protein-protein interactions may contribute to the observed change in energy transfer. A direct relationship between an increased degree of protein-protein interactions and phosphoenzyme formation is confirmed by investigations using a reconstituted preparation containing a mixed popu Lation of Ca-ATPase polypeptide chains labeled either with IAEDANS or with DABMI. These results suggest a phosphorylation dependent change in either the affinity or orientation of Ca-ATPase polypeptide chains with respect

  10. Development and implementation of a miniaturized high-throughput time-resolved fluorescence energy transfer assay to identify small molecule inhibitors of polo-like kinase 1.

    PubMed

    Sharlow, Elizabeth R; Leimgruber, Stephanie; Shun, Tong Ying; Lazo, John S

    2007-12-01

    Polo-like kinase (Plk) 1 is a key enzyme involved in regulating the mammalian cell cycle that is also a validated anticancer drug target. Nonetheless, there are relatively few readily available potent and selective small molecule inhibitors of Plk1. To increase the availability of pharmacologically valuable Plk1 inhibitors, we describe herein the development, variability assessment, validation, and implementation of a 384-well automated, miniaturized high-throughput time-resolved fluorescence energy transfer screening assay designed to identify Plk1 kinase inhibitors. Using a small molecule library of pharmaceutically active compounds to gauge high-throughput assay robustness and reproducibility, we found nine general kinase inhibitors, including H-89, which was selected as the minimum control. We then interrogated a 97,101 compound library from the National Institutes of Health repository for small molecule inhibitors of Plk1 kinase activity. The initial primary hit rate in a single 10 microM concentration format was 0.21%. Hit compounds were subjected to concentration-response confirmation and interference assays. Identified in the screen were seven compounds with 50% inhibitory concentration (IC50) values below 1 microM, 20 compounds with IC50 values between 1 microM and 5 microM, and eight compounds with IC50 values between 5 and 10 microM, which could be assigned to seven distinct chemotype classes. Hit compounds were also examined for their ability to inhibit other kinases such as protein kinase D, focal adhesion kinase, rho-associated coiled coil protein kinase 2, c-jun NH2-terminal kinase 3, and protein kinase A via experimentation or data-mining. These compounds should be useful as probes for the biological activity of Plk1 and as leads for the development of new selective inhibitors of Plk1. PMID:18181689

  11. Spatial proximity of the HIV-1 nucleocapsid protein zinc fingers investigated by time-resolved fluorescence and fluorescence resonance energy transfer.

    PubMed

    Mély, Y; Jullian, N; Morellet, N; De Rocquigny, H; Dong, C Z; Piémont, E; Roques, B P; Gérard, D

    1994-10-11

    The three-dimensional structure of peptides encompassing the two zinc-saturated finger motifs of the nucleocapsid protein NCp7 of HIV-1 has been reported by several groups. Whereas the folded structures of the finger motifs were in good agreement, discrepancies existed concerning their spatial relationship since the fingers were found either close to each other [Morellet, N., Jullian, N., De Rocquigny, H., Maigret, B., Darlix, J. L., & Roques, B. P. (1992) Embo J. 11, 3059-3065] or independently folded [Omichinski, J. G., Clore, G. M., Sakaguchi, K., Appella, E., & Gronenborn, A. M. (1991) FEBS Lett. 292, 25-30, Summers, M. F., Henderson, L. E., Chance, M. R., Bess, J. W., Jr., South, T. L., Blake, P. R., Sagi, I., Perez-Alvarado, G., Sowder, R.C., III, Hare, D.R., & Arthur, L. O. (1992) Protein Sci. 1, 563-574]. As in the interacting finger model, Phe16 in the NH2-terminal finger and Trp37 in the COOH-terminal finger were found to be spatially close, the fluorescence properties of the aromatic residues at positions 16 and 37 in the wild-type and two conservatively substituted (12-53) NCp7 peptides were investigated and compared with those of three negative control derivatives where the finger motifs were not in close contact. Direct distance measurements by Tyr-Trp fluorescence resonance energy transfer of the former derivatives yielded a 7-12 A interchromophore distance range which is clearly inconsistent with the 12.5-18 A range measured for the negative controls and thus a random orientation of the zinc finger motifs.(ABSTRACT TRUNCATED AT 250 WORDS)

  12. Spatial proximity of the HIV-1 nucleocapsid protein zinc fingers investigated by time-resolved fluorescence and fluorescence resonance energy transfer.

    PubMed

    Mély, Y; Jullian, N; Morellet, N; De Rocquigny, H; Dong, C Z; Piémont, E; Roques, B P; Gérard, D

    1994-10-11

    The three-dimensional structure of peptides encompassing the two zinc-saturated finger motifs of the nucleocapsid protein NCp7 of HIV-1 has been reported by several groups. Whereas the folded structures of the finger motifs were in good agreement, discrepancies existed concerning their spatial relationship since the fingers were found either close to each other [Morellet, N., Jullian, N., De Rocquigny, H., Maigret, B., Darlix, J. L., & Roques, B. P. (1992) Embo J. 11, 3059-3065] or independently folded [Omichinski, J. G., Clore, G. M., Sakaguchi, K., Appella, E., & Gronenborn, A. M. (1991) FEBS Lett. 292, 25-30, Summers, M. F., Henderson, L. E., Chance, M. R., Bess, J. W., Jr., South, T. L., Blake, P. R., Sagi, I., Perez-Alvarado, G., Sowder, R.C., III, Hare, D.R., & Arthur, L. O. (1992) Protein Sci. 1, 563-574]. As in the interacting finger model, Phe16 in the NH2-terminal finger and Trp37 in the COOH-terminal finger were found to be spatially close, the fluorescence properties of the aromatic residues at positions 16 and 37 in the wild-type and two conservatively substituted (12-53) NCp7 peptides were investigated and compared with those of three negative control derivatives where the finger motifs were not in close contact. Direct distance measurements by Tyr-Trp fluorescence resonance energy transfer of the former derivatives yielded a 7-12 A interchromophore distance range which is clearly inconsistent with the 12.5-18 A range measured for the negative controls and thus a random orientation of the zinc finger motifs.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7918429

  13. Time-resolved spectral studies of blue-green fluorescence of artichoke (Cynara cardunculus L. Var. Scolymus) leaves: identification of chlorogenic acid as one of the major fluorophores and age-mediated changes.

    PubMed

    Morales, Fermín; Cartelat, Aurélie; Alvarez-Fernández, Ana; Moya, Ismael; Cerovic, Zoran G

    2005-12-14

    Synchrotron radiation and the time-correlated single-photon counting technique were used to investigate the spectral and time-resolved characteristics of blue-green fluorescence (BGF) of artichoke leaves. Leaves emitted BGF under ultraviolet (UV) excitation; the abaxial side was much more fluorescent than the adaxial side, and in both cases, the youngest leaves were much more fluorescent than the oldest ones. The BGF of artichoke leaves was dominated by the presence of hydroxycinnamic acids. A decrease in the percentage of BGF attributable to the very short kinetic component (from 42 to 20%), in the shape of the BGF excitation spectra, and chlorogenic acid concentrations indicate that there is a loss of hydroxycinnamic acid with leaf age. Studies on excitation, emission, and synchronized fluorescence spectra of leaves and trichomes and chlorogenic acid contents indicate that chlorogenic acid is one of the main blue-green fluorophores in artichoke leaves. Results of the present study indicate that 20-42% (i.e., the very short kinetic component) of the overall BGF is emitted by chlorogenic acid. Time-resolved BGF measurements could be a means to extract information on chlorogenic acid fluorescence from the overall leaf BGF.

  14. Investigation of temporal profiles at the symmetrical points of the target in tissue phantoms by time-resolved fluorescence diffuse optical tomography

    NASA Astrophysics Data System (ADS)

    Locharoenrat, Kitsakorn

    2013-06-01

    We have performed a target size dependence of fluorescence temporal profile from an Indocyanine Green (ICG) target filled in a phantom by Fluorescence Diffuse Optical Tomography (FDOT) method. From the results of experiment and statistical analysis, we have found that the geometry effect of target size was one of the key parameters in temporal profile determining the resolution of the optical reconstruction images.

  15. Complexity of Lipid Domains and Rafts in Giant Unilamellar Vesicles Revealed by Combining Imaging and Microscopic and Macroscopic Time-Resolved Fluorescence

    PubMed Central

    de Almeida, Rodrigo F. M.; Borst, JanWillem; Fedorov, Alexander; Prieto, Manuel; Visser, Antonie J. W. G.

    2007-01-01

    The application of fluorescence lifetime imaging microscopy to study gel/fluid and raftlike lipid domains in giant unilamellar vesicles (GUVs) is demonstrated here. Different regions of the ternary dipalmitoylphosphatidylcholine/dioleoylphosphatidylcholine/cholesterol phase diagram were studied. The head-labeled phospholipid Rhodamine-dioleoylphosphatidylethanolamine (Rhod-DOPE) was used as a fluorescent probe. Gel/fluid and liquid-ordered (lo)/liquid-disordered (ld) phase separation were clearly visualized upon two-photon excitation. Fluorescence intensity decays in different regions of a GUV were also obtained with the microscope in fixed laser-beam configuration. The ensemble behavior of the system was studied by obtaining fluorescence intensity decays of Rhod-DOPE in nongiant vesicle suspensions. The fingerprints for gel/fluid coexistence and for the presence of lo raftlike phase, based on fluorescence lifetime imaging microscopy histograms and images, and on the fluorescence intensity decay parameters of Rhod-DOPE, are presented. The presence of three lipid phases in one single GUV is detected unequivocally. From the comparison of lifetime parameters, it can be concluded that the lo phase is formed in the binary dipalmitoylphosphatidylcholine/cholesterol but not in the dioleoylphosphatidylcholine/cholesterol mixture. The domains apparent in fluorescence intensity images have a more complex substructure revealed by analysis of the lifetime data. The potential applications of this combined imaging/microscopic/macroscopic methodology are discussed. PMID:17449668

  16. A Global Instrumentation Approach To Resolution Of Ground-State And Excited-State Processes In Time-Resolved Fluorescence Spectroscopy

    NASA Astrophysics Data System (ADS)

    Ross, J. B. Alexander; Laws, William R.; Wyssbrod, Herman R.

    1988-06-01

    The application of global analysis to time-dependent fluorescence data provides a power-ful technique for investigating complex, heterogeneous systems by defining common parameters to elements of a data set where the data has been obtained as a function of an independent variable. Another, related approach is to incorporate information obtained from other physical techniques into the analysis of time-dependent fluorescence data. The information from the independent method is used to define a linkage among the parameters within one element of the time-dependent fluorescence data set; the linkage establishes certain parameters as dependent variables. We call this a linked-function analysis, which can be used in the analysis of a single data set element or globally in the analysis of multiple data set elemets. This approach has been applied to time-domain fluorescence spectroscopy combi n ned with 1H-NMR spectroscopy to resolve ground-state and excited-state processes in the fluorescence decay of aromatic amino residues in peptides.

  17. Multiemission wavelength picosecond time-resolved fluorescence decay data obtained on the millisecond time scale: application to protein:DNA interactions and protein-folding reactions

    NASA Astrophysics Data System (ADS)

    Beechem, Joseph M.

    1992-04-01

    One of the major aspects of fluorescence spectroscopy which differentiates this technique from many other spectroscopic approaches is the inherent multidimensional nature of the data. For instance, the basic pulsed-laser fluorescence data set is characterized by fluorescence versus: emission wavelength, polarization state (parallel and perpendicular intensities), time of emission (picoseconds to nanoseconds), and time of biological reaction (milliseconds to minutes). Usually, this six-dimensional data set is obtained piecemeal, single dimension at a time; often complete data sets are not even collected. This is especially true of the biological time scale axis. Data acquisition times for picosecond decay data are typically seconds to minutes, and, therefore, it has not been generally possible to perform this experiment in a kinetic mode. What is described in this report is the construction of a parallel multichannel time-correlated single-photon counting (TCSPC) fluorometer which is capable of simultaneous collection of: fluorescence vs. picosecond to nanosecond time vs. emission wavelength vs. polarization state vs. millisecond to second time. Use is made of two multi-anode microchannel plate detectors, each obtaining data at two different polarization states, six different emission wavelengths, along 12 independent TCSPC channels. This instrument is interfaced to a three-syringe stepper motor controlled stop-flow apparatus, and picosecond decay data along all of these channels is stored and collected by two 33 MHz 80486 computers at rates approaching 1200 - 12000 data sets per second.

  18. Revealing the radiative and non-radiative relaxation rates of the fluorescent dye Atto488 in a λ/2 Fabry-Pérot-resonator by spectral and time resolved measurements.

    PubMed

    Konrad, Alexander; Metzger, Michael; Kern, Andreas M; Brecht, Marc; Meixner, Alfred J

    2016-08-14

    Using a Fabry-Pérot-microresonator with controllable cavity lengths in the λ/2-regime, we show the controlled modification of the vibronic relaxation dynamics of a fluorescent dye molecule in the spectral and time domain. By altering the photonic mode density around the fluorophores we are able to shape the fluorescence spectrum and enhance specifically the probability of the radiative transitions from the electronic excited state to distinct vibronic excited states of the electronic ground state. Analysis and correlation of the spectral and time resolved measurements by a theoretical model and a global fitting procedure allows us to reveal quantitatively the spectrally distributed radiative and non-radiative relaxation dynamics of the respective dye molecule under ambient conditions at the ensemble level.

  19. Revealing the radiative and non-radiative relaxation rates of the fluorescent dye Atto488 in a λ/2 Fabry-Pérot-resonator by spectral and time resolved measurements

    NASA Astrophysics Data System (ADS)

    Konrad, Alexander; Metzger, Michael; Kern, Andreas M.; Brecht, Marc; Meixner, Alfred J.

    2016-07-01

    Using a Fabry-Pérot-microresonator with controllable cavity lengths in the λ/2-regime, we show the controlled modification of the vibronic relaxation dynamics of a fluorescent dye molecule in the spectral and time domain. By altering the photonic mode density around the fluorophores we are able to shape the fluorescence spectrum and enhance specifically the probability of the radiative transitions from the electronic excited state to distinct vibronic excited states of the electronic ground state. Analysis and correlation of the spectral and time resolved measurements by a theoretical model and a global fitting procedure allows us to reveal quantitatively the spectrally distributed radiative and non-radiative relaxation dynamics of the respective dye molecule under ambient conditions at the ensemble level.Using a Fabry-Pérot-microresonator with controllable cavity lengths in the λ/2-regime, we show the controlled modification of the vibronic relaxation dynamics of a fluorescent dye molecule in the spectral and time domain. By altering the photonic mode density around the fluorophores we are able to shape the fluorescence spectrum and enhance specifically the probability of the radiative transitions from the electronic excited state to distinct vibronic excited states of the electronic ground state. Analysis and correlation of the spectral and time resolved measurements by a theoretical model and a global fitting procedure allows us to reveal quantitatively the spectrally distributed radiative and non-radiative relaxation dynamics of the respective dye molecule under ambient conditions at the ensemble level. Electronic supplementary information (ESI) available. See DOI: 10.1039/C6NR02380K

  20. Oligomerization of epidermal growth factor receptors on A431 cells studied by time-resolved fluorescence imaging microscopy. A stereochemical model for tyrosine kinase receptor activation

    PubMed Central

    1995-01-01

    The aggregation states of the epidermal growth factor receptor (EGFR) on single A431 human epidermoid carcinoma cells were assessed with two new techniques for determining fluorescence resonance energy transfer: donor photobleaching fluorescence resonance energy transfer (pbFRET) microscopy and fluorescence lifetime imaging microscopy (FLIM). Fluorescein-(donor) and rhodamine-(acceptor) labeled EGF were bound to the cells and the extent of oligomerization was monitored by the spatially resolved FRET efficiency as a function of the donor/acceptor ratio and treatment conditions. An average FRET efficiency of 5% was determined after a low temperature (4 degrees C) incubation with the fluorescent EGF analogs for 40 min. A subsequent elevation of the temperature for 5 min caused a substantial increase of the average FRET efficiency to 14% at 20 degrees C and 31% at 37 degrees C. In the context of a two-state (monomer/dimer) model for the EGFR, these FRET efficiencies were consistent with minimal average receptor dimerizations of 13, 36, and 69% at 4, 20, and 37 degrees C, respectively. A431 cells were pretreated with the monoclonal antibody mAb 2E9 that specifically blocks EGF binding to the predominant population of low affinity EGFR (15). The average FRET efficiency increased dramatically to 28% at 4 degrees C, indicative of a minimal receptor dimerization of 65% for the subpopulation of high affinity receptors. These results are in accordance with prior studies indicating that binding of EGF leads to a fast and temperature- dependent microclustering of EGFR, but suggest in addition that the high affinity functional subclass of receptors on quiescent A431 cells are present in a predimerized or oligomerized state. We propose that the transmission of the external ligand-binding signal to the cytoplasmic domain is effected by a concerted relative rotational rearrangement of the monomeric units comprising the dimeric receptor, thereby potentiating a mutual activation of

  1. Time-resolved fluorescence spectroscopy investigation of the effect of 4-hydroxynonenal on endogenous NAD(P)H in living cardiac myocytes

    NASA Astrophysics Data System (ADS)

    Chorvatova, Alzbeta; Aneba, Swida; Mateasik, Anton; Chorvat, Dusan; Comte, Blandine

    2013-06-01

    Lipid peroxidation is a major biochemical consequence of the oxidative deterioration of polyunsaturated lipids in cell membranes and causes damage to membrane integrity and loss of protein function. 4-hydroxy-2-nonenal (HNE), one of the most reactive products of n-6 polyunsaturated fatty acid peroxidation of membrane phospholipids, has been shown to be capable of affecting both nicotinamide adenine dinucleotide (phosphate) reduced [NAD(P)H] as well as NADH production. However, the understanding of its effects in living cardiac cells is still lacking. Our goal was to therefore investigate HNE effects on NAD(P)H noninvasively in living cardiomyocytes. Spectrally resolved lifetime detection of endogenous fluorescence, an innovative noninvasive technique, was employed. Individual fluorescence components were resolved by spectral linear unmixing approach. Gathered results revealed that HNE reduced the amplitude of both resolved NAD(P)H components in a concentration-dependent manner. In addition, HNE increased flavoprotein fluorescence and responsiveness of the NAD(P)H component ratio to glutathione reductase (GR) inhibitor. HNE also increased the percentage of oxidized nucleotides and decreased maximal NADH production. Presented data indicate that HNE provoked an important cell oxidation by acting on NAD(P)H regulating systems in cardiomyocytes. Understanding the precise role of oxidative processes and their products in living cells is crucial for finding new noninvasive tools for biomedical diagnostics of pathophysiological states.

  2. Reorientational motion of a cross-link junction in a poly(dimethylsiloxane) network measured by time-resolved fluorescence depolarization

    SciTech Connect

    Stein, A.D. ); Hoffman, D.A. ); Frank, C.W. ); Fayer, M.D. )

    1992-02-15

    The reorientational dynamics of a cross-link junction in poly(dimethylsiloxane) networks, measured by the fluorescence anisotropy decay of a chromophore tagged to the cross-link, have been investigated over a range of temperatures from {ital T}{sub {ital g}}+75 to {ital T}{sub {ital g}}+150. The probe chromophore, 1-dimethylamino-5-sulfonylnaphthalene amide (dansyl amide), is pendant to a trifunctional silane that acts as a cross-linking molecule. In cyclohexanol, the fluorescence anisotropy decay is in agreement with Debye--Stokes--Einstein hydrodynamic theory (rotational diffusion) demonstrating that the cross-linker can be used as a probe of orientational relaxation. The fluorescence anisotropy decays at a rapid rate in an end-linked poly(dimethyl siloxane) network reflecting fast reorientational motion of the cross-link junction. This reorientation appears diffusive and has a temperature dependence in accord with the Williams--Landel--Ferry equation. A model is proposed that suggests that reorientation and translational motion of the cross-link occur simultaneously and are both coupled to fluctuations of the polymer chain ends.

  3. Time-Resolved Fluorescence Anisotropy of Bicyclo[1.1.1]pentane/Tolane-Based Molecular Rods Included in Tris(o-phenylenedioxy)cyclotriphosphazene (TPP)

    PubMed Central

    2015-01-01

    We examine the fluorescence anisotropy of rod-shaped guests held inside the channels of tris(o-phenylenedioxy)cyclotriphosphazene (TPP) host nanocrystals, characterized by powder X-ray diffraction and solid state NMR spectroscopy. We address two issues: (i) are light polarization measurements on an aqueous colloidal solution of TPP nanocrystals meaningful, or is depolarization by scattering excessive? (ii) Can measurements of the rotational mobility of the included guests be performed at low enough loading levels to suppress depolarization by intercrystallite energy transfer? We find that meaningful measurements are possible and demonstrate that the long axis of molecular rods included in TPP channels performs negligible vibrational motion. PMID:25937858

  4. A study of the time-resolved fluorescence spectrum and red edge effect of ANF in a room-temperature ionic liquid.

    PubMed

    Hu, Zhonghan; Margulis, Claudio J

    2006-06-15

    In a recent article, we have analyzed using molecular dynamics simulations the steady-state red edge effect (REE) observed by Samanta and co-workers when the fluorescent probe 2-amino-7-nitrofluorene (ANF) is photoexcited at different wavelengths in 1-butyl-3-methylimidazolium ([BMIM+]) hexafluorophosphate ([PF6-]). In this letter, we predict the time- and wavelength-dependent emission spectra of ANF in the same ionic solvent. From the analysis of our simulated data, we are able to derive an approximate time scale for reorganization of the solvent around the solute probe. The effect that slow varying local liquid environments have on the overall time-dependent signal is also discussed.

  5. Intramolecular charge transfer of 4-(dimethylamino)benzonitrile probed by time-resolved fluorescence and transient absorption: No evidence for two ICT states and a pisigma( *) reaction intermediate.

    PubMed

    Zachariasse, Klaas A; Druzhinin, Sergey I; Kovalenko, Sergey A; Senyushkina, Tamara

    2009-12-14

    For the double exponential fluorescence decays of the locally excited (LE) and intramolecular charge transfer (ICT) states of 4-(dimethylamino)benzonitrile (DMABN) in acetonitrile (MeCN) the same times tau(1) and tau(2) are observed. This means that the reversible LE<==>ICT reaction, starting from the initially excited LE state, can be adequately described by a two state mechanism. The most important factor responsible for the sometimes experimentally observed differences in the nanosecond decay time, with tau(1)(LE)fluorescence response functions with a time resolution of 0.5 ps/channel in 1200 channels reliable kinetic and thermodynamic data can be obtained. The arguments presented in the literature in favor of a pisigma( *) state with a bent CN group as an intermediate in the ICT reaction of DMABN are discussed. From the appearance of an excited state absorption (ESA) band in the spectral region between 700 and 800 nm in MeCN for N,N-dimethylanilines with CN, Br, F, CF(3), and C(=O)OC(2)H(2) p-substituents, it is concluded that this ESA band cannot be attributed to a pisigma( *) state, as only the C-C[Triple Bond]N group can undergo the required 120 degrees bending. PMID:20001042

  6. Dynamic Solvent Control of a Reaction in Ionic Deep Eutectic Solvents: Time-Resolved Fluorescence Measurements of Reactive and Nonreactive Dynamics in (Choline Chloride + Urea) Melts.

    PubMed

    Das, Anuradha; Biswas, Ranjit

    2015-08-01

    Dynamic fluorescence anisotropy and Stokes shift measurements of [f choline chloride + (1 - f) urea)] deep eutectic solvents at f = 0.33 and 0.40 have been carried out using a dipolar solute, coumarin 153 (C153), in the temperature range 298 ≤ T ≤ 333 K. Subsequently, measured time-dependent solvent response is utilized to investigate the dynamic solvent control on the measured rates of photoexcited intramolecular charge transfer (ICT) reactions of two molecules, 4-(1-azetidinyl)benzonitrile (P4C) and 4-(1-pyrrolidinyl)benzonitrile (P5C), occurring in these media. Measured average reaction time scales (⟨τ(rxn)⟩) exhibit the following dependence on average solvation times scales (⟨τ(s)⟩): ⟨τ(rxn)⟩ ∝ ⟨τ(s)⟩(α) with α = 0.5 and 0.35 for P4C and P5C, respectively. Such a strong dynamic solvent control of ⟨τ(rxn)⟩, particularly for P4C, is different from earlier observations with these ICT molecules in conventional molecular solvents. Excitation wavelength-dependent fluorescence emissions of C153 and trans-2-[4-(dimethylamino)styryl]-benzothiazole (DMASBT), which differ widely in average fluorescence lifetimes (⟨τ(life)⟩), suggest the presence of substantial spatial heterogeneity in these systems. Dynamic heterogeneity is reflected via the following fractional viscosity (η) dependences of ⟨τ(s)⟩ and ⟨τ(r)⟩ (⟨τ(r)⟩ being solute's average rotation time): ⟨τx⟩ ∝ (η/T)(p) with 0.7 ≤ p ≤ 0.9. Different correlations between ⟨τ(s)⟩ and ⟨τ(r)⟩ emerge at different temperature regimes, indicating variable frictional coupling at low and high temperatures. Estimated dynamic Stokes shifts in these media vary between ∼1200 and ∼1600 cm(-1), more than 50% of which possess a time scale much faster than the temporal resolution (∼75 ps) employed in these measurements. Estimated activation energy for η is closer to that for ⟨τ(r)⟩ than that for ⟨τ(s)⟩, suggesting ⟨τ(s)⟩ being more decoupled

  7. Probing the Aggregation Behavior of Neat Imidazolium-Based Alkyl Sulfate (Alkyl = Ethyl, Butyl, Hexyl, and Octyl) Ionic Liquids through Time Resolved Florescence Anisotropy and NMR and Fluorescence Correlation Spectroscopy Study.

    PubMed

    Majhi, Debashis; Pabbathi, Ashok; Sarkar, Moloy

    2016-01-14

    Aggregation behavior of a series of neat 1-ethyl 3-methylimidazolium alkyl sulfate (alkyl = ethyl, butyl, hexyl, and octyl) ionic liquids has been investigated through combined time-resolved fluorescence spectroscopy, 1-D and 2-D NMR spectroscopy, and fluorescence correlation spectroscopy (FCS). Interestingly, experimentally measured rotational relaxation times (τr) for ethyl, butyl, hexyl and octyl systems are measured to be 2.25, 1.64, 1.36, and 1.32 times higher than the estimated (from Stokes-Einstein-Debye theory) values for the same respective systems. This indicates that the emitting species is not the monomeric imidazolium moiety rather an associated species, and volume of the rotating fluorescing species decreases even though the length of the alkyl moiety on the anions is increased. The shift in the (1)H proton signal as well as a change in the width of the same signal upon dilution of the neat ionic liquids indicates that ionic liquids exist in the aggregated form. Further investigation through the 2D-ROESY experiment shows that interaction between imidazolium and sulfate is relatively stronger in the ethyl system than that of the longer octyl system. FCS measurements independently show that the hydrodynamic volume decreases with an increase in the anion chain length. The NMR and FCS results are consistent with the findings of the fluorescence anisotropy study. PMID:26654730

  8. A study of the time-resolved fluorescence spectrum and red edge effect of ANF in a room-temperature ionic liquid.

    PubMed

    Hu, Zhonghan; Margulis, Claudio J

    2006-06-15

    In a recent article, we have analyzed using molecular dynamics simulations the steady-state red edge effect (REE) observed by Samanta and co-workers when the fluorescent probe 2-amino-7-nitrofluorene (ANF) is photoexcited at different wavelengths in 1-butyl-3-methylimidazolium ([BMIM+]) hexafluorophosphate ([PF6-]). In this letter, we predict the time- and wavelength-dependent emission spectra of ANF in the same ionic solvent. From the analysis of our simulated data, we are able to derive an approximate time scale for reorganization of the solvent around the solute probe. The effect that slow varying local liquid environments have on the overall time-dependent signal is also discussed. PMID:16771357

  9. Time-resolved non-contact fluorescence diffuse optical tomography measurements with ultra-fast time-correlated single photon counting avalanche photodiodes

    NASA Astrophysics Data System (ADS)

    Bérubé-Lauzière, Yves; Robichaud, Vincent; Lapointe, Éric

    2007-07-01

    The design and fabrication of time-correlated single photon counting (TCSPC) avalanche photodiodes (APDs) and associated quenching circuits have made significant progresses in recent years. APDs with temporal resolutions comparable to microchannel plate photomultiplier tubes (MCP-PMTs) are now available. MCP-PMTs were until these progresses the best TCSPC detectors with timing resolutions down to 30ps. APDs can now achieve these resolutions at a fraction of the cost. Work is under way to make the manufacturing of TCSPC APDs compatible with standard electronics fabrication practices. This should allow to further reduce their cost and render them easier to integrate in complex multi-channel TCSPC electronics, as needed in diffuse optical tomography (DOT) systems. Even if their sensitive area is much smaller than that of the ubiquitous PMT used in TCSPC, we show that with appropriate selection of optical components, TCSPC APDs can be used in time-domain DOT. To support this, we present experimental data and calculations clearly demonstrating that comparable measurements can be obtained with APDs and PMTs. We are, to our knowledge, the first group using APDs in TD DOT, in particular in non-contact TD fluorescence DOT.

  10. Comparison of microenvironments of aqueous sodium dodecyl sulfate micelles in the presence of inorganic and organic salts: a time-resolved fluorescence anisotropy approach.

    PubMed

    Dutt, G B

    2005-11-01

    Microenvironments of aqueous sodium dodecyl sulfate (SDS) micelles was examined in the presence of additives such as sodium chloride and p-toluidine hydrochloride (PTHC) by monitoring the fluorescence anisotropy decays of two hydrophobic probes, 2,5-dimethyl-1,4-dioxo-3,6-diphenylpyrrolo[3,4-c]pyrrole (DMDPP) and coumarin 6 (C6). It has been well-established that SDS micelles undergo a sphere-to-rod transition and that their mean hydrodynamic radius increases from 19 to 100 A upon the addition of 0.0-0.7 M NaCl at 298 K. A similar size and shape transition is induced by PTHC at concentrations that are 20 times lower compared to that of NaCl. This study was undertaken to find out how the microviscosity of the micelles is influenced under these circumstances. It was noticed that the microviscosity of the SDS/NaCl system increased by approximately 45%, whereas there was a less than 10% variation in the microviscosity of the SDS/PTHC system. The large increase in the microviscosity of the former system with salt concentration has been rationalized on the basis of the high concentration of sodium ions in the headgroup region of the micelles and their ability to strongly coordinate with the water present in this region, which decreases the mobility of the probe molecules.

  11. Comparison of microenvironments of aqueous sodium dodecyl sulfate micelles in the presence of inorganic and organic salts: a time-resolved fluorescence anisotropy approach.

    PubMed

    Dutt, G B

    2005-11-01

    Microenvironments of aqueous sodium dodecyl sulfate (SDS) micelles was examined in the presence of additives such as sodium chloride and p-toluidine hydrochloride (PTHC) by monitoring the fluorescence anisotropy decays of two hydrophobic probes, 2,5-dimethyl-1,4-dioxo-3,6-diphenylpyrrolo[3,4-c]pyrrole (DMDPP) and coumarin 6 (C6). It has been well-established that SDS micelles undergo a sphere-to-rod transition and that their mean hydrodynamic radius increases from 19 to 100 A upon the addition of 0.0-0.7 M NaCl at 298 K. A similar size and shape transition is induced by PTHC at concentrations that are 20 times lower compared to that of NaCl. This study was undertaken to find out how the microviscosity of the micelles is influenced under these circumstances. It was noticed that the microviscosity of the SDS/NaCl system increased by approximately 45%, whereas there was a less than 10% variation in the microviscosity of the SDS/PTHC system. The large increase in the microviscosity of the former system with salt concentration has been rationalized on the basis of the high concentration of sodium ions in the headgroup region of the micelles and their ability to strongly coordinate with the water present in this region, which decreases the mobility of the probe molecules. PMID:16262297

  12. TIME-RESOLVED VIBRATIONAL SPECTROSCOPY

    SciTech Connect

    Andrei Tokmakoff, MIT; Paul Champion, Northeastern University; Edwin J. Heilweil, NIST; Keith A. Nelson, MIT; Larry Ziegler, Boston University

    2009-05-14

    This document contains the Proceedings from the 14th International Conference on Time-Resolved Vibrational Spectroscopy, which was held in Meredith, NH from May 9-14, 2009. The study of molecular dynamics in chemical reaction and biological processes using time-resolved spectroscopy plays an important role in our understanding of energy conversion, storage, and utilization problems. Fundamental studies of chemical reactivity, molecular rearrangements, and charge transport are broadly supported by the DOE’s Office of Science because of their role in the development of alternative energy sources, the understanding of biological energy conversion processes, the efficient utilization of existing energy resources, and the mitigation of reactive intermediates in radiation chemistry. In addition, time-resolved spectroscopy is central to all five of DOE’s grand challenges for fundamental energy science. The Time-Resolved Vibrational Spectroscopy conference is organized biennially to bring the leaders in this field from around the globe together with young scientists to discuss the most recent scientific and technological advances. The latest technology in ultrafast infrared, Raman, and terahertz spectroscopy and the scientific advances that these methods enable were covered. Particular emphasis was placed on new experimental methods used to probe molecular dynamics in liquids, solids, interfaces, nanostructured materials, and biomolecules.

  13. Rate constant for the reaction of OH with methyl iodide, a re-determination by flash photolysis of water vapour and time resolved resonance fluorescence of OH

    NASA Astrophysics Data System (ADS)

    Zhang, Shaoliang; Strekowski, Rafal; Zetzsch, Cornelius

    2010-05-01

    Methyl iodide is a major source gas for atmospheric iodine, and it is mainly emitted from the ocean. Aqueous-phase reactions, such as hydrolysis and exchange reactions with chloride control its emissions to the atmosphere, where its lifetime is limited to less than a week, mainly by photolysis. A minor contribution to the loss processes in the troposphere is the gas-phase reaction with OH radicals, that has been investigated by several authors. On the other hand, this reaction turned out to be uncertain in spite of interest in nuclear safety after the International Phebus Fission Product programme, initiated in 1988. Some of the most important observed phenomena with regard to the chemistry of iodine were not predicted, clearly showing the need for carrying out rate constant determinations for the reactions of I2 and CH3I with OH, which is a major oxidant product from the air radiolysis under accident conditions. We have measured the rate constant for the reaction OH + CH3I - H2O + CH2I in He at 260 mbar in the temperature range from 298 to 362 K. OH radicals were produced by flash photolysis of H2O in the vacuum-UV at wavelengths > 115 nm using a Xe flash lamp with a MgF2 window. Time profiles of OH radicals are monitored by resonance fluorescence of the A2 Σ - X2 Π transition at 308 nm, induced by the emission from a microwave discharge of a flow of He and H2O, a few Torr each. The signal is monitored by photon counting and multichannel scaling, collecting the counts from 50 flashes each, obtaind by pulsed photolysis of various mixtures of H2O and CH3I under slow-flow conditions. Decays of OH in the presence of CH3I are observed to be exponential, and the decay rates are found to be linearly dependent on the concentration of CH3I. Rate constants, k ± 2σ (in units of 10-14 cm3 s-1) of 4.14±0.20, 6.33±0.68, 7.31±1.18 and 8.24±1.60 at 298, 326, 352 and 362 K, respectively, are obtained from linear regressions and lead to an Arrhenius expression of k = 1.5

  14. Sensing cell metabolism by time-resolved autofluorescence.

    PubMed

    Wu, Yicong; Zheng, Wei; Qu, Jianan Y

    2006-11-01

    We built a time-resolved confocal fluorescence spectroscopy system equipped with the multichannel time-correlated single-photon-counting technique. The instrument provides a unique approach to study the fluorescence sensing of cell metabolism via analysis of the wavelength- and time-resolved intracellular autofluorescence. The experiments on monolayered cell cultures show that with UV excitation at 365 nm the time-resolved autofluorescence decays, dominated by free-bound reduced nicotinamide adenine dinucleotide signals, are sensitive indicators for cell metabolism. However, the sensitivity decreases with the increase of excitation wavelength possibly due to the interference from free-bound flavin adenine dinucleotide fluorescence. The results demonstrate that time-resolved autofluorescence can be potentially used as an important contrast mechanism to detect epithelial precancer. PMID:17041655

  15. Sensing cell metabolism by time-resolved autofluorescence

    NASA Astrophysics Data System (ADS)

    Wu, Yicong; Zheng, Wei; Qu, Jianan Y.

    2006-11-01

    We built a time-resolved confocal fluorescence spectroscopy system equipped with the multichannel time-correlated single-photon-counting technique. The instrument provides a unique approach to study the fluorescence sensing of cell metabolism via analysis of the wavelength- and time-resolved intracellular autofluorescence. The experiments on monolayered cell cultures show that with UV excitation at 365 nm the time-resolved autofluorescence decays, dominated by free-bound reduced nicotinamide adenine dinucleotide signals, are sensitive indicators for cell metabolism. However, the sensitivity decreases with the increase of excitation wavelength possibly due to the interference from free-bound flavin adenine dinucleotide fluorescence. The results demonstrate that time-resolved autofluorescence can be potentially used as an important contrast mechanism to detect epithelial precancer.

  16. Time-resolved molecular imaging

    NASA Astrophysics Data System (ADS)

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

    2016-06-01

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

  17. Complexation of Cm(III) and Eu(III) with a hydrophilic 2,6-bis(1,2,4-triazin-3-yl)-pyridine studied by time-resolved laser fluorescence spectroscopy.

    PubMed

    Ruff, Christian M; Müllich, Udo; Geist, Andreas; Panak, Petra J

    2012-12-28

    The complexation of Cm(III) and Eu(III) with 2,6-bis(5,6-di(sulfophenyl)-1,2,4-triazin-3-yl)pyridine (aq-BTP) is studied in water at pH 3.0 applying time-resolved laser fluorescence spectroscopy. With increasing ligand concentration [M(H(2)O)(9-3n)(aq-BTP)(n)] (M = Cm(III)/Eu(III), n = 1, 2, 3) complex species are spectroscopically identified. The conditional stability constants of the M(III) 1 : 3 complex species with aq-BTP are log β(03) = 12.2 for Cm(III) and log β(03) = 10.2 for Eu(III). The complexation reaction is enthalpy- and entropy-driven for both metal ions, while the enthalpy change ΔH(03) is 9.7 kJ mol(-1) more negative for Cm(III); changes in ΔS(03) are marginal. The difference in ΔG(03) of -12.7 kJ mol(-1) between the formation of the [M(aq-BTP)(3)] complexes agrees with aq-BTP's selectivity in liquid-liquid extraction studies. PMID:23018544

  18. Independent flexible motion of submolecular domains of the Ca2+,Mg2+-ATPase of sarcoplasmic reticulum measured by time-resolved fluorescence depolarization of site-specifically attached probes.

    PubMed

    Suzuki, S; Kawato, S; Kouyama, T; Kinosita, K; Ikegami, A; Kawakita, M

    1989-09-19

    The Ca2+-transporting ATPase of rabbit skeletal muscle sarcoplasmic reticulum was site-specifically labeled with either N-(1-anilinonaphth-4-yl)maleimide (ANM) or 5-[[(iodoacetamido)-ethyl]amino]naphthalene-1-sulfonate (IAEDANS), and the segmental motion of submolecular domains of the ATPase molecule was examined by means of time-resolved and steady-state fluorescence anisotropy measurements. The ANM-binding domain showed wobbling with a rotational relaxation time phi = 69 ns in the absence of free Ca2+ without any independent wobbling of the ANM moiety. The IAEDANS-binding domain showed a significantly slower wobbling with phi = 190 ns in the absence of Ca2+. The present results demonstrated for the first time that the ATPase molecule is composed of distinct domains whose mobilities are considerably different from each other. The binding of Ca2+ to the transport site increased the segmental motion of ANM-labeled domain, leading to a phi value of 65 ns. Solubilization of the ANM-labeled SR membranes by deoxycholate led to a further increase in the segmental flexibility (phi = 48 ns in the absence of free Ca2+), indicating that the mobility of the ANM-binding domain was considerably restricted through interaction with the membrane. The mobility of the ANM-binding domain of solubilized ATPase was also increased to some extent upon binding of Ca2+.

  19. Complexation of Cm(III) and Eu(III) with a hydrophilic 2,6-bis(1,2,4-triazin-3-yl)-pyridine studied by time-resolved laser fluorescence spectroscopy.

    PubMed

    Ruff, Christian M; Müllich, Udo; Geist, Andreas; Panak, Petra J

    2012-12-28

    The complexation of Cm(III) and Eu(III) with 2,6-bis(5,6-di(sulfophenyl)-1,2,4-triazin-3-yl)pyridine (aq-BTP) is studied in water at pH 3.0 applying time-resolved laser fluorescence spectroscopy. With increasing ligand concentration [M(H(2)O)(9-3n)(aq-BTP)(n)] (M = Cm(III)/Eu(III), n = 1, 2, 3) complex species are spectroscopically identified. The conditional stability constants of the M(III) 1 : 3 complex species with aq-BTP are log β(03) = 12.2 for Cm(III) and log β(03) = 10.2 for Eu(III). The complexation reaction is enthalpy- and entropy-driven for both metal ions, while the enthalpy change ΔH(03) is 9.7 kJ mol(-1) more negative for Cm(III); changes in ΔS(03) are marginal. The difference in ΔG(03) of -12.7 kJ mol(-1) between the formation of the [M(aq-BTP)(3)] complexes agrees with aq-BTP's selectivity in liquid-liquid extraction studies.

  20. Protein stabilization by osmolytes from hyperthermophiles: effect of mannosylglycerate on the thermal unfolding of recombinant nuclease a from Staphylococcus aureus studied by picosecond time-resolved fluorescence and calorimetry.

    PubMed

    Faria, Tiago Q; Lima, João C; Bastos, Margarida; Maçanita, António L; Santos, Helena

    2004-11-19

    2-O-alpha-Mannosylglycerate, a negatively charged osmolyte widely distributed among (hyper)thermophilic microorganisms, is known to provide notable protection to proteins against thermal denaturation. To study the mechanism responsible for protein stabilization, pico-second time-resolved fluorescence spectroscopy was used to characterize the thermal unfolding of a model protein, Staphylococcus aureus recombinant nuclease A (SNase), in the presence or absence of mannosylglycerate. The fluorescence decay times are signatures of the protein state, and the pre-exponential coefficients are used to evaluate the molar fractions of the folded and unfolded states. Hence, direct determination of equilibrium constants of unfolding from molar fractions was carried out. Van't Hoff plots of the equilibrium constants provided reliable thermodynamic data for SNase unfolding. Differential scanning calorimetry was used to validate this thermodynamic analysis. The presence of 0.5 m potassium mannosylglycerate caused an increase of 7 degrees C in the SNase melting temperature and a 2-fold increase in the unfolding heat capacity. Despite the considerable degree of stabilization rendered by this solute, the nature and population of protein states along unfolding were not altered in the presence of mannosylglycerate, denoting that the unfolding pathway of SNase was unaffected. The stabilization of SNase by mannosylglycerate arises from decreased unfolding entropy up to 65 degrees C and from an enthalpy increase above this temperature. In molecular terms, stabilization is interpreted as resulting from destabilization of the denatured state caused by preferential exclusion of the solute from the protein hydration shell upon unfolding, and stabilization of the native state by specific interactions. The physiological significance of charged solutes in hyperthermophiles is discussed.

  1. Eicosapentaenoic acid plays a role in stabilizing dynamic membrane structure in the deep-sea piezophile Shewanella violacea: a study employing high-pressure time-resolved fluorescence anisotropy measurement.

    PubMed

    Usui, Keiko; Hiraki, Toshiki; Kawamoto, Jun; Kurihara, Tatsuo; Nogi, Yuichi; Kato, Chiaki; Abe, Fumiyoshi

    2012-03-01

    Shewanella violacea DSS12 is a psychrophilic piezophile that optimally grows at 30MPa. It contains a substantial amount of eicosapentaenoic acid (EPA) in the membrane. Despite evidence linking increased fatty acid unsaturation and bacterial growth under high pressure, little is known of how the physicochemical properties of the membrane are modulated by unsaturated fatty acids in vivo. By means of the newly developed system performing time-resolved fluorescence anisotropy measurement under high pressure (HP-TRFAM), we demonstrate that the membrane of S. violacea is highly ordered at 0.1MPa and 10°C with the order parameter S of 0.9, and the rotational diffusion coefficient D(w) of 5.4μs(-1) for 1-[4-(trimethylamino)pheny]-6-phenyl-1,3,5-hexatriene in the membrane. Deletion of pfaA encoding the omega-3 polyunsaturated fatty acid synthase caused disorder of the membrane and enhanced the rotational motion of acyl chains, in concert with a 2-fold increase in the palmitoleic acid level. While the wild-type membrane was unperturbed over a wide range of pressures with respect to relatively small effects of pressure on S and D(w), the ΔpfaA membrane was disturbed judging from the degree of increased S and decreased D(w). These results suggest that EPA prevents the membrane from becoming hyperfluid and maintains membrane stability against significant changes in pressure. Our results counter the generally accepted concept that greater fluidity is a membrane characteristic of microorganisms that inhabit cold, high-pressure environments. We suggest that retaining a certain level of membrane physical properties under high pressure is more important than conferring membrane fluidity alone. PMID:22037146

  2. Time-resolved imaging of latent fingerprints with nanosecond resolution

    NASA Astrophysics Data System (ADS)

    Seah, L. K.; Dinish, U. S.; Ong, S. K.; Chao, Z. X.; Murukeshan, V. M.

    2004-07-01

    Imaging of latent fingerprints using time-resolved (TR) method offers a broader platform to eliminate the unwanted background emission. In this paper, a novel TR imaging technique is demonstrated and implemented, which facilitates the detection of latent fingerprints with nanosecond resolution. Simulated experiments were carried out with two overlapping fingerprints treated with two fluorescent powders having different lifetimes in nanosecond range. The dependence of the fluorescence emission intensity in nanosecond resolution of TR imaging is also revealed.

  3. Combined time-resolved laser fluorescence spectroscopy and extended X-ray absorption fine structure spectroscopy study on the complexation of trivalent actinides with chloride at T = 25-200 °C.

    PubMed

    Skerencak-Frech, Andrej; Fröhlich, Daniel R; Rothe, Jörg; Dardenne, Kathy; Panak, Petra J

    2014-01-21

    The complexation of trivalent actinides (An(III)) with chloride is studied in the temperature range from 25 to 200 °C by spectroscopic methods. Time-resolved laser fluorescence spectroscopy (TRLFS) is applied to determine the thermodynamic data of Cm(III)-Cl(-) complexes, while extended X-ray absorption fine structure spectroscopy (EXAFS) is used to determine the structural data of the respective Am(III) complexes. The experiments are performed in a custom-built high-temperature cell which is modified for the respective spectroscopic technique. The TRLFS results show that at 25 °C the speciation is dominated mainly by the Cm(3+) aquo ion. Only a minor fraction of the CmCl(2+) complex is present in solution. As the temperature increases, the fraction of this species decreases further. Simultaneously, the fraction of the CmCl2(+) complex increases strongly with the temperature. Also, the CmCl3 complex is formed to a minor extent at T > 160 °C. The conditional stability constant log β'2 is determined as a function of the temperature and extrapolated to zero ionic strength with the specific ion interaction theory approach. The log β°2(T) values increase by more than 3 orders of magnitude in the studied temperature range. The temperature dependency of log β°2 is fitted by the extended van't Hoff equation to determine ΔrH°m, ΔrS°m, and ΔrC°p,m. The EXAFS results support these findings. The results confirm the absence of americium(III) chloride complexes at T = 25 and 90 °C ([Am(III)] = 10(-3) m, [Cl(-)] = 3.0 m), and the spectra are described by 9-10 oxygen atoms at a distance of 2.44-2.48 Å. At T = 200 °C two chloride ligands are present in the inner coordination sphere of Am(III) at a distance of 2.78 Å.

  4. Time-resolved Raman spectroscopy for in situ planetary mineralogy.

    PubMed

    Blacksberg, Jordana; Rossman, George R; Gleckler, Anthony

    2010-09-10

    Planetary mineralogy can be revealed through a variety of remote sensing and in situ investigations that precede any plans for eventual sample return. We briefly review those techniques and focus on the capabilities for on-surface in situ examination of Mars, Venus, the Moon, asteroids, and other bodies. Over the past decade, Raman spectroscopy has continued to develop as a prime candidate for the next generation of in situ planetary instruments, as it provides definitive structural and compositional information of minerals in their natural geological context. Traditional continuous-wave Raman spectroscopy using a green laser suffers from fluorescence interference, which can be large (sometimes saturating the detector), particularly in altered minerals, which are of the greatest geophysical interest. Taking advantage of the fact that fluorescence occurs at a later time than the instantaneous Raman signal, we have developed a time-resolved Raman spectrometer that uses a streak camera and pulsed miniature microchip laser to provide picosecond time resolution. Our ability to observe the complete time evolution of Raman and fluorescence spectra in minerals makes this technique ideal for exploration of diverse planetary environments, some of which are expected to contain strong, if not overwhelming, fluorescence signatures. We discuss performance capability and present time-resolved pulsed Raman spectra collected from several highly fluorescent and Mars-relevant minerals. In particular, we have found that conventional Raman spectra from fine grained clays, sulfates, and phosphates exhibited large fluorescent signatures, but high quality spectra could be obtained using our time-resolved approach.

  5. Time-Resolved Photoluminescence and Photovoltaics

    SciTech Connect

    Metzger, W. K.; Ahrenkiel, R. K.; Dippo, P.; Geisz, J.; Wanlass, M. W.; Kurtz, S.

    2005-01-01

    The time-resolved photoluminescence (TRPL) technique and its ability to characterize recombination in bulk photovoltaic semiconductor materials are reviewed. Results from a variety of materials and a few recent studies are summarized and compared.

  6. Time-resolved transillumination and optical tomography

    NASA Astrophysics Data System (ADS)

    de Haller, Emmanuel B.

    1996-01-01

    In response to an invitation by the editor-in-chief, I would like to present the current status of time-domain imaging. With exciting new photon diffusion techniques being developed in the frequency domain and promising optical coherence tomography, time-resolved transillumination is in constant evolution and the subject of passionate discussions during the numerous conferences dedicated to this subject. The purpose of time-resolved optical tomography is to provide noninvasive, high-resolution imaging of the interior of living bodies by the use of nonionizing radiation. Moreover, the use of visible to near-infrared wavelength yields metabolic information. Breast cancer screening is the primary potential application for time-resolved imaging. Neurology and tissue characterization are also possible fields of applications. Time- resolved transillumination and optical tomography should not only improve diagnoses, but the welfare of the patient. As no overview of this technique has yet been presented to my knowledge, this paper briefly describes the various methods enabling time-resolved transillumination and optical tomography. The advantages and disadvantages of these methods, as well as the clinical challenges they face are discussed. Although an analytic and computable model of light transport through tissues is essential for a meaningful interpretation of the transillumination process, this paper will not dwell on the mathematics of photon propagation.

  7. Detection of colorectal cancer using time-resolved autofluorescence spectrometer

    NASA Astrophysics Data System (ADS)

    Fu, Sheng; Kwek, Leong-Chuan; Chia, Teck-Chee; Lim, Chu-Sing; Tang, Choong-Leong; Ang, Wuan-Suan; Zhou, Miao-Chang; Loke, Po-Ling

    2006-04-01

    As we know Quantum mechanics is a mathematical theory that can describe the behavior of objects that are at microscopic level. Time-resolved autofluorescence spectrometer monitors events that occur during the lifetime of the excited state. This time ranges from a few picoseconds to hundreds of nanoseconds. That is an extremely important advance as it allows environmental parameters to be monitored in a spatially defined manner in the specimen under study. This technique is based on the application of Quantum Mechanics. This principle is applied in our project as we are trying to use different fluorescence spectra to detect biological molecules commonly found in cancerous colorectal tissue and thereby differentiate the cancerous and non-cancerous colorectal polyps more accurately and specifically. In this paper, we use Fluorescence Lifetime Spectrometer (Edinburgh Instruments FL920) to measure decay time of autofluorescence of colorectal cancerous and normal tissue sample. All specimens are from Department of Colorectal Surgery, Singapore General Hospital. The tissues are placed in the time-resolved autofluorescence instrument, which records and calculates the decay time of the autofluorescence in the tissue sample at the excitation and emission wavelengths pre-determined from a conventional spectrometer. By studying the decay time,τ, etc. for cancerous and normal tissue, we aim to present time-resolved autofluorescence as a feasible technique for earlier detection of malignant colorectal tissues. By using this concept, we try to contribute an algorithm even an application tool for real time early diagnosis of colorectal cancer for clinical services.

  8. Time-resolved optical diffusion tomography

    NASA Astrophysics Data System (ADS)

    Appledorn, C. Robert; Kruger, Robert A.; Liu, Pingyu

    1994-05-01

    A mathematical model is proposed describing time-resolved output measurements obtained on the surface of a diffusely scattering body due to an input pulse of near-IR light at a different location also on the surface. Such measurements can be obtained using a pulsed near-IR laser coupled with a CCD streak camera. The scattering body is assumed to exhibit homogenous scattering and spatially varying absorption. Using this model, an iterative algorithm is derived using maximum likelihood methods that allows the reconstruction of the spatial absorption pattern from a set of time-resolved tomographic measurements. The methodology places no restrictions upon the time-of-arrival of the detected photons, thus permitting the entire time-resolved signal to be used in the reconstruction process. The reconstruction algorithm is easily initialized and preliminary results indicate that stable reconstructions can be performed.

  9. Time resolved thermal lens in edible oils

    NASA Astrophysics Data System (ADS)

    Albuquerque, T. A. S.; Pedreira, P. R. B.; Medina, A. N.; Pereira, J. R. D.; Bento, A. C.; Baesso, M. L.

    2003-01-01

    In this work time resolved thermal lens spectrometry is applied to investigate the optical properties of the following edible oils: soya, sunflower, canola, and corn oils. The experiments were performed at room temperature using the mode mismatched thermal lens configuration. The results showed that when the time resolved procedure is adopted the technique can be applied to investigate the photosensitivity of edible oils. Soya oil presented a stronger photochemical reaction as compared to the other investigated samples. This observation may be relevant for future studies evaluating edible oils storage conditions and also may contribute to a better understanding of the physical and chemical properties of this important foodstuff.

  10. Steady-state and time-resolved Thioflavin-T fluorescence can report on morphological differences in amyloid fibrils formed by Aβ(1-40) and Aβ(1-42)

    SciTech Connect

    Lindberg, David J.; Wranne, Moa S.; Gilbert Gatty, Mélina; Westerlund, Fredrik; Esbjörner, Elin K.

    2015-03-06

    Thioflavin-T (ThT) is one of the most commonly used dyes for amyloid detection, but the origin of its fluorescence enhancement is not fully understood. Herein we have characterised the ThT fluorescence response upon binding to the Aβ(1-40) and Aβ(1-42) variants of the Alzheimer's-related peptide amyloid-β, in order to explore how the photophysical properties of this dye relates to structural and morphological properties of two amyloid fibril types formed by peptides with a high degree of sequence homology. We show that the steady-state ThT fluorescence is 1.7 times more intense with Aβ(1-40) compared to Aβ(1-42) fibrils in concentration matched samples prepared under quiescent conditions. By measuring the excited state lifetime of bound ThT, we also demonstrate a distinct difference between the two fibril isoforms, with Aβ(1-42) fibrils producing a longer ThT fluorescence lifetime compared to Aβ(1-40). The substantial steady-state intensity difference is therefore not explained by differences in fluorescence quantum yield. Further, we find that the ThT fluorescence intensity, but not the fluorescence lifetime, is dependent on the fibril preparation method (quiescent versus agitated conditions). We therefore propose that the fluorescence lifetime is inherent to each isoform and sensitively reports on fibril microstructure in the protofilament whereas the total fluorescence intensity relates to the amount of exposed β-sheet in the mature Aβ fibrils and hence to differences in their morphology. Our results highlight the complexity of ThT fluorescence, and demonstrate its extended use in amyloid fibril characterisation. - Highlights: • ThT emission is more intense with Aβ(1-40) fibrils than with Aβ(1-42) fibrils. • Aβ(1-42) fibrils induce longer ThT fluorescence lifetimes and higher quantum yield. • ThT emission intensity in Aβ fibril samples reports on fibril morphology. • The ThT fluorescence lifetime is a characteristic feature of each A

  11. Comparison of the rate constants for energy transfer in the light-harvesting protein, C-phycocyanin, calculated from Foerster`s theory and experimentally measured by time-resolved fluorescence spectroscopy

    SciTech Connect

    Debreczeny, M.P.

    1994-05-01

    We have measured and assigned rate constants for energy transfer between chromophores in the light-harvesting protein C-phycocyanin (PC), in the monomeric and trimeric aggregation states, isolated from Synechococcus sp. PCC 7002. In order to compare the measured rate constants with those predicted by Fdrster`s theory of inductive resonance in the weak coupling limit, we have experimentally resolved several properties of the three chromophore types ({beta}{sub 155} {alpha}{sub 84}, {beta}{sub 84}) found in PC monomers, including absorption and fluorescence spectra, extinction coefficients, fluorescence quantum yields, and fluorescence lifetimes. The cpcB/C155S mutant, whose PC is missing the {beta}{sub 155} chromophore, was, useful in effecting the resolution of the chromophore properties and in assigning the experimentally observed rate constants for energy transfer to specific pathways.

  12. Deflection evaluation using time-resolved radiography

    SciTech Connect

    Fry, D.A.; Lucero, J.P.

    1990-01-01

    Time-resolved radiography is the creation of an x-ray image for which both the start-exposure and stop-exposure times are known with respect to the event under study. The combination of image and timing are used to derive information about the event. We have applied time-resolved radiography to evaluate motions of explosive-driven events. In the particular application discussed here, our intent is to measure maximum deflections of the components involved. Exposures are made during the time just before to just after the event of interest occurs. A smear or blur of motion out to its furthest extent is recorded on the image. Comparison of the dynamic images with static images allows deflection measurements to be made. 2 figs.

  13. Time resolved astronomy with the SALT

    NASA Astrophysics Data System (ADS)

    Buckley, D. A. H.; Crawford, S.; Gulbis, A. A. S.; McPhate, J.; Nordsieck, K. H.; Potter, S. B.; O'Donoghue, D.; Siegmund, O. H. W.; Schellart, P.; Spark, M.; Welsh, B. Y.; Zietsman, E.

    2010-07-01

    While time resolved astronomical observations are not new, the extension of such studies to sub-second time resolution is and has resulted in the opening of a new observational frontier, High Time Resolution Astronomy (HTRA). HTRA studies are well suited to objects like compact binary stars (CVs and X-ray binaries) and pulsars, while asteroseismology of pulsating stars, occultations, transits and the study of transients, will all benefit from such HTRA studies. HTRA has been a SALT science driver from the outset and the first-light instruments, namely the UV-VIS imager, SALTICAM, and the multi-purpose Robert Stobie Spectrograph (RSS), both have high time resolution modes. These are described, together with some observational examples. We also discuss the commissioning observations with the photon counting Berkeley Visible Image Tube camera (BVIT) on SALT. Finally we describe the software tools, developed in Python, to reduce SALT time resolved observations.

  14. Bayesian approach to time-resolved tomography.

    PubMed

    Myers, Glenn R; Geleta, Matthew; Kingston, Andrew M; Recur, Benoit; Sheppard, Adrian P

    2015-07-27

    Conventional X-ray micro-computed tomography (μCT) is unable to meet the need for real-time, high-resolution, time-resolved imaging of multi-phase fluid flow. High signal-to-noise-ratio (SNR) data acquisition is too slow and results in motion artefacts in the images, while fast acquisition is too noisy and results in poor image contrast. We present a Bayesian framework for time-resolved tomography that uses priors to drastically reduce the required amount of experiment data. This enables high-quality time-resolved imaging through a data acquisition protocol that is both rapid and high SNR. Here we show that the framework: (i) encompasses our previous, algorithms for imaging two-phase flow as limiting cases; (ii) produces more accurate results from imperfect (i.e. real) data, where it can be compared to our previous work; and (iii) is generalisable to previously intractable systems, such as three-phase flow. PMID:26367664

  15. The differences in short- and long-term varicella-zoster virus (VZV) immunoglobulin G levels following varicella vaccination of healthcare workers measured by VZV fluorescent-antibody-to-membrane-antigen assay (FAMA), VZV time-resolved fluorescence immunoassay and a VZV purified glycoprotein enzyme immunoassay.

    PubMed

    Maple, P A C; Haedicke, J; Quinlivan, M; Steinberg, S P; Gershon, A A; Brown, K E; Breuer, J

    2016-08-01

    Healthcare workers (HCWs) reporting no history of varicella frequently receive varicella vaccination (vOka) if they test varicella-zoster virus (VZV) immunoglobulin G (IgG) negative. In this study, the utilities of VZV-IgG time-resolved fluorescence immunoassay (VZV-TRFIA) and a commercial VZV-IgG purified glycoprotein enzyme immunoassay (gpEIA) currently used in England for confirming VZV immunity have been compared to the fluorescent-antibody-to-membrane-antigen assay (FAMA). A total of 110 HCWs received two doses of vOka vaccine spaced 6 weeks apart and sera collected pre-vaccination (n = 100), at 6 weeks post-completion of vaccination (n = 86) and at 12-18 months follow-up (n = 73) were analysed. Pre-vaccination, by FAMA, 61·0% sera were VZV IgG negative, and compared to FAMA the sensitivities of VZV-TRFIA and gpEIA were 74·4% [95% confidence interval (CI) 57·9-87·0] and 46·2% (95% CI 30·1-62·8), respectively. Post-completion of vaccination the seroconversion rate by FAMA was 93·7% compared to rates of 95·8% and 70·8% determined by VZV-TRFIA and gpEIA, respectively. At 12-18 months follow-up seropositivity rates by FAMA, VZV-TRFIA and gpEIA were 78·1%, 74·0% and 47·9%, respectively. Compared to FAMA the sensitivities of VZV-TRFIA and gpEIA for measuring VZV IgG following vaccination were 96·4% (95% CI 91·7-98·8) and 74·6% (95% CI 66·5-81·6), respectively. Using both FAMA and VZV-TRFIA to identify healthy adult VZV susceptibles and measure seroconversion showed that vOka vaccination of HCWs is highly immunogenic.

  16. Time-resolved multiphoton imaging of basal cell carcinoma

    NASA Astrophysics Data System (ADS)

    Cicchi, R.; Sestini, S.; De Giorgi, V.; Stambouli, D.; Carli, P.; Massi, D.; Pavone, F. S.

    2007-02-01

    We investigated human cutaneous basal cell carcinoma ex-vivo samples by combined time resolved two photon intrinsic fluorescence and second harmonic generation microscopy. Morphological and spectroscopic differences were found between malignant skin and corresponding healthy skin tissues. In comparison with normal healthy skin, cancer tissue showed a different morphology and a mean fluorescence lifetime distribution slightly shifted towards higher values. Topical application of delta-aminolevulinic acid to the lesion four hours before excision resulted in an enhancement of the fluorescence signal arising from malignant tissue, due to the accumulation of protoporphyrines inside tumor cells. Contrast enhancement was prevalent at tumor borders by both two photon fluorescence microscopy and fluorescence lifetime imaging. Fluorescence-based images showed a good correlation with conventional histopathological analysis, thereby supporting the diagnostic accuracy of this novel method. Combined morphological and lifetime analysis in the study of ex-vivo skin samples discriminated benign from malignant tissues, thus offering a reliable, non-invasive tool for the in-vivo analysis of inflammatory and neoplastic skin lesions.

  17. Structural kinetics of myosin by transient time-resolved FRET

    PubMed Central

    Nesmelov, Yuri E.; Agafonov, Roman V.; Negrashov, Igor V.; Blakely, Sarah E.; Titus, Margaret A.; Thomas, David D.

    2011-01-01

    For many proteins, especially for molecular motors and other enzymes, the functional mechanisms remain unsolved due to a gap between static structural data and kinetics. We have filled this gap by detecting structure and kinetics simultaneously. This structural kinetics experiment is made possible by a new technique, (TR)2FRET (transient time-resolved FRET), which resolves protein structural states on the submillisecond timescale during the transient phase of a biochemical reaction. (TR)2FRET is accomplished with a fluorescence instrument that uses a pulsed laser and direct waveform recording to acquire an accurate subnanosecond time-resolved fluorescence decay every 0.1 ms after stopped flow. To apply this method to myosin, we labeled the force-generating region site specifically with two probes, mixed rapidly with ATP to initiate the recovery stroke, and measured the interprobe distance by (TR)2FRET with high resolution in both space and time. We found that the relay helix bends during the recovery stroke, most of which occurs before ATP is hydrolyzed, and two structural states (relay helix straight and bent) are resolved in each nucleotide-bound biochemical state. Thus the structural transition of the force-generating region of myosin is only loosely coupled to the ATPase reaction, with conformational selection driving the motor mechanism. PMID:21245357

  18. Time Resolved Deposition Measurements in NSTX

    SciTech Connect

    C.H. Skinner; H. Kugel; A.L. Roquemore; J. Hogan; W.R. Wampler; the NSTX team

    2004-08-03

    Time-resolved measurements of deposition in current tokamaks are crucial to gain a predictive understanding of deposition with a view to mitigating tritium retention and deposition on diagnostic mirrors expected in next-step devices. Two quartz crystal microbalances have been installed on NSTX at a location 0.77m outside the last closed flux surface. This configuration mimics a typical diagnostic window or mirror. The deposits were analyzed ex-situ and found to be dominantly carbon, oxygen, and deuterium. A rear facing quartz crystal recorded deposition of lower sticking probability molecules at 10% of the rate of the front facing one. Time resolved measurements over a 4-week period with 497 discharges, recorded 29.2 {micro}g/cm{sup 2} of deposition, however surprisingly, 15.9 {micro}g/cm{sup 2} of material loss occurred at 7 discharges. The net deposited mass of 13.3 {micro}g/cm{sup 2} matched the mass of 13.5 {micro}g/cm{sup 2} measured independently by ion beam analysis. Monte Carlo modeling suggests that transient processes are likely to dominate the deposition.

  19. Time-resolved resonance Raman spectroscopy: exploring reactive intermediates.

    PubMed

    Sahoo, Sangram Keshari; Umapathy, Siva; Parker, Anthony W

    2011-10-01

    The study of reaction mechanisms involves systematic investigations of the correlation between structure, reactivity, and time. The challenge is to be able to observe the chemical changes undergone by reactants as they change into products via one or several intermediates such as electronic excited states (singlet and triplet), radicals, radical ions, carbocations, carbanions, carbenes, nitrenes, nitrinium ions, etc. The vast array of intermediates and timescales means there is no single "do-it-all" technique. The simultaneous advances in contemporary time-resolved Raman spectroscopic techniques and computational methods have done much towards visualizing molecular fingerprint snapshots of the reactive intermediates in the microsecond to femtosecond time domain. Raman spectroscopy and its sensitive counterpart resonance Raman spectroscopy have been well proven as means for determining molecular structure, chemical bonding, reactivity, and dynamics of short-lived intermediates in solution phase and are advantageous in comparison to commonly used time-resolved absorption and emission spectroscopy. Today time-resolved Raman spectroscopy is a mature technique; its development owes much to the advent of pulsed tunable lasers, highly efficient spectrometers, and high speed, highly sensitive multichannel detectors able to collect a complete spectrum. This review article will provide a brief chronological development of the experimental setup and demonstrate how experimentalists have conquered numerous challenges to obtain background-free (removing fluorescence), intense, and highly spectrally resolved Raman spectra in the nanosecond to microsecond (ns-μs) and picosecond (ps) time domains and, perhaps surprisingly, laid the foundations for new techniques such as spatially offset Raman spectroscopy. PMID:21986070

  20. Water-soluble poly(2,7-dibenzosilole) as an ultra-bright fluorescent label for antibody-based flow cytometry.

    PubMed

    Wang, Xin; Hu, Yi-Zhen; Chen, Aimei; Wu, Yexin; Aggeler, Robert; Low, Quentin; Kang, Hee Chol; Gee, Kyle R

    2016-03-14

    A series of novel water-soluble PEGylated dibenzosilole-based conjugated polymers were prepared as ultra-bright fluorescent labels for biomolecules. Due to their superior solubility and brightness, antibody conjugates labeled with functionalized polymers showed significantly enhanced signal and sensitivity relative to traditional fluorophores in functional flow cytometry applications. PMID:26888307

  1. An ultra-sensitive monoclonal antibody-based fluorescent microsphere immunochromatographic test strip assay for detecting aflatoxin M1 in milk

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A rapid lateral flow fluorescent microspheres immunochromatography test strip (FMs-ICTS) has been developed for the detection of aflatoxin M1 (AFM1) residues in milk. For this purpose, an ultra-sensitive anti-AFM1 monoclonal antibody (MAb) 1D3 was prepared and identified. The IC50 value of the MA...

  2. Time-resolved force distribution analysis

    PubMed Central

    2013-01-01

    Background Biomolecules or other complex macromolecules undergo conformational transitions upon exposure to an external perturbation such as ligand binding or mechanical force. To follow fluctuations in pairwise forces between atoms or residues during such conformational changes as observed in Molecular Dynamics (MD) simulations, we developed Time-Resolved Force Distribution Analysis (TRFDA). Results The implementation focuses on computational efficiency and low-memory usage and, along with the wide range of output options, makes possible time series analysis of pairwise forces variation in long MD simulations and for large molecular systems. It also provides an exact decomposition of pairwise forces resulting from 3- and 4-body potentials and a unified treatment of pairwise forces between atoms or residues. As a proof of concept, we present a stress analysis during unfolding of ubiquitin in a force-clamp MD simulation. Conclusions TRFDA can be used, among others, in tracking signal propagation at atomic level, for characterizing dynamical intermolecular interactions (e.g. protein-ligand during flexible docking), in development of force fields and for following stress distribution during conformational changes. PMID:24499624

  3. Time-resolved tribo-thermography

    NASA Astrophysics Data System (ADS)

    Dinwiddie, Ralph B.; Blau, Peter J.

    1999-03-01

    Wear of coated surfaces tends to progress through a series of stages in which damage accumulates until the coating fails to protect its substrate. Depending on the coating system and the contact conditions, these stages can sometimes be detected as a series of discrete periods of changing frictional behavior, or they can occur quite rapidly, leading to rapid removal of the coating. A new technique has been developed to capture magnified infrared (IR) images of a selected location on a moving wear surface and to synchronize these cycle-by-cycle images with the instantaneous friction force that occurs at the same location. A pin-on-disk tribometer has been used to demonstrate the principle, but other kinds of test geometries can also be used. Contrast in the IR images derives not only from the surface temperatures but also from the emissivity of surface features. A spatial calibration of the system allows the measurement of the width of the wear path as a function of time. By studying a series of captured and friction- synchronized images, it is possible to observe the detailed progression of wear and the corresponding frictional transitions in a limitless variety of materials. Examples of several different materials, including, steel, aluminum, brass, and paint, will be used to illustrate the application of time-resolved microscopic tribo-thermography to coatings research.

  4. Time resolved ion beam induced charge collection

    SciTech Connect

    SEXTON,FREDERICK W.; WALSH,DAVID S.; DOYLE,BARNEY L.; DODD,PAUL E.

    2000-04-01

    Under this effort, a new method for studying the single event upset (SEU) in microelectronics has been developed and demonstrated. Called TRIBICC, for Time Resolved Ion Beam Induced Charge Collection, this technique measures the transient charge-collection waveform from a single heavy-ion strike with a {minus}.03db bandwidth of 5 GHz. Bandwidth can be expanded up to 15 GHz (with 5 ps sampling windows) by using an FFT-based off-line waveform renormalization technique developed at Sandia. The theoretical time resolution of the digitized waveform is 24 ps with data re-normalization and 70 ps without re-normalization. To preserve the high bandwidth from IC to the digitizing oscilloscope, individual test structures are assembled in custom high-frequency fixtures. A leading-edge digitized waveform is stored with the corresponding ion beam position at each point in a two-dimensional raster scan. The resulting data cube contains a spatial charge distribution map of up to 4,096 traces of charge (Q) collected as a function of time. These two dimensional traces of Q(t) can cover a period as short as 5 ns with up to 1,024 points per trace. This tool overcomes limitations observed in previous multi-shot techniques due to the displacement damage effects of multiple ion strikes that changed the signal of interest during its measurement. This system is the first demonstration of a single-ion transient measurement capability coupled with spatial mapping of fast transients.

  5. Time-resolved local strain tracking microscopy for cell mechanics

    NASA Astrophysics Data System (ADS)

    Aydin, O.; Aksoy, B.; Akalin, O. B.; Bayraktar, H.; Alaca, B. E.

    2016-02-01

    A uniaxial cell stretching technique to measure time-resolved local substrate strain while simultaneously imaging adherent cells is presented. The experimental setup comprises a uniaxial stretcher platform compatible with inverted microscopy and transparent elastomer samples with embedded fluorescent beads. This integration enables the acquisition of real-time spatiotemporal data, which is then processed using a single-particle tracking algorithm to track the positions of fluorescent beads for the subsequent computation of local strain. The present local strain tracking method is demonstrated using polydimethylsiloxane (PDMS) samples of rectangular and dogbone geometries. The comparison of experimental results and finite element simulations for the two sample geometries illustrates the capability of the present system to accurately quantify local deformation even when the strain distribution is non-uniform over the sample. For a regular dogbone sample, the experimentally obtained value of local strain at the center of the sample is 77%, while the average strain calculated using the applied cross-head displacement is 48%. This observation indicates that considerable errors may arise when cross-head measurement is utilized to estimate strain in the case of non-uniform sample geometry. Finally, the compatibility of the proposed platform with biological samples is tested using a unibody PDMS sample with a well to contain cells and culture media. HeLa S3 cells are plated on collagen-coated samples and cell adhesion and proliferation are observed. Samples with adherent cells are then stretched to demonstrate simultaneous cell imaging and tracking of embedded fluorescent beads.

  6. Examination of laser microbeam cell lysis in a PDMS microfluidic channel using time-resolved imaging.

    PubMed

    Quinto-Su, Pedro A; Lai, Hsuan-Hong; Yoon, Helen H; Sims, Christopher E; Allbritton, Nancy L; Venugopalan, Vasan

    2008-03-01

    We use time-resolved imaging to examine the lysis dynamics of non-adherent BAF-3 cells within a microfluidic channel produced by the delivery of single highly-focused 540 ps duration laser pulses at lambda = 532 nm. Time-resolved bright-field images reveal that the delivery of the pulsed laser microbeam results in the formation of a laser-induced plasma followed by shock wave emission and cavitation bubble formation. The confinement offered by the microfluidic channel constrains substantially the cavitation bubble expansion and results in significant deformation of the PDMS channel walls. To examine the cell lysis and dispersal of the cellular contents, we acquire time-resolved fluorescence images of the process in which the cells were loaded with a fluorescent dye. These fluorescence images reveal cell lysis to occur on the nanosecond to microsecond time scale by the plasma formation and cavitation bubble dynamics. Moreover, the time-resolved fluorescence images show that while the cellular contents are dispersed by the expansion of the laser-induced cavitation bubble, the flow associated with the bubble collapse subsequently re-localizes the cellular contents to a small region. This capacity of pulsed laser microbeam irradiation to achieve rapid cell lysis in microfluidic channels with minimal dilution of the cellular contents has important implications for their use in lab-on-a-chip applications. PMID:18305858

  7. Time-resolved spectral imaging: better photon economy, higher accuracy

    NASA Astrophysics Data System (ADS)

    Fereidouni, Farzad; Reitsma, Keimpe; Blab, Gerhard A.; Gerritsen, Hans C.

    2015-03-01

    Lifetime and spectral imaging are complementary techniques that offer a non-invasive solution for monitoring metabolic processes, identifying biochemical compounds, and characterizing their interactions in biological tissues, among other tasks. Newly developed instruments that perform time-resolved spectral imaging can provide even more information and reach higher sensitivity than either modality alone. Here we report a multispectral lifetime imaging system based on a field-programmable gate array (FPGA), capable of operating at high photon count rates (12 MHz) per spectral detection channel, and with time resolution of 200 ps. We performed error analyses to investigate the effect of gate width and spectral-channel width on the accuracy of estimated lifetimes and spectral widths. Temporal and spectral phasors were used for analysis of recorded data, and we demonstrated blind un-mixing of the fluorescent components using information from both modalities. Fractional intensities, spectra, and decay curves of components were extracted without need for prior information. We further tested this approach with fluorescently doubly-labeled DNA, and demonstrated its suitability for accurately estimating FRET efficiency in the presence of either non-interacting or interacting donor molecules.

  8. Time-resolved local strain tracking microscopy for cell mechanics.

    PubMed

    Aydin, O; Aksoy, B; Akalin, O B; Bayraktar, H; Alaca, B E

    2016-02-01

    A uniaxial cell stretching technique to measure time-resolved local substrate strain while simultaneously imaging adherent cells is presented. The experimental setup comprises a uniaxial stretcher platform compatible with inverted microscopy and transparent elastomer samples with embedded fluorescent beads. This integration enables the acquisition of real-time spatiotemporal data, which is then processed using a single-particle tracking algorithm to track the positions of fluorescent beads for the subsequent computation of local strain. The present local strain tracking method is demonstrated using polydimethylsiloxane (PDMS) samples of rectangular and dogbone geometries. The comparison of experimental results and finite element simulations for the two sample geometries illustrates the capability of the present system to accurately quantify local deformation even when the strain distribution is non-uniform over the sample. For a regular dogbone sample, the experimentally obtained value of local strain at the center of the sample is 77%, while the average strain calculated using the applied cross-head displacement is 48%. This observation indicates that considerable errors may arise when cross-head measurement is utilized to estimate strain in the case of non-uniform sample geometry. Finally, the compatibility of the proposed platform with biological samples is tested using a unibody PDMS sample with a well to contain cells and culture media. HeLa S3 cells are plated on collagen-coated samples and cell adhesion and proliferation are observed. Samples with adherent cells are then stretched to demonstrate simultaneous cell imaging and tracking of embedded fluorescent beads. PMID:26931864

  9. PREFACE: Time-resolved scanning tunnelling microscopy Time-resolved scanning tunnelling microscopy

    NASA Astrophysics Data System (ADS)

    Zandvliet, Harold J. W.; Lin, Nian

    2010-07-01

    out the potential landscape of the system (often a molecule or an atom) under study [4, 5]. However, the dynamical processes might also be induced by the tunnelling process itself [6, 7]. In the field of molecular science, excited single molecule experiments have been especially performed [8]. As a nice example, we refer to the work of Sykes' group [9] on thioether molecular rotors. In addition, several groups explore the possibility of combining time-resolved scanning tunnelling microscopy with optical techniques [10, 11]. Although the majority of studies that have been performed so far focus on rather simple systems under nearly ideal and well-defined conditions, we anticipate that time-resolved scanning tunnelling microscopy can also be applied in other research areas, such as biology and soft condensed matter, where the experimental conditions are often less ideal. We hope that readers will enjoy this collection of papers and that it will trigger them to further explore the possibilities of this simple, but powerful technique. References [1] Besenbacher F, Laegsgaard E and Stengaard I 2005 Mater. Today 8 26 [2] van Houselt A and Zandvliet H J W 2010 Rev. Mod. Phys. 82 1593 [3] Tringides M C and Hupalo M 2010 J. Phys.: Condens. Matter 22 264002 [4] Ronci F, Colonna S, Cricenti A and Le Lay G 2010 J. Phys.: Condens. Matter 22 264003 [5] van Houselt A, Poelsema B and Zandvliet H J W 2010 J. Phys.: Condens. Matter 22 264004 [6] Sprodowski C, Mehlhorn M and Morgenstern K 2010 J. Phys.: Condens. Matter 22 264005 [7] Saedi A, Poelsema B and Zandvliet H J W 2010 J. Phys.: Condens. Matter 22 264007 [8] Sloan P A 2010 J. Phys.: Condens. Matter 22 264001 [9] Jewell A D, Tierney H L, Baber A E, Iski E V, Laha M M and Sykes E C H 2010 J. Phys.: Condens. Matter 22 264006 [10] Riedel D 2010 J. Phys.: Condens. Matter 22 264009 [11] Terada Y, Yoshida S, Takeuchi O and Shigekawa H 2010 J. Phys.: Condens. Matter 22 264008

  10. High-speed detector for time-resolved diffraction studies

    PubMed Central

    Singh, Bipin; Miller, Stuart R.; Bhandari, Harish B.; Graceffa, Rita; Irving, Thomas C.; Nagarkar, Vivek V.

    2013-01-01

    There are a growing number of high brightness synchrotron sources that require high-frame-rate detectors to provide the time-scales required for performing time-resolved diffraction experiments. We report on the development of a very high frame rate CMOS X-ray detector for time-resolved muscle diffraction and time-resolved solution scattering experiments. The detector is based on a low-afterglow scintillator, provides a megapixel resolution with frame rates of up to 120,000 frames per second, an effective pixel size of 64 µm, and can be adapted for various X-ray energies. The paper describes the detector design and initial results of time-resolved diffraction experiments on a synchrotron beamline. PMID:24489595

  11. High-speed detector for time-resolved diffraction studies

    NASA Astrophysics Data System (ADS)

    Singh, Bipin; Miller, Stuart R.; Bhandari, Harish B.; Graceffa, Rita; Irving, Thomas C.; Nagarkar, Vivek V.

    2013-03-01

    There are a growing number of high brightness synchrotron sources that require high-frame-rate detectors to provide the time-scales required for performing time-resolved diffraction experiments. We report on the development of a very high frame rate CMOS X-ray detector for time-resolved muscle diffraction and time-resolved solution scattering experiments. The detector is based on a low-afterglow scintillator, provides a megapixel resolution with frame rates of up to 120,000 frames per second, an effective pixel size of 64 um, and can be adapted for various X-ray energies. The paper describes the detector design and initial results of time-resolved diffraction experiments on a synchrotron beamline.

  12. Millifluidics for Chemical Synthesis and Time-resolved Mechanistic Studies

    PubMed Central

    Krishna, Katla Sai; Biswas, Sanchita; Navin, Chelliah V.; Yamane, Dawit G.; Miller, Jeffrey T.; Kumar, Challa S.S.R.

    2013-01-01

    Procedures utilizing millifluidic devices for chemical synthesis and time-resolved mechanistic studies are described by taking three examples. In the first, synthesis of ultra-small copper nanoclusters is described. The second example provides their utility for investigating time resolved kinetics of chemical reactions by analyzing gold nanoparticle formation using in situ X-ray absorption spectroscopy. The final example demonstrates continuous flow catalysis of reactions inside millifluidic channel coated with nanostructured catalyst. PMID:24327099

  13. Millifluidics for chemical synthesis and time-resolved mechanistic studies.

    PubMed

    Krishna, Katla Sai; Biswas, Sanchita; Navin, Chelliah V; Yamane, Dawit G; Miller, Jeffrey T; Kumar, Challa S S R

    2013-01-01

    Procedures utilizing millifluidic devices for chemical synthesis and time-resolved mechanistic studies are described by taking three examples. In the first, synthesis of ultra-small copper nanoclusters is described. The second example provides their utility for investigating time resolved kinetics of chemical reactions by analyzing gold nanoparticle formation using in situ X-ray absorption spectroscopy. The final example demonstrates continuous flow catalysis of reactions inside millifluidic channel coated with nanostructured catalyst. PMID:24327099

  14. Seventh international conference on time-resolved vibrational spectroscopy

    SciTech Connect

    Dyer, R.B.; Martinez, M.A.D.; Shreve, A.; Woodruff, W.H.

    1997-04-01

    The International Conference on Time-Resolved Vibrational Spectroscopy (TRVS) is widely recognized as the major international forum for the discussion of advances in this rapidly growing field. The 1995 conference was the seventh in a series that began at Lake Placid, New York, 1982. Santa Fe, New Mexico, was the site of the Seventh International Conference on Time-Resolved Vibrational Spectroscopy, held from June 11 to 16, 1995. TRVS-7 was attended by 157 participants from 16 countries and 85 institutions, and research ranging across the full breadth of the field of time-resolved vibrational spectroscopy was presented. Advances in both experimental capabilities for time-resolved vibrational measurements and in theoretical descriptions of time-resolved vibrational methods continue to occur, and several sessions of the conference were devoted to discussion of these advances and the associated new directions in TRVS. Continuing the interdisciplinary tradition of the TRVS meetings, applications of time-resolved vibrational methods to problems in physics, biology, materials science, and chemistry comprised a large portion of the papers presented at the conference.

  15. Latent fingerprint and trace explosives detection by photoluminescence and time-resolved imaging

    NASA Astrophysics Data System (ADS)

    Bouldin, Kimberly Kay

    Latent fingerprint detection by photoluminescence is a well-developed field. Many development techniques exist and are currently being employed in forensic laboratories to detect fingerprints by making them luminescent. However, in forensic science, time-resolved imaging techniques, designed to suppress background fluorescence that interferes with fingerprint detectability, are to date not used outside of the research laboratory, and the chemistry necessary to use time-resolved imaging for fingerprint detection is somewhat limited. For this reason, the first section of this dissertation deals with fingerprint detection methods that have direct application to time-resolved imaging techniques. Trace explosive detection field methods based on chemical reactions have until recently utilized only colorimetric products. To increase the sensitivity of such detection, a field explosive test kit which produces a product that is both colorimetric and luminescent is studied. Detection sensitivity can be gained by taking advantage of the luminescence of these products, something that has not been done to date. When the appropriate chemistry is chosen for explosive detection, time-resolved imaging techniques may again be applicable. This dissertation thus looks at possibilities of taking trace explosives detection to this next level.

  16. Time-resolved experiments in the frequency domain using synchrotron radiation (invited)

    NASA Astrophysics Data System (ADS)

    De Stasio, Gelsomina; Giusti, A. M.; Parasassi, T.; Ravagnan, G.; Sapora, O.

    1992-01-01

    PLASTIQUE is the only synchrotron radiation beam line in the world that performs time-resolved fluorescence experiments in frequency domain. These experiments are extremely valuable sources of information on the structure and the dynamics of molecules. This technique measures fluorescence lifetimes with picosecond resolution in the near UV spectral range. Such accurate measurements are rendered possible by taking phase and modulation data, and by the advantages of the cross-correlation technique. A successful experiment demonstrated the radiation damage induced by low doses of radiation on rabbit blood cell membranes.

  17. Protein-ligand interactions probed by time-resolved crystallography

    SciTech Connect

    Schmidt, M.; Ihee, H.; Pahl, R.; Srajer, V.

    2005-03-09

    Time-resolved (TR) crystallography is a unique method for determining the structures of intermediates in biomolecular reactions. The technique reached its mature stage with the development of the powerful third-generation synchrotron X-ray sources, and the advances in data processing and analysis of time-resolved Laue crystallographic data. A time resolution of 100 ps has been achieved and relatively small structural changes can be detected even from only partial reaction initiation. The remaining challenge facing the application of this technique to a broad range of biological systems is to find an efficient and rapid, system-specific method for the reaction initiation in the crystal. Other frontiers for the technique involve the continued improvement in time resolution and further advances in methods for determining intermediate structures and reaction mechanisms. The time-resolved technique, combined with trapping methods and computational approaches, holds the promise for a complete structure-based description of biomolecular reactions.

  18. Time-resolved EPR spectroscopy in a Unix environment.

    PubMed

    Lacoff, N M; Franke, J E; Warden, J T

    1990-02-01

    A computer-aided time-resolved electron paramagnetic resonance (EPR) spectrometer implemented under version 2.9 BSD Unix was developed by interfacing a Varian E-9 EPR spectrometer and a Biomation 805 waveform recorder to a PDP-11/23A minicomputer having MINC A/D and D/A capabilities. Special problems with real-time data acquisition in a multiuser, multitasking Unix environment, addressing of computer main memory for the control of hardware devices, and limitation of computer main memory were resolved, and their solutions are presented. The time-resolved EPR system and the data acquisition and analysis programs, written entirely in C, are described. Furthermore, the benefits of utilizing the Unix operating system and the C language are discussed, and system performance is illustrated with time-resolved EPR spectra of the reaction center cation in photosystem 1 of green plant photosynthesis.

  19. A time-resolved image sensor for tubeless streak cameras

    NASA Astrophysics Data System (ADS)

    Yasutomi, Keita; Han, SangMan; Seo, Min-Woong; Takasawa, Taishi; Kagawa, Keiichiro; Kawahito, Shoji

    2014-03-01

    This paper presents a time-resolved CMOS image sensor with draining-only modulation (DOM) pixels for tube-less streak cameras. Although the conventional streak camera has high time resolution, the device requires high voltage and bulky system due to the structure with a vacuum tube. The proposed time-resolved imager with a simple optics realize a streak camera without any vacuum tubes. The proposed image sensor has DOM pixels, a delay-based pulse generator, and a readout circuitry. The delay-based pulse generator in combination with an in-pixel logic allows us to create and to provide a short gating clock to the pixel array. A prototype time-resolved CMOS image sensor with the proposed pixel is designed and implemented using 0.11um CMOS image sensor technology. The image array has 30(Vertical) x 128(Memory length) pixels with the pixel pitch of 22.4um. .

  20. Alteration of time-resolved autofluorescence properties of rat aorta, induced by diabetes mellitus

    NASA Astrophysics Data System (ADS)

    Uherek, M.; Uličná, O.; Vančová, O.; Muchová, J.; Ďuračková, Z.; Šikurová, L.; Chorvát, D.

    2016-10-01

    Changes in autofluorescence properties of isolated rat aorta, induced by diabetes mellitus, were detected using time-resolved fluorescence spectroscopy with pulsed ultraviolet (UV) laser excitation. We demonstrated that time-resolved spectroscopy was able to detect changes in aorta tissues related to diabetes and unambiguously discriminate diabetic (τ 1 0.63  ±  0.05 ns, τ 2 3.66  ±  0.10 ns) samples from the control (τ 1 0.76  ±  0.03 ns, τ 2 4.48  ±  0.15 ns) group. We also report changes in the ratio of relative amplitudes of the two lifetime component in aorta tissue during diabetes, most likely related to the pseudohypoxic state with altered NADH homeostasis.

  1. Time-resolved photon emission from layered turbid media

    NASA Astrophysics Data System (ADS)

    Hielscher, Andreas H.; Liu, Hanli; Chance, Britton; Tittel, Frank K.; Jacques, Steven L.

    1996-02-01

    We present numerical and experimental results of time-resolved emission profiles from various layered turbid media. Numerical solutions determined by time-resolved Monte Carlo simulations are compared with measurements on layered-tissue phantoms made from gelatin. In particular, we show that in certain cases the effects of the upper layers can be eliminated. As a practical example, these results are used to analyze in vivo measurements on the human head. This demonstrates the influence of skin, skull, and meninges on the determination of the blood oxygenation in the brain.

  2. Time-Resolved X-Ray Crystallography of Heme Proteins

    SciTech Connect

    Srajer, Vukica; Royer, Jr., William E.

    2008-04-29

    Heme proteins, with their natural photosensitivity, are excellent systems for the application of time-resolved crystallographic methods. Ligand dissociation can be readily initiated by a short laser pulse with global structural changes probed at the atomic level by X-rays in real time. Third-generation synchrotrons provide 100-ps X-ray pulses of sufficient intensity for monitoring very fast processes. Successful application of such time-resolved crystallographic experiments requires that the structural changes being monitored are compatible with the crystal lattice. These techniques have recently permitted observing for the first time allosteric transitions in real time for a cooperative dimeric hemoglobin.

  3. Time-resolved x-ray crystallography of heme proteins

    PubMed Central

    Royer, William E.

    2012-01-01

    Heme proteins, with their natural photosensitivity, are excellent systems for the application of time-resolved crystallographic methods. Ligand dissociation can be readily initiated by a short laser pulse with global structural changes probed at the atomic level by X-rays in real time. Third generation synchrotrons provide 100ps X-ray pulses of sufficient intensity for monitoring very fast processes. Successful application of such time-resolved crystallographic experiments requires that the structural changes being monitored are compatible with the crystal lattice. These techniques have permitted observing allosteric transitions in real time for a cooperative dimeric hemoglobin. PMID:18433638

  4. Time-resolved diffuse optical spectroscopy: a differential absorption approach

    NASA Astrophysics Data System (ADS)

    Taroni, Paola; Bassi, Andrea; Spinelli, Lorenzo; Cubeddu, Rinaldo; Pifferi, Antonio

    2009-07-01

    A method was developed to estimate spectral changes of the absorption properties of turbid media from time-resolved reflectance/transmittance measurements. It was derived directly from the microscopic Beer-Lambert law, and tested against simulations and phantom measurements.

  5. A high sensitivity time-resolved microfluorimeter for real-time cell biology

    NASA Astrophysics Data System (ADS)

    Martin-Fernandez, M. L.; Tobin, M. J.; Clarke, D. T.; Gregory, C. M.; Jones, G. R.

    1996-10-01

    We describe an instrument based on the novel combination of synchrotron radiation, a high sensitivity time-resolved microfluorimeter, and a multiframe single photon counting data acquisition system. This instrument has been designed specifically to measure kinetic events in live cells using fluorescence resonance energy transfer, and is capable of rapidly collecting multiple consecutive decay profiles from a small number of fluorophores. The low irradiance on the samples (<10 mW/cm2) greatly reduces probe photobleaching and specimen photodamage during prolonged exposures. The Daresbury Synchrotron Radiation Source provides fully wavelength tunable light pulses that have a full width half-maximum of 160 ps at a repetition rate of 3.125 MHz, with the high temporal stability required for continuous measurements over periods of hours. A very low limit of detection (<104 molecules/mW/cm2) is accomplished by combining a high-gain single photon counting detection system with a low fluorescence background optical layout. The latter is achieved by the inclusion of collimating optics, a reflecting objective, and a specially designed beam stop situated in the epi-fluorescence light-path. A typical irradiance of 8 mW/cm2 on a sample of ˜105 fluorescein molecules gives, in under 20 s, a fluorescence decay profile with a peak height of 104 counts, over 400 channels, and a signal to background ratio better than 40. The data acquisition system has been developed to have a real-time time-resolved fluorescence collection capability (denoted as TR2) so that fluorescence lifetime data can be continually collected throughout a changing process. To illustrate the potential of this instrument, we present the results of a TR2 experiment in which lifetime measurements of fluorescence resonance energy transfer are used to monitor the degree of clustering of epidermal growth factor receptors during endocytosis, over a period of about 1 h, with a 5 s resolution.

  6. Time-resolved Förster-resonance-energy-transfer DNA assay on an active CMOS microarray

    PubMed Central

    Schwartz, David Eric; Gong, Ping; Shepard, Kenneth L.

    2008-01-01

    We present an active oligonucleotide microarray platform for time-resolved Förster resonance energy transfer (TR-FRET) assays. In these assays, immobilized probe is labeled with a donor fluorophore and analyte target is labeled with a fluorescence quencher. Changes in the fluorescence decay lifetime of the donor are measured to determine the extent of hybridization. In this work, we demonstrate that TR-FRET assays have reduced sensitivity to variances in probe surface density compared with standard fluorescence-based microarray assays. Use of an active array substrate, fabricated in a standard complementary metal-oxide-semiconductor (CMOS) process, provides the additional benefits of reduced system complexity and cost. The array consists of 4096 independent single-photon avalanche diode (SPAD) pixel sites and features on-chip time-to-digital conversion. We demonstrate the functionality of our system by measuring a DNA target concentration series using TR-FRET with semiconductor quantum dot donors. PMID:18515059

  7. The analysis of time-resolved optical waveguide absorption spectroscopy based on positive matrix factorization.

    PubMed

    Liu, Ping; Li, Zhu; Li, Bo; Shi, Guolong; Li, Minqiang; Yu, Daoyang; Liu, Jinhuai

    2013-08-01

    Time-resolved optical waveguide absorption spectroscopy (OWAS) makes use of an evanescent field to detect the polarized absorption spectra of sub-monomolecular adlayers. This technique is suitable for the investigation of kinetics at the solid/liquid interface of dyes, pigments, fluorescent molecules, quantum dots, metallic nanoparticles, and proteins with chromophores. In this work, we demonstrate the application of positive matrix factorization (PMF) to analyze time-resolved OWAS for the first time. Meanwhile, PCA is researched to compare with PMF. The absorption/desorption kinetics of Rhodamine 6G (R6G) onto a hydrophilic glass surface and the dynamic process of Meisenheimer complex between Cysteine and TNT are selected as samples to verify experimental system and analytical methods. The results are shown that time-resolved OWAS can well record the absorption/desorption of R6G onto a hydrophilic glass surface and the dynamic formation process of Meisenheimer complexes. The feature of OWAS extracted by PMF is dynamic and consistent with the results analyzed by the traditional function of time/wavelength-absorbance. Moreover, PMF prevents the negative factors from occurring, avoids contradicting physical reality, and makes factors more easily interpretable. Therefore, we believe that PMF will provide a valuable analysis route to allow processing of increasingly large and complex data sets.

  8. Kerr-gated time-resolved Raman spectroscopy of equine cortical bone tissue.

    PubMed

    Morris, Michael D; Matousek, Pavel; Towrie, Michael; Parker, Anthony W; Goodship, Allen E; Draper, Edward R C

    2005-01-01

    Picosecond time-resolved Raman spectroscopy in equine cortical bone tissue is demonstrated. Using 400-nm pulsed laser excitation (1 ps at 1 kHz) it is shown that Kerr cell gating with a 4-ps window provides simultaneously time-resolved rejection of fluorescence and time-resolved Raman scatter enabling depth profiling through tissue. The Raman shifts are the same as those observed by conventional cw Raman spectroscopy using deep-red or near-infrared lasers. The time decay of Raman photons is shown to fit an inverse square root of time function, suggesting propagation by a diffusive mechanism. Using polystyrene behind a bone specimen, it is shown that the 400-nm laser light penetrates at least 0.31 mm below the surface of a fully mineralized bone tissue specimen and generates observable bone Raman scatter (approximately 415 to 430 nm) through most of this depth. These novel results demonstrate great promise for in vivo applications for studying diseased bone tissue, and ways to optimize the setup are discussed.

  9. New time-resolved micro-photoluminescence spectroscopy of natural and synthetic analogue minerals

    NASA Astrophysics Data System (ADS)

    Panczer, G.; Ollier, N.; Champagnon, B.; Gaft, M.

    2003-04-01

    Minerals as well as geomaterials often present light emissions under UV or visible excitations. This property called photoluminescence is due to low concentration impurities such as the rare earths, the transition elements and the lanthanides. The induced color is used for ore prospection but only spectroscopic analyses indicate the nature of the emitted centers. However natural samples contained numerous luminescent centers simultaneously and with regular steady-state measurements (such as in cathodoluminescence) all the emissions are often over lapping. In order to record the contributions of each separate center, it is possible to use time-resolved measurements based on the decay time of the emissions and using pulsed laser excitation. Some characteristic examples will be presented on apatites, zircons as well as gemstones. Geomaterials present as well micro scale heterogeneities (growth zoning, inclusions, devitrification, microphases...). Precise identification and optical effects of such heterogeneities have to be taken into account. To reach the microscale using photo luminescence studies, a microscope has be modified to allowed pulsed laser injection (from UV to visible), beam focus with micro scale resolution on the sample (<10 μm), as well as time resolved collection of micro fluorescence. Such equipment allows now undertaking time-resolved measurements of microphases. Applications on geomaterials will be presented.

  10. Time-resolved spatially offset Raman spectroscopy for depth analysis of diffusely scattering layers.

    PubMed

    Iping Petterson, Ingeborg E; Dvořák, Patrick; Buijs, Joost B; Gooijer, Cees; Ariese, Freek

    2010-12-01

    The objective of this study is to use time-resolved (TR) Raman spectroscopy, spatially offset Raman spectroscopy (SORS), and a combination of these approaches to obtain high quality Raman spectra from materials hidden underneath an opaque layer. Both TR Raman and SORS are advanced techniques that allow for an increased relative selectivity of photons from deeper layers within a sample. Time-resolved detection reduces fluorescence background, and the selectivity for the second layer is improved. By combining this with spatially offset excitation we additionally increased selectivity for deeper layers. Test samples were opaque white polymer blocks of several mm thicknesses. Excitation was carried out with a frequency-doubled Ti:sapphire laser at 460 nm, 3 ps pulse width and 76 MHz repetition rate. Detection was either with a continuous-wave CCD camera or in time-resolved mode using an intensified CCD camera with a 250 ps gate width. The Raman photons were collected in backscatter mode, with or without lateral offset. By measuring the delay of the Raman signal from the second layer (polyethylene terephthalate/PET/Arnite), the net photon migration speeds through Teflon, polythene, Delrin and Nylon were determined. Raman spectra could be obtained from a second layer of PET through Teflon layers up to 7 mm of thickness. The ability to obtain chemical information through layers of diffusely scattering materials has powerful potential for biomedical applications.

  11. Heterodyne-detected and ultrafast time-resolved second-harmonic generation for sensitive measurements of charge transfer.

    PubMed

    Wilcox, Daniel E; Sykes, Matthew E; Niedringhaus, Andrew; Shtein, Max; Ogilvie, Jennifer P

    2014-07-15

    In organic photovoltaics many key ultrafast processes occur at the interface between electron donor and acceptor molecules. Traditional ultrafast spectroscopies, such as pump-probe or time-resolved fluorescence, are not ideal for studying the interface because most of their signal is from the bulk material. Time-resolved second-harmonic generation (TRSHG) spectroscopy solves this problem by only generating signal from the interface. We demonstrate an optically heterodyned TRSHG to reduce the impact of stray light, enhance sensitivity, and detect the full complex signal field.

  12. Time-resolved FRET and PCT in cationic conjugated polymer/dye-labeled DNA complex

    NASA Astrophysics Data System (ADS)

    Kim, Inhong; Kim, Jihoon; Kim, Bumjin; Kang, Mijeong; Woo, Han Young; Kyhm, Kwangseuk

    2011-12-01

    The energy transfer mechanism between cationic conjugated polyelectrolytes and a single stranded DNA labeled with fluorescein was investigated in terms of Förster resonance energy transfer (FRET) and photo-induced charge transfer (PCT) by time-resolved fluorescence. Both FRET and PCT rate efficiencies were obtained by phenomenological coupled rate equations, which are in excellent agreement with experiments. We found the total energy transfer in the complex is maximized as a consequence of FRET and PCT at an optimum distance 32.7Å.

  13. Exploiting time-resolved magnetic field effects for determining radical ion reaction rates

    NASA Astrophysics Data System (ADS)

    Bessmertnykh, A. O.; Borovkov, V. I.; Bagryansky, V. A.; Molin, Yu N.

    2016-07-01

    The capabilities of the method of time-resolved magnetic field effect in determining the rates of charge transfer reactions between radical ions and molecules on a nanosecond time scale have been investigated. The approach relies on the electron spin coherence in radical pair's partners generated by ionizing radiation. The spin evolution of the pair is sensitive to the reaction since the latter results in changing magnetic interactions of the unpaired electron. This process can be monitored by magnetic-field-sensitive fluorescence from an irradiated sample that is illustrated using reactions involving alkane radical cations. The accuracy and limitations of the approach are discussed.

  14. Planetary Surface Exploration Using Time-Resolved Laser Spectroscopy on Rovers and Landers

    NASA Astrophysics Data System (ADS)

    Blacksberg, Jordana; Alerstam, Erik; Maruyama, Yuki; Charbon, Edoardo; Rossman, George

    2013-04-01

    Planetary surface exploration using laser spectroscopy has become increasingly relevant as these techniques become a reality on Mars surface missions. The ChemCam instrument onboard the Curiosity rover is currently using laser induced breakdown spectroscopy (LIBS) on a mast-mounted platform to measure elemental composition of target rocks. The RLS Raman Spectrometer is included on the payload for the ExoMars mission to be launched in 2018 and will identify minerals and organics on the Martian surface. We present a next-generation instrument that builds on these widely used techniques to provide a means for performing both Raman spectroscopy and LIBS in conjunction with microscopic imaging. Microscopic Raman spectroscopy with a laser spot size smaller than the grains of interest can provide surface mapping of mineralogy while preserving morphology. A very small laser spot size (~ 1 µm) is often necessary to identify minor phases that are often of greater interest than the matrix phases. In addition to the difficulties that can be posed by fine-grained material, fluorescence interference from the very same material is often problematic. This is particularly true for many of the minerals of interest that form in environments of aqueous alteration and can be highly fluorescent. We use time-resolved laser spectroscopy to eliminate fluorescence interference that can often make it difficult or impossible to obtain Raman spectra. As an added benefit, we have found that with small changes in operating parameters we can include microscopic LIBS using the same hardware. This new technique relies on sub-ns, high rep-rate lasers with relatively low pulse energy and compact solid state detectors with sub-ns time resolution. The detector technology that makes this instrument possible is a newly developed Single-Photon Avalanche Diode (SPAD) sensor array based on Complementary Metal-Oxide Semiconductor (CMOS) technology. The use of this solid state time-resolved detector offers a

  15. Time resolved FRET measurement in various heterogeneous media using merocyanine dye as a donor

    NASA Astrophysics Data System (ADS)

    Kedia, Niraja; Bagchi, Sanjib

    2015-06-01

    Ultrafast fluorescence resonance energy transfer (FRET) from a merocyanine dye to a Rhodamine 6G (R6G) molecule in micelles formed by the surfactants SDS and DTAB and also in a catanionic vesicle formed by SDS and DTAB has been studied by picosecond time resolved emission spectroscopy. Here the dye acts as a donor molecule and R6G acts as the acceptor molecule. Multiple timescales of FRET have been detected, namely, an ultrafast component of 100-500 ps and relatively long component (1800-3300 ps). The different time scales are attributed to different donor-acceptor distances.

  16. Time resolved FRET measurement in various heterogeneous media using merocyanine dye as a donor.

    PubMed

    Kedia, Niraja; Bagchi, Sanjib

    2015-06-15

    Ultrafast fluorescence resonance energy transfer (FRET) from a merocyanine dye to a Rhodamine 6G (R6G) molecule in micelles formed by the surfactants SDS and DTAB and also in a catanionic vesicle formed by SDS and DTAB has been studied by picosecond time resolved emission spectroscopy. Here the dye acts as a donor molecule and R6G acts as the acceptor molecule. Multiple timescales of FRET have been detected, namely, an ultrafast component of 100-500 ps and relatively long component (1800-3300 ps). The different time scales are attributed to different donor-acceptor distances.

  17. Application of time-resolved luminescence spectroscopy to a remote uranyl sensor

    NASA Astrophysics Data System (ADS)

    Varineau, Pierre T.; Duesing, Richard W., Jr.; Wangen, Larry E.

    1992-03-01

    Time-resolved luminescence spectroscopy is an effective method for the determination of a wide range of uranyl concentrations in aqueous samples. We have applied this technique to the development of a remote-sensing device using fiber optic cables coupled with a microflow cell to probe for uranyl in aqueous samples. This sensor incorporates a Nafion membrane through which UO22+ can diffuse into a reaction/analysis chamber containing phosphoric acid, a reagent that enhances the uranyl luminescence intensity and lifetime. With this device, anionic and fluorescing organic interferences could be eliminated, allowing for the determination of uranyl over a concentration range of 10-4 to 10-9 M.

  18. Time-Resolved Rayleigh Scattering Measurements in Hot Gas Flows

    NASA Technical Reports Server (NTRS)

    Mielke, Amy F.; Elam, Kristie A.; Sung, Chih-Jen

    2008-01-01

    A molecular Rayleigh scattering technique is developed to measure time-resolved gas velocity, temperature, and density in unseeded gas flows at sampling rates up to 32 kHz. A high power continuous-wave laser beam is focused at a point in an air flow field and Rayleigh scattered light is collected and fiber-optically transmitted to the spectral analysis and detection equipment. The spectrum of the light, which contains information about the temperature and velocity of the flow, is analyzed using a Fabry-Perot interferometer. Photomultipler tubes operated in the photon counting mode allow high frequency sampling of the circular interference pattern to provide time-resolved flow property measurements. Mean and rms velocity and temperature fluctuation measurements in both an electrically-heated jet facility with a 10-mm diameter nozzle and also in a hydrogen-combustor heated jet facility with a 50.8-mm diameter nozzle at NASA Glenn Research Center are presented.

  19. Benchtop time-resolved magneto-optical Kerr magnetometer.

    PubMed

    Barman, Anjan; Kimura, T; Otani, Y; Fukuma, Y; Akahane, K; Meguro, S

    2008-12-01

    We present here the construction and application of a compact benchtop time-resolved Kerr magnetometer to measure the magnetization precession in magnetic thin films and lithographically patterned elements. As opposed to very expensive femtosecond lasers this system is built upon a picosecond pulsed injection diode laser and electronic pulse and delay generators. The precession is triggered by the electronic pulses of controlled duration and shape, which is launched onto the sample by a microstrip line. We used polarized optical pulses synchronous to the electronic pulses to measure the magneto-optical Kerr rotation. The system is integrated in a conventional upright microscope configuration with separate illumination, imaging, and magneto-optical probe paths. The system offers high stability, relative ease of alignment, sample changing, and a long range of time delay. We demonstrate the measurements of time-resolved dynamics of a Permalloy microwire and microdot using this system, which showed dynamics at two different time scales. PMID:19123577

  20. Time resolved optical tomography of the human forearm

    NASA Astrophysics Data System (ADS)

    Hillman, Elizabeth M. C.; Hebden, Jeremy C.; Schweiger, Martin; Dehghani, Hamid; Schmidt, Florian E. W.; Delpy, David T.; Arridge, Simon R.

    2001-04-01

    A 32-channel time-resolved optical imaging instrument has been developed principally to study functional parameters of the new-born infant brain. As a prelude to studies on infants, the device and image reconstruction methodology have been evaluated on the adult human forearm. Cross-sectional images were generated using time-resolved measurements of transmitted light at two wavelengths. All data were acquired using a fully automated computer-controlled protocol. Images representing the internal scattering and absorbing properties of the arm are presented, as well as images that reveal physiological changes during a simple finger flexion exercise. The results presented in this paper represent the first simultaneous tomographic reconstruction of the internal scattering and absorbing properties of a clinical subject using purely temporal data, with additional co-registered difference images showing repeatable absorption changes at two wavelengths in response to exercise.

  1. Theory of time-resolved inelastic x-ray diffraction

    SciTech Connect

    Lorenz, Ulf; Moeller, Klaus B.; Henriksen, Niels E.

    2010-02-15

    Starting from a general theory of time-resolved x-ray scattering, we derive a convenient expression for the diffraction signal based on a careful analysis of the relevant inelastic scattering processes. We demonstrate that the resulting inelastic limit applies to a wider variety of experimental conditions than similar, previously derived formulas, and it directly allows the application of selection rules when interpreting diffraction signals. Furthermore, we present a simple extension to systems simultaneously illuminated by x rays and a laser beam.

  2. Ultrafast Time-Resolved Electron Diffraction with Megavolt Electron Beams

    SciTech Connect

    Hastings, J.B.; Rudakov, F.M.; Dowell, D.H.; Schmerge, J.F.; Cardoza, J.D.; Castro, J.M.; Gierman, S.M.; Loos, H.; Weber, P.M.; /Brown U.

    2006-10-24

    An rf photocathode electron gun is used as an electron source for ultrafast time-resolved pump-probe electron diffraction. We observed single-shot diffraction patterns from a 160 nm Al foil using the 5.4 MeV electron beam from the Gun Test Facility at the Stanford Linear Accelerator. Excellent agreement with simulations suggests that single-shot diffraction experiments with a time resolution approaching 100 fs are possible.

  3. Ultrafast time resolved vibrational spectroscopy in liquid systems

    NASA Astrophysics Data System (ADS)

    Seifert, G.; Hofmann, M.; Weidlich, K.; Graener, H.

    1996-04-01

    The ultrafast dynamics of small molecules in the liquid phase can successfully be studied tracing the relaxation pathways of vibrational excess energy. Two complementing experimental techniques, picosecond IR double resonance spectroscopy and time resolved incoherent Anti-Stokes Raman spectroscopy, are very powerful tools for such studies. The capabilities of investigations combining these methods are discussed on the example of new experimental data on liquid dichloromethane (CH2Cl2).

  4. Time-resolved photoconductivity of PbSe nanocrystal arrays.

    PubMed

    Murphy, James E; Beard, Matthew C; Nozik, Arthur J

    2006-12-21

    We report the sub-picosecond photoconductivity dynamics of chemically treated PbSe nanocrystal arrays utilizing time-resolved terahertz spectroscopy (TRTS). TRTS allows both the degree of interdot electronic coupling and the carrier dynamics to be extracted simultaneously. The following capping ligands bonded to the quantum dot surface were studied: hydrazine, ethylenediamine, butlyamine, and aniline. In addition, the arrays were treated with NaOH. We find that the treatments affect both the degree of electronic coupling and the carrier dynamics.

  5. The RATIO method for time-resolved Laue crystallography

    PubMed Central

    Coppens, Philip; Pitak, Mateusz; Gembicky, Milan; Messerschmidt, Marc; Scheins, Stephan; Benedict, Jason; Adachi, Shin-ichi; Sato, Tokushi; Nozawa, Shunsuke; Ichiyanagi, Kohei; Chollet, Matthieu; Koshihara, Shin-ya

    2009-01-01

    A RATIO method for analysis of intensity changes in time-resolved pump–probe Laue diffraction experiments is described. The method eliminates the need for scaling the data with a wavelength curve representing the spectral distribution of the source and removes the effect of possible anisotropic absorption. It does not require relative scaling of series of frames and removes errors due to all but very short term fluctuations in the synchrotron beam. PMID:19240334

  6. Time resolved optical spectra from MIG welding arc ignitions

    SciTech Connect

    Eriksen, P.

    1985-03-01

    Optical radiation from MIG (GMAW) welding arc ignitions has been measured with a rapid scan spectrometer. The time resolved spectral measurements reveal a substantial overshoot of ultraviolet radiation during the ignition phase of a 200 A aluminum arc. Calculations which follow the ACGIH guidelines show that, at a welding current of 300 A, the unprotected eye at a distance of 0.5 m may suffer a flash after the reception of radiation from only one ignition.

  7. Time-resolved MR angiography with limited projections.

    PubMed

    Huang, Yuexi; Wright, Graham A

    2007-08-01

    A method for reconstruction of time-resolved MRI called highly-constrained backprojection (HYPR) has been developed. To evaluate the HYPR reconstruction in relation to data sparsity and temporal dynamics, computer simulations were performed, investigating signal modulations under different situations that reflect dynamic contrast-enhanced MR angiography (MRA). In vivo studies were also performed with gadolinium diethylenetriamine pentaacetic acid (Gd-DTPA) for abdominal MRA in a canine model to demonstrate the application of HYPR for three-dimensional (3D) time-resolved MRA. When contrast dynamics vary over space, large vessels (e.g., veins) tend to introduce signal interference to small vessels (e.g., arteries) in HYPR, particularly when the vessels are in close proximity. The enhancement of background tissue signals may also alter the arterial and venous temporal profiles in HYPR. However, the artifacts are manifest as intensity modulation rather than structural interference, and therefore have little impact on structural diagnosis. Increasing the number of projections per time point increases temporal blur while reducing corruption of temporal behavior from adjacent tissues. Uniformly interleaved acquisition order, such as the bit-reversed order, is important to reduce artifacts. With high signal-to-noise ratio (SNR) and limited artifacts, HYPR reconstruction has potential to greatly improve time-resolved MRA in clinical practice.

  8. Time-resolved photoelectron spectroscopy: from wavepackets to observables.

    PubMed

    Wu, Guorong; Hockett, Paul; Stolow, Albert

    2011-11-01

    Time-resolved photoelectron spectroscopy (TRPES) is a powerful tool for the study of intramolecular dynamics, particularly excited state non-adiabatic dynamics in polyatomic molecules. Depending on the problem at hand, different levels of TRPES measurements can be performed: time-resolved photoelectron yield; time- and energy-resolved photoelectron yield; time-, energy-, and angle-resolved photoelectron yield. In this pedagogical overview, a conceptual framework for time-resolved photoionization measurements is presented, together with discussion of relevant theory for the different aspects of TRPES. Simple models are used to illustrate the theory, and key concepts are further amplified by experimental examples. These examples are chosen to show the application of TRPES to the investigation of a range of problems in the excited state dynamics of molecules: from the simplest vibrational wavepacket on a single potential energy surface; to disentangling intrinsically coupled electronic and nuclear motions; to identifying the electronic character of the intermediate states involved in non-adiabatic dynamics by angle-resolved measurements in the molecular frame, the most complete measurement.

  9. [System of ns time-resolved spectroscopy diagnosis and radioprotection].

    PubMed

    Yao, Wei-Bo; Guo, Jian-Ming; Zhang, Yong-min; Tang, Jun-Ping; Cheng, Liang; Xu, Qi-fuo

    2014-06-01

    Cathode plasma of high current electron beam diode is an important research on high power microwave and strong pulsed radio accelerator. It is a reliable method to study cathode plasma by diagnosing the cathode plasma parameters with non-contact spectroscopy measurement system. The present paper introduced the work principle, system composition and performance of the nanosecond (ns) time-resolved spectroscopy diagnosis system. Furthermore, it introduced the implementing method and the temporal relation of lower jitter synchronous trigger system. Simultaneously, the authors designed electromagnetic and radio shield room to protect the diagnosis system due to the high electromagnetic and high X-ray and γ-ray radiation, which seriously interferes with the system. Time-resolved spectroscopy experiment on brass (H62) cathode shows that, the element and matter composition of cathode plasma is clearly increase with the increase in the diode pulsed voltage and current magnitude. The spectroscopy diagnosis system could be of up to 10 ns time resolve capability. It's least is 2 ns. Synchronous trigger system's jitter is less than 4 ns. The spectroscopy diagnosis system will open a new way to study the cathode emission mechanism in depth. PMID:25358142

  10. Time-resolved luminescence of nanocrystalline inorganic complex oxides

    NASA Astrophysics Data System (ADS)

    Pankratov, V.; Millers, D.; Grigorjeva, L.; Lojkowski, W.; Kareiva, A.

    2007-12-01

    Two types of complex nanosized oxides - cerium doped Y3Al5O12 (YAG) and CaWO4- have been studied by means of time-resolved luminescence spectroscopy. Comparative study of time-resolve luminescence characteristics of cerium doped YAG single crystal, nanopowders and nanoceramic as well as for CaWO4 macro- and nanocrystals has been done. Two components in the decay kinetic of Ce3+ related emission in YAG nanocrystals were detected and it was suggested that a different energy transfer rate to volume and surface Ce3+ ions takes place. It is shown that the segregation of Ce3+ ions near nanoparticles surface and/or dislocation lines plays a crucial role in degradation of light yield of cerium related luminescence in YAG nanocrystals. Time-resolved properties of sol-gel synthesized CaWO4 nanocrystals depends strongly on the synthesis rout. It was shown that shallow traps have a strong influence on the luminescence decay times of nanosized CaWO4.

  11. Studies of Minerals, Organic and Biogenic Materials through Time-Resolved Raman Spectroscopy

    NASA Technical Reports Server (NTRS)

    Garcia, Christopher S.; Abedin, M. Nurul; Ismail, Syed; Sharma, Shiv K.; Misra, Anupam K.; Nyugen, Trac; Elsayed-Ali, hani

    2009-01-01

    A compact remote Raman spectroscopy system was developed at NASA Langley Research center and was previously demonstrated for its ability to identify chemical composition of various rocks and minerals. In this study, the Raman sensor was utilized to perform time-resolved Raman studies of various samples such as minerals and rocks, Azalea leaves and a few fossil samples. The Raman sensor utilizes a pulsed 532 nm Nd:YAG laser as excitation source, a 4-inch telescope to collect the Raman-scattered signal from a sample several meters away, a spectrograph equipped with a holographic grating, and a gated intensified CCD (ICCD) camera system. Time resolved Raman measurements were carried out by varying the gate delay with fixed short gate width of the ICCD camera, allowing measurement of both Raman signals and fluorescence signals. Rocks and mineral samples were characterized including marble, which contain CaCO3. Analysis of the results reveals the short (approx.10-13 s) lifetime of the Raman process, and shows that Raman spectra of some mineral samples contain fluorescence emission due to organic impurities. Also analyzed were a green (pristine) and a yellow (decayed) sample of Gardenia leaves. It was observed that the fluorescence signals from the green and yellow leaf samples showed stronger signals compared to the Raman lines. Moreover, it was also observed that the fluorescence of the green leaf was more intense and had a shorter lifetime than that of the yellow leaf. For the fossil samples, Raman shifted lines could not be observed due the presence of very strong short-lived fluorescence.

  12. Relationship between time-resolved and non-time-resolved Beer-Lambert law in turbid media.

    PubMed

    Nomura, Y; Hazeki, O; Tamura, M

    1997-06-01

    The time-resolved Beer-Lambert law proposed for oxygen monitoring using pulsed light was extended to the non-time-resolved case in a scattered medium such as living tissues with continuous illumination. The time-resolved Beer-Lambert law was valid for the phantom model and living tissues in the visible and near-infrared regions. The absolute concentration and oxygen saturation of haemoglobin in rat brain and thigh muscle could be determined. The temporal profile of rat brain was reproduced by Monte Carlo simulation. When the temporal profiles of rat brain under different oxygenation states were integrated with time, the absorbance difference was linearly related to changes in the absorption coefficient. When the simulated profiles were integrated, there was a linear relationship within the absorption coefficient which was predicted for fractional inspiratory oxygen concentration from 10 to 100% and, in the case beyond the range of the absorption coefficient, the deviation from linearity was slight. We concluded that an optical pathlength which is independent of changes in the absorption coefficient is a good approximation for near-infrared oxygen monitoring.

  13. A Protein L -Based Immunodiagnostic Approach Utilizing Time-Resolved Förster Resonance Energy Transfer

    PubMed Central

    Hepojoki, Satu; Nurmi, Visa; Vaheri, Antti; Hedman, Klaus; Vapalahti, Olli; Hepojoki, Jussi

    2014-01-01

    Chelated lanthanides such as europium (Eu) have uniquely long fluorescence emission half-lives permitting their use in time-resolved fluorescence (TRF) assays. In Förster resonance energy transfer (FRET) a donor fluorophore transfers its emission energy to an acceptor fluorophore if in sufficiently close proximity. The use of time-resolved (TR) FRET minimizes the autofluorescence of molecules present in biological samples. In this report, we describe a homogenous immunoassay prototype utilizing TR-FRET for detection of antibodies in solution. The assay is based on labeled protein L, a bacterial protein that binds to immunoglobulin (Ig) light chain, and labeled antigen, which upon association with the same Ig molecule produce a TR-FRET active complex. We show that the approach is functional and can be utilized for both mono- and polyvalent antigens. We also compare the assay performance to that of another homogenous TR-FRET immunoassay reported earlier. This novel assay may have wide utility in infectious disease point-of-care diagnostics. PMID:25181527

  14. Time-to-digital converter card for multichannel time-resolved single-photon counting applications

    NASA Astrophysics Data System (ADS)

    Tamborini, Davide; Portaluppi, Davide; Tisa, Simone; Tosi, Alberto

    2015-03-01

    We present a high performance Time-to-Digital Converter (TDC) card that provides 10 ps timing resolution and 20 ps (rms) timing precision with a programmable full-scale-range from 160 ns to 10 μs. Differential Non-Linearity (DNL) is better than 1.3% LSB (rms) and Integral Non-Linearity (INL) is 5 ps rms. Thanks to the low power consumption (400 mW) and the compact size (78 mm x 28 mm x 10 mm), this card is the building block for developing compact multichannel time-resolved instrumentation for Time-Correlated Single-Photon Counting (TCSPC). The TDC-card outputs the time measurement results together with the rates of START and STOP signals and the number of valid TDC conversions. These additional information are needed by many TCSPC-based applications, such as: Fluorescence Lifetime Imaging (FLIM), Time-of-Flight (TOF) ranging measurements, time-resolved Positron Emission Tomography (PET), single-molecule spectroscopy, Fluorescence Correlation Spectroscopy (FCS), Diffuse Optical Tomography (DOT), Optical Time-Domain Reflectometry (OTDR), quantum optics, etc.

  15. Time-resolved luminescence resonance energy transfer imaging of protein-protein interactions in living cells.

    PubMed

    Rajapakse, Harsha E; Miller, Lawrence W

    2012-01-01

    Lanthanide-based or luminescence resonance energy transfer (LRET) microscopy can be used to sensitively image interactions between reporter-labeled proteins in living mammalian cells. With LRET, luminescent lanthanide complexes are used as donors, conventional fluorophores are used as acceptors, and donor-sensitized acceptor emission occurs at time scales that reflect the long (~ms) lanthanide emission lifetime. These long-lived signals can be separated from short-lifetime (~ns) sample autofluorescence and directly excited acceptor fluorescence by using pulsed light to excite the specimen and by implementing a short delay (>100 ns) before detection, thereby increasing measurement sensitivity. As practical implementation of time-resolved LRET microscopy requires several potentially unfamiliar experimental techniques, we explicitly describe herein methods to label proteins in living mammalian cells with luminescent terbium complexes, image interactions between terbium-labeled proteins and green fluorescent protein fusions, and quantitatively analyze LRET images. PMID:22289461

  16. Diffusion of Drug Delivery Nanoparticles into Biogels Using Time-Resolved MicroMRI.

    PubMed

    Wang, Xiaoling; Chen, Ying; Xue, Lian; Pothayee, Nipon; Zhang, Rui; Riffle, Judy S; Reineke, Theresa M; Madsen, Louis A

    2014-11-01

    Nanoparticle-based therapeutic agents can in some cases provide selective delivery to tumors, yet this field would greatly benefit from more detailed understanding of particle transport into and within tumor tissue. To provide fundamental information for optimizing interstitial transport of polymeric nanoparticles, we have developed a quantitative approach employing real-time analysis of nanoparticle diffusion into bulk biological hydrogels using microMRI. We use two distinct imaging approaches to probe the migration of two novel "theranostic" polymeric agents (combining drug delivery and contrast agent functions) into bulk hydrogels. Theranostic agent diffusion measured using time-resolved MRI agrees well with diffusion measured for simple probe particles using fluorescence spectroscopies. Furthermore, compared with established fluorescence techniques, which are restricted by sample thickness, our approach provides a three-dimensional diffusion rate and concentration distribution of nanoparticles over macroscopic distances in biological media. These results carry implications for in vivo tracking of theranostic nanoparticles into tumor interstitium. PMID:26278755

  17. Lucas–Kanade fluid trajectories for time-resolved PIV

    NASA Astrophysics Data System (ADS)

    Yegavian, Robin; Leclaire, Benjamin; Champagnat, Frédéric; Illoul, Cédric; Losfeld, Gilles

    2016-08-01

    We introduce a new method for estimating fluid trajectories in time-resolved PIV. It relies on a Lucas–Kanade paradigm and consists in a simple and direct extension of a two-frame estimation with FOLKI-PIV (Champagnat et al 2011 Exp. Fluids 50 1169–82). The so-called Lucas–Kanade Fluid Trajectories (LKFT) are assumed to be polynomial in time, and are found as the minimizer of a global functional, in which displacements are sought so as to match the intensities of a series of images pairs in the sequence, in the least-squares sense. All pairs involve the central image, similar to other recent time-resolved approaches (FTC (Lynch and Scarano 2013 Meas. Sci. Technol. 24 035305) and FTEE (Jeon et al 2014 Exp. Fluids 55 1–16)). As switching from a two-frame to a time-resolved objective simply amounts to adding terms in a functional, no significant additional algorithmic element is required. Similar to FOLKI-PIV the method is very well suited for GPU acceleration, which is an important feature as computational complexity increases with the image sequence size. Tests on synthetic data exhibiting peak-locking show that increasing the image sequence size strongly reduces both associated bias and random error, and that LKFT has a remaining total error comparable to that of FTEE on this case. Results on case B of the third PIV challenge (Stanislas et al 2008 Exp. Fluids 45 27–71) also show its ability to drastically reduce the error in situations with low signal-to-noise ratio. These results are finally confirmed on experimental images acquired in the near-field of a low Reynolds number jet. Strong reductions in peak-locking, spatial and temporal noise compared to two-frame estimation are also observed, on the displacement components themselves, as well as on spatial or temporal derivatives, such as vorticity and material acceleration.

  18. Lucas-Kanade fluid trajectories for time-resolved PIV

    NASA Astrophysics Data System (ADS)

    Yegavian, Robin; Leclaire, Benjamin; Champagnat, Frédéric; Illoul, Cédric; Losfeld, Gilles

    2016-08-01

    We introduce a new method for estimating fluid trajectories in time-resolved PIV. It relies on a Lucas-Kanade paradigm and consists in a simple and direct extension of a two-frame estimation with FOLKI-PIV (Champagnat et al 2011 Exp. Fluids 50 1169-82). The so-called Lucas-Kanade Fluid Trajectories (LKFT) are assumed to be polynomial in time, and are found as the minimizer of a global functional, in which displacements are sought so as to match the intensities of a series of images pairs in the sequence, in the least-squares sense. All pairs involve the central image, similar to other recent time-resolved approaches (FTC (Lynch and Scarano 2013 Meas. Sci. Technol. 24 035305) and FTEE (Jeon et al 2014 Exp. Fluids 55 1-16)). As switching from a two-frame to a time-resolved objective simply amounts to adding terms in a functional, no significant additional algorithmic element is required. Similar to FOLKI-PIV the method is very well suited for GPU acceleration, which is an important feature as computational complexity increases with the image sequence size. Tests on synthetic data exhibiting peak-locking show that increasing the image sequence size strongly reduces both associated bias and random error, and that LKFT has a remaining total error comparable to that of FTEE on this case. Results on case B of the third PIV challenge (Stanislas et al 2008 Exp. Fluids 45 27-71) also show its ability to drastically reduce the error in situations with low signal-to-noise ratio. These results are finally confirmed on experimental images acquired in the near-field of a low Reynolds number jet. Strong reductions in peak-locking, spatial and temporal noise compared to two-frame estimation are also observed, on the displacement components themselves, as well as on spatial or temporal derivatives, such as vorticity and material acceleration.

  19. Time-resolved Absolute Velocity Quantification with Projections

    PubMed Central

    Langham, Michael C.; Jain, Varsha; Magland, Jeremy F.; Wehrli, Felix W.

    2010-01-01

    Quantitative information on time-resolved blood velocity along the femoral/popliteal artery can provide clinical information on peripheral arterial disease and complement MR angiography since not all stenoses are hemodynamically significant. The key disadvantages of the most widely used approach to time-resolve pulsatile blood flow by cardiac-gated velocity-encoded gradient-echo imaging are gating errors and long acquisition time. Here we demonstrate a rapid non-triggered method that quantifies absolute velocity on the basis of phase difference between successive velocity-encoded projections after selectively removing the background static tissue signal via a reference image. The tissue signal from the reference image’s center k-space line is isolated by masking out the vessels in the image domain. The performance of the technique, in terms of reproducibility and agreement with results obtained with conventional phase contrast (PC)-MRI was evaluated at 3T field strength with a variable-flow rate phantom and in vivo of the triphasic velocity waveforms at several segments along the femoral and popliteal arteries. Additionally, time-resolved flow velocity was quantified in five healthy subjects and compared against gated PC-MRI results. To illustrate clinical feasibility the proposed method was shown to be able to identify hemodynamic abnormalities and impaired reactivity in a diseased femoral artery. For both phantom and in vivo studies, velocity measurements were within 1.5 cm/s and the coefficient of variation was less than 5% in an in vivo reproducibility study. In five healthy subjects, the average differences in mean peak velocities and their temporal locations were within 1 cm/s and 10 ms compared to gated PC-MRI. In conclusion, the proposed method provides temporally-resolved arterial velocity with a temporal resolution of 20 ms with minimal post-processing. PMID:20677235

  20. Rapid high-resolution spin- and time-resolved ARPES

    NASA Astrophysics Data System (ADS)

    Lin, Chiu-Yun; Gotlieb, Kenneth; Jozwiak, Chris; Hussain, Zahid; Bostwick, Aaron; Lanzara, Alessandra; Advanced Light Source, Lawrence Berkeley National Laboratory Collaboration; Graduate Group in Applied Science; Technology, University of California, Berkeley Collaboration

    2015-03-01

    A high-efficiency spin- and angle-resolved photoemission spectroscopy (spin-ARPES) spectrometer, coupled with a lab-based 6 eV laser, will be presented in this talk. Combining time-of-flight(TOF) energy measurements with low-energy exchange scattering spin polarimetry, spin-TOF ARPES achieves unprecedented measurements of near-EF physics rapidly. In addition, the successful integration of the spectrometer with the pulsed laser system demonstrates its potential for simultaneous spin- and time-resolved ARPES with pump-probe based measurements.

  1. A compact electron gun for time-resolved electron diffraction

    SciTech Connect

    Robinson, Matthew S.; Lane, Paul D.; Wann, Derek A.

    2015-01-15

    A novel compact time-resolved electron diffractometer has been built with the primary goal of studying the ultrafast molecular dynamics of photoexcited gas-phase molecules. Here, we discuss the design of the electron gun, which is triggered by a Ti:Sapphire laser, before detailing a series of calibration experiments relating to the electron-beam properties. As a further test of the apparatus, initial diffraction patterns have been collected for thin, polycrystalline platinum samples, which have been shown to match theoretical patterns. The data collected demonstrate the focusing effects of the magnetic lens on the electron beam, and how this relates to the spatial resolution of the diffraction pattern.

  2. Time-Resolved Conformational Dynamics in Hydrocarbon Chains

    SciTech Connect

    Minitti, Michael P.; Weber, Peter M.

    2007-06-22

    Internal rotation about carbon-carbon bonds allows N,N-dimethyl-2-butanamine (DM2BA) and N,N-dimethyl-3-hexanamine (DM3HA) to assume multiple conformeric structures. We explore the equilibrium composition and dynamics between such conformeric structures using Rydberg fingerprint spectroscopy. Time constants for conformeric interconversion of DM2BA (at 1.79 eV of internal energy) are 19 and 66 ps, and for DM3HA (1.78 eV) 23 and 41 ps. For the first time, a time-resolved and quantitative view of conformational dynamics of flexible hydrocarbon molecules at high temperatures is revealed.

  3. Femtosecond time-resolved electronic relaxation dynamics in tetrathiafulvalene

    SciTech Connect

    Staedter, D.; Polizzi, L.; Thiré, N.; Mairesse, Y.; Mayer, P.; Blanchet, V.

    2015-05-21

    In the present paper, the ultrafast electronic relaxation of tetrathiafulvalene (TTF) initiated around 4 eV is studied by femtosecond time-resolved velocity-map imaging. The goal is to investigate the broad double structure observed in the absorption spectrum at this energy. By monitoring the transients of the parent cation and its fragments and by varying the pump and the probe wavelengths, two internal conversions and intramolecular vibrational relaxation are detected both on the order of a few hundred of femtoseconds. Photoelectron images permit the assignment of a dark electronic state involved in the relaxation. In addition, the formation of the dimer of TTF has been observed.

  4. Time-resolved multispectral imaging of combustion reaction

    NASA Astrophysics Data System (ADS)

    Huot, Alexandrine; Gagnon, Marc-André; Jahjah, Karl-Alexandre; Tremblay, Pierre; Savary, Simon; Farley, Vincent; Lagueux, Philippe; Guyot, Éric; Chamberland, Martin; Marcotte, Fréderick

    2015-05-01

    Thermal infrared imaging is a field of science that evolves rapidly. Scientists have used for years the simplest tool: thermal broadband cameras. This allows to perform target characterization in both the longwave (LWIR) and midwave (MWIR) infrared spectral range. Infrared thermal imaging is used for a wide range of applications, especially in the combustion domain. For example, it can be used to follow combustion reactions, in order to characterize the injection and the ignition in a combustion chamber or even to observe gases produced by a flare or smokestack. Most combustion gases such as carbon dioxide (CO2) selectively absorb/emit infrared radiation at discrete energies, i.e. over a very narrow spectral range. Therefore, temperatures derived from broadband imaging are not reliable without prior knowledge about spectral emissivity. This information is not directly available from broadband images. However, spectral information is available using spectral filters. In this work, combustion analysis was carried out using Telops MS-IR MW camera which allows multispectral imaging at a high frame rate. A motorized filter wheel allowing synchronized acquisitions on eight (8) different channels was used to provide time-resolved multispectral imaging of combustion products of a candle in which black powder has been burnt to create a burst. It was then possible to estimate the temperature by modeling spectral profile derived from information obtained with the different spectral filters. Comparison with temperatures obtained using conventional broadband imaging illustrates the benefits of time-resolved multispectral imaging for the characterization of combustion processes.

  5. Time-resolved multispectral imaging of combustion reactions

    NASA Astrophysics Data System (ADS)

    Huot, Alexandrine; Gagnon, Marc-André; Jahjah, Karl-Alexandre; Tremblay, Pierre; Savary, Simon; Farley, Vincent; Lagueux, Philippe; Guyot, Éric; Chamberland, Martin; Marcotte, Frédérick

    2015-10-01

    Thermal infrared imaging is a field of science that evolves rapidly. Scientists have used for years the simplest tool: thermal broadband cameras. These allow to perform target characterization in both the longwave (LWIR) and midwave (MWIR) infrared spectral range. Infrared thermal imaging is used for a wide range of applications, especially in the combustion domain. For example, it can be used to follow combustion reactions, in order to characterize the injection and the ignition in a combustion chamber or even to observe gases produced by a flare or smokestack. Most combustion gases, such as carbon dioxide (CO2), selectively absorb/emit infrared radiation at discrete energies, i.e. over a very narrow spectral range. Therefore, temperatures derived from broadband imaging are not reliable without prior knowledge of spectral emissivity. This information is not directly available from broadband images. However, spectral information is available using spectral filters. In this work, combustion analysis was carried out using a Telops MS-IR MW camera, which allows multispectral imaging at a high frame rate. A motorized filter wheel allowing synchronized acquisitions on eight (8) different channels was used to provide time-resolved multispectral imaging of combustion products of a candle in which black powder has been burnt to create a burst. It was then possible to estimate the temperature by modeling spectral profiles derived from information obtained with the different spectral filters. Comparison with temperatures obtained using conventional broadband imaging illustrates the benefits of time-resolved multispectral imaging for the characterization of combustion processes.

  6. Time-resolved biofilm deformation measurements using optical coherence tomography.

    PubMed

    Blauert, Florian; Horn, Harald; Wagner, Michael

    2015-09-01

    The interaction of shear stress with the biofilm leads to a dynamic deformation, which is related to the structural and material characteristics of biofilms. We show how optical coherence tomography can be used as an imaging technique to investigate the time-resolved deformation on the biofilm mesoscale as well as to estimate mechanical properties of the biofilm. For the first time time-resolved deformation from cross-sectional views of the inner biofilm structure could be shown. Changes in the biofilm structure and rheological properties were calculated from cross sections in real-time and time-lapsed measurements. Heterotrophic biofilms were grown in a flow cell set-up at low shear stress of τw  = 0.01 Pa. By applying higher shear stress elastic and viscoelastic behavior of biofilms were quantified. Deformation led to a change in biofilm conformation and allowed to estimate rheological properties. Assuming an ideal wall shear stress calculation, the shear modulus G = 29.7 ± 1.7 Pa and the Young's modulus E = 36.0 ± 2.6 Pa were estimated.

  7. Formulation for Time-resolved Aerodynamic Damping in Dynamic Stall

    NASA Astrophysics Data System (ADS)

    Corke, Thomas; Bowles, Patrick; Coleman, Dusty; Thomas, Flint

    2012-11-01

    A new Hilbert transform formulation of the equation of motion for a pitching airfoil in a uniform stream yields a time resolved aerodynamic damping factor, Ξ (t) = (√{ (Cm2 (t) +C m2 } /αmax) sinψ (t) , where Cm (t) is the instantaneous pitch moment coefficient, and C m (t) is the Hilbert transform of Cm (t) , αmax is the pitching amplitude, and ψ (t) is the time-resolved phase difference between the aerodynamic pitch moment and the instantaneous angle of attack. A Ξ (t) < 0 indicates unstable pressure loading that can be considered a necessary condition to excite stall flutter in an elastic airfoil. This will be illustrated in experiments with conditions producing ``light'' dynamic stall for a range of Mach numbers from 0.3-0.6. These reveal large negative excursions of Ξ (t) during the pitch-up portion of the cycle that correlates with the formation and convection of the dynamic stall vortex. The fact that the cycle-integrated damping coefficient is positive in all these cases underscores how the traditional diagnostic masks much of the physics that underlies the destabilizing effect of the dynamic stall process. This new insight can explain instances of transient limit-cycle growth of helicopter rotor vibrations. Supported by Bell Helicopter.

  8. Time Resolved FTIR Analysis of Tailpipe Exhaust for Several Automobiles

    NASA Astrophysics Data System (ADS)

    White, Allen R.; Allen, James; Devasher, Rebecca B.

    2011-06-01

    The automotive catalytic converter reduces or eliminates the emission of various chemical species (e.g. CO, hydrocarbons, etc.) that are the products of combustion from automobile exhaust. However, these units are only effective once they have reached operating temperature. The design and placement of catalytic converters has changed in order to reduce both the quantity of emissions and the time that is required for the converter to be effective. In order to compare the effectiveness of catalytic converters, time-resolved measurements were performed on several vehicles, including a 2010 Toyota Prius, a 2010 Honda Fit, a 1994 Honda Civic, and a 1967 Oldsmobile 442 (which is not equipped with a catalytic converter but is used as a baseline). The newer vehicles demonstrate bot a reduced overall level of CO and hydrocarbon emissions but are also effective more quickly than older units. The time-resolved emissions will be discussed along with the impact of catalytic converter design and location on the measured emissions.

  9. Time-resolved luminescent lateral flow assay technology.

    PubMed

    Song, Xuedong; Knotts, Michael

    2008-09-26

    We here report a detection technology that integrates highly sensitive time-resolved luminescence technique into lateral flow assay platform to achieve excellent detection performance with low cost. We have developed very bright, surface-functionalized and mono-dispersed phosphorescent nanoparticles of long lifetime under ambient conditions. The phosphorescent nanoparticles have been used to conjugate with monoclonal antibody for C-reactive protein (CRP), an inflammatory biomarker. Lateral flow immunoassay devices have been developed using the conjugate for highly sensitive detection of CRP. The CRP assay can achieve a detection sensitivity of <0.2 ngmL(-1) in serum with a linear response from 0.2 to 200 ngmL(-1) CRP. We have also developed a low cost time-resolved luminescence reader for the lateral flow immunoassay (LFIA) devices. The reader does not use expensive band pass filter and still provide very low detection background and high detection sensitivity on solid substrates such as nitrocellulose membranes. The reader can detect less than 2.5 ng phosphorescent particles captured on a nitrocellulose membrane strip with more than three orders of magnitude linear detection dynamic range. The technology should find a number of applications, ranging from clinical diagnostics, detection of chemical and biological warfare agents, to food and environmental monitoring. PMID:18790120

  10. Time-resolved spectroscopy of the Mercury 6 3P1 state

    NASA Technical Reports Server (NTRS)

    Halstead, J. A.; Reeves, R. R.

    1981-01-01

    The time-resolved fluorescence was observed from the Hg 6 3P1 state under the influence of the earth's magnetic field and with applied fields of up to 14 G. Modulation of the fluorescence decay signal was observed as a function of both time and space and can be interpreted in terms of a classical precession of the excited atom about the magnetic field or as quantum beats resulting from interference between coherently populated Zeeman sublevels. This modulation was studied for each of the five resolvable components of the hyperfine structure separately. The fluorescence from the even isotopes was determined to be almost completely modulated while the fluorescence from the odd isotopes was only partially modulated. The frequency of modulation of the fluorescence from the mercury-202 isotope was observed as a function of the applied magnetic field and a value for the Lande factor of 1.46 + or - 0.03 was obtained. This is within experimental error of the accepted value of 1.486. In addition, the frequency of modulation as a function of applied magnetic field was determined for each of the three resolvable components with more than one contributing isotopic hyperfine line. An investigation of the effect of radiation trapping on the degree modulation was also made.

  11. Ultrafast time-resolved spectroscopy of the light-harvesting complex 2 (LH2) from the photosynthetic bacterium Thermochromatium tepidum

    SciTech Connect

    Niedzwiedzki, Dariusz M.; Fuciman, Marcel; Kobayashi, Masayuki; Frank, Harry A.; Blankenship, Robert E.

    2011-10-08

    The light-harvesting complex 2 from the thermophilic purple bacterium Thermochromatium tepidum was purified and studied by steady-state absorption and fluorescence, sub-nanosecond-time-resolved fluorescence and femtosecond time-resolved transient absorption spectroscopy. The measurements were performed at room temperature and at 10 K. The combination of both ultrafast and steady-state optical spectroscopy methods at ambient and cryogenic temperatures allowed the detailed study of carotenoid (Car)-to-bacteriochlorophyll (BChl) as well BChl-to-BChl excitation energy transfer in the complex. The studies show that the dominant Cars rhodopin (N = 11) and spirilloxanthin (N = 13) do not play a significant role as supportive energy donors for BChl a. This is related with their photophysical properties regulated by long π-electron conjugation. On the other hand, such properties favor some of the Cars, particularly spirilloxanthin (N = 13) to play the role of the direct quencher of the excited singlet state of BChl.

  12. Time-Resolved Nucleic Acid Hybridization Beacons Utilizing Unimolecular and Toehold-Mediated Strand Displacement Designs.

    PubMed

    Massey, Melissa; Ancona, Mario G; Medintz, Igor L; Algar, W Russ

    2015-12-01

    Nucleic acid hybridization probes are sought after for numerous assay and imaging applications. These probes are often limited by the properties of fluorescent dyes, prompting the development of new probes where dyes are paired with novel or nontraditional luminescent materials. Luminescent terbium complexes are an example of such a material, and these complexes offer several unique spectroscopic advantages. Here, we demonstrate two nonstem-loop designs for light-up nucleic acid hybridization beacons that utilize time-resolved Förster resonance energy transfer (TR-FRET) between a luminescent Lumi4-Tb cryptate (Tb) donor and a fluorescent reporter dye, where time-resolved emission from the dye provides an analytical signal. Both designs are based on probe oligonucleotides that are labeled at their opposite termini with Tb and a fluorescent reporter dye. In one design, a probe is partially blocked with a quencher dye-labeled oligonucleotide, and target hybridization is signaled through toehold-mediated strand displacement and loss of a competitive FRET pathway. In the other design, the intrinsic folding properties of an unblocked probe are utilized in combination with a temporal mechanism for signaling target hybridization. This temporal mechanism is based on a recently elucidated "sweet spot" for TR-FRET measurements and exploits distance control over FRET efficiencies to shift the Tb lifetime within or outside the time-gated detection window for measurements. Both the blocked and unblocked beacons offer nanomolar (femtomole) detection limits, response times on the order of minutes, multiplexing through the use of different reporter dyes, and detection in complex matrices such as serum and blood. The blocked beacons offer better mismatch selectivity, whereas the unblocked beacons are simpler in design. The temporal mechanism of signaling utilized with the unblocked beacons also plays a significant role with the blocked beacons and represents a new and effective

  13. Pose estimation using time-resolved inversion of diffuse light.

    PubMed

    Raviv, Dan; Barsi, Christopher; Naik, Nikhil; Feigin, Micha; Raskar, Ramesh

    2014-08-25

    We present a novel approach for evaluation of position and orientation of geometric shapes from scattered time-resolved data. Traditionally, imaging systems treat scattering as unwanted and are designed to mitigate the effects. Instead, we show here that scattering can be exploited by implementing a system based on a femtosecond laser and a streak camera. The result is accurate estimation of object pose, which is a fundamental tool in analysis of complex scenarios and plays an important role in our understanding of physical phenomena. Here, we experimentally show that for a given geometry, a single incident illumination point yields enough information for pose estimation and tracking after multiple scattering events. Our technique can be used for single-shot imaging behind walls or through turbid media.

  14. Multidimensional Time-Resolved Spectroscopy of Vibrational Coherence in Biopolyenes

    NASA Astrophysics Data System (ADS)

    Buckup, Tiago; Motzkus, Marcus

    2014-04-01

    Multidimensional femtosecond time-resolved vibrational coherence spectroscopy allows one to investigate the evolution of vibrational coherence in electronic excited states. Methods such as pump-degenerate four-wave mixing and pump-impulsive vibrational spectroscopy combine an initial ultrashort laser pulse with a nonlinear probing sequence to reinduce vibrational coherence exclusively in the excited states. By carefully exploiting specific electronic resonances, one can detect vibrational coherence from 0 cm-1 to over 2,000 cm-1 and map its evolution. This review focuses on the observation and mapping of high-frequency vibrational coherence for all-trans biological polyenes such as β-carotene, lycopene, retinal, and retinal Schiff base. We discuss the role of molecular symmetry in vibrational coherence activity in the S1 electronic state and the interplay of coupling between electronic states and vibrational coherence.

  15. Time-resolved molecular transport across living cell membranes.

    PubMed

    Zeng, Jia; Eckenrode, Heather M; Dounce, Susan M; Dai, Hai-Lung

    2013-01-01

    It is shown that the nonlinear optical phenomenon known as second-harmonic generation can be used for label-free, time-resolved study of the transport of molecules through living cell membranes. The adsorption and transport of a 300-Da molecular-mass hydrophobic ion at the Escherichia coli membrane is observed. Remarkably, at low ion concentrations, the second-harmonic generation technique clearly exposes a multistep molecular transport process: Transport of the molecular ion across the outer and cytoplasmic membranes of the Gram-negative bacteria is recorded, in sequence, in time. Fitting of the data to a multiprocess kinematic model reveals that the transport of this hydrophobic ion through the outer membrane is much faster than through the cytoplasmic membrane, likely reflecting the effectiveness of ion transport porins. The observations illustrate an experimental means for studying the interactions of small molecules with cell membranes.

  16. FXR LIA Optimization - Time-resolved OTR Emittance Measurement

    SciTech Connect

    Jacob, J; Ong, M; Wargo, P; LeSage, G

    2005-07-21

    The Flash X-Ray Radiography (FXR) facility at Lawrence Livermore National Laboratory utilizes a high current, long pulse linear induction accelerator to produce high doses of x-ray radiation. Accurate characterization of the transverse beam emittance is required in order to facilitate accelerator modeling and tuning efforts and, ultimately, to optimize the final focus spot size, yielding higher resolution radiographs. In addition to conventional magnet scan, pepper-pot, and multiple screen techniques, optical transition radiation (OTR) has been proven as a useful emittance measurement diagnostic and is particularly well suited to the FXR accelerator. We shall discuss the time-resolved emittance characterization of an induction linac electron beam using OTR, and we will present our experimental apparatus and analysis software. We shall also develop the theoretical background of beam emittance and transition radiation.

  17. Spectral characteristics of time resolved magnonic spin Seebeck effect

    SciTech Connect

    Etesami, S. R.; Chotorlishvili, L.; Berakdar, J.

    2015-09-28

    Spin Seebeck effect (SSE) holds promise for new spintronic devices with low-energy consumption. The underlying physics, essential for a further progress, is yet to be fully clarified. This study of the time resolved longitudinal SSE in the magnetic insulator yttrium iron garnet concludes that a substantial contribution to the spin current stems from small wave-vector subthermal exchange magnons. Our finding is in line with the recent experiment by S. R. Boona and J. P. Heremans [Phys. Rev. B 90, 064421 (2014)]. Technically, the spin-current dynamics is treated based on the Landau-Lifshitz-Gilbert equation also including magnons back-action on thermal bath, while the formation of the time dependent thermal gradient is described self-consistently via the heat equation coupled to the magnetization dynamics.

  18. Time-resolved air monitoring using Fourier absorption spectroscopy

    SciTech Connect

    Biermann, H.W.

    1995-12-31

    Two categories where spectroscopic techniques excel are the capabilities to perform air analyses in situ and to obtain data at very high time resolutions. Because of these features, the Department of Pesticide Regulation augmented its extensive air monitoring capabilities with a Fourier transform infrared (FTIR) spectrometer using open-path optical systems for time resolved ambient air monitoring. A description of the instrumentation and the data analysis procedures will be presented based on two data sets obtained with this FTIR system. In one case, a 100 m folded optical path was used to measure methyl bromide concentrations after fumigation in a warehouse with a time resolution of 15 min and a detection limit of 0.2 ppm. And trying to assess the capability of this FTIR spectrometer to determine flux, water vapor concentrations were measured with a four-meter path length at a time resolution of 0.6 seconds.

  19. Time-resolved thermal transport in compositionally modulated metal films

    SciTech Connect

    Clemens, B.M.; Eesley, G.L.; Paddock, C.A.

    1988-01-15

    We report on investigations of one-dimensional thermal transport in compositionally modulated metal films produced with a systematic variation in atomic lattice mismatch. In the case of Ni-Cu, Ni-Mo, Ni-Ti, and Ni-Zr, we observe the relative effects of interfacial disorder on thermal diffusion. Our observations demonstrate the thermal impedance of a single metal-metal interface and indicate that thermal diffusion in a bilayer film is strongly influenced by the interface between contacting metal pairs. This study is made possible by picosecond time-resolved thermoreflectance measurements which probe thermal transport perpendicular to the film plane. This technique can impact on our understanding of electron scattering and transport across metallic boundaries, and it provides a means of inferring electrical transport properties.

  20. Revealing Deactivation Pathways Hidden in Time-Resolved Photoelectron Spectra

    PubMed Central

    Ruckenbauer, Matthias; Mai, Sebastian; Marquetand, Philipp; González, Leticia

    2016-01-01

    Time-resolved photoelectron spectroscopy is commonly employed with the intention to monitor electronic excited-state dynamics occurring in a neutral molecule. With the help of theory, we show that when excited-state processes occur on similar time scales the different relaxation pathways are completely obscured in the total photoionization signal recorded in the experiment. Using non-adiabatic molecular dynamics and Dyson norms, we calculate the photoionization signal of cytosine and disentangle the transient contributions originating from the different deactivation pathways of its tautomers. In the simulations, the total signal from the relevant keto and enol tautomers can be decomposed into contributions either from the neutral electronic state populations or from the distinct mechanistic pathways across the multiple potential surfaces. The lifetimes corresponding to these contributions cannot be extracted from the experiment, thereby illustrating that new experimental setups are necessary to unravel the intricate non-adiabatic pathways occurring in polyatomic molecules after irradiation by light. PMID:27762396

  1. Photon-Counting Arrays for Time-Resolved Imaging.

    PubMed

    Antolovic, I Michel; Burri, Samuel; Hoebe, Ron A; Maruyama, Yuki; Bruschini, Claudio; Charbon, Edoardo

    2016-01-01

    The paper presents a camera comprising 512 × 128 pixels capable of single-photon detection and gating with a maximum frame rate of 156 kfps. The photon capture is performed through a gated single-photon avalanche diode that generates a digital pulse upon photon detection and through a digital one-bit counter. Gray levels are obtained through multiple counting and accumulation, while time-resolved imaging is achieved through a 4-ns gating window controlled with subnanosecond accuracy by a field-programmable gate array. The sensor, which is equipped with microlenses to enhance its effective fill factor, was electro-optically characterized in terms of sensitivity and uniformity. Several examples of capture of fast events are shown to demonstrate the suitability of the approach. PMID:27367697

  2. Multidimensional time-resolved spectroscopy of vibrational coherence in biopolyenes.

    PubMed

    Buckup, Tiago; Motzkus, Marcus

    2014-01-01

    Multidimensional femtosecond time-resolved vibrational coherence spectroscopy allows one to investigate the evolution of vibrational coherence in electronic excited states. Methods such as pump-degenerate four-wave mixing and pump-impulsive vibrational spectroscopy combine an initial ultrashort laser pulse with a nonlinear probing sequence to reinduce vibrational coherence exclusively in the excited states. By carefully exploiting specific electronic resonances, one can detect vibrational coherence from 0 cm(-1) to over 2,000 cm(-1) and map its evolution. This review focuses on the observation and mapping of high-frequency vibrational coherence for all-trans biological polyenes such as β-carotene, lycopene, retinal, and retinal Schiff base. We discuss the role of molecular symmetry in vibrational coherence activity in the S1 electronic state and the interplay of coupling between electronic states and vibrational coherence.

  3. Nonselective and polarization effects in time-resolved optogalvanic spectroscopy

    NASA Astrophysics Data System (ADS)

    Zhechev, D.; Steflekova, V.

    2016-02-01

    Three interfering effects in optogalvanic (OG) spectroscopy are identified in a hollow cathode discharge (HCD) - OG detector. The laser beam is found to generate two nonselective processes, namely photoelectron emission (PE) from the cathode surface with a sub-breakdown bias applied, and nonresonant space ionization. The convolution of these galvanic contributions was determined experimentally as an instrumental function and a deconvolution procedure to determine the actual OG signal was developed. Specific plasma conductance is detected dependent on the polarization of the laser beam irradiating. Linearly/circularly polarized light beam is found to induce OG signals differ in amplitude (and their shape parameters in the time-resolved OG signals (TROGS)). The phenomena coherence and specific conductance are found to be in causal relationship. The additional conductance due to coherent states of atoms manifests itself as an intrinsic instrumental property of OG detector.

  4. Time-resolved spectroscopy of low-dimensional semiconductor structures

    NASA Astrophysics Data System (ADS)

    Murphy, Joseph R.

    This dissertation is a survey of ultrafast time-resolved optical measurements conducted on a variety of low-dimensional semiconductor systems to further the understanding of the dynamic behavior in the following systems: ZnMnTe/ZnSe quantum dots, ZnTe/ZnMnSe quantum dots, InGaAs quantum wells, CdMnSe colloidal quantum dots, multi-shell CdSe/CdMnS/CdS colloidal nanoplatelets, and graphene and graphene-related solutions and films. Using time-resolved photoluminescence to study epitaxially-grown ZnTe and ZnMnTe quantum dots in corresponding ZnMnSe and ZnSe matrices, the location dependence of manganese ions in respect to magnetic polaron formation is shown. The structure with manganese ions located in the matrix exhibited magnetic polaron behavior consistent with previous literature, whereas the structure with the magnetic ions located within the quantum dots exhibited unconventional magnetic polaron properties. These properties, including temperature and magnetic field insensitivity, were explained through the use of a model that predicted an increased internal magnetic field due to a decreased effective volume of the magnetic polaron and a higher effective temperature due to laser heating. Magneto-time-resolved photoluminescence measurements on a system of colloidal CdMnSe quantum dots show that the magnetic polaron properties differ significantly from the epitaxially grown quantum dots. First the timescales at which the magnetic polaron forms and the polarization saturates are different by more than an order of magnitude, and second, the magnetic polaron energy exhibited step-like behavior as the strength of the externally applied magnetic field is increased. The field dependent MP formation energy that is observed experimentally is explained as due to the breaking of the antiferromagnetic coupling of Mn dimers within the QDs. This model is further verified by the observation of quantized behavior in the Zeeman energy splitting. Through the use of magneto

  5. Time Resolved Single Wire Aluminum Optical Spectroscopy Experiments

    NASA Astrophysics Data System (ADS)

    Blesener, Kate; Pikuz, Sergey; Shelkovenko, Tatiana; Blesener, Isaac; Chalenski, David; Hammer, David; Maron, Yitzhak; Bernshtam, Vladimir

    2010-11-01

    We are exploring the conditions of plasmas generated by current-driven explosions of single fine aluminum wires, including temperatures, electron density, ionization state, and potentially magnetic field, using time-resolved emission spectroscopy at visible wavelengths. The experiments are being carried out with 15μm to 75μm Al wires driven by the 10kA, 500ns rise time LCP3 pulser. To determine the magnetic field, a new diagnostic method is being developed which makes use of Zeeman-effect-produced differences in the line shapes of two fine structure components of a multiplet that are equally broadened by Stark and Doppler effects. This method has been demonstrated at the Weizmann Institute of Science in laser-produced plasmas with lower energy densities [1]. [4pt] [1] E. Stambulchik, et al. Phys. Rev. Lett. 98, 225001 (2007).

  6. Time-resolved phase-sensitive second harmonic generation spectroscopy.

    PubMed

    Nowakowski, Paweł J; Woods, David A; Bain, Colin D; Verlet, Jan R R

    2015-02-28

    A methodology based on time-resolved, phase-sensitive second harmonic generation (SHG) for probing the excited state dynamics of species at interfaces is presented. It is based on an interference measurement between the SHG from the sample and a local oscillator generated at a reference together with a lock-in measurement to remove the large constant offset from the interference. The technique is characterized by measuring the phase and excited state dynamics of the dye malachite green at the water/air interface. The key attributes of the technique are that the observed signal is directly proportional to sample concentration, in contrast to the quadratic dependence from non-phase sensitive SHG, and that the real and imaginary parts of the 2nd order non-linear susceptibility can be determined independently. We show that the method is highly sensitive and can provide high quality excited state dynamics in short data acquisition times. PMID:25725724

  7. Towards time-resolved serial crystallography in a microfluidic device.

    PubMed

    Pawate, Ashtamurthy S; Šrajer, Vukica; Schieferstein, Jeremy; Guha, Sudipto; Henning, Robert; Kosheleva, Irina; Schmidt, Marius; Ren, Zhong; Kenis, Paul J A; Perry, Sarah L

    2015-07-01

    Serial methods for crystallography have the potential to enable dynamic structural studies of protein targets that have been resistant to single-crystal strategies. The use of serial data-collection strategies can circumvent challenges associated with radiation damage and repeated reaction initiation. This work utilizes a microfluidic crystallization platform for the serial time-resolved Laue diffraction analysis of macroscopic crystals of photoactive yellow protein (PYP). Reaction initiation was achieved via pulsed laser illumination, and the resultant electron-density difference maps clearly depict the expected pR(1)/pR(E46Q) and pR(2)/pR(CW) states at 10 µs and the pB1 intermediate at 1 ms. The strategies presented here have tremendous potential for extension to chemical triggering methods for reaction initiation and for extension to dynamic, multivariable analyses.

  8. Time-resolving electron temperature diagnostic for ALCATOR C

    NASA Astrophysics Data System (ADS)

    Fairfax, S. A.

    1984-05-01

    A diagnostic that provides time-resolved central electron temperatures was designed, built, and tested on the ALCATOR C Tokamak. The diagnostic uses an array of fixed-wavelength X-ray crystal monochromators to sample the X-ray continuum and determine the absolute electron temperature. The resolution and central energy of each channel were chosen to exclude any contributions from impurity line radiation. The need for such a diagnostic tool, the design methodology, and the results with typical ALCATOR C plasmas are described. Sawtooth (m = 1) temperature oscillations were observed after pellet fueling of the plasma. This is the first time that such oscillations were observed with an X-ray temperature diagnostic.

  9. Time-resolved phase-sensitive second harmonic generation spectroscopy

    NASA Astrophysics Data System (ADS)

    Nowakowski, Paweł J.; Woods, David A.; Bain, Colin D.; Verlet, Jan R. R.

    2015-02-01

    A methodology based on time-resolved, phase-sensitive second harmonic generation (SHG) for probing the excited state dynamics of species at interfaces is presented. It is based on an interference measurement between the SHG from the sample and a local oscillator generated at a reference together with a lock-in measurement to remove the large constant offset from the interference. The technique is characterized by measuring the phase and excited state dynamics of the dye malachite green at the water/air interface. The key attributes of the technique are that the observed signal is directly proportional to sample concentration, in contrast to the quadratic dependence from non-phase sensitive SHG, and that the real and imaginary parts of the 2nd order non-linear susceptibility can be determined independently. We show that the method is highly sensitive and can provide high quality excited state dynamics in short data acquisition times.

  10. CCD time-resolved photometry of faint cataclysmic variables. I

    NASA Technical Reports Server (NTRS)

    Howell, Steve; Szkody, Paula

    1988-01-01

    CCD time-resolved V and B differential light curves are presented for the dwarf novae AR And, FS Aur, TT Boo, UZ Boo, AF Cam, AL Com, AW Gem, X Leo, RZ Leo, CW Mon, SW UMa, and TW Vir. The time-series observations ranged from 2 to 6 hours and have accuracies of 0.025 mag or better for the majority of the runs. Except for AR And, X Leo, CW Mon, and TW Vir, the periods are below the cataclysmic-variable period gap (about 2 hours), and the systems are potential SU UMa stars. Photometric periods for five of the stars are the first such determinations, while those for the other seven generally confirm previous spectroscopic or photometric observations. In several cases, the photometric modulations are large amplitude (up to 0.5 mag). The results on AL Com and SW UMa indicate they may be magnetic variables.

  11. Time-resolved doubly bent crystal x-ray spectrometer

    SciTech Connect

    Hockaday, M.P.; Wilke, M.D.; Blake, R.L.; Vaninetti, J.; Gray, N.T.; Nedrow, P.T.

    1988-08-01

    X-ray spectroscopy is an essential tool in high-temperature plasma research. We describe a time-resolved x-ray spectrometer suitable for measuring spectra in harsh environments common to many very high-energy density laboratory plasma sources. The spectrometer consisted of a doubly curved Si(111) crystal diffraction element, a WL-1201 (ZnO:Ga) phosphor, a coherent fiber-optic array, and two visible streak cameras. The spectrometer design described here has a minimum time resolution of 1.3 ns with 2.8-eV spectral resolution over a 200-eV-wide bandpass in the 6--7-keV region of the spectrum. Complete system spectral throughput calibrations were done at the Cornell High Energy Synchrotron (CHESS). Details of the design and calibration results are presented.

  12. Time-resolved doubly bent crystal x-ray spectrometer

    SciTech Connect

    Hockaday, M.P.; Wilke, M.D.; Blake, R.L.; Vaninetti, J.; Gray, N.T.; Nedrow, P.T.

    1988-01-01

    X-ray spectroscopy is an essential tool in high temperature plasma research. We describe a time-resolved x-ray spectrometer suitable for measuring spectra in harsh environments common to many very high energy density laboratory plasma sources. The spectrometer consisted of a doubly curved Si(111) crystal diffraction element, a WL-1201 (ZnO:Ga) phosphor, a coherent fiber optic array, and two visible streak cameras. The spectrometer design described here has a minimum time resolution of 1.3 ns with 2.8 eV spectral resolution over a 200 eV wide bandpass in the 6-7 keV region of the spectrum. Complete system spectral throughput calibrations were done at the Cornell High Energy Synchrotron (CHESS). Details of the design and calibration results are presented. 5 refs., 5 figs.

  13. Time resolved PIV measurement of fluid dynamics in agitated vessels

    NASA Astrophysics Data System (ADS)

    Jasikova, D.; Kotek, M.; Kopecky, V.

    2015-01-01

    Here we present the results obtained by TR PIV measurements focused on detailed flow analysis in the selected region. The investigated area was placed 3mm above the blades axis and 5mm far from the blade edge. The captured images were firstly analysed on the mean velocity distribution and the intensity of turbulence {UV} statistics. Here we used the time resolved technique for the experimental study of the flow field in the agitated vessel. The results of the application POD and ODP algorithm on the captured datasets uncovered the existence of unsteady structures in the area that was assumed to be stable. The existence of these structures is bringing a novel view on the mixing process.

  14. Towards time-resolved serial crystallography in a microfluidic device

    PubMed Central

    Pawate, Ashtamurthy S.; Šrajer, Vukica; Schieferstein, Jeremy; Guha, Sudipto; Henning, Robert; Kosheleva, Irina; Schmidt, Marius; Ren, Zhong; Kenis, Paul J. A.; Perry, Sarah L.

    2015-01-01

    Serial methods for crystallography have the potential to enable dynamic structural studies of protein targets that have been resistant to single-crystal strategies. The use of serial data-collection strategies can circumvent challenges associated with radiation damage and repeated reaction initiation. This work utilizes a microfluidic crystallization platform for the serial time-resolved Laue diffraction analysis of macroscopic crystals of photoactive yellow protein (PYP). Reaction initiation was achieved via pulsed laser illumination, and the resultant electron-density difference maps clearly depict the expected pR1/pRE46Q and pR2/pRCW states at 10 µs and the pB1 intermediate at 1 ms. The strategies presented here have tremendous potential for extension to chemical triggering methods for reaction initiation and for extension to dynamic, multivariable analyses. PMID:26144226

  15. Photon-Counting Arrays for Time-Resolved Imaging

    PubMed Central

    Antolovic, I. Michel; Burri, Samuel; Hoebe, Ron A.; Maruyama, Yuki; Bruschini, Claudio; Charbon, Edoardo

    2016-01-01

    The paper presents a camera comprising 512 × 128 pixels capable of single-photon detection and gating with a maximum frame rate of 156 kfps. The photon capture is performed through a gated single-photon avalanche diode that generates a digital pulse upon photon detection and through a digital one-bit counter. Gray levels are obtained through multiple counting and accumulation, while time-resolved imaging is achieved through a 4-ns gating window controlled with subnanosecond accuracy by a field-programmable gate array. The sensor, which is equipped with microlenses to enhance its effective fill factor, was electro-optically characterized in terms of sensitivity and uniformity. Several examples of capture of fast events are shown to demonstrate the suitability of the approach. PMID:27367697

  16. Frequency domain approach for time-resolved pump-probe microscopy using intensity modulated laser diodes.

    PubMed

    Miyazaki, J; Kawasumi, K; Kobayashi, T

    2014-09-01

    We present a scheme for time-resolved pump-probe microscopy using intensity modulated laser diodes. The modulation frequencies of the pump and probe beams are varied up to 500 MHz with fixed frequency detuning typically set at 15 kHz. The frequency response of the pump-probe signal is detected using a lock-in amplifier referenced at the beat frequency. This frequency domain method is capable of characterizing the nanosecond to picosecond relaxation dynamics of sample species without the use of a high speed detector or a high frequency lock-in amplifier. Furthermore, as the pump-probe signal is based on the nonlinear interaction between the two laser beams and the sample, our scheme provides better spatial resolution than the conventional diffraction-limited optical microscopes. Time-resolved pump-probe imaging of fluorescence beads and aggregates of quantum dots demonstrates that this method is useful for the microscopic analysis of optoelectronic devices. The system is implemented using compact and low-cost laser diodes, and thus has a broad range of applications in the fields of photochemistry, optical physics, and biological imaging.

  17. Time-Resolved Luminescence Nanothermometry with Nitrogen-Vacancy Centers in Nanodiamonds.

    PubMed

    Tzeng, Yan-Kai; Tsai, Pei-Chang; Liu, Hsiou-Yuan; Chen, Oliver Y; Hsu, Hsiang; Yee, Fu-Goul; Chang, Ming-Shien; Chang, Huan-Cheng

    2015-06-10

    Measuring temperature in nanoscale spatial resolution either at or far from equilibrium is of importance in many scientific and technological applications. Although negatively charged nitrogen-vacancy (NV(-)) centers in diamond have recently emerged as a promising nanometric temperature sensor, the technique has been applied only under steady state conditions so far. Here, we present a three-point sampling method that allows real-time monitoring of the temperature changes over ±100 K and a pump-probe-type experiment that enables the study of nanoscale heat transfer with a temporal resolution of better than 10 μs. The utility of the time-resolved luminescence nanothermometry was demonstrated with 100 nm fluorescent nanodiamonds spin-coated on a glass substrate and submerged in gold nanorod solution heated by a near-infrared laser, and the validity of the measurements was verified with finite-element numerical simulations. The combined theoretical and experimental approaches will be useful to implement time-resolved temperature sensing in laser processing of materials and even for devices in operation at the nanometer scale. PMID:25951304

  18. Simultaneous Velocity Discrimination Method of Two-Phase Flows Using Time Resolved Stereo PIV and PTV

    NASA Astrophysics Data System (ADS)

    Vanderwerker, P. B.; Chen, Y.; Torregrosa, M. M.; Diez, F. J.; Photos, S.; Troolin, D.

    2007-03-01

    Multiphase jets laden with particles appear in many engineering and environmental processes. Typical examples are sprays containing liquid fuel drops in combustion processes, air jets laden with coal particles in a power plant, and the dispersion of harmful substances like soot and pollutants from steady exhaust flows, among others. Studies of particle-laden turbulent flows suggest that particle distribution is not uniform but preferential. In order to understand the mechanism of particle dispersion, time resolved simultaneous 3D velocity measurements of the disperse phase and of the fluid flow were made. Two-phase discrimination algorithms were developed based upon the filtering methodology proposed by Khalitov & Longmire (2002), allowing for complete separation of the two-phases in stereo PIV images. The different filtering methods studied include separation of the two-phases using: (1) particle size discrimination, (2) particle intensity discrimination, (3) particle size and intensity discrimination, and (4) fluorescent particles for one of the two-phases. This methodology also enables time-resolved instantaneous 3D velocity fields using PTV and PIV on the disperse phase and fluid flow phase respectively. These allow visualization of 3D turbulent coherent structure evolution in the fluid as well as the evolution of the dispersed phase.

  19. Toward picosecond time-resolved X-ray absorption studies of interfacial photochemistry

    NASA Astrophysics Data System (ADS)

    Gessner, Oliver; Mahl, Johannes; Neppl, Stefan

    2016-05-01

    We report on the progress toward developing a novel picosecond time-resolved transient X-ray absorption spectroscopy (TRXAS) capability for time-domain studies of interfacial photochemistry. The technique is based on the combination of a high repetition rate picosecond laser system with a time-resolved X-ray fluorescent yield setup that may be used for the study of radiation sensitive materials and X-ray spectroscopy compatible photoelectrochemical (PEC) cells. The mobile system is currently deployed at the Advanced Light Source (ALS) and may be used in all operating modes (two-bunch and multi-bunch) of the synchrotron. The use of a time-stamping technique enables the simultaneous recording of TRXAS spectra with delays between the exciting laser pulses and the probing X-ray pulses spanning picosecond to nanosecond temporal scales. First results are discussed that demonstrate the viability of the method to study photoinduced dynamics in transition metal-oxide semiconductor (SC) samples under high vacuum conditions and at SC-liquid electrolyte interfaces during photoelectrochemical water splitting. Opportunities and challenges are outlined to capture crucial short-lived intermediates of photochemical processes with the technique. This work was supported by the Department of Energy Office of Science Early Career Research Program.

  20. Two-Photon Absorption and Time-Resolved Stimulated Emission Depletion Spectroscopy of a New Fluorenyl Derivative

    PubMed Central

    Bondar, Mykhailo V.; Morales, Alma R.; Yue, Xiling; Luchita, Gheorghe; Przhonska, Olga V.; Kachkovsky, Olexy D.

    2012-01-01

    The synthesis, comprehensive linear photophysical characterization, two-photon absorption (2PA), steady-state and time-resolved stimulated emission depletion properties of a new fluorene derivative, (E)-1-(2-(di-p-tolylamino)-9,9-diethyl-9H-fluoren-7-yl)-3-(thiophen-2-yl)prop-2-en-1-one (1), are reported. The primary linear spectral properties, including excitation anisotropy, fluorescence lifetimes, and photostability, were investigated in a number of aprotic solvents at room temperature. The degenerate 2PA spectra of 1 were obtained with an open aperture Z-scan and two-photon induced fluorescence methods, using a 1-kHz femtosecond laser system, and maximum 2PA cross-sections of ~400–600 GM were obtained. The nature of the electronic absorption processes in 1 was investigated by DFT-based quantum chemical methods implemented in the Gaussian 09 program. The one- and two-photon stimulated emission spectra of 1 were measured over a broad spectral range using a femtosecond pump probe–based fluorescence quenching technique, while a new methodology for time-resolved fluorescence emission spectroscopy is proposed. An effective application of 1 in fluorescence bioimaging was demonstrated via one- and two-photon fluorescence microscopy images of HCT 116 cells containing the dye encapsulated micelles. PMID:22887914

  1. New method for measuring time-resolved spectra of lanthanide emission using square-wave excitation

    SciTech Connect

    Qin, Feng; Zhao, Hua; Cai, Wei; Duan, Qianqian; Zhang, Zhiguo; Cao, Wenwu

    2013-11-15

    A method using modulated continuous wave (CW) visible laser to measure time-resolved fluorescence spectra of trivalent rare-earth ions has been developed. Electro-optic modulator was used to modulate the CW pumping laser with a rise time of 2 μs. CW Nd{sup 3+} lasers were used as examples to present the method. Upconversion dynamic process of Ho{sup 3+} was studied utilizing a 532 nm CW laser. Quantum cutting dynamic process from Tb{sup 3+} to Yb{sup 3+} was analyzed by a 473 nm CW laser. This method can be applied to any CW laser such as He-Ne laser, Ar{sup +} laser, Kr{sup +} laser, Ti:sapphire laser, etc.

  2. Space- and time-resolved protein dynamics in single bacterial cells observed on a chip.

    PubMed

    Greif, Dominik; Pobigaylo, Nataliya; Frage, Benjamin; Becker, Anke; Regtmeier, Jan; Anselmetti, Dario

    2010-09-15

    Life cell imaging of bacterial cells over long times is very challenging because of the small dimensions and the need for a liquid environment assuring cell viability. In order to obtain space- and time-resolved information about protein dynamics, high resolution time-lapse fluorescence images (TLFI) of single bacterial cells were recorded in a poly(dimethylsiloxane) (PDMS) microfluidic chip. A new gradient coating technique was applied to ensure cell loading. As a proof-of-concept, we monitored the evenly distributed cytoplasmic protein GcrA as well as the asymmetric localization of the DivK protein in cells of S. meliloti over at least two division cycles. Localization of DivK was characterized by dividing each bacterial cell into 4 sections with dimensions closely above the optical limit of resolution. This approach of generating spatio-temporal resolved information of protein dynamics in single bacterial cells is applicable to many problems.

  3. New method for measuring time-resolved spectra of lanthanide emission using square-wave excitation.

    PubMed

    Qin, Feng; Zhao, Hua; Duan, Qianqian; Cai, Wei; Zhang, Zhiguo; Cao, Wenwu

    2013-11-01

    A method using modulated continuous wave (CW) visible laser to measure time-resolved fluorescence spectra of trivalent rare-earth ions has been developed. Electro-optic modulator was used to modulate the CW pumping laser with a rise time of 2 μs. CW Nd(3+) lasers were used as examples to present the method. Upconversion dynamic process of Ho(3+) was studied utilizing a 532 nm CW laser. Quantum cutting dynamic process from Tb(3+) to Yb(3+) was analyzed by a 473 nm CW laser. This method can be applied to any CW laser such as He-Ne laser, Ar(+) laser, Kr(+) laser, Ti:sapphire laser, etc.

  4. Time resolved optical biopsy spectroscopy of normal, benign and malignant tissues from NADH and FAD changes

    NASA Astrophysics Data System (ADS)

    Masilamani, V.; Das, B. B.; Secor, J.; AlSalhi, M.; Amer, S. B.; Farhat, K.; Rabah, D.; Alfano, R. R.

    2012-01-01

    Histo pathological examination is the gold standard to discriminate between benign and malignant growth of tissue. But this is invasive and stressful. Hence many non invasive imaging techniques, such as CT, MRI, PET, etc are employed, each having certain advantages and disadvantages. In this context optical biopsy is a newly emerging technique, since it employs non-ionizing radiation like light or laser, which could be shined directly or launched through optical fiber to reach any part of the body. This paper reports results of time resolved emission spectra of 24 excised tissue sample (normal control=12; benign=4; malignant=8) of breast and prostate, employing a 390nm, 100 fs, Ti-Sapphire laser pulses. The fluorescence decay times were measured using streak camera and fitted for single and bi- exponential decays with reliability of 97%. Our results show the distinct difference between normal, benign and malignant tissues attributed changes of NADH and FAD levels.

  5. Time-resolved delayed luminescence image microscopy using an europium ion chelate complex.

    PubMed Central

    Marriott, G.; Heidecker, M.; Diamandis, E. P.; Yan-Marriott, Y.

    1994-01-01

    Improvements and extended applications of time-resolved delayed luminescence imaging microscopy (TR-DLIM) in cell biology are described. The emission properties of europium ion complexed to a fluorescent chelating group capable of labeling proteins are exploited to provide high contrast images of biotin labeled ligands through detection of the delayed emission. The streptavidin-based macromolecular complex (SBMC) employs streptavidin cross-linked to thyroglobulin multiply labeled with the europium-fluorescent chelate. The fluorescent chelate is efficiently excited with 340-nm light, after which it sensitizes europium ion emission at 612 nm hundreds of microseconds later. The SBMC complex has a high quantum yield orders of magnitude higher than that of eosin, a commonly used delayed luminescent probe, and can be readily seen by the naked eye, even in specimens double-labeled with prompt fluorescent probes. Unlike triplet-state phosphorescent probes, sensitized europium ion emission is insensitive to photobleaching and quenching by molecular oxygen; these properties have been exploited to obtain delayed luminescence images of living cells in aerated medium thus complementing imaging studies using prompt fluorescent probes. Since TR-DLIM has the unique property of rejecting enormous signals that originate from scattered light, autofluorescence, and prompt fluorescence it has been possible to resolve double emission images of living amoeba cells containing an intensely stained lucifer yellow in pinocytosed vesicles and membrane surface-bound SBMC-labeled biotinylated concanavalin A. Images of fixed cells represented in terms of the time decay of the sensitized emission show the lifetime of the europium ion emission is sensitive to the environment in which it is found. Through the coupling of SBMC to streptavidin,a plethora of biotin-based tracer molecules are available for immunocytochemical studies. Images FIGURE 1 FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 5 FIGURE 6 FIGURE 7

  6. Electron-transfer acceleration investigated by time resolved infrared spectroscopy.

    PubMed

    Vlček, Antonín; Kvapilová, Hana; Towrie, Michael; Záliš, Stanislav

    2015-03-17

    Ultrafast electron transfer (ET) processes are important primary steps in natural and artificial photosynthesis, as well as in molecular electronic/photonic devices. In biological systems, ET often occurs surprisingly fast over long distances of several tens of angströms. Laser-pulse irradiation is conveniently used to generate strongly oxidizing (or reducing) excited states whose reactions are then studied by time-resolved spectroscopic techniques. While photoluminescence decay and UV-vis absorption supply precise kinetics data, time-resolved infrared absorption (TRIR) and Raman-based spectroscopies have the advantage of providing additional structural information and monitoring vibrational energy flows and dissipation, as well as medium relaxation, that accompany ultrafast ET. We will discuss three cases of photoinduced ET involving the Re(I)(CO)3(N,N) moiety (N,N = polypyridine) that occur much faster than would be expected from ET theories. [Re(4-N-methylpyridinium-pyridine)(CO)3(N,N)](2+) represents a case of excited-state picosecond ET between two different ligands that remains ultrafast even in slow-relaxing solvents, beating the adiabatic limit. This is caused by vibrational/solvational excitation of the precursor state and participation of high-frequency quantum modes in barrier crossing. The case of Re-tryptophan assemblies demonstrates that excited-state Trp → *Re(II) ET is accelerated from nanoseconds to picoseconds when the Re(I)(CO)3(N,N) chromophore is appended to a protein, close to a tryptophan residue. TRIR in combination with DFT calculations and structural studies reveals an interaction between the N,N ligand and the tryptophan indole. It results in partial electronic delocalization in the precursor excited state and likely contributes to the ultrafast ET rate. Long-lived vibrational/solvational excitation of the protein Re(I)(CO)3(N,N)···Trp moiety, documented by dynamic IR band shifts, could be another accelerating factor. The last

  7. Time-resolved neutron imaging at ANTARES cold neutron beamline

    NASA Astrophysics Data System (ADS)

    Tremsin, A. S.; Dangendorf, V.; Tittelmeier, K.; Schillinger, B.; Schulz, M.; Lerche, M.; Feller, W. B.

    2015-07-01

    In non-destructive evaluation with X-rays light elements embedded in dense, heavy (or high-Z) matrices show little contrast and their structural details can hardly be revealed. Neutron radiography, on the other hand, provides a solution for those cases, in particular for hydrogenous materials, owing to the large neutron scattering cross section of hydrogen and uncorrelated dependency of neutron cross section on the atomic number. The majority of neutron imaging experiments at the present time is conducted with static objects mainly due to the limited flux intensity of neutron beamline facilities and sometimes due to the limitations of the detectors. However, some applications require the studies of dynamic phenomena and can now be conducted at several high intensity beamlines such as the recently rebuilt ANTARES beam line at the FRM-II reactor. In this paper we demonstrate the capabilities of time resolved imaging for repetitive processes, where different phases of the process can be imaged simultaneously and integrated over multiple cycles. A fast MCP/Timepix neutron counting detector was used to image the water distribution within a model steam engine operating at 10 Hz frequency. Within <10 minutes integration the amount of water was measured as a function of cycle time with a sub-mm spatial resolution, thereby demonstrating the capabilities of time-resolved neutron radiography for the future applications. The neutron spectrum of the ANTARES beamline as well as transmission spectra of a Fe sample were also measured with the Time Of Flight (TOF) technique in combination with a high resolution beam chopper. The energy resolution of our setup was found to be ~ 0.8% at 5 meV and ~ 1.7% at 25 meV. The background level (most likely gammas and epithermal/fast neutrons) of the ANTARES beamline was also measured in our experiments and found to be on the scale of 3% when no filters are installed in the beam. Online supplementary data available from stacks.iop.org/jinst/10

  8. Steady-state and time-resolved investigations on pyrene-based chemosensors.

    PubMed

    Fernández-Lodeiro, Javier; Núñez, Cristina; de Castro, Catherine S; Bértolo, Emilia; Seixas de Melo, J Sérgio; Capelo, José Luis; Lodeiro, Carlos

    2013-01-01

    Two novel fluorescent probes bearing a single (P) and two (a podand-like structure, L) pyrene units derived from 1,5-bis(2-aminophenoxy)-3-oxopentane have been synthesized and investigated in dioxane using UV-vis absorption, and steady-state and time-resolved (in a picosecond time scale) emission spectroscopy; in the gas phase, matrix-assisted laser desorption ionization mass spectrometry was employed. In dioxane, the absorption and emission spectra of P present a unique band with maxima at 361 and 392 nm, which have been associated with the monomer absorption and emission bands, respectively. In dioxane, for compound L, an additional band with a maximum at ∼525 nm is observed; upon the addition of water, an emissive band (with maxima varying from 405 to 490 nm) appears in both P and L spectra; this is discussed in terms of the emission of a species with charge character. Upon metal addition (Cu(2+), Zn(2+), and Ag(+)) to P, a gradual quenching effect of the monomer emission is observed and found to be more pronounced with Cu(2+). In the case of L, upon the addition of metal cations, the long emission band (∼550 nm) decreases and the monomer emission band increases (with an isoemissive point at ∼450 nm) and no evidence for the intermediate band (at ∼405-490 nm) now exists. Time-resolved data in dioxane/water mixtures showed that for P and L these two fit double- and triple-exponential decay laws, respectively. With P, this has been attributed to a two-state system, which involves the monomer and a charged species, with its emission maxima varying with the polarity of the media (here mirrored by its dielectric constant), which can potentially be addressed to an exciplex-like species, whereas with L, it has been attributed to a three-state system involving, in addition to these two species, an excimer. From absorption and fluorescence excitation and time-resolved data, evidence is given for the presence of intramolecular dimer formation in the ground state

  9. Time-resolved pump-probe experiments at the LCLS.

    PubMed

    Glownia, James M; Cryan, J; Andreasson, J; Belkacem, A; Berrah, N; Blaga, C I; Bostedt, C; Bozek, J; DiMauro, L F; Fang, L; Frisch, J; Gessner, O; Gühr, M; Hajdu, J; Hertlein, M P; Hoener, M; Huang, G; Kornilov, O; Marangos, J P; March, A M; McFarland, B K; Merdji, H; Petrovic, V S; Raman, C; Ray, D; Reis, D A; Trigo, M; White, J L; White, W; Wilcox, R; Young, L; Coffee, R N; Bucksbaum, P H

    2010-08-16

    The first time-resolved x-ray/optical pump-probe experiments at the SLAC Linac Coherent Light Source (LCLS) used a combination of feedback methods and post-analysis binning techniques to synchronize an ultrafast optical laser to the linac-based x-ray laser. Transient molecular nitrogen alignment revival features were resolved in time-dependent x-ray-induced fragmentation spectra. These alignment features were used to find the temporal overlap of the pump and probe pulses. The strong-field dissociation of x-ray generated quasi-bound molecular dications was used to establish the residual timing jitter. This analysis shows that the relative arrival time of the Ti:Sapphire laser and the x-ray pulses had a distribution with a standard deviation of approximately 120 fs. The largest contribution to the jitter noise spectrum was the locking of the laser oscillator to the reference RF of the accelerator, which suggests that simple technical improvements could reduce the jitter to better than 50 fs.

  10. Time-resolved electric-field-induced second harmonic

    NASA Astrophysics Data System (ADS)

    Meshulam, Guilia; Berkovic, Garry; Kotler, Zvi

    2001-12-01

    One limitation of using electric field induced second harmonic (EFISH) to determine the molecular first hyperpolarizability (beta) of nonlinear optical molecules lies in the fact that part of the second harmonic signal comes from the second hyperpolarizability (gamma) produced by mixing two optical fields with the DC field. In analyzing EFISH results, the second hyperpolarizability contribution of the studied molecules is generally neglected. We present a modified time resolved EFISH technique that allows us, in a single experiment, to determine separately the beta and the gamma contributions. We study para-nitro aniline dissolved in Glycerol, a highly viscous solvent, and apply the DC field via a high voltage pulse with a fast rise time of approximately 40 nsec. As a result, the orientation of the molecules under the applied electric field is slow relative to the build-up of the field, enabling us to directly measure only the DC induced second harmonic (gamma contribution), at the beginning of the HV pulse. The pure beta contribution is determined from the difference between this signal and the conventional EFISH signal at the plateau of the HV pulse. Our result confirm that the gamma contribution is indeed less than 10% of the total.

  11. A Clinical Tissue Oximeter Using NIR Time-Resolved Spectroscopy.

    PubMed

    Fujisaka, Shin-ichi; Ozaki, Takeo; Suzuki, Tsuyoshi; Kamada, Tsuyoshi; Kitazawa, Ken; Nishizawa, Mitsunori; Takahashi, Akira; Suzuki, Susumu

    2016-01-01

    The tNIRS-1, a new clinical tissue oximeter using NIR time-resolved spectroscopy (TRS), has been developed. The tNIRS-1 measures oxygenated, deoxygenated and total hemoglobin and oxygen saturation in living tissues. Two-channel TRS measurements are obtained using pulsed laser diodes (LD) at three wavelengths, multi-pixel photon counters (MPPC) for light detection, and time-to-digital converters (TDC) for time-of-flight photon measurements. Incorporating advanced semiconductor devices helped to make the design of this small-size, low-cost and low-power TRS instrument possible. In order to evaluate the correctness and reproducibility of measurement data obtained with the tNIRS-1, a study using blood phantoms and healthy volunteers was conducted to compare data obtained from a conventional SRS device and data from an earlier TRS system designed for research purposes. The results of the study confirmed the correctness and reproducibility of measurement data obtained with the tNIRS-1. Clinical evaluations conducted in several hospitals demonstrated a high level of usability in clinical situations and confirmed the efficacy of measurement data obtained with the tNIRS-1.

  12. Time resolved strain dependent morphological study of electrically conducting nanocomposites

    NASA Astrophysics Data System (ADS)

    Khan, Imran; Mitchell, Geoffrey; Mateus, Artur; Kamma-Lorger, Christina S.

    2015-10-01

    An efficient and reliable method is introduced to understand the network behaviour of nano-fillers in a polymeric matrix under uniaxial strain coupled with small angle x-ray scattering measurements. The nanoparticles (carbon nanotubes) are conductive and the particles form a percolating network that becomes apparent source of electrical conduction and consequently the samples behave as a bulk conductor. Polyurethane based nanocomposites containing 2% w/w multiwall carbon nanotubes are studied. The electrical conductivity of the nanocomposite was (3.28×10-5s/m).The sample was able to be extended to an extension ratio of 1.7 before fracture. A slight variation in the electrical conductivity is observed under uniaxial strain which we attribute to the disturbance of conductive pathways. Further, this work is coupled with in- situ time resolved small angle x-ray scattering measurements using a synchrotron beam line to enable its measurements to be made during the deformation cycle. We use a multiscale structure to model the small angle x-ray data. The results of the analysis are interpreted as the presence of aggregates which would also go some way towards understanding why there is no alignment of the carbon nanotubes.

  13. Time-resolved infrared spectroscopic techniques as applied to channelrhodopsin

    PubMed Central

    Ritter, Eglof; Puskar, Ljiljana; Bartl, Franz J.; Aziz, Emad F.; Hegemann, Peter; Schade, Ulrich

    2015-01-01

    Among optogenetic tools, channelrhodopsins, the light gated ion channels of the plasma membrane from green algae, play the most important role. Properties like channel selectivity, timing parameters or color can be influenced by the exchange of selected amino acids. Although widely used, in the field of neurosciences for example, there is still little known about their photocycles and the mechanism of ion channel gating and conductance. One of the preferred methods for these studies is infrared spectroscopy since it allows observation of proteins and their function at a molecular level and in near-native environment. The absorption of a photon in channelrhodopsin leads to retinal isomerization within femtoseconds, the conductive states are reached in the microsecond time scale and the return into the fully dark-adapted state may take more than minutes. To be able to cover all these time regimes, a range of different spectroscopical approaches are necessary. This mini-review focuses on time-resolved applications of the infrared technique to study channelrhodopsins and other light triggered proteins. We will discuss the approaches with respect to their suitability to the investigation of channelrhodopsin and related proteins. PMID:26217670

  14. A Clinical Tissue Oximeter Using NIR Time-Resolved Spectroscopy.

    PubMed

    Fujisaka, Shin-ichi; Ozaki, Takeo; Suzuki, Tsuyoshi; Kamada, Tsuyoshi; Kitazawa, Ken; Nishizawa, Mitsunori; Takahashi, Akira; Suzuki, Susumu

    2016-01-01

    The tNIRS-1, a new clinical tissue oximeter using NIR time-resolved spectroscopy (TRS), has been developed. The tNIRS-1 measures oxygenated, deoxygenated and total hemoglobin and oxygen saturation in living tissues. Two-channel TRS measurements are obtained using pulsed laser diodes (LD) at three wavelengths, multi-pixel photon counters (MPPC) for light detection, and time-to-digital converters (TDC) for time-of-flight photon measurements. Incorporating advanced semiconductor devices helped to make the design of this small-size, low-cost and low-power TRS instrument possible. In order to evaluate the correctness and reproducibility of measurement data obtained with the tNIRS-1, a study using blood phantoms and healthy volunteers was conducted to compare data obtained from a conventional SRS device and data from an earlier TRS system designed for research purposes. The results of the study confirmed the correctness and reproducibility of measurement data obtained with the tNIRS-1. Clinical evaluations conducted in several hospitals demonstrated a high level of usability in clinical situations and confirmed the efficacy of measurement data obtained with the tNIRS-1. PMID:26782242

  15. Time resolved EUV spectra from Zpinching capillary discharge plasma

    NASA Astrophysics Data System (ADS)

    Jancarek, Alexandr; Nevrkla, Michal; Nawaz, Fahad

    2015-09-01

    We developed symmetrically charged driver to obtain high voltage, high current Z-pinching capillary discharge. Plasma is created by up to 70 kA, 29 ns risetime current pulse passing through a 5 mm inner diameter, 224 mm long capillary filled with gas to initial pressure in the range of 1 kPa. Due to the low inductance design of the driver, the pinch is observable directly from the measured current curve. Time-integrated and time-resolved spectra of discharge plasma radiation are recorded together with the capillary current and analyzed. The most encouraging spectra were captured in the wavelength range 8.3 ÷ 14 nm. This spectral region contains nitrogen Balmer series lines including potentially lasing NVII 2 - 3 transition. Spectral lines are identified in the NIST database using the FLY kinetic code. The line of 13.38 nm wavelength, transition NVII 2 - 3, was observed in gated, and also in time-integrated spectra for currents >60 kA. This work has been supported by the Ministry of Education, Youth and Sports of the Czech Republic grants LG13029.

  16. Time-resolved neurite mechanics by thermal fluctuation assessments.

    PubMed

    Gárate, Fernanda; Betz, Timo; Pertusa, María; Bernal, Roberto

    2015-12-30

    In the absence of simple noninvasive measurements, the knowledge of temporal and spatial variations of axons mechanics remains scarce. By extending thermal fluctuation spectroscopy (TFS) to long protrusions, we determine the transverse amplitude thermal fluctuation spectra that allow direct and simultaneous access to three key mechanics parameters: axial tension, bending flexural rigidity and plasma membrane tension. To test our model, we use PC12 cell protrusions-a well-know biophysical model of axons-in order to simplify the biological system under scope. For instance, axial and plasma membrane tension are found in the range of nano Newton and tens of pico Newtons per micron respectively. Furthermore, our results shows that the TFS technique is capable to distinguish quasi-identical protrusions. Another advantage of our approach is the time resolved nature of the measurements. Indeed, in the case of long term experiments on PC12 protrusions, TFS has revealed large temporal, correlated variations of the protrusion mechanics, displaying extraordinary feedback control over the axial tension in order to maintain a constant tension value.

  17. Time-resolved FRET strategy to screen GPCR ligand library.

    PubMed

    Oueslati, Nadia; Hounsou, Candide; Belhocine, Abderazak; Rodriguez, Thieric; Dupuis, Elodie; Zwier, Jurriaan M; Trinquet, Eric; Pin, Jean-Philippe; Durroux, Thierry

    2015-01-01

    Screening chemical libraries to find specific drugs for G protein-coupled receptors is still of major interest. Indeed, because of their major roles in all physiological functions, G protein-coupled receptors remain major targets for drug development programs. Currently, interest in GPCRs as drug targets has been boosted by the discovery of biased ligands, thus allowing the development of drugs not only specific for one target but also for the specific signaling cascade expected to have the therapeutic effect. Such molecules are then expected to display fewer side effects. To reach such a goal, there is much interest in novel, efficient, simple, and direct screening assays that may help identify any drugs interacting with the target, these being then analyzed for their biased activity. Here, we present an efficient strategy to screen ligands on their binding properties. The method described is based on time-resolved FRET between a receptor and a ligand. This method has already been used to develop new assays called Tag-lite(®) binding assays for numerous G protein-coupled receptors, proving its broad application and its power.

  18. Ultrafast surface dynamics probed with time resolved photoemission

    NASA Astrophysics Data System (ADS)

    Dell'Angela, M.; Hieke, F.; Sorgenfrei, F.; Gerken, N.; Beye, M.; Gerasimova, N.; Redlin, H.; Wurth, W.

    2016-01-01

    Time resolved core level photoemission (trXPS) allows real-time atom-specific investigation of ultrafast surface dynamics. Core levels contain information on the chemical state and the structure of the surface as well as the local charge distribution around specific atoms. Monitoring their evolution after optically exciting the surface, can give valuable information on the electronic (few picosecond time scale) and lattice dynamics (several picosecond timescale). We have performed a trXPS experiment at the free-electron laser FLASH at DESY in Hamburg on a clean Ir(111) surface measuring the temporal evolution of the 4f core levels of Ir(111) after optically exciting the sample. The spectral changes due to X-ray and optical laser induced space charge effects which occur in trXPS experiments with high fluence pump and probe pulses have been fully characterized and controlled during the measurements. At early time scales after the optical excitation we observe time-dependent energy shifts and intensity changes which can be partially attributed to the formation of sidebands. Furthermore, we can clearly identify contributions which result from a change in the surface electron density which then relaxes on a time scale on the order of 2 ps.

  19. Time resolved spectroscopic NMR imaging using hyperpolarized 129Xe.

    PubMed

    Han, S; Kühn, H; Häsing, F W; Münnemann, K; Blümich, B; Appelt, S

    2004-04-01

    We have visualized the melting and dissolution processes of xenon (Xe) ice into different solvents using the methods of nuclear magnetic resonance (NMR) spectroscopy, imaging, and time resolved spectroscopic imaging by means of hyperpolarized 129Xe. Starting from the initial condition of a hyperpolarized solid Xe layer frozen on top of an ethanol (ethanol/water) ice block we measured the Xe phase transitions as a function of time and temperature. In the pure ethanol sample, pieces of Xe ice first fall through the viscous ethanol to the bottom of the sample tube and then form a thin layer of liquid Xe/ethanol. The xenon atoms are trapped in this liquid layer up to room temperature and keep their magnetization over a time period of 11 min. In the ethanol/water mixture (80 vol%/20%), most of the polarized Xe liquid first stays on top of the ethanol/water ice block and then starts to penetrate into the pores and cracks of the ethanol/water ice block. In the final stage, nearly all the Xe polarization is in the gas phase above the liquid and trapped inside the pores. NMR spectra of homogeneous samples of pure ethanol containing thermally polarized Xe and the spectroscopic images of the melting process show that very high concentrations of hyperpolarized Xe (about half of the density of liquid Xe) can be stored or delivered in pure ethanol. PMID:15040986

  20. Time-resolved photoluminescence of SiOx encapsulated Si

    NASA Astrophysics Data System (ADS)

    Kalem, Seref; Hannas, Amal; Österman, Tomas; Sundström, Villy

    Silicon and its oxide SiOx offer a number of exciting electrical and optical properties originating from defects and size reduction enabling engineering new electronic devices including resistive switching memories. Here we present the results of photoluminescence dynamics relevant to defects and quantum confinement effects. Time-resolved luminescence at room temperature exhibits an ultrafast decay component of less than 10 ps at around 480 nm and a slower component of around 60 ps as measured by streak camera. Red shift at the initial stages of the blue luminescence decay confirms the presence of a charge transfer to long lived states. Time-correlated single photon counting measurements revealed a life-time of about 5 ns for these states. The same quantum structures emit in near infrared close to optical communication wavelengths. Nature of the emission is described and modeling is provided for the luminescence dynamics. The electrical characteristics of metal-oxide-semiconductor devices were correlated with the optical and vibrational measurement results in order to have better insight into the switching mechanisms in such resistive devices as possible next generation RAM memory elements. ``This work was supported by ENIAC Joint Undertaking and Laser-Lab Europe''.

  1. Time-Resolved Hard X-Ray Spectrometer

    SciTech Connect

    Kenneth Moya; Ian McKennaa; Thomas Keenana; Michael Cuneob

    2007-03-01

    Wired array studies are being conducted at the SNL Z accelerator to maximize the x-ray generation for inertial confinement fusion targets and high energy density physics experiments. An integral component of these studies is the characterization of the time-resolved spectral content of the x-rays. Due to potential spatial anisotropy in the emitted radiation, it is also critical to diagnose the time-evolved spectral content in a space-resolved manner. To accomplish these two measurement goals, we developed an x-ray spectrometer using a set of high-speed detectors (silicon PIN diodes) with a collimated field-of-view that converged on a 1-cm-diameter spot at the pinch axis. Spectral discrimination is achieved by placing high Z absorbers in front of these detectors. We built two spectrometers to permit simultaneous different angular views of the emitted radiation. Spectral data have been acquired from recent Z shots for the radial and polar views. UNSPEC1 has been adapted to analyze and unfold the measured data to reconstruct the x-ray spectrum. The unfold operator code, UFO2, is being adapted for a more comprehensive spectral unfolding treatment.

  2. Time-resolved hard x-ray spectrometer

    NASA Astrophysics Data System (ADS)

    Moy, Kenneth; Cuneo, Michael; McKenna, Ian; Keenan, Thomas; Sanford, Thomas; Mock, Ray

    2006-08-01

    Wired array studies are being conducted at the SNL Z accelerator to maximize the x-ray generation for inertial confinement fusion targets and high energy density physics experiments. An integral component of these studies is the characterization of the time-resolved spectral content of the x-rays. Due to potential spatial anisotropy in the emitted radiation, it is also critical to diagnose the time-evolved spectral content in a space-resolved manner. To accomplish these two measurement goals, we developed an x-ray spectrometer using a set of high-speed detectors (silicon PIN diodes) with a collimated field-of-view that converged on a 1-cm-diameter spot at the pinch axis. Spectral discrimination is achieved by placing high Z absorbers in front of these detectors. We built two spectrometers to permit simultaneous different angular views of the emitted radiation. Spectral data have been acquired from recent Z shots for the radial and axial (polar) views. UNSPEC 1 has been adapted to analyze and unfold the measured data to reconstruct the x-ray spectrum. The unfold operator code, UFO2, is being adapted for a more comprehensive spectral unfolding treatment.

  3. Femtosecond time-resolved MeV electron diffraction

    SciTech Connect

    Zhu, Pengfei; Zhu, Y.; Hidaka, Y.; Wu, L.; Cao, J.; Berger, H.; Geck, J.; Kraus, R.; Pjerov, S.; Shen, Y.; Tobey, R. I.; Hill, J. P.; Wang, X. J.

    2015-06-02

    We report the experimental demonstration of femtosecond electron diffraction using high-brightness MeV electron beams. High-quality, single-shot electron diffraction patterns for both polycrystalline aluminum and single-crystal 1T-TaS2 are obtained utilizing a 5 fC (~3 × 104 electrons) pulse of electrons at 2.8 MeV. The high quality of the electron diffraction patterns confirms that electron beam has a normalized emittance of ~50 nm rad. The transverse and longitudinal coherence length is ~11 and ~2.5 nm, respectively. The timing jitter between the pump laser and probe electron beam was found to be ~100 fs (rms). The temporal resolution is demonstrated by observing the evolution of Bragg and superlattice peaks of 1T-TaS2 following an 800 nm optical pump and was found to be 130 fs. Lastly, our results demonstrate the advantages of MeV electrons, including large elastic differential scattering cross-section and access to high-order reflections, and the feasibility of ultimately realizing below 10 fs time-resolved electron diffraction.

  4. Femtosecond time-resolved MeV electron diffraction

    DOE PAGESBeta

    Zhu, Pengfei; Zhu, Y.; Hidaka, Y.; Wu, L.; Cao, J.; Berger, H.; Geck, J.; Kraus, R.; Pjerov, S.; Shen, Y.; et al

    2015-06-02

    We report the experimental demonstration of femtosecond electron diffraction using high-brightness MeV electron beams. High-quality, single-shot electron diffraction patterns for both polycrystalline aluminum and single-crystal 1T-TaS2 are obtained utilizing a 5 fC (~3 × 104 electrons) pulse of electrons at 2.8 MeV. The high quality of the electron diffraction patterns confirms that electron beam has a normalized emittance of ~50 nm rad. The transverse and longitudinal coherence length is ~11 and ~2.5 nm, respectively. The timing jitter between the pump laser and probe electron beam was found to be ~100 fs (rms). The temporal resolution is demonstrated by observing themore » evolution of Bragg and superlattice peaks of 1T-TaS2 following an 800 nm optical pump and was found to be 130 fs. Lastly, our results demonstrate the advantages of MeV electrons, including large elastic differential scattering cross-section and access to high-order reflections, and the feasibility of ultimately realizing below 10 fs time-resolved electron diffraction.« less

  5. Versatile portable fluorometer for time-resolved luminescence analysis

    NASA Astrophysics Data System (ADS)

    Chen, Guoying

    2005-06-01

    A robust, filter-based portable fluorometer was designed, prototyped, and tested for time-resolved luminescence (TRL) analysis. Its flexible optical design allows interchangeable configurations to support three measurement modes: liquid-phase TRL using a sample cuvette, solid-matrix TRL using a sorbent strip, and evanescent-field TRL using a quartz-rod waveguide. A xenon flashlamp is used as the light source and a photomultiplier tube (PMT) as the photodetector. A gating technique was implemented to overcome PMT saturation by the intense xenon lamp flash, therefore higher gains can be set to measure weak luminescence signals. The TRL signal is digitized at a 4μs time resolution and a 12bit amplitude resolution. Individual flashes were monitored by a photodiode and its current was integrated to compensate for source light fluctuation. Using tetracycline as a model analyte, a 0.025ppb limit of detection (LOD) with a typical 2% relative standard deviation, and a 3 orders of magnitude (0.5-300ppb) linear dynamic range (r2=0.9996) were achieved.

  6. Fielding of a Time-Resolved Tomographic Diagnostic

    SciTech Connect

    Daniel Frayer, Brian Cox, Wendi Dreesen, Douglas Johnson, Mike Jones, Morris Kaufman

    2008-09-11

    A diagnostic instrument has been developed for the acquisition of high-speed time-resolved images at the Dual-Axis Radiographic Hydrodynamic Test (DARHT) Facility at Los Alamos National Laboratory. The instrument was developed in order to create time histories of the electron beam. Four discrete optical subsystems view Cerenkov light generated at an x-ray target inside of a vacuum envelope. Each system employs cylindrical optics to image light in one direction and collapse light in the orthogonal direction. Each of the four systems images and collapses in unique axes, thereby capturing unique information. Light along the imaging axis is relayed via optical fiber to streak cameras. A computer is used to reconstruct the original image from the four optically collapsed images. Due to DARHT’s adverse environment, the instrument can be operated remotely to adjust optical parameters and contains a subsystem for remote calibration. The instrument was deployed and calibrated, and has been used to capture and reconstruct images. Matters of alignment, calibration, control, resolution, and adverse conditions will be discussed.

  7. Time-resolved pump-probe experiments at the LCLS

    SciTech Connect

    Glownia, James; Cryan, J.; Andreasson, J.; Belkacem, A.; Berrah, N.; Blaga, C.L.; Bostedt, C.; Bozek, J.; DiMauro, L.F.; Fang, L.; Frisch, J.; Gessner, O.; Guhr, M.; Hajdu, J.; Hertlein, M.P.; Hoener, M.; Huang, G.; Kornilov, O.; Marangos, J.P.; March, A.M.; McFarland, B.K.; /SLAC /Stanford U., Phys. Dept. /SLAC /IRAMIS, Saclay /Stanford U., Phys. Dept. /Georgia Tech /Argonne /Kansas State U. /SLAC /Stanford U., Phys. Dept. /SLAC /Stanford U., Appl. Phys. Dept. /Stanford U., Appl. Phys. Dept. /SLAC /LBNL /Argonne /SLAC /SLAC /Stanford U., Appl. Phys. Dept. /Stanford U., Phys. Dept.

    2011-08-12

    The first time-resolved x-ray/optical pump-probe experiments at the SLAC Linac Coherent Light Source (LCLS) used a combination of feedback methods and post-analysis binning techniques to synchronize an ultrafast optical laser to the linac-based x-ray laser. Transient molecular nitrogen alignment revival features were resolved in time-dependent x-ray-induced fragmentation spectra. These alignment features were used to find the temporal overlap of the pump and probe pulses. The strong-field dissociation of x-ray generated quasi-bound molecular dications was used to establish the residual timing jitter. This analysis shows that the relative arrival time of the Ti:Sapphire laser and the x-ray pulses had a distribution with a standard deviation of approximately 120 fs. The largest contribution to the jitter noise spectrum was the locking of the laser oscillator to the reference RF of the accelerator, which suggests that simple technical improvements could reduce the jitter to better than 50 fs.

  8. Pre-denaturing transitions in human serum albumin probed using time-resolved phosphorescence.

    PubMed

    Sagoo, Kulwinder; Hirsch, Richard; Johnston, Pamela; McLoskey, David; Hungerford, Graham

    2014-04-24

    The investigation of protein dynamics has long been of interest, since protein interactions and functions can be determined by their structure and changes in conformation. Although fluorescence, occurring on the nanosecond timescale, from intrinsic fluorescent amino acids has been extensively used, in order to fully access conformational changes longer timescales are required. Phosphorescence enables processes on the microsecond to second timescale to be accessed. However, at room temperature this emission can be weak and non trivial to measure. It requires the removal of oxygen - a common triplet state quencher and appropriate instrumentation. In this work we make use of a chemical deoxygenator to study room temperature phosphorescence from tryptophan in human serum albumin excited using a pulsed UV light emitting diode. This is extended to monitor the phosphorescence emission upon increasing temperature, allowing pre-denaturing transitions to be observed. Time-resolved data are analysed, both as the sum of exponential decays and using a distribution analysis based on non extensive decay kinetics. These results are compared to a fluorescence study and both the average lifetime and contribution of the different emitting components were found to give more dramatic changes on the phosphorescence timescale.

  9. Miniaturized time-resolved Raman spectrometer for planetary science based on a fast single photon avalanche diode detector array.

    PubMed

    Blacksberg, Jordana; Alerstam, Erik; Maruyama, Yuki; Cochrane, Corey J; Rossman, George R

    2016-02-01

    We present recent developments in time-resolved Raman spectroscopy instrumentation and measurement techniques for in situ planetary surface exploration, leading to improved performance and identification of minerals and organics. The time-resolved Raman spectrometer uses a 532 nm pulsed microchip laser source synchronized with a single photon avalanche diode array to achieve sub-nanosecond time resolution. This instrument can detect Raman spectral signatures from a wide variety of minerals and organics relevant to planetary science while eliminating pervasive background interference caused by fluorescence. We present an overview of the instrument design and operation and demonstrate high signal-to-noise ratio Raman spectra for several relevant samples of sulfates, clays, and polycyclic aromatic hydrocarbons. Finally, we present an instrument design suitable for operation on a rover or lander and discuss future directions that promise great advancement in capability.

  10. Time-resolved fluoroimmunoassay of plasma daidzein and genistein.

    PubMed

    Wang, G J; Lapcík, O; Hampl, R; Uehara, M; Al-Maharik, N; Stumpf, K; Mikola, H; Wähälä, K; Adlercreutz, H

    2000-06-01

    We present a method for the determination of the phytoestrogens daidzein and genistein in plasma (serum). These weakly estrogenic isoflavones occur in soybeans and in smaller amounts in some other beans and plants. It has been suggested that they may afford protection against prostate and breast cancer. The method is based on time-resolved fluoroimmunoassay (TR-FIA) using a europium chelate as a label. After synthesis of 4'-O-carboxymethyl-daidzein and 4'-O-carboxymethyl-genistein the compounds are coupled to bovine serum albumin (BSA), then used as antigens to immunize rabbits. The tracers with the europium chelate are synthesized using the same 4'-O-derivative of the isoflavones. After enzymatic hydrolysis and ether extraction the immunoassay is carried out using the VICTOR 1420 multilabel counter (Wallac Oy, Turku, Finland). The antisera cross-reacted to some extent with some isoflavonoids but not with flavonoids. The cross-reactivity seems not to influence the results, which were highly specific for both compounds. The correlation coefficients between the TR-FIA methods and the reference method based on isotope dilution gas chromatography-mass spectrometry were high; r-values were about 0.95-0.99 depending on concentration. The intra-assay coefficients of variation (CV%) for daidzein and genistein at three different concentrations vary 3.2-4.5 and 3.2-4.1, respectively. The inter-assay CVs vary 5.0-6.3 and 4.5-5.3, respectively. The working ranges of the daidzein and genistein assays are 1.0-216 and 1.7-370 nmol/l, respectively. The plasma values (n = 80) of daidzein and genistein are very low in Finnish subjects (mean for daidzein, 3.8+/-6.8 and for genistein, 3.2+/-7.6 nmol/l; median value for daidzein 1.5 and for genistein 1.4 nmol/l). PMID:10802284

  11. Time-resolved microrheology of actively remodeling actomyosin networks

    NASA Astrophysics Data System (ADS)

    Silva, Marina Soares e.; Stuhrmann, Björn; Betz, Timo; Koenderink, Gijsje H.

    2014-07-01

    Living cells constitute an extraordinary state of matter since they are inherently out of thermal equilibrium due to internal metabolic processes. Indeed, measurements of particle motion in the cytoplasm of animal cells have revealed clear signatures of nonthermal fluctuations superposed on passive thermal motion. However, it has been difficult to pinpoint the exact molecular origin of this activity. Here, we employ time-resolved microrheology based on particle tracking to measure nonequilibrium fluctuations produced by myosin motor proteins in a minimal model system composed of purified actin filaments and myosin motors. We show that the motors generate spatially heterogeneous contractile fluctuations, which become less frequent with time as a consequence of motor-driven network remodeling. We analyze the particle tracking data on different length scales, combining particle image velocimetry, an ensemble analysis of the particle trajectories, and finally a kymograph analysis of individual particle trajectories to quantify the length and time scales associated with active particle displacements. All analyses show clear signatures of nonequilibrium activity: the particles exhibit random motion with an enhanced amplitude compared to passive samples, and they exhibit sporadic contractile fluctuations with ballistic motion over large (up to 30 μm) distances. This nonequilibrium activity diminishes with sample age, even though the adenosine triphosphate level is held constant. We propose that network coarsening concentrates motors in large clusters and depletes them from the network, thus reducing the occurrence of contractile fluctuations. Our data provide valuable insight into the physical processes underlying stress generation within motor-driven actin networks and the analysis framework may prove useful for future microrheology studies in cells and model organisms.

  12. Time-resolved White-light Interferometry for Ultrafast Metrology

    NASA Astrophysics Data System (ADS)

    Mingareev, I.; Wortmann, D.; Brand, A.; Horn, A.

    2010-10-01

    The material modification in the volume of transparent dielectrics using tightly focused fs-laser radiation is an important topic for many research groups all over the world. A wide range of applications like the writing of waveguides, micro-structuring by material modification and subsequent etching, or the micro-welding of glass is based on the localized melting and quenching in a different state. Time-resolved white-light interferometry is adopted for the measurement of the optical phase changes in processed materials. A modified Mach-Zehnder interferometer setup combined with microscope objectives is used. The white light is generated by focusing ultrafast laser radiation (tp = 80 fs) in a sapphire crystal using a micro-lens array to minimize temporal and spatial fluctuations in the white-light continuum. Lateral and coaxial pump-probe measurements of the phase changes during material processing are performed using one or two coupled ultrafast laser sources at different repetition rates (frep = 1 kHz-1 MHz) or by adopting single pulses. The temporal delay between the pump and the probe can be adjusted in the range τ⩽1.8 μs in dependence on the repetition rate of the pump radiation. The optical phase shift and therefore the refractive index of the material is calculated from the interference images. The knowledge of the refractive index during the modification process with a temporal resolution in the ps-range and a spatial resolution of several microns leads to a better understanding of the initial processes for the permanent material modifications.

  13. Time-resolved study of Higgs mode in superconductors

    NASA Astrophysics Data System (ADS)

    Shimano, Ryo

    The behavior of superconductors far from equilibrium has been intensively studied over decades. Goals of these studies are the elucidation of bosonic fluctuations essential for the pairing mechanisms, the manifestation of competing orders or hidden phases, and the optical manipulation of superconductivity. The study of collective modes is crucially important for these perspectives as it provides the information on the dynamics of order parameters in non-equilibirium states. Generally, collective modes in ordered phases associated with spontaneous symmetry breaking are classified into 1) gapless phase modes and 2) gapped amplitude modes. In superconductors, the phase mode is eaten by gauge field, according to the Anderson-Higgs mechanism. The remaining amplitude mode is recently termed as Higgs mode from its analogy to the Higgs boson in particle physics. Despite its long history of investigation, unambiguous observation of Higgs mode has remained elusive. This is because the Higgs mode does not have a charge nor electric dipole and therefore it does not couple directly to the electromagnetic field. Here we report on our recent observation of Higgs mode in s-wave superconductors by using THz-pump and THz-probe spectroscopy technique. After nonadiabatic excitation near the superconducting gap energy with monocycle THz pulses, Higgs mode was observed as oscillations in the transmission of THz probe pulse. The resonant nonlinear coupling between the Higgs mode and coherent radiation field was also discovered, resulting in an efficient third order harmonic generation of the incident THz radiation. The extension of experiments to multiband superconductors and unconventional superconductors will be discussed. Time-resolved study of Higgs mode in superconductors.

  14. Time-resolved diagnostics for concrete target response

    SciTech Connect

    Baum, D.W.; Kuklo, R.M.; Reaugh, J.E.; Simonson, S.C.

    1996-05-01

    In order to facilitate the design of advanced penetrating weapons for defeating land targets, the interaction of concrete with high-velocity penetrators needs to be better characterized. To aid in this effort, three new types of time-resolved diagnostics are being developed and have been used in two experiments and one demonstration: fiber optic arrays to localize penetrators in space and time, Fabry-Perot velocimetry to record the concrete particle velocity, which is related to the pressure, at specific locations within concrete targets, and micropower impulse radar to provide a non-intrusive measure of the penetrator position-time history in a target. The two experiments used the fiber optic array and the Fabry-Perot velocimeter to diagnose the response of concrete to penetration by a Viper shaped charge jet. The results were analyzed using the CALE continuum mechanics simulation program, for which a preliminary model of the material properties of concrete was developed. The fiber optic arrays recorded the bow shock at locations 6.4 and 16.9 cm from the front surfaces. The Fabry-Perot velocimeter measured a free-surface velocity of 0.13 km/s at a distance of 3 cm and obliquity 70{degree} from the jet, which was moving at an interface velocity of 4.0 km/s at a depth of 29 cm. These values imply a pressure of about 6.6 kbar at that location. The demonstration used micropower impulse radar with a pulse repetition frequency of 0.25 MHz and a cell size of 30 ps to detect and record the motion of a metal penetrator simulant moving inside a cylindrical concrete target.

  15. Comparison between two time-resolved approaches for prostate cancer diagnosis: high rate imager vs. photon counting system

    NASA Astrophysics Data System (ADS)

    Boutet, J.; Debourdeau, M.; Laidevant, A.; Hervé, L.; Dinten, J.-M.

    2010-02-01

    Finding a way to combine ultrasound and fluorescence optical imaging on an endorectal probe may improve early detection of prostate cancer. A trans-rectal probe adapted to fluorescence diffuse optical tomography measurements was developed by our team. This probe is based on a pulsed NIR laser source, an optical fiber network and a time-resolved detection system. A reconstruction algorithm was used to help locate and quantify fluorescent prostate tumors. In this study, two different kinds of time-resolved detectors are compared: High Rate Imaging system (HRI) and a photon counting system. The HRI is based on an intensified multichannel plate and a CCD Camera. The temporal resolution is obtained through a gating of the HRI. Despite a low temporal resolution (300ps), this system allows a simultaneous acquisition of the signal from a large number of detection fibers. In the photon counting setup, 4 photomultipliers are connected to a Time Correlated Single Photon Counting (TCSPC) board, providing a better temporal resolution (0.1 ps) at the expense of a limited number of detection fibers (4). At last, we show that the limited number of detection fibers of the photon counting setup is enough for a good localization and dramatically improves the overall acquisition time. The photon counting approach is then validated through the localization of fluorescent inclusions in a prostate-mimicking phantom.

  16. Time-resolved spectroscopy of the pulsating CV GW Lib

    NASA Astrophysics Data System (ADS)

    van Spaandonk, L.; Steeghs, D.; Marsh, T. R.; Torres, M. A. P.

    2010-01-01

    We present time-resolved optical spectroscopy of the dwarf nova GW Librae during its rare 2007 April superoutburst and compare these with quiescent epochs. The data provide the first opportunity to track the evolution of the principal spectral features. In the early stages of the outburst, the optically thick disc dominates the optical and the line components show clear orbital radial velocity excursions. In the course of several weeks, optically thin regions become more prominent as strong emission lines replace the broad disc absorption. Post-outburst spectroscopy covering the I band illustrates the advantages of CaII relative to the commonly used Balmer lines when attempting to constrain binary parameters. Due to the lower ionization energy combined with smaller thermal and shear broadening of these lines, a sharp emission component is seen to be moving in between the accretion disc peaks in the CaII line. No such component is visible in the Balmer lines. We interpret this as an emission component originating on the hitherto unseen mass donor star. This emission component has a mean velocity of ~ -15 +/- 5 kms-1 which is associated with the systemic velocity γ, and a velocity semi-amplitude of Kem = 82.2 +/- 4.9 kms-1. Doppler tomography reveals an asymmetric accretion disc, with the S-wave mapping to a sharp spot in the tomogram with a velocity consistent to what is obtained with line profile fitting. A centre of symmetry analysis of the disc component suggests a very small value for the WD orbital velocity K1 as is also inferred from double Gaussian fits to the spectral lines. While our conservative dynamical limits place a hard upper limit on the binary mass ratio of q < 0.23, we favour a significantly lower value near q ~ 0.06. Pulsation modelling suggests a white dwarf mass ~1Msolar. This, paired with a low-mass donor, near the empirical sequence of an evolved cataclysmic variable close to the period bounce, appears to be consistent with all the

  17. Time-Resolved Spectroscopy of Active Binary Stars

    NASA Technical Reports Server (NTRS)

    Brown, Alexander

    2000-01-01

    This NASA grant covered EUVE observing and data analysis programs during EUVE Cycle 5 GO observing. The research involved a single Guest Observer project 97-EUVE-061 "Time-Resolved Spectroscopy of Active Binary Stars". The grant provided funding that covered 1.25 months of the PI's salary. The activities undertaken included observation planning and data analysis (both temporal and spectral). This project was awarded 910 ksec of observing time to study seven active binary stars, all but one of which were actually observed. Lambda-And was observed on 1997 Jul 30 - Aug 3 and Aug 7-14 for a total of 297 ksec; these observations showed two large complex flares that were analyzed by Osten & Brown (1999). AR Psc, observed for 350 ksec on 1997 Aug 27 - Sep 13, showed only relatively small flares that were also discussed by Osten & Brown (1999). EUVE observations of El Eri were obtained on 1994 August 24-28, simultaneous with ASCA X-ray spectra. Four flares were detected by EUVE with one of these also observed simultaneously, by ASCA. The other three EUVE observations were of the stars BY Dra (1997 Sep 22-28), V478 Lyr (1998 May 18-27), and sigma Gem (1998 Dec 10-22). The first two stars showed a few small flares. The sigma Gem data shows a beautiful complete flare with a factor of ten peak brightness compared to quiescence. The flare rise and almost all the decay phase are observed. Unfortunately no observations in other spectral regions were obtained for these stars. Analysis of the lambda-And and AR Psc observations is complete and the results were published in Osten & Brown (1999). Analysis of the BY Dra, V478 Lyr and sigma Gem EUVE data is complete and will be published in Osten (2000, in prep.). The El Eri EUV analysis is also completed and the simultaneous EUV/X-ray study will be published in Osten et al. (2000, in prep.). Both these latter papers will be submitted in summer 2000. All these results will form part of Rachel Osten's PhD thesis.

  18. Frame-Transfer Gating Raman Spectroscopy for Time-Resolved Multiscalar Combustion Diagnostics

    NASA Technical Reports Server (NTRS)

    Nguyen, Quang-Viet; Fischer, David G.; Kojima, Jun

    2011-01-01

    Accurate experimental measurement of spatially and temporally resolved variations in chemical composition (species concentrations) and temperature in turbulent flames is vital for characterizing the complex phenomena occurring in most practical combustion systems. These diagnostic measurements are called multiscalar because they are capable of acquiring multiple scalar quantities simultaneously. Multiscalar diagnostics also play a critical role in the area of computational code validation. In order to improve the design of combustion devices, computational codes for modeling turbulent combustion are often used to speed up and optimize the development process. The experimental validation of these codes is a critical step in accepting their predictions for engine performance in the absence of cost-prohibitive testing. One of the most critical aspects of setting up a time-resolved stimulated Raman scattering (SRS) diagnostic system is the temporal optical gating scheme. A short optical gate is necessary in order for weak SRS signals to be detected with a good signal- to-noise ratio (SNR) in the presence of strong background optical emissions. This time-synchronized optical gating is a classical problem even to other spectroscopic techniques such as laser-induced fluorescence (LIF) or laser-induced breakdown spectroscopy (LIBS). Traditionally, experimenters have had basically two options for gating: (1) an electronic means of gating using an image intensifier before the charge-coupled-device (CCD), or (2) a mechanical optical shutter (a rotary chopper/mechanical shutter combination). A new diagnostic technology has been developed at the NASA Glenn Research Center that utilizes a frame-transfer CCD sensor, in conjunction with a pulsed laser and multiplex optical fiber collection, to realize time-resolved Raman spectroscopy of turbulent flames that is free from optical background noise (interference). The technology permits not only shorter temporal optical gating (down

  19. In vivo time-resolved autofluorescence measurements on human skin

    NASA Astrophysics Data System (ADS)

    Katika, Kamal M.; Pilon, Laurent; Dipple, Katrina; Levin, Seymour; Blackwell, Jennifer; Berberoglu, Halil

    2006-02-01

    In this paper we present preliminary results obtained from fluorescence lifetime measurements on human skin using time-correlated single photon counting (TCSPC) techniques. Human skin was exposed to light from a pulsed LED of 700 ps pulse width at a wavelength of 375 nm and fluorescence decays were recorded at four different emission wavelengths (442, 460, 478 and 496 nm) using a photomultiplier tube (PMT) coupled to a monochromator. Measurements were carried out on the left and right palms of subjects recruited for the study after obtaining consent using a UCLA IRB approved consent form. The subjects recruited consisted of 18 males and 17 females with different skin complexions and ages ranging from 10 to 70 years. In addition, a set of experiments were also performed on various locations including the palm, the arm and the cheek of a Caucasian subject. The fluorescence decays thus obtained were fit to a three-exponential decay model in all cases and were approximately 0.4, 2.7 and 9.4 ns, respectively. The variations in these lifetimes with location, gender, skin complexion and age are studied. It is speculated that the shorter lifetimes correspond to free and bound NADH while the longer lifetime is due to AGE crosslinks.

  20. [Time-resolved optical studies of charge relaxation and charge transfer at electrode interfaces

    SciTech Connect

    Not Available

    1992-01-01

    Key components were identified in a quantitative model of carrier relaxation in semiconductor electrodes: nonlinear aspects of nonradiative and radiative recombination, effect of space charge field on carrier dynamics, self-absorption effects in direct gas semiconductors, and influence of surface state population kinetics on charge carrier recombination. For CdSe, the first three are operative (no direct proof of the last one). A realistic kinetic model for carrier recombination in the bulk of CdSe was used which includes important nonlinear effects, both radiative and nonradiative. The change in interfacial recombination velocity with the chemical nature of the sinterface was studied (n-CdSe/silane interfaces). Temperature effect (278 to 328 K) on fluorescence decay of n-CdSe in contact with 0.5 M KOH was found to be weak. An analytical solution was obtained for time-resolved fluoresence from electrodes under potential bias, and is being tested. Fluorescence work on a different material, CdS, indicate different recombination kinetics; this material was used to directly pump an optical transition of a surface state.

  1. [Time-resolved optical studies of charge relaxation and charge transfer at electrode interfaces

    SciTech Connect

    Not Available

    1992-12-31

    Key components were identified in a quantitative model of carrier relaxation in semiconductor electrodes: nonlinear aspects of nonradiative and radiative recombination, effect of space charge field on carrier dynamics, self-absorption effects in direct gas semiconductors, and influence of surface state population kinetics on charge carrier recombination. For CdSe, the first three are operative (no direct proof of the last one). A realistic kinetic model for carrier recombination in the bulk of CdSe was used which includes important nonlinear effects, both radiative and nonradiative. The change in interfacial recombination velocity with the chemical nature of the sinterface was studied (n-CdSe/silane interfaces). Temperature effect (278 to 328 K) on fluorescence decay of n-CdSe in contact with 0.5 M KOH was found to be weak. An analytical solution was obtained for time-resolved fluoresence from electrodes under potential bias, and is being tested. Fluorescence work on a different material, CdS, indicate different recombination kinetics; this material was used to directly pump an optical transition of a surface state.

  2. In vivo flow cytometry and time-resolved near-IR angiography and lymphography

    NASA Astrophysics Data System (ADS)

    Galanzha, Ekaterina I.; Tuchin, Valery V.; Brock, Robert W.; Zharov, Vladimir P.

    2007-05-01

    Integration of photoacoustic and photothermal techniques with high-speed, high-resolution transmission and fluorescence microscopy shows great potential for in vivo flow cytometry and indocyanine green (ICG) near-infrared (IR) angiography of blood and lymph microvessels. In particular, the capabilities of in vivo flow cytometry using rat mesentery and nude mouse ear models are demonstrated for real-time quantitative detection of circulating and migrating individual blood and cancer cells in skin, mesentery, lymph nodes, liver, kidney; studying vascular dynamics with a focus on lymphatics; monitoring cell traffic between blood and lymph systems; high-speed imaging of cell deformability in flow; and label-free real-time monitoring of single cell extravasation from blood vessel lumen into tissue. As presented, the advantages of ICG IR-angiography include estimation of time resolved dye dynamics (appearance and clearance) in blood and lymph microvessels using fluorescent and photoacoustic modules of the integrated technique. These new approaches are important for monitoring and quantifying metastatic and apoptotic cells; comparative measurements of plasma and cell velocities; analysis of immune responses; monitoring of circulating macromolecules, chylomicrons, bacteria, viruses and nanoparticles; molecular imaging. In the future, we believe that the integrated technique presented will have great potential for translation to early disease diagnoses (e.g. cancer) or assessment of innovative therapeutic interventions in humans.

  3. High-sensitivity detection of PSA by time-resolved fluorometry with Europium chelate

    NASA Astrophysics Data System (ADS)

    Nahm, Kie B.; Jeong, Jin H.; Kim, Byoung C.; Kim, Jae H.; Kim, Young M.; Jeong, Dong S.; Oh, Sang W.; Choi, Eui Y.; Ko, Dong S.

    2006-01-01

    Prostate-specific antigen (PSA) is an androgen-dependent glycoprotein protease (M.W. 33 kDa) and a member of kallikrein super-family of serine protease, and has chymotrypsin-like enzymatic activity. It is synthesized by the prostate epithelial cells and found in the prostate gland and seminal plasma as a major protein. It is widely used as a clinical marker for diagnosis, screening, monitoring and prognosis of prostate cancer. In normal male adults, the concentration of PSA in the blood is below 4 ng/ml and this value increases in patients with the prostate cancer or the benign prostatic hyperplasia (BPH) due to its leakage into the circulatory system. As such, systematic monitoring of the PSA level in the blood can provide critical information about the progress of the prostatic disease. We have fabricated a bread-board time resolved fluorescence system that could detect a concentration of Prostate Specific Antigen t-PSA) at clinically meaningful level in plasma as well as in whole blood sample. We chose Europium chelates as the fluorescence markers to attach to the PSA for its long decay lifetime and relative photostability. We have simplified the electronic circuits considerably by employing a MCS. With this setup, we have successfully proved that PSA concentration of 4pg/mL can be detected with acceptable reliability.

  4. Time-Resolved Energy-Dispersive XAFS Station for Wide-Energy Range at SPring-8

    SciTech Connect

    Kato, K.; Uruga, T.; Tanida, H.; Yokota, S.; Imai, Y.; Irie, T.

    2007-01-19

    A time-resolved energy-dispersive XAFS (DXAFS) station has been constructed at the bending magnet beamline BL28B2 at SPring-8 to study the local structural changes of materials during chemical reactions and functional processes. The bending magnet source at SPring-8 has a high photon flux above 50 keV. The purpose of this station is to measure DXAFS spectra in a wide energy range from 7 to 50 keV covering K-edges of lanthanides. Its main components are a polychromator with a bent silicon crystal, a mirror to reject higher harmonics, and a position-sensitive detector (PSD). To correspond to a wide energy range, polychromators for Bragg and Laue geometry were developed for the energy range below and above 12 keV, respectively. The PSD used is CCD coupled with a fluorescent screen and lens system. The fluorescent materials and their thickness were optimized for measurement in the x-ray range. Good quality spectra of Ce K-edge (40.5 keV) were obtained with exposures of 360 ms for the standard samples. The present status of the system and some experimental examples are presented in this report.

  5. Time Resolved Measurements of Primary Biogenic Aerosol Particles in Amazonia

    NASA Astrophysics Data System (ADS)

    Wollny, A. G.; Garland, R.; Pöschl, U.

    2009-04-01

    Biogenic aerosols are ubiquitous in the Earth's atmosphere and they influence atmospheric chemistry and physics, the biosphere, climate, and public health. They play an important role in the spread of biological organisms and reproductive materials, and they can cause or enhance human, animal, and plant diseases. Moreover, they influence the Earth's energy budget by scattering and absorbing radiation, and they can initiate the formation of clouds and precipitation as cloud condensation and ice nuclei. The composition, abundance, and origin of biogenic aerosol particles and components are, however, still not well understood and poorly quantified. Prominent examples of primary biogenic aerosol particles, which are directly emitted from the biosphere to the atmosphere, are pollen, bacteria, fungal spores, viruses, and fragments of animals and plants. During the Amazonian Aerosol Characterization Experiment (AMAZE-08) a large number of aerosol and gas-phase measurements were taken on a remote site close to Manaus, Brazil, during a period of five weeks in February and March 2008. This presented study is focused on data from an ultraviolet aerodynamic particle sizer (UVAPS, TSI inc.) that has been deployed for the first time in Amazonia. In this instrument, particle counting and aerodynamic sizing over the range of 0.5-20 µm are complemented by the measurement of UV fluorescence at 355 nm (excitation) and 420-575 nm (emission), respectively. Fluorescence at these wavelengths is characteristic for reduced pyridine nucleotides (e.g., NAD(P)H) and for riboflavin, which are specific for living cells. Thus particles exhibiting fluorescence signals can be regarded as "viable aerosols" or "fluorescent bioparticles" (FBAP), and their concentration can be considered as lower limit for the actual abundance of primary biogenic aerosol particles. Data from the UVAPS were averaged over 5 minute time intervals. The presence of bioparticles in the observed size range has been

  6. Detection of early metabolic alterations in the ocular fundus of diabetic patients by time-resolved autofluorescence of endogenous fluorophores

    NASA Astrophysics Data System (ADS)

    Schweitzer, D.; Klemm, M.; Quick, S.; Deutsch, L.; Jentsch, S.; Hammer, M.; Dawczynski, J.; Kloos, C. H.; Mueller, U. A.

    2011-07-01

    Measurements of time-resolved autofluorescence (FLIM) at the human ocular fundus of diabetic patients permit the detection of early pathologic alterations before signs of diabetic retinopathy are visible. The measurements were performed by the Jena Fluorescence Lifetime Laser Scanner Ophthalmoscope applying time-correlated single photon counting (TCSPC) in two spectral channels (K1: 490-560 nm, K2:560-700ps). The fluorescence was excited by 70 ps pulses (FWHM) at 448 nm. The decay of fluorescence intensity was triple-exponentially approximated. The frequency of amplitudes, lifetimes, and relative contributions was compared in fields of the same size and position in healthy subjects and in diabetic patients. The most sensitive parameter was the lifetime T2 in the short-wavelength channel, which corresponds to the neuronal retina. The changes in lifetime point to a loss of free NADH and an increased contribution of protein-bound NADH in the pre-stage of diabetic retinopathy.

  7. Low-temperature and time-resolved spectroscopic characterization of the LOV2 domain of Avena sativa phototropin 1

    NASA Astrophysics Data System (ADS)

    Gauden, Magdalena; Crosson, Sean; van Stokkum, I. H. M.; van Grondelle, Rienk; Moffat, Keith; Kennis, John T. M.

    2004-09-01

    The phototropins are plant blue-light receptors that base their light-dependent action on the reversible formation of a covalent bond between a flavin mononucleotide (FMN) cofactor and a conserved cysteine residue in light, oxygen or voltage (LOV) domains. The spectroscopic properties of the LOV2 domain of phototropin 1 of Avena sativa (oat) have been investigated by means of low-temperature absorption and fluorescence spectroscopy and by time-resolved fluorescence spectroscopy. The low-temperature absorption spectrum of the LOV2 domain showed a fine structure around 473 nm, indicating heterogeneity in the flavin binding pocket. The fluorescence quantum yield of the flavin cofactor increased from 0.13 to 0.41 upon cooling the sample from room temperature to 77 K. A pronounced phosphorescence emission around 600 nm was observed in the LOV2 domain between 77 and 120 K, allowing for an accurate positioning of the flavin triplet state in the LOV2 domain at 16900 cm-1. Fluorescence from the cryotrapped covalent adduct state was extremely weak, with a fluorescence spectrum showing a maximum at 440 nm. Time-resolved fluorescence experiments utilizing a synchroscan streak camera revealed a singlet-excited state lifetime of the LOV2 domain of 2.4 ns. FMN dissolved in aqueous solution showed a pH-dependent lifetime ranging between 2.9 ns at pH 2.0 to 4.7 ns at pH 8.0. No spectral shifting of the flavin emission was observed in the LOV2 domain nor in FMN in aqueous solution.

  8. Low-temperature and time-resolved spectroscopic characterization of the LOV2 domain of Avena sativa phototropin

    SciTech Connect

    Gauden, Magdalena; Crosson, Sean; van Stokkum, I.H.; Grondelle, Rienkvan; Moffat, Keith; Kennis, John T.

    2004-12-13

    The phototropins are plant blue-light receptors that base their light-dependent action on the reversible formation of a covalent bond between a flavin mononucleotide (FMN) cofactor and a conserved cysteine residue in light, oxygen or voltage (LOV) domains. The spectroscopic properties of the LOV2 domain of phototropin 1 of Avena sativa (oat) have been investigated by means of low-temperature absorption and fluorescence spectroscopy and by time-resolved fluorescence spectroscopy. The low-temperature absorption spectrum of the LOV2 domain showed a fine structure around 473 nm, indicating heterogeneity in the flavin binding pocket. The fluorescence quantum yield of the flavin cofactor increased from 0.13 to 0.41 upon cooling the sample from room temperature to 77 K. A pronounced phosphorescence emission around 600 nm was observed in the LOV2 domain between 77 and 120 K, allowing for an accurate positioning of the flavin triplet state in the LOV2 domain at 16900 cm{sup -1}. Fluorescence from the cryotrapped covalent adduct state was extremely weak, with a fluorescence spectrum showing a maximum at 440 nm. Time-resolved fluorescence experiments utilizing a synchroscan streak camera revealed a singlet-excited state lifetime of the LOV2 domain of 2.4 ns. FMN dissolved in aqueous solution showed a pH-dependent lifetime ranging between 2.9 ns at pH 2.0 to 4.7 ns at pH 8.0. No spectral shifting of the flavin emission was observed in the LOV2 domain nor in FMN in aqueous solution.

  9. Bright, highly water-soluble triazacyclononane europium complexes to detect ligand binding with time-resolved FRET microscopy.

    PubMed

    Delbianco, Martina; Sadovnikova, Victoria; Bourrier, Emmanuel; Mathis, Gérard; Lamarque, Laurent; Zwier, Jurriaan M; Parker, David

    2014-09-26

    Luminescent europium complexes are used in a broad range of applications as a result of their particular emissive properties. The synthesis and application of bright, highly water-soluble, and negatively charged sulfonic- or carboxylic acid derivatives of para-substituted aryl-alkynyl triazacyclononane complexes are described. Introduction of the charged solubilizing moieties suppresses cellular uptake or adsorption to living cells making them applicable for labeling and performing assays on membrane receptors. These europium complexes are applied to monitor fluorescent ligand binding on cell-surface proteins with time-resolved Förster resonance energy transfer (TR-FRET) assays in plate-based format and using TR-FRET microscopy.

  10. Simultaneous Quantification of Anticardiolipin IgG and IgM by Time Resolved Fluoroimmunoassay

    PubMed Central

    Liu, Jie; Li, Mei; Ye, Yan; Chen, Yu

    2016-01-01

    The autoimmune disease antiphospholipid syndrome (APS) is characterized by the presence of anticardiolipin antibodies (aCL), along with anti-β2-glycoprotein I (β2GPI) antibodies and lupus anticoagulant (LA). In this study, we developed a time-resolved fluoroimmunoassay (TRFIA) system for simultaneous quantification of aCL IgG and IgM. A 96-well microtiter plate precoated with the complex of cardiolipin from bovine heart and bovine β2GPI was incubated with the anticardiolipin IgG and IgM standard substance or serum, and the conjugate of Eu3+-labeled anti-human IgG and Sm3+-labeled anti-human IgM was pipetted to the wells to form a tipical double-antibody-sandwich immunoreactions; finally the fluorescent intensity of Eu3+ and Sm3+ was detected to reflect the quantity of anticardiolipin IgG and IgM. This assay showed a good relationship between fluorescence intensities and the concentration of anticardiolipin antibody(aCL) IgG and IgM, with a low-end sensitivity of 0.1 U/ml for IgG and 0.1 U/ml for IgM, respectively. The intra- and inter-assay coefficients of variation (CV) of the calibrators was 3.0% and 4.51% for IgG, and 2.76% and 4.45% for IgM. The average recovery was 100.38% for aCL IgG and 100.45% for aCL IgM. For serum samples, the results of our method showed a good correlation with those obtained with ELISA kit. Simultaneous detection of aCL-IgG and aCL-IgM in the same reaction well can optimize assay performance by avoiding potential influence of different reaction conditions-timing, and well-to-well difference in concentration and characteristics of cardiolipin antigen. The results of a combo aCL-IgG and aCL-IgM assay for the same sample are more consistent and more reliable. This dual-label time-resolved fluoroimmunoassay is sensitive for detecting aCL IgG and IgM across a wide concentration range with stable reagents and may assist in the clinical diagnosis of antiphospholipid syndrome. PMID:27661084

  11. Time-resolved lidar fluorosensor for sea pollution detection

    NASA Technical Reports Server (NTRS)

    Ferrario, A.; Pizzolati, P. L.; Zanzottera, E.

    1986-01-01

    A contemporary time and spectral analysis of oil fluorescence is useful for the detection and the characterization of oil spills on the sea surface. Nevertheless the fluorosensor lidars, which were realized up to now, have only partial capability to perform this double analysis. The main difficulties are the high resolution required (of the order of 1 nanosecond) and the complexity of the detection system for the recording of a two-dimensional matrix of data for each laser pulse. An airborne system whose major specifications were: time range, 30 to 75 ns; time resolution, 1 ns; spectral range, 350 to 700 nm; and spectral resolution, 10 nm was designed and constructed. The designed system of a short pulse ultraviolet laser source and a streak camera based detector are described.

  12. Sensitive time-resolved fluoroimmunoassay for quantitative determination of clothianidin in agricultural samples.

    PubMed

    Li, Ming; Sheng, Enze; Yuan, Yulong; Liu, Xiaofeng; Hua, Xiude; Wang, Minghua

    2014-05-01

    Europium (Eu(3+))-labeled antibody was used as a fluorescent label to develop a highly sensitive time-resolved fluoroimmunoassay (TRFIA) for determination of clothianidin residues in agricultural samples. Toward this goal, the Eu(3+)-labeled polyclonal antibody and goat anti-rabbit antibody were prepared for developing and evaluating direct competitive TRFIA (dc-TRFIA) and indirect competitive TRFIA (ic-TRFIA). Under optimal conditions, the half-maximal inhibition concentration (IC50) and the limit of detection (LOD, IC10) of clothianidin were 9.20 and 0.0909 μg/L for the dc-TRFIA and 2.07 and 0.0220 μg/L for the ic-TRFIA, respectively. The ic-TRFIA has no obvious cross-reactivity with the analogues of clothianidin except for dinotefuran. The average recoveries of clothianidin from spiked water, soil, cabbage, and rice samples were estimated to range from 74.1 to 115.9 %, with relative standard deviations of 3.3 to 11.7 %. The results of TRFIA for the blind samples were largely consistent with gas chromatography (R (2) = 0.9902). The optimized ic-TRFIA might become a sensitive and satisfactory analytical method for the quantitative monitoring of clothianidin residues in agricultural samples.

  13. Time-Resolved Luminescence Nanothermometry with Nitrogen-Vacancy Centers in Nanodiamonds

    NASA Astrophysics Data System (ADS)

    Tsai, Pei-Chang; Chen, Oliver Y.; Tzeng, Yan-Kai; Liu, Hsiou-Yuan; Hsu, Hsiang; Huang, Shaio-Chih; Chen, Jeson; Yee, Fu-Ghoul; Chang, Huan-Cheng; Chang, Ming-Shien

    2016-05-01

    Measuring thermal properties with nanoscale spatial resolution either at or far from equilibrium is gaining importance in many scientific and engineering applications. Although negatively charged nitrogen-vacancy (NV-) centers in diamond have recently emerged as promising nanometric temperature sensors, most previous measurements were performed under steady state conditions. Here we employ a three-point sampling method which not only enables real-time detection of temperature changes over +/-100 K with a sensitivity of 2 K/(Hz)1/2, but also allows the study of nanometer scale heat transfer with a temporal resolution of better than 1 μs with the use of a pump-probe-type experiment. In addition to temperature sensing, we further show that nanodiamonds conjugated with gold nanorods, as optically-activated dual-functional nanoheaters and nanothermometers, are useful for highly localized hyperthermia treatment. We experimentally demonstrated time-resolved fluorescence nanothermometry, and the validity of the measurements was verified with finite-element numerical simulations. The approaches provided here will be useful for probing dynamical thermal properties on nanodevices in operation.

  14. Structural dynamics of the myosin relay helix by time-resolved EPR and FRET

    PubMed Central

    Agafonov, Roman V.; Negrashov, Igor V.; Tkachev, Yaroslav V.; Blakely, Sarah E.; Titus, Margaret A.; Thomas, David D.; Nesmelov, Yuri E.

    2009-01-01

    We have used two complementary time-resolved spectroscopic techniques, dipolar electron–electron resonance and fluorescence resonance energy transfer to determine conformational changes in a single structural element of the myosin motor domain, the relay helix, before and after the recovery stroke. Two double-Cys mutants were labeled with optical probes or spin labels, and interprobe distances were determined. Both methods resolved two distinct structural states of myosin, corresponding to straight and bent conformations of the relay helix. The bent state was occupied only upon nucleotide addition, indicating that relay helix, like the entire myosin head, bends in the recovery stroke. However, saturation of myosin with nucleotide, producing a single biochemical state, did not produce a single structural state. Both straight and bent structural states of the relay helix were occupied when either ATP (ADP.BeFx) or ADP.Pi (ADP.AlF4) analogs were bound at the active site. A greater population was found in the bent structural state when the posthydrolysis analog ADP.AlF4 was bound. We conclude that the bending of the relay helix in the recovery stroke does not require ATP hydrolysis but is favored by it. A narrower interprobe distance distribution shows ordering of the relay helix, despite its bending, during the recovery stroke, providing further insight into the dynamics of this energy-transducing structural transition. PMID:19966224

  15. Host Sensitized Luminescence and Time-Resolved Spectroscopy of YVO4: Ho3+ Nanocrystals

    NASA Astrophysics Data System (ADS)

    Mahata, Manoj Kumar; Koppe, Tristan; Hofsäss, Hans; Kumar, Kaushal; Vetter, Ulrich

    Rare earth doped phosphors have attracted much interest because of their high chemical durability and wide range of attractive applications. In this work, Ho3+ doped tetragonal YVO4 nanocrystals have been synthesized via a facile co-precipitation method. The phosphor was characterized by various methods including X-ray diffraction, photoluminescence, cathodoluminescence, time-resolved spectroscopy measurements. The frequency upconversion emission in the synthesized phosphor has been investigated under 800 nm laser excitation. UV-excited photoluminescence (PL) and cathodoluminescence (CL) measurements were performed at room temperature (300 K). A broad band which arises at ∼ 370-600 nm is attributed to the relaxation of VO43- groups from conduction band to valence band. Under UV-excitation, the presence of a sharp band at 550 nm due to the intra-4f transitions of the trivalent holmium ions suggests energy transfer from YVO4 host to RE ions. Luminescence measurements show that this material is suitable for field emission displays (FED) and fluorescent lamps. Also the conversion of UV radiation as well as IR radiation into the visible region suggests the application of this material for photon harvesting in solar cells.

  16. Time-Resolved Imaging Reveals Heterogeneous Landscapes of Nanomolar Ca2+ in Neurons and Astroglia

    PubMed Central

    Zheng, Kaiyu; Bard, Lucie; Reynolds, James P.; King, Claire; Jensen, Thomas P.; Gourine, Alexander V.; Rusakov, Dmitri A.

    2015-01-01

    Summary Maintaining low intracellular calcium is essential to the functioning of brain cells, yet the phenomenology and mechanisms involved remain an enigma. We have advanced a two-photon excitation time-resolved imaging technique, which exploits high sensitivity of the OGB-1 fluorescence lifetime to nanomolar Ca2+ concentration ([Ca2+]) and enables a high data acquisition rate in situ. The [Ca2+] readout is not affected by dye concentration, light scattering, photobleaching, micro-viscosity, temperature, or the main known concomitants of cellular activity. In quiescent tissue, standard whole-cell configuration has little effect on resting [Ca2+] inside neuronal dendrites or inside astroglia dye-filled via gap junctions. Mapping basal [Ca2+] in neurons and astrocytes with submicron resolution unveils heterogeneous concentration landscapes that depend on age and preceding activity. The rich information content represented by such landscapes in acute slices and in vivo promises to unveil the hitherto unexplored, potentially fundamental aspects of brain cell physiology. Video Abstract PMID:26494277

  17. Time-Resolved Imaging of Single HIV-1 Uncoating In Vitro and in Living Cells.

    PubMed

    Francis, Ashwanth C; Marin, Mariana; Shi, Jiong; Aiken, Christopher; Melikyan, Gregory B

    2016-06-01

    Disassembly of the cone-shaped HIV-1 capsid in target cells is a prerequisite for establishing a life-long infection. This step in HIV-1 entry, referred to as uncoating, is critical yet poorly understood. Here we report a novel strategy to visualize HIV-1 uncoating using a fluorescently tagged oligomeric form of a capsid-binding host protein cyclophilin A (CypA-DsRed), which is specifically packaged into virions through the high-avidity binding to capsid (CA). Single virus imaging reveals that CypA-DsRed remains associated with cores after permeabilization/removal of the viral membrane and that CypA-DsRed and CA are lost concomitantly from the cores in vitro and in living cells. The rate of loss is modulated by the core stability and is accelerated upon the initiation of reverse transcription. We show that the majority of single cores lose CypA-DsRed shortly after viral fusion, while a small fraction remains intact for several hours. Single particle tracking at late times post-infection reveals a gradual loss of CypA-DsRed which is dependent on reverse transcription. Uncoating occurs both in the cytoplasm and at the nuclear membrane. Our novel imaging assay thus enables time-resolved visualization of single HIV-1 uncoating in living cells, and reveals the previously unappreciated spatio-temporal features of this incompletely understood process. PMID:27322072

  18. Time-Resolved Imaging of Single HIV-1 Uncoating In Vitro and in Living Cells

    PubMed Central

    Francis, Ashwanth C.; Marin, Mariana; Shi, Jiong; Aiken, Christopher; Melikyan, Gregory B.

    2016-01-01

    Disassembly of the cone-shaped HIV-1 capsid in target cells is a prerequisite for establishing a life-long infection. This step in HIV-1 entry, referred to as uncoating, is critical yet poorly understood. Here we report a novel strategy to visualize HIV-1 uncoating using a fluorescently tagged oligomeric form of a capsid-binding host protein cyclophilin A (CypA-DsRed), which is specifically packaged into virions through the high-avidity binding to capsid (CA). Single virus imaging reveals that CypA-DsRed remains associated with cores after permeabilization/removal of the viral membrane and that CypA-DsRed and CA are lost concomitantly from the cores in vitro and in living cells. The rate of loss is modulated by the core stability and is accelerated upon the initiation of reverse transcription. We show that the majority of single cores lose CypA-DsRed shortly after viral fusion, while a small fraction remains intact for several hours. Single particle tracking at late times post-infection reveals a gradual loss of CypA-DsRed which is dependent on reverse transcription. Uncoating occurs both in the cytoplasm and at the nuclear membrane. Our novel imaging assay thus enables time-resolved visualization of single HIV-1 uncoating in living cells, and reveals the previously unappreciated spatio-temporal features of this incompletely understood process. PMID:27322072

  19. Simultaneous detection of imidacloprid and parathion by the dual-labeled time-resolved fluoroimmunoassay.

    PubMed

    Shi, Haiyan; Sheng, Enze; Feng, Lu; Zhou, Liangliang; Hua, Xiude; Wang, Minghua

    2015-10-01

    A highly sensitive direct dual-labeled time-resolved fluoroimmunoassay (TRFIA) to detect parathion and imidacloprid simultaneously in food and environmental matrices was developed. Europium (Eu(3+)) and samarium (Sm(3+)) were used as fluorescent labels by coupling separately with L1-Ab and A1P1-Ab. Under optimal assay conditions, the half-maximal inhibition concentration (IC50) and limit of detection (LOD, IC10) were 10.87 and 0.025 μg/L for parathion and 7.08 and 0.028 μg/L for imidacloprid, respectively. The cross-reactivities (CR) were negligible except for methyl-parathion (42.4 %) and imidaclothiz (103.4 %). The average recoveries of imidacloprid ranged from 78.9 to 104.2 % in water, soil, rice, tomato, and Chinese cabbage with a relative standard deviation (RSD) of 2.4 to 11.6 %, and those of parathion were from 81.5 to 110.9 % with the RSD of 3.2 to 10.5 %. The results of TRFIA for the authentic samples were validated by comparison with gas chromatography (GC) analyses, and satisfactory correlations (parathion: R (2) = 0.9918; imidacloprid: R (2) = 0.9908) were obtained. The results indicate that the dual-labeled TRFIA is convenient and reliable to detect parathion and imidacloprid simultaneously in food and environmental matrices. PMID:25994268

  20. A multi-analytical investigation of semi-conductor pigments with time-resolved spectroscopy and imaging

    NASA Astrophysics Data System (ADS)

    Nevin, A.; Cesaratto, A.; D'Andrea, C.; Valentini, Gianluca; Comelli, D.

    2013-05-01

    We present the non-invasive study of historical and modern Zn- and Cd-based pigments with time-resolved fluorescence spectroscopy, fluorescence multispectral imaging and fluorescence lifetime imaging (FLIM). Zinc oxide and Zinc sulphide are semiconductors which have been used as white pigments in paintings, and the luminescence of these pigments from trapped states is strongly dependent on the presence of impurities and crystal defects. Cadmium sulphoselenide pigments vary in hue from yellow to deep red based on their composition, and are another class of semiconductor pigments which emit both in the visible and the near infrared. The Fluorescence lifetime of historical and modern pigments has been measured using both an Optical Multichannel Analyser (OMA) coupled with a Nd:YAG nslaser, and a streak camera coupled with a ps-laser for spectrally-resolved fluorescence lifetime measurements. For Znbased pigments we have also employed Fluorescence Lifetime Imaging (FLIM) for the measurement of luminescence. A case study of FLIM applied to the analysis of the painting by Vincent Van Gogh on paper - "Les Bretonnes et le pardon de Pont-Aven" (1888) is presented. Through the integration of complementary, portable and non-invasive spectroscopic techniques, new insights into the optical properties of Zn- and Cd-based pigments have been gained which will inform future analysis of late 19th] and early 20th C. paintings.

  1. In Situ Planetary Mineralogy Using Simultaneous Time Resolved Fluorescence and Raman Spectroscopy

    NASA Technical Reports Server (NTRS)

    Blacksberg, J.; Rossman , G.R.

    2011-01-01

    Micro-Raman spectroscopy is one of the primary methods of mineralogical analysis in the laboratory, and more recently in the field. Because of its versatility and ability to interrogate rocks in their natural form it is one of the front runners for the next generation of in situ instruments designed to explore adverse set of solar system bodies (e.g. Mars, Venus, the Moon, and other primitive bodies such as asteroids and the Martian moons Phobos and Deimos), as well as for pre-selection of rock and soil samples for potential cache and return missions.

  2. Antibody-based fiberoptics biosensor for the carcinogen benzo(a)pyrene

    SciTech Connect

    Vo-Dinh, T.; Tromberg, B.J.; Griffin, G.D.; Ambrose, K.R.; Sepaniak, M.J.; Gardenhire, E.M.

    1987-07-01

    A new antibody-based fiberoptics biosensor was used to detect the important carcinogen benzo(a)pyrene (BaP). The fiberoptics sensor utilizes anti-BaP antibodies covalently bound to its tip. A helium cadmium laser was used as the excitation source to induce fluorescence from BaP conjugated to the bound anti-BaP antibodies. The fiberoptics device can detect 1 femtomole of BaP in a 5-..mu..L sample drop.

  3. An inexpensive technique for the time resolved laser induced plasma spectroscopy

    NASA Astrophysics Data System (ADS)

    Ahmed, Rizwan; Ahmed, Nasar; Iqbal, J.; Baig, M. Aslam

    2016-08-01

    We present an efficient and inexpensive method for calculating the time resolved emission spectrum from the time integrated spectrum by monitoring the time evolution of neutral and singly ionized species in the laser produced plasma. To validate our assertion of extracting time resolved information from the time integrated spectrum, the time evolution data of the Cu II line at 481.29 nm and the molecular bands of AlO in the wavelength region (450-550 nm) have been studied. The plasma parameters were also estimated from the time resolved and time integrated spectra. A comparison of the results clearly reveals that the time resolved information about the plasma parameters can be extracted from the spectra registered with a time integrated spectrograph. Our proposed method will make the laser induced plasma spectroscopy robust and a low cost technique which is attractive for industry and environmental monitoring.

  4. Flow cytometry using Brillouin imaging and sensing via time-resolved optical (BISTRO) measurements.

    PubMed

    Meng, Zhaokai; Petrov, Georgi I; Yakovlev, Vladislav V

    2015-11-01

    A novel concept of Brillouin imaging and sensing via time-resolved optical (BISTRO) measurements is introduced for flow cytometry applications. The system affords robust, maintenance-free and high-speed elasticity-sensitive measurements. PMID:26347908

  5. Wavelet-based fast time-resolved magnetic sensing with electronic spins in diamond

    NASA Astrophysics Data System (ADS)

    Xu, Nanyang; Jiang, Fengjian; Tian, Yu; Ye, Jianfeng; Shi, Fazhan; Lv, Haijiang; Wang, Ya; Wrachtrup, Jörg; Du, Jiangfeng

    2016-04-01

    Time-resolved magnetic sensing is of great importance from fundamental studies to applications in physical and biological sciences. Recently, the nitrogen-vacancy defect center in diamond has been developed as a promising sensor of magnetic fields under ambient conditions. However, methods to reconstruct time-resolved magnetic fields with high sensitivity are not yet fully developed. Here, we propose and demonstrate a sensing method based on spin echo and Haar wavelet transformation. Our method is exponentially faster in reconstructing time-resolved magnetic fields with comparable sensitivity than existing methods. It is also easier to implement in experiments. Furthermore, the wavelet's unique features enable our method to extract information from the whole signal with only part of the measuring sequences. We then explore this feature for a fast detection of simulated nerve impulses. These results will be useful to time-resolved magnetic sensing with quantum probes at nanoscale.

  6. Time-resolved temperature and O atom measurements in nanosecond pulse discharges in combustible mixtures

    NASA Astrophysics Data System (ADS)

    Lanier, Suzanne; Bowman, Sherrie; Burnette, David; Adamovich, Igor V.; Lempert, Walter R.

    2014-11-01

    The paper presents results of time-resolved rotational temperature measurements, by pure rotational coherent anti-Stokes Raman spectroscopy and absolute O atom number density measurements, by two-photon absorption laser induced fluorescence. The experiments were conducted in nanosecond pulse discharges in H2-O2-Ar and C2H4-O2-Ar mixtures, initially at room temperature, operated at a high pulse repetition rate of 40 kHz, in a plane-to-plane double dielectric barrier geometry at a pressure of 40 Torr. Intensified charge-coupled device images show that O2-Ar and H2-O2-Ar plasmas remain diffuse and volume-filling during the entire burst. Images taken in C2H4-O2-Ar plasma demonstrate significant discharge filamentation and constriction along the center plane and in the corners of the test section. The experimental results demonstrate high accuracy of pure rotational psec CARS for thermometry measurements at low partial pressures of oxygen in nonequilibrium plasmas. The results are compared with kinetic modeling calculations, using two different H2-O2 chemistry and C2H4-O2 chemistry mechanisms. In H2-O2-Ar mixtures, the kinetic modeling predictions are in fairly good agreement with the data, predicting temperature rise and O atom accumulation in long discharge bursts, up to 450 pulses. The results show that adding hydrogen to the mixture results in an additional temperature rise, due to its partial oxidation by radicals generated in the plasma, essentially without chain branching. In C2H4-O2-Ar mixtures, the model consistently underpredicts both temperature and O atom number density. The most likely reason for the difference between the experimental data and model predictions is discharge filamentation developing when ethylene is added to the O2-Ar mixture, at fairly low temperatures.

  7. Time-resolved digital holographic microscopy of laser-induced forward transfer process

    PubMed Central

    Ma, H.; Venugopalan, V.

    2014-01-01

    We develop a method for time-resolved digital holographic microscopy to obtain time-resolved 3-D deformation measurements of laser induced forward transfer (LIFT) processes. We demonstrate nanometer axial resolution and nanosecond temporal resolution of our method which is suitable for measuring dynamic morphological changes in LIFT target materials. Such measurements provide insight into the early dynamics of the LIFT process and a means to examine the effect of laser and material parameters on LIFT process dynamics. PMID:24748724

  8. [A method for time-resolved laser-induced breakdown spectroscopy measurement].

    PubMed

    Pan, Cong-Yuan; Han, Zhen-Yu; Li, Chao-Yang; Yu, Yun-Si; Wang, Sheng-Bo; Wang, Qiu-Ping

    2014-04-01

    Laser-Induced Breakdown Spectroscopy (LIBS) is strongly time related. Time-resolved LIBS measurement is an important technique for the research on laser induced plasma evolution and self-absorption of the emission lines. Concerning the temporal characteristics of LIBS spectrum, a method is proposed in the present paper which can achieve micros-scale time-resolved LIBS measurement by using general ms-scale detector. By setting different integration delay time of the ms-scale spectrum detector, a series of spectrum are recorded. And the integration delay time interval should be longer than the worst temporal precision. After baseline correction and spectrum fitting, the intensity of the character line was obtained. Calculating this intensity with differential method at a certain time interval and then the difference value is the time-resolved line intensity. Setting the plasma duration time as X-axis and the time-resolved line intensity as Y-axis, the evolution curve of the character line intensity can be plotted. Character line with overlap-free and smooth background should be a priority to be chosen for analysis. Using spectrometer with ms-scale integration time and a control system with temporal accuracy is 0.021 micros, experiments carried out. The results validate that this method can be used to characterize the evolution of LIBS characteristic lines and can reduce the cost of the time-resolved LIBS measurement system. This method makes high time-resolved LIBS spectrum measurement possible with cheaper system.

  9. Time-resolved crystallography and protein design: signalling photoreceptors and optogenetics.

    PubMed

    Moffat, Keith

    2014-07-17

    Time-resolved X-ray crystallography and solution scattering have been successfully conducted on proteins on time-scales down to around 100 ps, set by the duration of the hard X-ray pulses emitted by synchrotron sources. The advent of hard X-ray free-electron lasers (FELs), which emit extremely intense, very brief, coherent X-ray pulses, opens the exciting possibility of time-resolved experiments with femtosecond time resolution on macromolecular structure, in both single crystals and solution. The X-ray pulses emitted by an FEL differ greatly in many properties from those emitted by a synchrotron, in ways that at first glance make time-resolved measurements of X-ray scattering with the required accuracy extremely challenging. This opens up several questions which I consider in this brief overview. Are there likely to be chemically and biologically interesting structural changes to be revealed on the femtosecond time-scale? How shall time-resolved experiments best be designed and conducted to exploit the properties of FELs and overcome challenges that they pose? To date, fast time-resolved reactions have been initiated by a brief laser pulse, which obviously requires that the system under study be light-sensitive. Although this is true for proteins of the visual system and for signalling photoreceptors, it is not naturally the case for most interesting biological systems. To generate more biological targets for time-resolved study, can this limitation be overcome by optogenetic, chemical or other means?

  10. The primary photophysics of the Avena sativa phototropin 1 LOV2 domain observed with time-resolved emission spectroscopy.

    PubMed

    van Stokkum, Ivo H M; Gauden, Magdalena; Crosson, Sean; van Grondelle, Rienk; Moffat, Keith; Kennis, John T M

    2011-01-01

    The phototropins are blue-light receptors that base their light-dependent action on the reversible formation of a covalent bond between a flavin mononucleotide (FMN) cofactor and a conserved cysteine in light, oxygen or voltage (LOV) domains. The primary reactions of the Avena sativa phototropin 1 LOV2 domain were investigated by means of time-resolved and low-temperature fluorescence spectroscopy. Synchroscan streak camera experiments revealed a fluorescence lifetime of 2.2 ns in LOV2. A weak long-lived component with emission intensity from 600 to 650 nm was assigned to phosphorescence from the reactive FMN triplet state. This observation allowed determination of the LOV2 triplet state energy level at physiological temperature at 16600 cm(-1). FMN dissolved in aqueous solution showed pH-dependent fluorescence lifetimes of 2.7 ns at pH 2 and 3.9-4.1 ns at pH 3-8. Here, too, a weak phosphorescence band was observed. The fluorescence quantum yield of LOV2 increased from 0.13 to 0.41 upon cooling the sample from 293 to 77 K. A pronounced phosphorescence emission around 600 nm was observed in the LOV2 domain between 77 and 120 K in the steady-state emission.

  11. Wide-field time-resolved luminescence imaging and spectroscopy to decipher obliterated documents in forensic science

    NASA Astrophysics Data System (ADS)

    Suzuki, Mototsugu; Akiba, Norimitsu; Kurosawa, Kenji; Kuroki, Kenro; Akao, Yoshinori; Higashikawa, Yoshiyasu

    2016-01-01

    We applied a wide-field time-resolved luminescence (TRL) method with a pulsed laser and a gated intensified charge coupled device (ICCD) for deciphering obliterated documents for use in forensic science. The TRL method can nondestructively measure the dynamics of luminescence, including fluorescence and phosphorescence lifetimes, which prove to be useful parameters for image detection. First, we measured the TRL spectra of four brands of black porous-tip pen inks on paper to estimate their luminescence lifetimes. Next, we acquired the TRL images of 12 obliterated documents at various delay times and gate times of the ICCD. The obliterated contents were revealed in the TRL images because of the difference in the luminescence lifetimes of the inks. This method requires no pretreatment, is nondestructive, and has the advantage of wide-field imaging, which makes it is easy to control the gate timing. This demonstration proves that TRL imaging and spectroscopy are powerful tools for forensic document examination.

  12. Time-resolved photoluminescence spectroscopy and imaging: new approaches to the analysis of cultural heritage and its degradation.

    PubMed

    Nevin, Austin; Cesaratto, Anna; Bellei, Sara; D'Andrea, Cosimo; Toniolo, Lucia; Valentini, Gianluca; Comelli, Daniela

    2014-04-02

    Applications of time-resolved photoluminescence spectroscopy (TRPL) and fluorescence lifetime imaging (FLIM) to the analysis of cultural heritage are presented. Examples range from historic wall paintings and stone sculptures to 20th century iconic design objects. A detailed description of the instrumentation developed and employed for analysis in the laboratory or in situ is given. Both instruments rely on a pulsed laser source coupled to a gated detection system, but differ in the type of information they provide. Applications of FLIM to the analysis of model samples and for the in-situ monitoring of works of art range from the analysis of organic materials and pigments in wall paintings, the detection of trace organic substances on stone sculptures, to the mapping of luminescence in late 19th century paintings. TRPL and FLIM are employed as sensors for the detection of the degradation of design objects made in plastic. Applications and avenues for future research are suggested.

  13. Time-Resolved Photoluminescence Spectroscopy and Imaging: New Approaches to the Analysis of Cultural Heritage and Its Degradation

    PubMed Central

    Nevin, Austin; Cesaratto, Anna; Bellei, Sara; D'Andrea, Cosimo; Toniolo, Lucia; Valentini, Gianluca; Comelli, Daniela

    2014-01-01

    Applications of time-resolved photoluminescence spectroscopy (TRPL) and fluorescence lifetime imaging (FLIM) to the analysis of cultural heritage are presented. Examples range from historic wall paintings and stone sculptures to 20th century iconic design objects. A detailed description of the instrumentation developed and employed for analysis in the laboratory or in situ is given. Both instruments rely on a pulsed laser source coupled to a gated detection system, but differ in the type of information they provide. Applications of FLIM to the analysis of model samples and for the in-situ monitoring of works of art range from the analysis of organic materials and pigments in wall paintings, the detection of trace organic substances on stone sculptures, to the mapping of luminescence in late 19th century paintings. TRPL and FLIM are employed as sensors for the detection of the degradation of design objects made in plastic. Applications and avenues for future research are suggested. PMID:24699285

  14. Radiative lifetime measurements of some Tm I and Tm II levels by time-resolved laser spectroscopy

    NASA Astrophysics Data System (ADS)

    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. Monoclonal antibody-based therapy for neuroblastoma.

    PubMed

    Cheung, N K

    2000-11-01

    Dose-intensive combination chemotherapy can improve the clinical response of many pediatric solid tumors. However, cure remains elusive. Stage 4 neuroblastoma stands out as an exception. Part of this success is a result of antibody-based strategies, which include immunomagnetic purging of autologous marrow prior to autologous marrow transplantation and immunotherapy directed at minimal residual disease. It is striking that treatment with monoclonal antibodies, even when targeted at a single antigen, namely, ganglioside G(D2), can affect long-term progression-free survival among these patients. The potential role of the idiotype network in tumor control can be exploited clinically. The genetic engineering of these antibodies into novel forms holds great promise for more specific and effective targeting possibilities, including the delivery of cytokines and cells. Preclinical results are also promising. It is expected that the availability of novel antibodies directed at a broader spectrum of pediatric solid tumors will facilitate the successful application of this approach to more patients. Experience with metastatic neuroblastoma has provided proof of this principle. It is likely that other tumors will fall.

  16. A New Approach of Oil Spill Detection Using Time-Resolved LIF Combined with Parallel Factors Analysis for Laser Remote Sensing.

    PubMed

    Liu, Deqing; Luan, Xiaoning; Guo, Jinjia; Cui, Tingwei; An, Jubai; Zheng, Ronger

    2016-08-23

    In hope of developing a method for oil spill detection in laser remote sensing, a series of refined and crude oil samples were investigated using time-resolved fluorescence in conjunction with parallel factors analysis (PARAFAC). The time resolved emission spectra of those investigated samples were taken by a laser remote sensing system on a laboratory basis with a detection distance of 5 m. Based on the intensity-normalized spectra, both refined and crude oil samples were well classified without overlapping, by the approach of PARAFAC with four parallel factors. Principle component analysis (PCA) has also been operated as a comparison. It turned out that PCA operated well in classification of broad oil type categories, but with severe overlapping among the crude oil samples from different oil wells. Apart from the high correct identification rate, PARAFAC has also real-time capabilities, which is an obvious advantage especially in field applications. The obtained results suggested that the approach of time-resolved fluorescence combined with PARAFAC would be potentially applicable in oil spill field detection and identification.

  17. A New Approach of Oil Spill Detection Using Time-Resolved LIF Combined with Parallel Factors Analysis for Laser Remote Sensing.

    PubMed

    Liu, Deqing; Luan, Xiaoning; Guo, Jinjia; Cui, Tingwei; An, Jubai; Zheng, Ronger

    2016-01-01

    In hope of developing a method for oil spill detection in laser remote sensing, a series of refined and crude oil samples were investigated using time-resolved fluorescence in conjunction with parallel factors analysis (PARAFAC). The time resolved emission spectra of those investigated samples were taken by a laser remote sensing system on a laboratory basis with a detection distance of 5 m. Based on the intensity-normalized spectra, both refined and crude oil samples were well classified without overlapping, by the approach of PARAFAC with four parallel factors. Principle component analysis (PCA) has also been operated as a comparison. It turned out that PCA operated well in classification of broad oil type categories, but with severe overlapping among the crude oil samples from different oil wells. Apart from the high correct identification rate, PARAFAC has also real-time capabilities, which is an obvious advantage especially in field applications. The obtained results suggested that the approach of time-resolved fluorescence combined with PARAFAC would be potentially applicable in oil spill field detection and identification. PMID:27563899

  18. A New Approach of Oil Spill Detection Using Time-Resolved LIF Combined with Parallel Factors Analysis for Laser Remote Sensing

    PubMed Central

    Liu, Deqing; Luan, Xiaoning; Guo, Jinjia; Cui, Tingwei; An, Jubai; Zheng, Ronger

    2016-01-01

    In hope of developing a method for oil spill detection in laser remote sensing, a series of refined and crude oil samples were investigated using time-resolved fluorescence in conjunction with parallel factors analysis (PARAFAC). The time resolved emission spectra of those investigated samples were taken by a laser remote sensing system on a laboratory basis with a detection distance of 5 m. Based on the intensity-normalized spectra, both refined and crude oil samples were well classified without overlapping, by the approach of PARAFAC with four parallel factors. Principle component analysis (PCA) has also been operated as a comparison. It turned out that PCA operated well in classification of broad oil type categories, but with severe overlapping among the crude oil samples from different oil wells. Apart from the high correct identification rate, PARAFAC has also real-time capabilities, which is an obvious advantage especially in field applications. The obtained results suggested that the approach of time-resolved fluorescence combined with PARAFAC would be potentially applicable in oil spill field detection and identification. PMID:27563899

  19. The kinetic dose limit in room-temperature time-resolved macromolecular crystallography

    SciTech Connect

    Schmidt, M.; Srajer, V.; Purwar, N.; Tripathi, S.

    2012-05-24

    Protein X-ray structures are determined with ionizing radiation that damages the protein at high X-ray doses. As a result, diffraction patterns deteriorate with the increased absorbed dose. Several strategies such as sample freezing or scavenging of X-ray-generated free radicals are currently employed to minimize this damage. However, little is known about how the absorbed X-ray dose affects time-resolved Laue data collected at physiological temperatures where the protein is fully functional in the crystal, and how the kinetic analysis of such data depends on the absorbed dose. Here, direct evidence for the impact of radiation damage on the function of a protein is presented using time-resolved macromolecular crystallography. The effect of radiation damage on the kinetic analysis of time-resolved X-ray data is also explored.

  20. Time-resolved electron beam energy spectrum diagnostics for Vanderbilt FEL

    NASA Astrophysics Data System (ADS)

    Feng, Bibo; Kozub, John A.; Gabella, William E.

    2002-06-01

    A fast electron energy spectrometer has been built using a photodiode array measuring the backward optical transition radiation from a thin film of aluminum. The resolution of the electron energy spectrometer is about 0.2% with a time resolution of 50 ns. The maximum energy spread that can be measured is 6.4%. We present the measurements of the time-resolved electron beam energy spectrum on the Mark III linear accelerator at Vanderbilt University, while lasing at different wavelengths and while not lasing. We also discuss the effects of different parameters, such as cathode heating, alpha magnet strength and RF phase, on the electron energy spectrum and optical spectrum. The diagnostics of time-resolved electron energy spectrum and time-resolved laser spectrum provide the technology to understand the physical process of the FEL interaction. Based on these diagnostics, the FEL facility can realize some special modes of operation, such as macropulse chirping and macropulse two color lasing.

  1. The time-resolved D-SERS vibrational spectra of pesticide thiram.

    PubMed

    Li, Pan; Liu, Honglin; Yang, Liangbao; Liu, Jinhuai

    2013-12-15

    Time-resolved dynamic-SERS (D-SERS) can observe the process of chemical reaction between target and substrate and changes of adsorptive forms for analytes. In this paper, the vibrational spectra of pesticide thiram adsorbed on Au nanoparticles and intensity alternation of SERS spectra depended on different laser powers have been systematically investigated using the method of D-SERS. The Raman intensities of b2 and a1 modes of thiram related to the standard band appear different regulars with the extending time. Meanwhile, due to SERS vibrational spectra of pesticide thiram at different concentrations exhibit different SERS signals, the results of time-resolve D-SERS demonstrate the breakdown of band and different adsorptive forms of molecule on Au substrate. The continuous time-resolved the spectroscopic method offers the fingerprints of target molecules and provides great practical potentials for the continuous assessment and identification of pesticide or other probe molecules.

  2. Cluster mass fraction and size distribution determined by fs-time-resolved measurements

    NASA Astrophysics Data System (ADS)

    Gao, Xiaohui; Wang, Xiaoming; Shim, Bonggu; Arefiev, Alexey; Tushentsov, Mikhail; Breizman, Boris; Downer, Mike

    2009-11-01

    Characterization of supersonic gas jets is important for accurate interpretation and control of laser-cluster experiments. While average size and total atomic density can be found by standard Rayleigh scatter and interferometry, cluster mass fraction and size distribution are usually difficult to measure. Here we determine the cluster fraction and the size distribution with fs-time-resolved refractive index and absorption measurements in cluster gas jets after ionization and heating by an intense pump pulse. The fs-time-resolved refractive index measured with frequency domain interferometer (FDI) shows different contributions from monomer plasma and cluster plasma in the time domain, enabling us to determine the cluster fraction. The fs-time-resolved absorption measured by a delayed probe shows the contribution from clusters of various sizes, allowing us to find the size distribution.

  3. Fractal dimension of time-resolved autofluorescence discriminates tumour from healthy tissues in the oral cavity.

    PubMed

    Klatt, Jan; Gerich, Carola E; Gröbe, Alexander; Opitz, Jörg; Schreiber, Jürgen; Hanken, Henning; Salomon, Georg; Heiland, Max; Kluwe, Lan; Blessmann, Marco

    2014-09-01

    Early detection and complete resection of oral carcinomas is of crucial importance for patient survival. This could be significantly improved by developing a non-invasive, sensitive and real-time detection technique. Time-resolved autofluorescence measurement is state-of-the-art technology originally developed for non-destructive inspection of material. In this study, we measured time-resolved autofluorescence in tumours and healthy tissues of the oral cavity ex vivo and calculated the corresponding fractal dimension which was significantly higher in tumours than in healthy tissues (1.8 vs. 1.6, P < 0.001, unpaired t-test) with non-overlapping 95% confidential intervals 1.88-1.84 and 1.57-1.69, respectively. Very high specificity (86%) could be reached at 100% sensitivity. The area under the curve was 99%, further suggesting the superior prediction potential of fractal dimension based on time-resolved autofluorescence spectra.

  4. Disentangling Multichannel Photodissociation Dynamics in Acetone by Time-Resolved Photoelectron-Photoion Coincidence Spectroscopy.

    PubMed

    Maierhofer, Paul; Bainschab, Markus; Thaler, Bernhard; Heim, Pascal; Ernst, Wolfgang E; Koch, Markus

    2016-08-18

    For the investigation of photoinduced dynamics in molecules with time-resolved pump-probe photoionization spectroscopy, it is essential to obtain unequivocal information about the fragmentation behavior induced by the laser pulses. We present time-resolved photoelectron-photoion coincidence (PEPICO) experiments to investigate the excited-state dynamics of isolated acetone molecules triggered by two-photon (269 nm) excitation. In the complex situation of different relaxation pathways, we unambiguously identify three distinct pump-probe ionization channels. The high selectivity of PEPICO detection allows us to observe the fragmentation behavior and to follow the time evolution of each channel separately. For channels leading to fragment ions, we quantitatively obtain the fragment-to-parent branching ratio and are able to determine experimentally whether dissociation occurs in the neutral molecule or in the parent ion. These results highlight the importance of coincidence detection for the interpretation of time-resolved photochemical relaxation and dissociation studies if multiple pathways are present.

  5. Feasibility experiments on time-resolved fluorosensing applied to oil slicks

    NASA Technical Reports Server (NTRS)

    Camagni, P.; Colombo, G.; Koechler, C.; Pedrini, A.; Omenetto, N.; Rossi, G.

    1986-01-01

    The introduction of time resolved observations can provide a very penetrating tool in the practice of laser fluorosensing. The investigations have demonstrated a relevance of multispectral, time resolved analysis for oil fingerprinting. By comparative studies on a variety of crude oils and their most significant fractions, it was found that the process of time decay in a composite oil is characterized by a few steps, which are associated with specific components in the medium light range. The average decay times of these pure fractions are markedly differentiated as to absolute values and spectral spread; as a consequence, the corresponding parameters in the resultant crude are quite sensitive to the particular mixture of these components. Measurements of the time response give then a finer discrimination between oil classes, depending on the relative content of certain fractions. Experiments were pursued with an improved fluorosensor facility, in order to test the application of time resolved fluorosensing to remote samples on water.

  6. Apparatus and Techniques for Time-resolved Synchrotron X-ray Diffraction using Diamond Anvil Cells

    NASA Astrophysics Data System (ADS)

    Smith, J.; Sinogeikin, S. V.; Lin, C.; Rod, E.; Bai, L.; Shen, G.

    2015-12-01

    Complementary advances in synchrotron sources, x-ray optics, area detectors, and sample environment control have recently made possible many time-resolved experimental techniques for studying materials at extreme pressure and temperature conditions. The High Pressure Collaborative Access Team (HPCAT) at the Advanced Photon Source has made a sustained effort to assemble a powerful collection of high-pressure apparatus for time-resolved research, and considerable time has been invested in developing techniques for collecting high-quality time-resolved x-ray scattering data. Herein we present key aspects of the synchrotron beamline and ancillary equipment, including source considerations, rapid (de)compression apparatus, high frequency imaging detectors, and software suitable for processing large volumes of data. A number of examples are presented, including fast equation of state measurements, compression rate dependent synthesis of metastable states in silicon and germanium, and ultrahigh compression rates using a piezoelectric driven diamond anvil cell.

  7. Time-resolved materials science opportunities using synchrotron x-ray sources

    SciTech Connect

    Larson, B.C.; Tischler, J.Z.

    1995-06-01

    The high brightness, high intensity, and pulsed time-structure of synchrotron sources provide new opportunities for time-resolved x-ray diffraction investigations. With third generation synchrotron sources coming on line, high brilliance and high brightness are now available in x-ray beams with the highest flux. In addition to the high average flux, the instantaneous flux available in synchrotron beams is greatly enhanced by the pulsed time structure, which consists of short bursts of x-rays that are separated by {approximately}tens to hundreds of nanoseconds. Time-resolved one- and two-dimensional position sensitive detection techniques that take advantage of synchrotron radiation for materials science x-ray diffraction investigations are presented, and time resolved materials science applications are discussed in terms of recent diffraction and spectroscopy results and materials research opportunities.

  8. The kinetic dose limit in room-temperature time-resolved macromolecular crystallography

    PubMed Central

    Schmidt, M.; Šrajer, V.; Purwar, N.; Tripathi, S.

    2012-01-01

    Protein X-ray structures are determined with ionizing radiation that damages the protein at high X-ray doses. As a result, diffraction patterns deteriorate with the increased absorbed dose. Several strategies such as sample freezing or scavenging of X-ray-generated free radicals are currently employed to minimize this damage. However, little is known about how the absorbed X-ray dose affects time-resolved Laue data collected at physiological temperatures where the protein is fully functional in the crystal, and how the kinetic analysis of such data depends on the absorbed dose. Here, direct evidence for the impact of radiation damage on the function of a protein is presented using time-resolved macromolecular crystallography. The effect of radiation damage on the kinetic analysis of time-resolved X-ray data is also explored. PMID:22338689

  9. A high-sensitivity femtosecond to microsecond time-resolved infrared vibrational spectrometer.

    PubMed

    Towrie, Michael; Gabrielsson, Anders; Matousek, Pavel; Parker, Anthony W; Rodriguez, Ana Maria Blanco; Vlcek, Antonín

    2005-04-01

    We describe an apparatus that provides, for the first time, a seamless bridge between femtosecond and microsecond time-resolved Raman and infrared vibrational spectroscopy. The laser system comprises an actively Q-switched sub-nanosecond pulsed kilohertz laser electronically synchronized to an ultrafast titanium sapphire regenerative amplifier to within 0.2 ns. The ultrafast amplifier provides the stable probe light source enabling high-sensitivity infrared vibrational spectroscopy of transients. Time-resolved infrared spectra of the excited-state relaxation dynamics of metal carbonyl compounds are presented to illustrate the capability of the apparatus, and transient data is resolved from 1 picosecond to over 100 microseconds. The results are compared to conventional nanosecond Fourier transform infrared (FT-IR) and laser based flash photolysis time-resolved infrared technology.

  10. An instrument for time resolved monitoring of fast chemical transitions: Application to the kinetics of refolding of a globular protein

    NASA Astrophysics Data System (ADS)

    Ratner, Vladimir; Haas, Elisha

    1998-05-01

    The dynamics of kinetic intermediates of protein folding can be studied by time resolved measurements of nonradiative excitation energy transfer, in site-specific labeled protein derivatives, combined with fast mixing experiments. A new device based on the single pulse approach was developed. This experiment is performed over two time scales: the "chemical time scale" of the conformational changes (milliseconds), defined by the rates of changes of conformations in the sample, and the "spectroscopic time scale" (nanoseconds) defined by the lifetimes of the excited states of the fluorescent probes. The chemical process was synchronized by means of a fast mixing stopped flow device. The low cost laser used here is suitable for use with dyes with excitation wavelengths of 337 nm and higher. Up to 20 fluorescence decay curves per second, can be measured within a single stopped flow run. Each fluorescence decay curve is recorded within 250 ns or more. The time resolution (of the spectroscopic time scale) was 0.5 ns. The noise level is low enough to estimate distance distributions from energy transfer experiments, provided that the shortest changeable lifetime component of the fluorescence decay of the donor probes would not be lower than ˜4 ns. The amount of double labeled protein which should be used for each experiment in order to obtain a full data set, with time resolution of 10 ms during protein transition, is only fourfold more than the amount needed for a stopped flow study with steady state fluorescence monitoring. The results obtained for refolding of α-lactalbumin in the presence of 1,8-anilino-naphthalene sulfonic acid from the GuHCl induced denatured state, support the model in which the probe has two states. The first state, is characterized by a fluorescence lifetime of 14.2±0.5 ns and the second by a fluorescence lifetime of 0.5±0.4 ns or less. During refolding the dye is transferred from the first state to the second, at a rate that coincides with the

  11. Time-resolved crystallography and protein design: signalling photoreceptors and optogenetics

    PubMed Central

    Moffat, Keith

    2014-01-01

    Time-resolved X-ray crystallography and solution scattering have been successfully conducted on proteins on time-scales down to around 100 ps, set by the duration of the hard X-ray pulses emitted by synchrotron sources. The advent of hard X-ray free-electron lasers (FELs), which emit extremely intense, very brief, coherent X-ray pulses, opens the exciting possibility of time-resolved experiments with femtosecond time resolution on macromolecular structure, in both single crystals and solution. The X-ray pulses emitted by an FEL differ greatly in many properties from those emitted by a synchrotron, in ways that at first glance make time-resolved measurements of X-ray scattering with the required accuracy extremely challenging. This opens up several questions which I consider in this brief overview. Are there likely to be chemically and biologically interesting structural changes to be revealed on the femtosecond time-scale? How shall time-resolved experiments best be designed and conducted to exploit the properties of FELs and overcome challenges that they pose? To date, fast time-resolved reactions have been initiated by a brief laser pulse, which obviously requires that the system under study be light-sensitive. Although this is true for proteins of the visual system and for signalling photoreceptors, it is not naturally the case for most interesting biological systems. To generate more biological targets for time-resolved study, can this limitation be overcome by optogenetic, chemical or other means? PMID:24914168

  12. Time-resolved studies of particle effects in laser ablation inductively coupled plasma-mass spectrometry

    SciTech Connect

    Perdian, D.; Bajic, S.; Baldwin, D.; Houk, R.

    2007-11-13

    Time resolved signals in laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) are studied to determine the influence of experimental parameters on ICP-induced fractionation effects. Differences in sample composition and morphology, i.e., ablating brass, glass, or dust pellets, have a profound effect on the time resolved signal. Helium transport gas significantly decreases large positive signal spikes arising from large particles in the ICP. A binder for pellets also reduces the abundance and amplitude of spikes in the signal. MO{sup +} ions also yield signal spikes, but these MO{sup +} spikes generally occur at different times from their atomic ion counterparts.

  13. Four-wavelength time-resolved optical mammography in the 680-980-nm range

    NASA Astrophysics Data System (ADS)

    Pifferi, Antonio; Taroni, Paola; Torricelli, Alessandro; Messina, Fabrizio; Cubeddu, Rinaldo; Danesini, Gianmaria

    2003-07-01

    What is to our knowledge the first instrument for time-resolved optical mammography operating at wavelengths longer than 900 nm has been developed. It is a scanning system that relies on the acquisition of time-resolved transmittance curves at 683, 785, 912, and 975 nm, with a total measurement time of ~5 min for an entire image. Breast structures and lesions can be discriminated based on the different absorption and scattering properties at the four wavelengths, which reflect different contributions of oxyhemoglobin, deoxyhemoglobin, water, and lipids, as well as distinct structures. The system is currently used in a European clinical trial.

  14. Single shot, time-resolved measurement of the coherence properties of OCT swept source lasers.

    PubMed

    Butler, T; Slepneva, S; O'Shaughnessy, B; Kelleher, B; Goulding, D; Hegarty, S P; Lyu, H-C; Karnowski, K; Wojtkowski, M; Huyet, G

    2015-05-15

    A novel, time-resolved interferometric technique is presented that allows the reconstruction of the complex electric field output of a swept source laser in a single-shot measurement. The power of the technique is demonstrated by examining a short cavity swept source designed for optical coherence tomography (OCT) applications with a spectral width of over 100 nm. The novel analysis allows a time-resolved real-time characterization of the roll-off, optical spectrum, linewidth, and coherence properties of a dynamic, rapidly swept laser source.

  15. Combined single-pulse holography and time-resolved laser schlieren for flow visualization

    NASA Technical Reports Server (NTRS)

    Burner, A. W.; Goad, W. K.

    1981-01-01

    A pulsed ruby laser and continuous-wave argon ion laser were used in a combined setup at the Langley Expansion Tube for single pulse holography and time resolved laser schlieren with a common optical axis. The systems can be operated simultaneously for a single run. For a single frame, the pulsed holographic setup offers the options of shadowgraph, Schlieren, and interferometry from the reconstructed hologram as well as the advantage of post-run sensitivity adjustments. For flow establishment studies the time resolved laser Schlieren provides visualization of the flow field every 12.5 microns for up to 80 frames with an exposure time per frame of 5.4 microns.

  16. TRIASSIC: the Time-Resolved Industrial Alpha-Source Scanning Induced Current microscope

    NASA Astrophysics Data System (ADS)

    Pallone, Arthur

    Time-resolved ion beam induced current (TRIBIC) microscopy yields useful information such as carrier mobility and lifetimes in semiconductors and defect locations in devices; however, traditional TRIBIC uses large, expensive particle accelerators that require specialized training to operate and maintain. The time-resolved industrial alpha-source scanning induced current (TRIASSIC) microscope transforms TRIBIC by replacing the particle accelerator facility with an affordable, tabletop instrument suitable for use in research and education at smaller colleges and universities. I will discuss the development of, successes with, setbacks to and future directions for TRIASSIC.

  17. Expected resolution and detectability of adenocarcinoma tumors within human breast in time-resolved images

    NASA Astrophysics Data System (ADS)

    Gandjbakhche, Amir H.; Nossal, Ralph J.; Dadmarz, Roya; Schwartzentruber, Douglas; Bonner, Robert F.

    1995-04-01

    The prospects for time-resolved optical mammography rests on the ability to detect adenocarcinoma within the breast with sufficient resolution and specificity to compete with X-ray mammography. We characterized the optical properties of an unusually large (6 cm diameter) fresh adenocarcinoma and normal breast tissue (determined by histology to be predominantly adipose tissue) obtained from a patient undergoing mastectomy. Large specimens (5 mm thick and 3 cm wide) allowed the determination of absorption and scattering coefficients and their spatial heterogeneity as probed with a 1 mm diameter laser beam at 633 nm and 800 nm utilizing total reflectance and transmittance measure with integrating spheres. The difference between scattering coefficients of the malignant tumor and those of normal (principally adipose) breast tissue at 633 nm was much greater than the heterogeneity within each sample. This scattering difference is the principal source of contrast, particularly in time-resolved images. However, the high scattering coefficient of normal breast tissue at 633 nm limits the practicality of time-resolved mammography of a human breast compressed to 5 cm. Although the scattering coefficient of the normal breast tissue decreases at 800 nm, the differences between the optical properties of normal and abnormal breast tissue also are reduced. We used these empirical results in theoretical expressions obtained from random walk theory to quantify the expected resolution, contrast, and the detected intensity of 3, 6, and 9 mm tumors within otherwise homogeneous human breasts as a function of the gating-time of time-resolved optical mammography.

  18. Time Resolved Shadowgraph Images of Silicon during Laser Ablation:Shockwaves and Particle Generation

    SciTech Connect

    Liu, C.Y.; Mao, X.L.; Greif, R.; Russo, R.E.

    2006-05-06

    Time resolved shadowgraph images were recorded of shockwaves and particle ejection from silicon during laser ablation. Particle ejection and expansion were correlated to an internal shockwave resonating between the shockwave front and the target surface. The number of particles ablated increased with laser energy and was related to the crater volume.

  19. Time-resolved heat transfer and skin friction measurements in unsteady flow

    NASA Astrophysics Data System (ADS)

    Diller, T. E.; Telionis, D. P.

    A review of heat transfer and skin friction measurement methods is presented with particular emphasis on techniques that yield details of time-resolved properties. A description of the calibration methods necessary to insure accurate measurements is included. Examples of recent unsteady heat transfer and skin friction measurements with interpretations of the meaning and importance of the results are given.

  20. Femtosecond time-resolved photoelectron imaging on ultrafast electronic dephasing in an isolated molecule

    NASA Astrophysics Data System (ADS)

    Suzuki, Toshinori; Wang, Li; Kohguchi, Hiroshi

    1999-09-01

    Ultrafast dephasing in an intermediate case of molecular radiationless transition has been visualized for the first time by femtosecond time-resolved photoelectron imaging. The decay of photoexcited S1(n,π*) state of pyrazine in 100 ps and the corresponding build-up of triplet states were clearly observed.

  1. Dynamic structural science: recent developments in time-resolved spectroscopy and X-ray crystallography.

    PubMed

    Trincao, Jose; Hamilton, Michelle L; Christensen, Jeppe; Pearson, Arwen R

    2013-10-01

    To understand the mechanism of biological processes, time-resolved methodologies are required to investigate how functionality is linked to changes in molecular structure. A number of spectroscopic techniques are available that probe local structural rearrangements with high temporal resolution. However, for macromolecules, these techniques do not yield an overall high-resolution description of the structure. Time-resolved X-ray crystallographic methods exist, but, due to both instrument availability and stringent sample requirements, they have not been widely applied to macromolecular systems, especially for time resolutions below 1 s. Recently, there has been a resurgent interest in time-resolved structural science, fuelled by the recognition that both chemical and life scientists face many of the same challenges. In the present article, we review the current state-of-the-art in dynamic structural science, highlighting applications to enzymes. We also look to the future and discuss current method developments with the potential to widen access to time-resolved studies across discipline boundaries.

  2. Time-resolved VUV spectroscopy in the EXTRAP-T2 reversed field pinch

    NASA Astrophysics Data System (ADS)

    Hedqvist, Anders; Rachlew-Källne, Elisabeth

    1998-09-01

    Time-resolved VUV spectroscopy has been used to investigate the effects of impurities in a reversed field pinch operating with a resistive shell. Results of electron temperature, impurity ion densities, particle confinement time and 0741-3335/40/9/004/img1 together with a description of the interpretation and the equipment are presented.

  3. TIME-RESOLVED INFRARED SPECTROSCOPY IN THE U121R BEAMLINE AT THE NSLS

    SciTech Connect

    CARR,G.L.; LAVEIGNE,J.D.; LOBO,R.P.S.M.; REITZE,D.H.; TANNER,D.B.

    1999-07-19

    A facility for performing time-resolved infrared spectroscopy has been developed at the NSLS, primarily at beamline U12IR. The pulsed IR light from the synchrotron is used to perform pump-probe spectroscopy. The authors present here a description of the facility and results for the relaxation of photoexcitations in both a semiconductor and superconductor.

  4. Photophysical evaluation of a new functional terbium complex in FRET-based time-resolved homogenous fluoroassays.

    PubMed

    Cywiński, Piotr J; Nchimi Nono, Katia; Charbonnière, Loïc J; Hammann, Tommy; Löhmannsröben, Hans-Gerd

    2014-04-01

    A new functional luminescent lanthanide complex (LLC) has been synthesized with terbium as a central lanthanide ion and biotin as a functional moiety. Unlike in typical lanthanide complexes assembled via carboxylic moieties, in the presented complex, four phosphate groups are chelating the central lanthanide ion. This special chemical assembly enhances the complex stability in phosphate buffers conventionally used in biochemistry. The complex synthesis strategy and photophysical properties are described as well as the performance in time-resolved Förster Resonance Energy Transfer (FRET) assays. In those assays, this biotin-LLC transferred energy either to acceptor organic dyes (Cy5 or AF680) labelled on streptavidin or to quantum dots (QD655 or QD705) surface-functionalised with streptavidins. The permanent spatial donor-acceptor proximity is assured through strong and stable biotin-streptavidin binding. The energy transfer is evidenced from the quenching observed in donor emission and from a decrease in donor luminescence decay, both associated with simultaneous increase in acceptor intensity and in the decay time. The dye-based assays are realised in TRIS and in PBS, whereas QD-based systems are studied in borate buffer. The delayed emission analysis allows for quantifying the recognition process and for auto-fluorescence-free detection, which is particularly relevant for application in bioanalysis. In accordance with Förster theory, Förster-radii (R0) were found to be around 60 Å for organic dyes and around 105 Å for QDs. The FRET efficiency (η) reached 80% and 25% for dye and QD acceptors, respectively. Physical donor-acceptor distances (r) have been determined in the range 45-60 Å for organic dye acceptors, while for acceptor QDs between 120 Å and 145 Å. This newly synthesised biotin-LLC extends the class of highly sensitive analytical tools to be applied in the bioanalytical methods such as time-resolved fluoroimmunoassays (TR-FIA), luminescent

  5. Exploratory study on a statistical method to analyse time resolved data obtained during nanomaterial exposure measurements

    NASA Astrophysics Data System (ADS)

    Clerc, F.; Njiki-Menga, G.-H.; Witschger, O.

    2013-04-01

    Most of the measurement strategies that are suggested at the international level to assess workplace exposure to nanomaterials rely on devices measuring, in real time, airborne particles concentrations (according different metrics). Since none of the instruments to measure aerosols can distinguish a particle of interest to the background aerosol, the statistical analysis of time resolved data requires special attention. So far, very few approaches have been used for statistical analysis in the literature. This ranges from simple qualitative analysis of graphs to the implementation of more complex statistical models. To date, there is still no consensus on a particular approach and the current period is always looking for an appropriate and robust method. In this context, this exploratory study investigates a statistical method to analyse time resolved data based on a Bayesian probabilistic approach. To investigate and illustrate the use of the this statistical method, particle number concentration data from a workplace study that investigated the potential for exposure via inhalation from cleanout operations by sandpapering of a reactor producing nanocomposite thin films have been used. In this workplace study, the background issue has been addressed through the near-field and far-field approaches and several size integrated and time resolved devices have been used. The analysis of the results presented here focuses only on data obtained with two handheld condensation particle counters. While one was measuring at the source of the released particles, the other one was measuring in parallel far-field. The Bayesian probabilistic approach allows a probabilistic modelling of data series, and the observed task is modelled in the form of probability distributions. The probability distributions issuing from time resolved data obtained at the source can be compared with the probability distributions issuing from the time resolved data obtained far-field, leading in a

  6. Time-resolved analytical methods for liquid/solid interfaces. Progress report, November 1, 1993--October 31, 1994

    SciTech Connect

    Harris, J.M.

    1994-10-31

    A number of chemical phenomena that occur at the boundaries between insulating solids and liquids (adsorption, partition, monolayer self-assembly, catalysis, and chemical reactions) are important to energy-related analytical chemistry. These phenomena are central to the development and understanding of chromatographic methods, solid-phase extraction techniques, immobilized analytical reagents, and optical sensors. The goal of this program, therefore, is to develop surface-sensitive spectroscopies by which chemical kinetics at liquid/solid interfaces can be observed on time-scales from nanoseconds to seconds. In the second year of this program, the authors have used temperature-jump relaxation measurements to monitor adsorption/desorption kinetics at liquid/solid interfaces using Joule heating to compare the adsorption of ions from solution onto C1- and C4-derivatized silica surfaces. They completed a study of rate of migration of covalently-attached ligands on silica surfaces; from the temperature-dependence of the migration, the large energy barrier to migration was estimated. Surface heterogeneity of adsorption sites on silica was characterized by time-resolve fluorescence, and the chemical origins investigated by Si{sup 29} NMR spectroscopy. Surface-enhance Raman and fluorescence spectroscopies were modified to study adsorption and binding to silica surfaces. Molecular dynamics simulations were started to help better understand kinetic barriers to adsorption; ESR probe measurements were launched to measure and compare the chain mobility of silica-attached alkyl ligands.

  7. Nanosecond Motions Of Genetically-Engineered Antibodies: Structural Elements Controlling Segmental Flexibility Defined By Time-Resolved Emission Anisotropy

    NASA Astrophysics Data System (ADS)

    Wensel, Theodore G.; Schneider, William P.; Oi, Vernon T.; Stryel, Lubert

    1988-06-01

    Immunoglobulins are flexible proteins which display large-amplitude modes of motion on a nanosecond time scale. Different classes of antibodies differ markedly in their degree of segmental flexibility; a number of their essential biological functions are correlated with their nanosecond internal dynamics. These motions can be conveniently monitored by time-resolved fluorescence anisotropy measurements. An instrument built around a synch-pumped cavity-dumped dye laser and a fast time-to-digital convertor with histogramming memory has made it possible to obtain high-quality anisotropy in a few minutes on small amounts (ca. 100 pmol) of protein. Genetic engineering techniques have made it possible to construct a large number of immunoglobulins with identical binding sites for the fluorescent probe dansyllysine. These proteins differ in the heavy chain regions which are responsible for their biological effector functions and their segmental flexibility. We have analyzed a series of such constructs derived by genetic recombination between the genes coding for the mouse isotypes IgG1 and IgG2a. The results identify two regions responsible for their different degrees of segmental flexibility: the hinge region connecting the Fab and Fc portions of the antibodies, and a short stretch (residues 131-139) of sequence in the amino terminal half of the CH1 domain containing five amino acid substitutions.

  8. Improvement in fingerprint detection using Tb(III)-dipicolinic acid complex doped nanobeads and time resolved imaging.

    PubMed

    Hauser, Frank M; Knupp, Gerd; Officer, Simon

    2015-08-01

    This paper deals with the synthesis and application of lanthanide complex doped nanobeads used as a luminescent fingerprint powder. Due to their special optical properties, namely a long emission lifetime, sharp emission profiles and large Stokes shifts, luminescent lanthanide complexes are useful for discriminating against signals from background emissions. This is a big advantage because latent fingerprints placed on multicoloured fluorescent surfaces are difficult to develop with conventional powders. The complex of 2,6-dipicolinic acid (DPA) and terbium ([Tb(DPA)3](3-)) is used for this purpose. Using the Stöber process, this complex is incorporated into a silica matrix forming nanosized beads (230-630nm). It is shown that the [Tb(DPA)3](3-) is successfully incorporated into the beads and that these beads exhibit the wanted optical properties of the complex. A phenyl functionalisation is applied to increase the lipophilicity of the beads and finally the beads are used to develop latent fingerprints. A device for time resolved imaging was built to improve the contrast between developed fingerprint and different background signals, whilst still detecting the long lasting luminescence of the complex. The developed fingerprint powder is therefore promising to develop fingerprints on multicoloured fluorescent surfaces.

  9. Laser Detection Of Latent Fingerprints: Tris(2,2'-Bipyridyl)Ruthenium(II) Chloride Hexahydrate As A Staining Dye For Time-Resolved Imaging

    NASA Astrophysics Data System (ADS)

    Menzel, E. R.

    1988-04-01

    The compound tris(2,2'-bipyridyl)ruthenium(II) chloride hexahydrate is suitable for laser detection of latent fingerprints on difficult surfaces such as wood and masking tape, as well as surfaces such as polyethylene, metal, etc. The fingerprint treatment can Involve either dusting with powder blended with this compound or by solution staining. The compound displays a strong d-n phosphorescence with a lifetime of about 10-6 and is thus very well suited for time-resolved imaging to suppress background fluorescence.

  10. Developments in time-resolved high pressure x-ray diffraction using rapid compression and decompression

    SciTech Connect

    Smith, Jesse S.; Sinogeikin, Stanislav V.; Lin, Chuanlong; Rod, Eric; Bai, Ligang; Shen, Guoyin

    2015-07-15

    Complementary advances in high pressure research apparatus and techniques make it possible to carry out time-resolved high pressure research using what would customarily be considered static high pressure apparatus. This work specifically explores time-resolved high pressure x-ray diffraction with rapid compression and/or decompression of a sample in a diamond anvil cell. Key aspects of the synchrotron beamline and ancillary equipment are presented, including source considerations, rapid (de)compression apparatus, high frequency imaging detectors, and software suitable for processing large volumes of data. A number of examples are presented, including fast equation of state measurements, compression rate dependent synthesis of metastable states in silicon and germanium, and ultrahigh compression rates using a piezoelectric driven diamond anvil cell.

  11. Phosphorescent nanoparticles and their applications for time-resolved luminescent biological assays

    NASA Astrophysics Data System (ADS)

    Song, Xuedong; Huang, Lei; Knotts, Mike; Wu, Bin

    2009-02-01

    A new class of phosphorescent nanoparticles has been developed that use halogen-containing polymers and copolymers to encapsulate phosphorescent molecules. Their strong phosphorescence of long lifetime and large Stoke shift are not subject to oxygen quenching under ambient conditions due to the low oxygen permeability of the encapsulation matrix. The cross-linked phosphorescent particles are very stable and easily re-suspendable in aqueous media with surface functional groups to allow covalent tagging of biological recognition molecules such as antibodies. The conjugates can be used to provide very sensitive detection of analytes through time-resolved phosphorescence measurements. In addition to their applications for solution-based biological assays, those particles have also been demonstrated to be very useful for dry-chemistry-based time-resolved luminescent lateral flow assays.

  12. Sub-nanosecond time-resolved near-field scanning magneto-optical microscope.

    PubMed

    Rudge, J; Xu, H; Kolthammer, J; Hong, Y K; Choi, B C

    2015-02-01

    We report on the development of a new magnetic microscope, time-resolved near-field scanning magneto-optical microscope, which combines a near-field scanning optical microscope and magneto-optical contrast. By taking advantage of the high temporal resolution of time-resolved Kerr microscope and the sub-wavelength spatial resolution of a near-field microscope, we achieved a temporal resolution of ∼50 ps and a spatial resolution of <100 nm. In order to demonstrate the spatiotemporal magnetic imaging capability of this microscope, the magnetic field pulse induced gyrotropic vortex dynamics occurring in 1 μm diameter, 20 nm thick CoFeB circular disks has been investigated. The microscope provides sub-wavelength resolution magnetic images of the gyrotropic motion of the vortex core at a resonance frequency of ∼240 MHz. PMID:25725848

  13. Following [FeFe] Hydrogenase Active Site Intermediates by Time-Resolved Mid-IR Spectroscopy.

    PubMed

    Mirmohades, Mohammad; Adamska-Venkatesh, Agnieszka; Sommer, Constanze; Reijerse, Edward; Lomoth, Reiner; Lubitz, Wolfgang; Hammarström, Leif

    2016-08-18

    Time-resolved nanosecond mid-infrared spectroscopy is for the first time employed to study the [FeFe] hydrogenase from Chlamydomonas reinhardtii and to investigate relevant intermediates of the enzyme active site. An actinic 355 nm, 10 ns laser flash triggered photodissociation of a carbonyl group from the CO-inhibited state Hox-CO to form the state Hox, which is an intermediate of the catalytic proton reduction cycle. Time-resolved infrared spectroscopy allowed us to directly follow the subsequent rebinding of the carbonyl, re-forming Hox-CO, and determine the reaction half-life to be t1/2 ≈ 13 ± 5 ms at room temperature. This gives direct information on the dynamics of CO inhibition of the enzyme. PMID:27494400

  14. Estimating wide-angle, spatially varying reflectance using time-resolved inversion of backscattered light.

    PubMed

    Naik, Nikhil; Barsi, Christopher; Velten, Andreas; Raskar, Ramesh

    2014-05-01

    Imaging through complex media is a well-known challenge, as scattering distorts a signal and invalidates imaging equations. For coherent imaging, the input field can be reconstructed using phase conjugation or knowledge of the complex transmission matrix. However, for incoherent light, wave interference methods are limited to small viewing angles. On the other hand, time-resolved methods do not rely on signal or object phase correlations, making them suitable for reconstructing wide-angle, larger-scale objects. Previously, a time-resolved technique was demonstrated for uniformly reflecting objects. Here, we generalize the technique to reconstruct the spatially varying reflectance of shapes hidden by angle-dependent diffuse layers. The technique is a noninvasive method of imaging three-dimensional objects without relying on coherence. For a given diffuser, ultrafast measurements are used in a convex optimization program to reconstruct a wide-angle, three-dimensional reflectance function. The method has potential use for biological imaging and material characterization.

  15. Fourier-transform spectrophotometer for time-resolved emission measurements using a 100-point transient digitizer

    NASA Astrophysics Data System (ADS)

    Preses, Jack M.; Hall, Gregory E.; Muckerman, James T.; Sears, Trevor J.; Weston, Ralph E., Jr.; Guyot, Christian; Hanson, Jonathan C.; Flynn, George W.; Bernstein, Herbert J.

    1993-01-01

    An infrared time-resolved Fourier-transform emission spectrophotometer was constructed and its use demonstrated. The instrument is based on a commercial interferometer combined with a data acquisition system. Operation in a smooth scan mode and the use of a transient digitizer provides good time efficiency for data acquisition and reduces the need to maintain constant energy pulses for long periods of time. An entire 100-point time history of a single point of an interferogram is obtained from a single laser pulse and usable data can be obtained from 10 to 50 mirror scans. The experimental apparatus, data acquisition program, and data evaluation are reviewed. Time-resolved Fourier-transform spectroscopy is an efficient method of determining the dynamics of molecular reactions and relaxation.

  16. Towards Measurement of the Time-resolved Heat Release of Protein Conformation Dynamics

    NASA Technical Reports Server (NTRS)

    Puchalla, Jason; Adamek, Daniel; Austin, Robert

    2004-01-01

    We present a way to observe time-resolved heat release using a laminar flow diffusional mixer coupled with a highly sensitive infrared camera which measures the temperature change of the solvent. There are significant benefits to the use of laminar flow mixers for time-resolved calorimetry: (1) The thermal signal can be made position and time- stationary to allow for signal integration; (2) Extremely small volumes (nl/s) of sample are required for a measurement; (3) The same mixing environment can be observed spectroscopically to obtain state occupation information; (4) The mixer allows one to do out of equilibrium dynamic studies. The hope is that these measurements will allow us probe the non-equilibrium thermodynamics as a protein moves along a free energy trajectory from one state to another.

  17. Time-resolved backscattering of circularly and linearly polarized light in a turbid medium.

    PubMed

    Ni, Xiaohui; Alfano, R R

    2004-12-01

    Time-resolved backscattering profiles of circularly and linearly polarized light were measured from a turbid medium composed of small and large polystyrene sphere particles in water. It is shown that, based on the measurements of the time-resolved backscattered copolarized and cross-polarized components of the incident polarized light, either linearly or circularly polarized light can be used to effectively image an object that is deep inside a turbid medium composed of small particles, depending on the depolarization properties of the object itself. For large particles such as in tissue, fog, and clouds, the experimentally observed polarization memory effect on the backscattering temporal profiles suggests that a significant improvement in the image contrast can be achieved by use of circularly polarized light.

  18. Time-Resolved High-Spatial-Resolution Measurements of Underwater Laser Ionization and Filamentation

    NASA Astrophysics Data System (ADS)

    Jones, T. G.; Kaganovich, D.; Helle, M. H.; Penano, J.; Ting, A.; Gordon, D.

    2013-10-01

    Laser triggering and guiding of underwater electrical discharges are being investigated and developed at NRL for applications including advanced micromachining and low-frequency laser acoustic generation. As part of this development we recently made several high-spatial-resolution, time-resolved measurements of underwater optical filamentation and laser ionization. Using 2-laser pump-probe backlit imaging techniques, we were able to achieve time resolution as short as 35 fs and spatial resolution down to 1 micron. Shadowgraph images show few-micron diameter gas bubbles forming throughout the pump beam path in ps timescales. Microbubble numbers and density increased with pulse energy and time during the pump pulse. We also obtained time-resolved spectra of ns-laser-ionized water, revealing black-body radiation lasting more than 100 ns after the ionizing pulse. Results from ongoing underwater laser ionization, filamentation, and discharge-guiding experiments will be presented. This work is supported by NRL Base Funds.

  19. Following [FeFe] Hydrogenase Active Site Intermediates by Time-Resolved Mid-IR Spectroscopy.

    PubMed

    Mirmohades, Mohammad; Adamska-Venkatesh, Agnieszka; Sommer, Constanze; Reijerse, Edward; Lomoth, Reiner; Lubitz, Wolfgang; Hammarström, Leif

    2016-08-18

    Time-resolved nanosecond mid-infrared spectroscopy is for the first time employed to study the [FeFe] hydrogenase from Chlamydomonas reinhardtii and to investigate relevant intermediates of the enzyme active site. An actinic 355 nm, 10 ns laser flash triggered photodissociation of a carbonyl group from the CO-inhibited state Hox-CO to form the state Hox, which is an intermediate of the catalytic proton reduction cycle. Time-resolved infrared spectroscopy allowed us to directly follow the subsequent rebinding of the carbonyl, re-forming Hox-CO, and determine the reaction half-life to be t1/2 ≈ 13 ± 5 ms at room temperature. This gives direct information on the dynamics of CO inhibition of the enzyme.

  20. Time-resolved X-ray scattering program at the Advanced Photon Source

    SciTech Connect

    Rodricks, B.

    1994-08-01

    The Time-Resolved Scattering Program`s goal is the development of instruments and techniques for time-resolved studies. This entails the development of wide bandpass and focusing optics, high-speed detectors, mechanical choppers, and components for the measurement and creation of changes in samples. Techniques being developed are pump-probe experiments, single-bunch scattering experiments, high-speed white and pink beam Laue scattering, and nanosecond to microsecond synchronization of instruments. This program will be carried out primarily from a white-beam, bend-magnet source, experimental station, 1-BM-B, that immediately follows the first optics enclosure (1-BM-A). This paper will describe the experimental station and instruments under development to carry out the program.